C

:.u‘ -.~. “1.;
‘. "5 -
- .‘zww

fig ‘

“17-. -

31-? - "TM-“-

 

m

.i'i

 

'1.

 

.:
'2
t.

 

 

$ng ’ 3:1
‘ Win? .
1.2%.?“

,J... '
A H.”
var».-
~*.. . ‘

 

“.
2...-
fl

 

LIBRARY
Michigan State
University

 

 

 

This is to certify that the
dissertation entitled

EVALUATION OF THE EFFECT OF CHLORINE
DIOXIDE AND ALLYL-ISOTHIOCYANATE ON THE
GROWTH OF SALMONELLA TYPHIMURIUM AND
LISTERIA MONOCYTOGENES ON FRESH CHICKEN
BREAST AND EFFECT OF CHLORINE DIOXIDE
EXPOSURE ON THE PHYSICAL PROPERTIES OF
PLASTIC FILMS

presented by

Joongmin Shin

has been accepted towards fulfillment
of the requirements for the

Ph.D degree in school of packaging

 

fizz/4v W

Major Professor’s Signature
fl 7 '

Date

MSU is an ammative-action, equal-opportunity employer

 

 

PLACE IN RETURN BOX to remove this checkout from your record.
TO AVOID FINES return on or before date due.
MAY BE RECALLED with earlier due date if requested.

 

DATE DUE DATE DUE DATE DUE

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

6/07 p:/C|RC/DateDue.indd-p.1

EVALUATION OF THE EFFECT OF CHLORINE DIOXIDE AND ALLYL-
ISOTHIOCYANATE ON THE GROWTH OF SALMONELLA TYPHIMURIUM AND
LISTER/A MONOCYTOGENES ON FRESH CHICKEN BREAST AND EFFECT
OF CHLORINE DIOXIDE EXPOSURE ON THE PHYSICAL PROPERTIES OF
PLASTIC FILMS

By

Joongmin Shin

A DISSERTATION

Submitted to
Michigan State University
in partial fulfillment of the requirements
for the degree of

DOCTOR OF PHILOSOPHY
School of Packaging
2007

ABSTRACT

EVALUATION OF THE EFFECT OF CHLORINE DIOXIDE AND ALLYL-
ISOTHIOCYANATE ON THE GROWTH OF SALMONELLA TYPHIMURIUM AND
LISTER/A MONOCYTOGENES ON FRESH CHICKEN BREAST AND EFFECT
OF CHLORINE DIOXIDE EXPOSURE ON THE PHYSICAL PROPERTIES OF
PLASTIC FILMS

by
Joongmin Shin

Listeria monocytogenes and Salmonella Typhimun'um are major
pathogenic bacteria associated with poultry products. The consumption of
pathogen contaminated food can cause illness or even death. Controlled,
constant release of antimicrobial agents (antimicrobial packaging) on the surface
of poultry products has potential to inhibit the growth of pathogens and thus
enhance food safety.

In this study, two gas type antimicrobial agents, chlorine dioxide (CIOz)
and allyl-isothiocyanate (AITC), were used with conventional modified
atmosphere packaging (MAP) to determine their effectiveness against Listeria
monocytogenes and Salmonella Typhimun'um on fresh chicken breast at 4°C.
Several different constant release rate CIOz sachets and AITC canisters were
developed and placed into conventional MA packaging. Chicken breast was also
inoculated with the pathogenic bacteria, and gas flushed with 30%C02/70%N2.
The effects of CIOz and AITC on microbial growth, headspace gas composition,
color, and pH changes were evaluated periodically over 21 days.

Overall test results indicate that controlled, continuous release of ClOz or
AITC with MA packaging retarded the growth of Listeria monocytogenes and
Salmonella Typhimun'um, and thus would enhance the safety of fresh chicken
during storage. Some discoloration (L, a, b) was observed on the chicken breast
at high ClOz or AITC release rate.

The effect of chlorine dioxide (ClOz) gas exposure on physical properties
of selected plastic films was investigated. LDPE, PP, and PS were treated with
various concentrations of CIOz gas over selected treatment times. The control

and treated films were evaluated for their mechanical, barrier, and optical
properties. In the concentration range of 250 to 2000 ppm, the ClOz treatment
was shown to significantly affect the mechanical properties, oxygen transmission
rate, and appearance of PS. However, LDPE and PP did not show any significant
physical property changes associated with the treatment.

This dissertation is dedicated to my dearest family, Suwan Lee and Yewon Shin

ACKNOWLEDGEMENTS

I would like to express my deepest heartfelt thanks to Dr. Bruce Harte and
Dr. Susan Selke, my co-advisors. Dr. Bruce Harte always guided me right things.
His suggestions, encouragement, and support helped me in all times of research.
I could not successfully complete this research without his support. Dr. Susan
Selke gave me great educational advice and support through my graduate
program. I feel very lucky to meet them as my academic advisors.

I am also deeply grateful to my committee members, Dr. Elliot Ryser and
Dr. Maria Rubino. I am indebted to Dr. Elliot Ryser for all of his input and
collaboration as well as kindness to allow me to use his lab equipments to finish
the project. I also appreciate to Dr. Maria Rubino for her guidance and support for
this project. ‘

I would like to express my sincere appreciation to Yongwook Park at
Esaeng co. Ltd for financial support for this research. l have furthermore to thank
Yangjai Shin for his encouragement and support for this project.

I would like to thank Joel Tenney at ICA TriNova for his technical support
and materials I could use for this research

I am deeply thankful to my parents, father Euljai Shin and my mother,

Keehee Kim, who supported me through the difficult and the good times while
accomplishing my education. Without their support the study could not have been
completed. Especially, I would like to give my special thanks to my wife Suwan
Lee and my new born daughter, Yewon Shin. Their patient love and support

made this research possible to complete.

vi

TABLE OF CONTENTS

LIST OF TABLES ................................................................................................... x
LIST OF FIGURES .............................................................................................. xvi
INTRODUCTION .................................................................................................... 1
CHAPTER I LITERATURE REVIEW .................................................................... 6
1.1. Principle of antimicrobial packaging .................................................... 6

1.2 Types of antimicrobial packaging ......................................................... 8
1.2.1 Incorporation of antimicrobial agents into films ...................... 8

1.2.2. Edible films and coatings for antimicrobial packaging ........ 10

1.2.3. Photon excited polymer ....................................................... 11

1.2.4. Gas or volatile generating sachets ..................................... 11

1.3. Commercially used antimicrobial agents for food application ........... 12

1.3.1. Chlorine dioxide (CIOz) ....................................................... 12

1.3.2. Allyl isothiocyanate (AITC) ................................................. 14

1.3.3. Silver ions ........................................................................... 16

1.3.4. Ethanol ............................................................................... 17

1.3.5. Nisin ................................................................................... 18

1.3.6. Natural extract .................................................................... 18

1.3.7. Carbon dioxide (COz) ......................................................... 19

1.4. Regulation issues of antimicrobials for food packaging ..................... 20

1.5. Common pathogenic organisms in meat products ............................ 23

1.5.1. Salmonella Typhimurium ...................................................... 23

1.5.2. Listeria monocytogenes ....................................................... 25

1.5.3. Other foodborne pathogens ................................................. 26

1.6. Chicken shelf life ............................................................................... 27

1.7. Packaging of fresh meats .................................................................... 28

vii

1.8. Poultry meat market segment & packaging ........................................ 30

CHAPTER II MATERIALS AND METHODS ........................................................ 32
2.1. Culture preparation ........................................................................... 32

2.2. Determine headspace concentration of chlorine dioxide (00;) and its
antimicrobial effectiveness ....................................................................... 34

2.3. Headspace concentration of Allyl isothiocyanate (AITC) and its

antimicrobial effectiveness ....................................................................... 37
2.4 Design of constant antimicrobial releasing system ............................. 41
2.4.1. ClOz release system ........................................................... 41
2.4.2. AITC release system ........................................................... 41

2.5. Determination of the Cl02 and AITC release rate to inhibit microbial
growth on fresh chicken ............................................................................. 45

2.6. Evaluation of synergic effect of antimicrobials (CIOz and AITC) and
MAP combination ..................................................................................... 49

2.7. Fresh chicken packaged with antimicrobial and stored for 21 day....49

2.7.1 Sample and packaging ........................................................ 49

2.7.2. Microbial analysis ............................................................... 50

2.7.3. Headspace gas analysis .................................................... 50

2.7.4. pH measurement ................................................................ 50

2.7.5. Color measurement ............................................................ 51

2.8. Physical properties changes by ClOz treatment ................................ 52
2.8.1. Mechanical properties ......................................................... 53

2.8.2. Measurement of oxygen transmission rate (OTR) .............. 54

2.9. Statistical analysis ............................................................................... 55
CHAPTER III. RESULTS AND DISCUSSION ...................................................... 56
3,1. Inhibition performance of AITC and CIOZ .......................................... 56

viii

3.1.1. Inhibition effectiveness of AITC against Listeria

monocytogenes and Salmonella Typhimurium ............................. 56
3.1.2. Inhibition effectiveness of CD; against Listeria
monocytogenes and Salmonella Typhimurium ............................. 59
3.2. Development of AITC release system ............................................... 63
3.2.1 . AITC vapor pressure control ............................................... 63

3.2.2. AITC permeability of PE film and controlled release rate...70
3.3. Development of CIOz release system ................................................ 73

3.4. Determination of minimum release rate to inhibit target bacteria ...... 75
3.4.1. Target bacteria growth with AITC treatment ........................ 76
3.4.2. Target bacteria growth with Cl02 treatment ..................... 82

3.5. Comparison on the antimicrobial treatment with MAP and
antimicrobial packaging ............................................................................ 88

3.6. Fresh chicken package with antimicrobials for 21 day ....................... 94
3.6.1. Listeria monocytogenes with antimicrobial treatments ....... 94
3.6.2. Salmonella Typhimurium with antimicrobial treatments ..... 98

3.6.3. Total aerobic bacteria with microbial treatments .............. 102
3.6.4. pH of chicken breast during 21 days of storage ............... 106
3.6.5. Headspace gas analysis of the chicken packages ........... 110

3.6.6. Surface color on fresh chicken during 21 days of storag..113

3.7. Physical properties changes by CIOz treatments ............................. 123

3.7.1. Mechanical properties ....................................................... 123

3.7.2. Oxygen barrier properties ................................................. 125

3.7.3. Color ................................................................................. 126

CHAPTER IV. CONCLUSION AND FUTURE WORK ....................................... 128
APPENDICES .................................................................................................... 131
APPENDIX A Methods for analyzing chlorine dioxide and related
components in gas phases (ICA titration method) ................................... 131

ix

APPENDIX B Growth model system raw data at 37°C and 7°C ............. 134

APPENDIX C Summary tables of experimental AITC vapor pressures and
model verification .................................................................................... 145

APPENDIX D Raw data tables of microbial growth, headspace gas
composition, pH, and Color ..................................................................... 149

APPENDIX E Raw data tables of tensile strength (TS), elongation at break,
oxygen transmission rate, and color ........................................................ 172

BIBLIOGRAPHY ................................................................................................. 178

LIST OF TABLES
Table 1. List of antimicrobial agents that are approved for food additive (food
packaging) materials in USA ................................................................................ 21

Table 2. AITC vapor releasing system as related to AITC mole fraction in
triglycerdie mixture at 4°C .................................................................................... 69

Table 3. AITC vapor release system as related to AITC mole fractions in the
triglyceride mixture at 4°C .................................................................................... 72

Table 4. Total released ClOz amounts vs. experimental headspace concentration
as a function of time at 4°C .................................................................................. 74

Table 5. Inhibition effectiveness of AITC treatment on Salmonella Typhimurium,
Listeria monocytogenes, and total aerobic bacteria on fresh chicken at
7°C ....................................................................................................................... 78

Table 6. Inhibition effectiveness of CI02 on Salmonella Typhimurium, Listeria
monocytogenes, and total aerobic bacteria on fresh chicken at
7°C ....................................................................................................................... 87

Table 7. Comparison of inhibition effectiveness of AITC treatments on Salmonella
Typhimurium, Listeria monocytogenes, and total aerobic bacteria growth on fresh
chicken at 4°C ...................................................................................................... 92

Table 8. Comparison of inhibition effectiveness of CIOz treatments on Salmonella
Typhimurium, Listeria monocytogenes, and total aerobic bacteria growth on fresh
chicken at 4°C ...................................................................................................... 93
Table 9. Gas mixture sample, and antimicrobial release rates used .................... 94
Table 10. Growth of Listeria monocytogenes (log cfu/g) on fresh chicken breast

with constant release antimicrobial treatment (Cl02 and AITC) during 21 days of
storage at 4°C ...................................................................................................... 97

xi

Table 11. Growth of Salmonella Typhimurium (log cfu/g) on fresh chicken breast
with constant release antimicrobial treatments (CIOz and AITC) during 21 days of
storage at 4°C .................................................................................................... 101

Table 12. Total aerobic bacteria counts (log cfu/g) on fresh chicken breast with
constant release antimicrobials (CIOz and AITC) during 21 days of storage at 4°C.

........................................................................................................................... 105

Table 13. The pH values of fresh chicken breast during 21 days of storage, from
2 different local store .......................................................................................... 109

Table 14. Change in gas composition in the package headspace during storage
of chicken breasts (n=3) at 4°C .......................................................................... 111 '

Table 15. Color “L” values of fresh chicken breast during 21 days of storage (Day
0 “a” value was 53.181024) at 4°C .................................................................... 117

Table 16. Color “L” values of fresh chicken breast during 21 days of storage (Day
0 “L” value was 46.55: 0.99) at 4°C ................................................................... 118

Table 17. Color “a” values of fresh chicken breast during 21 days of storage (Day
0 “a” value were 2.68:0.16) at 4°C ..................................................................... 119

Table 18. Color “a” values of fresh chicken breast during 21 days of storage (Day
0 “a” value was 43110.14) at 4°C ..................................................................... 120

Table 19. Color “b” values of fresh chicken breast during 21 days of storage (Day
0 “b" value was 2.21:0.10) at 4°C ..................................................................... 121

Table 20. Color “b” values of fresh chicken breast during 21 days of storage (Day
0 “b” value was 1.39:0.05) at 4°C ..................................................................... 122

Table 21. Tensile strength and Elongation at break of packaging films (PP, LDPE,
and PS) .............................................................................................................. 126

xii

Table 22. Oxygen permeability of conventional packaging films (PS, LDPE, and
PP) after CIOZ treatment .................................................................................... 127

Table 23. Color changes of conventional packaging films (PS, LDPE, and PP)
after CIOZ treatment (n=3) ................................................................................. 127

Table 24. The reduction of Salmonella Typhimurium on agar plate (60 mm X 16
mm) in glass bottle (with soaked Allyl-isothiocyanate-oil mixture filter paper)
(37°C for 2 days) ................................................................................ 135

Table 25. The reduction of Listeria monocytogenes on agar plate (16 mm x 60
mm) in glass bottle (with soaked Allyl-isothiocyanate-oil mixture filter paper)
(37°C for 2 days) ................................................................................. 136

Table 26. The reduction of Salmonella Typhimurium on agar plate (60 mm X 16
mm) in glass bottle (with soaked Allylisothiocyanate-oil mixture filter paper) (7°C
for 7 days) ......................................................................................... 137

Table 27. The reduction of Listeria monocytogenes on agar plate (60 mm X 16
mm) in glass bottle (with soaked Allylisothiocyanate-oil mixture filter paper) (7°C
for 7 days) ......................................................................................... 138

Table 28. The reduction of Salmonella Typhimurium on agar plate (60 mm X 16
mm) in glass bottle (with chlorine dioxide treatment) (37°C for 2 days) ........... 139

Table 29. The reduction of Listeria monocytogene on agar plate (60 mm X 16
mm) in glass bottle (with chlorine dioxide treatment) (37°C for 2 days) ........... 140

Table 30. The reduction of Salmonella Typhimurium on agar plate (60 mm X 16
mm) in glass bottle (with chlorine dioxide treatment) (7°C for 7 days) ............ 141

Table 31. The reduction of Listeria monocytogene on agar plate (60 mm X 16
mm) in glass bottle (with chlorine dioxide treatment) (7°C for 7 days) ............. 142

Table 32. Headspace concentrations (ppm) of Chlorine dioxide (ClOz) with growth
medium (at 7 °C) .................................................................................. 143

xiii

Table 33. Headspace concentrations (ppm) of Chlorine dioxide (CIOz) with growth
medium (at 37 °C) ............................................................................... 143

Table 34. . Headspace concentrations (ng/ml) of Allyl isothiocyanate (AITC) with
growth medium (at 7 °C) ....................................................................... 144

Table 35. . Headspace concentrations (ng/ml) of Allyl isothiocyanate (AITC) with

growth medium (at 37 °C) ...................................................................... 144
Table 36. AITC vapor pressure based on mole faction at 4°C ....................... 146
Table 37. AITC vapor pressure based on mole faction at 7°C ....................... 146
Table 38. AITC vapor pressure based on mole faction at 22°C ..................... 147
Table 39. AITC vapor pressure based on mole faction at 37°C ..................... 147
Table 40. AITC perrneant profile through PE film as a function of time ........... 148

Table 41. Raw data of Salmonella Typhimurium growth (log cfu/g) on fresh
chicken based on AITC release rates at 7°C. ............................................ 150

Table 42. Raw data of Listeria monocytogenes growth (log cfu/g) on fresh
chicken based on AITC release rates at 7°C. ............................................ 151

Table 43. Raw data of total aerobic bacteria growth (log cfu/g) on fresh chicken
based on AITC release rates at 7°C ......................................................... 152

Table 44. Raw data of Salmonella Typhimurium growth (log cfu/g) on fresh
chicken based on CIOz release rates at 7°C .............................................. 153

Table 45. Raw data of Listeria monocytogenes growth (log cfu/g) on fresh
chicken depend on CIOz release rates at 7°C ............................................ 154

Table 46. Raw data of total aerobic bacteria growth (log cfu/g) on fresh chicken
depend on CIOz release rates at 7°C ...................................................... 155

xiv

Table 47. Raw data of Salmonella Typhimurium growth on fresh chicken using
AITC treatment with MAP and without MAP at 4°C .................................... 156

Table 48. Raw data of Listeria monocytogenes growth on fresh chicken using
AITC treatment with MAP and without MAP at 4°C .................................... 157

Table 49. Raw data of total aerobic bacteria growth on fresh chicken using AITC
treatment with MAP and without MAP at 4°C ............................................. 158

Table 50. Raw data of Salmonella Typhimurium growth on fresh chicken using
CIOz treatment with MAP and without MAP at 4°C ..................................... 159

Table 51. Raw data of Listeria monocytogenes growth on fresh chicken using
ClOz treatment with MAP and without MAP at 4°C ..................................... 160

Table 52. Raw data of total aerobic bacteria growth on fresh chicken using CIOz
treatment with MAP and without MAP at 4°C ............................................ 161

Table 53. Raw data of Listeria monocytogenes (log cfu/g) on fresh chicken breast
with constant release antimicrobial treatment during 21 days at 4°C .............. 162

Table 54. Raw data of Salmonella Typhimurium (log cfu/g) on fresh chicken
breast with constant release antimicrobial treatment during 21 days at 4°C ..... 163

Table 55. Raw data of total aerobic bacteria (log cfu/g) on fresh chicken breast
with constant release antimicrobial treatment during 21 days at 4°C ................. 164

Table 56. Raw data for pH values of fresh chicken breast for 21 days, 4°C (1°t
test set) ............................................................................................. 165

Table 57. Raw data for pH values of fresh chicken breast for 21 days, 4°C (2”t
test set) ............................................................................................ 166

Table 58. Raw data for color “L” values of fresh chicken breast for 21 days, 4°C
(1St test set) ....................................................................................... 167

XV

Table 59. Raw data for color “L” values of fresh chicken breast for 21 days, 4°C
(2"d test set) ...................................................................................... 168

Table 60. Raw data for color “a” values of fresh chicken breast for 21 days, 4°C
(1m test set) ...................................................................................... 169

Table 61. Raw data for color “a” values of fresh chicken breast for 21 days, 4°C
(2"d test set) ....................................................................................... 170

Table 62. Raw data for color “b” values of fresh chicken breast for 21 days, 4°C
(1St test set) ........................................................................................ 171

Table 63. Raw data for color “b” values of fresh chicken breast for 21 days, 4°C
(2"d test set) ....................................................................................... 173

Table 64. Raw data for Tensile strength and Elongation at break of packaging
films (PVC, LDPE, and PS) after CI02 treatment ......................................... 174

Table 65. Raw data for Oxygen permeability of packaging films (PVC, LDPE, and
PS) after CIOz treatment ........................................................................ 175

Table 66. Raw data for color changes of packaging films ((PVC, LDPE, and PS)
after ClOz treatment ............................................................................ 176

xvi

LIST OF FIGURES
Figure 1. An antimicrobial packaging system as part of a Hurdle technology ....... 7

Figure 2. Listeria / Salmonella culture preparation used for inoculation of the

chicken ................................................................................................................. 33
Figure 3. Schematic of canning jar used for CI02 concentration study ................ 34
Figure 4. Equilibrium CI02 gas generating system .............................................. 36

Figure 5. Photo of the detector tube and pump used to determine ClOz
concentration ........................................................................................................ 37

Figure 6. Schematic of canning jar used for AITC testing .................................... 38
Figure 7. Standard curve of AITC concentration vs. area in a hexane solution...39
Figure 8. Chromatogram of AITC in a Hexane- AITC solution ............................. 40
Figure 9. Schematic of AITC release canister ...................................................... 44

Figure 10. Quasi-isotatic cell used for measuring AITC permeability of the film
sample .................................................................................................................. 44

Figure 11. Photo of chicken breast cut to provide a standardized surface
area ...................................................................................................................... 45

Figure 12. Inoculated chicken sample with CIOZ and AITC sachets .................... 46

Figure 13. T-200 packaging machine used to pack the chicken breasts into the
trays ..................................................................................................................... 47

Figure 14. Schematic illustration of the thin agar overlay method ....................... 48

Figure 15. pH meter and assembly used to determine pH of chicken samples...51

xvii

Figure 16. Colorimeter used to measure the color of antimicrobial treated chicken
breast ................................................................................................................... 52

Figure 17. Schematics of CI02 film treatment system .......................................... 53

Figure 18. lnstron 4201 used for mechanical properties evaluation of packaging
films ...................................................................................................................... 54

Figure 19. Oxtran 8001 unit used for oxygen transmission testing of film
samples ................................................................................................................ 54

Figure 20. Reduction of Salmonella Typhimurium and Listeria monocytogenes on
agar due to AITC treatment (37°C for 2day) ......................................................... 58

Figure 21. Reduction of Salmonella Typhimurium and Listeria monocytogenes on
agar due to AITC treatment (7°C for 7 day) ......................................................... 58

Figure 22. AITC headspace concentrations in glass jars (950ml) with growth
media during storage ........................................................................................... 59

Figure 23. Reduction of Salmonella Typhimurium and Listeria monocytogenes on
agar due to CI02 treatments (37°C for 2 day) ..................................................... 62

Figure 24. Reduction of Salmonella Typhimurium and Listeria monocytogenes on
agar due to Cl02 treatments (7°C for 7 day) ....................................................... 62

Figure 25. CIOZ headspace concentrations in glass jar (950 ml) containing
growth medium ..................................................................................................... 63

Figure 26. Vapor pressure of AITC as a function of temperature ......................... 65

Figure 27. vapor pressures of AITC in a AITC-Triglyceride mixture at
Temperatures (4, 7, and 23 °C). ........................................................................ 66

Figure 28. Estimated 0 values as a function of temperature .............................. 69

xviii

Figure 29. Quasi-isotatic permeation curves for AITC and PE film at 23 °C as a
function of time ..................................................................................................... 71

Figure 30. Theoretical vs. experimental AITC headspace concentrations of the
different release systems ...................................................................................... 72

Figure 31. Effect of AITC release rate on Salmonella Typhimurium inoculated on
chicken breast and refrigerated at 7°C ................................................................ 78

Figure 32. Effect of AITC release rate on Listeria monocytogenes inoculated on
chicken breast and refrigerated at 7°C ................................................................ 79

Figure 33. Effect of AITC release rate on total aerobic bacteria on chicken breast
and refrigerated at 7°C ........................................................................................ 80

Figure 34. Effect of CID; release rate on Salmonella Typhimurium inoculated
onto chicken breast and refrigerated at 7°C ........................................................ 84

Figure 35. Effect of CI02 release rate on Listeria monocytogenes inoculated onto
chicken breast and refrigerated at 7°C ................................................................ 85

Figure 36. Effect of ClOz release rate on total aerobic bacteria on chicken breast
and refrigerated at 7°C ........................................................................................ 86

Figure 37. Comparison of inhibition effectiveness of antimicrobial treatments with
MAP and without MAP on Salmonella Typhiumruium on fresh chicken (after 8
days, at 4°C) ........................................................................................................ 89

Figure 38. . Comparison of inhibition effectiveness of antimicrobial treatments
with MAP and without MAP on Listeria monocytogenes on fresh chicken (after 8
days, at 4°C) ........................................................................................................ 90

Figure 39. . Comparison of inhibition effectiveness of antimicrobial treatments

with MAP and without MAP on total aerobic bacteria on fresh chicken (after 8
days, at 4°C) ........................................................................................................ 91

xix

Figure 40. Growth of Listeria monocytogenes on fresh chicken breast as a
function of storage time, at 4°C ............................................................................ 96

Figure 41. Growth of Salmonella Typhimurium on fresh chicken breast as a
function of storage time, at 4°C .......................................................................... 100

Figure 42. Total aerobic bacteria on fresh chicken breast as a function of storage
time at 4°C ......................................................................................................... 104

Figure 43. Appearance of chicken breast after 21 days at 4°C .......................... 116

Figure 44. Percent (%) elongation at break with different ClOz concentrations for
films studied ....................................................................................................... 124

Figure 45. Percent (%) tensile strength at different Cl02 concentrations used for
the films studied ................................................................................................. 125

XX

INTRODUCTION

Microbial contamination of food products is a major problem that can
reduce the shelf life of food. Microbial contamination of food usually occurs in
post processing during packaging and distribution. Natural or chemical
preservatives have been developed and can be added directly to food to extend
its shelf life. However, food additives are becoming less attractive and
consumers are demanding minimally processed and preservative free products
(Collins-Thompson and Hwang, 2000). Moreover, substances that are directly
added to the food may be neutralized on contact or may rapidly diffuse so that
their effectiveness is limited. Also, the addition of additives can cause an
undesirable taste in the food at high concentrations (Ahvenainen, 2003).

Antimicrobial packaging is an active packaging concept to control
microbial growth and enhance food safety. In this concept, the preservative
agent is released from the packaging instead of adding it directly to the product.
The application can be more effective than direct addition of the preservative,
because the packaging system can maintain the required minimum inhibition
level through constant release with lesser amounts of the active agents.

Chlorine dioxide (ClOz) and allyl isothiocyanate (AITC) are rapidly gaining
attention as antimicrobial agents because of their excellent microbial control
performance even at very low treatment levels (Brody, 2005).

Chlorine dioxide (ClOz) is a highly oxidizing greenish yellow gas. It reacts
with the proteins of cell membranes in microorganisms, and destroys them

through cell wall oxidation. Microorganisms are unable to mutate to resistant

forms due to the oxidation and destruction of the proteins within the cell (Du et al.,
2002). CI02 can be used to kill large populations of fungi, bacteria, viruses, and
algae at low concentrations (Clordisys, 2003), and is widely used in multiple
industries such as water purification, vegetable/fruit processing plants, and a
variety of other facilities. Its application as a food antimicrobial agent has not
been studied a lot but several studies have shown its potential. Du et al. (2002)
tested the bactericidal effects of chlorine dioxide against Listeria monocytogenes
on apple surfaces. At a dosage level of 4.0 mg/I, Listeria monocytogenes was
decreased by 3.9~6.6 log cfu. Han (2000) achieved 3 log and 6.45 log reduction
of Listeria monocytogenes on green peppers with 0.62 mg/liter of CI02 and 1.25
mg/l of CI02, respectively. Gas type ClOz treatment has proved more effective
than aqueous Cl02 treatment. Han et al. (2001) found that ClOz gas treatment
(0.3 and 3.0 mg/liter) was more effective than aqueous CI02 on green peppers
inoculated with Listeria monocytogenes. Singh et al. (2002) found that CI02 gas
was more effective than washing with aqueous CI02 for lettuce and baby carrots
inoculated with E. coli O157.

Allyl isothiocyanate (AITC) is a pungent volatile component in all plants
belonging to the family Cruciferae, such as Wasabi, black mustard, etc. For a
long time, it has been used as a food flavor fortifier, and its powerful antimicrobial
effect has been shown in many studies. lsshiki et al. (1992) tested fresh beef
and sliced raw tuna using AITC as an antimicrobial, and showed a 5 log
reduction in mold (Aspergillus niger and so on) . Farber (1991) applied AITC to

sandwiches and pizza, alone and in combination with MAP packaging. The shelf

life of the foods was increased by 30 days and 17 days, respectively. Kim et al.
(2002) found that AITC extract in combination with acetic acid completely
inhibited aerobic microbial growth in cooked rice.

Gas phase AITC requires less dosage than an aqueous solution to obtain
the same result. Sekiyama et al. (1996) showed that AITC vapor application
required 1/500-1/1000 of the minimum inhibitory concentration when the liquid
was mixed with agar. lsshiki et al. (1992) suppressed the growth of bacteria and
yeast with a very small amount of AITC gas (16 to 110 ng/ml).

Fresh poultry cuts like boneless, skinless breast have become very
popular and represent one of the fastest growing markets because of their
moderate cost, healthy nutrients and convenience (Greene, 1998). Consumption
of poultry meat in the US has an annual growth rate of 7% and the market
segment has reached $17.5 billion. Like other fresh or minimally processed food
products, microbial contamination is one of the biggest problems of the poultry
industry, in terms of shelf life and food safety. The conventional chill pack of
chicken has an approximate shelf life of 4-5 days (Sams, 2001). Microbial
contamination of the product may occur in post processing during handling,
packaging and distribution. Salmonella spp. and Listeria monocytogenes are
common pathogenic organisms associated with raw meat and poultry products
(Mead, 2004).

Salmonella Typhimurium is one species of salmonella isolated from
chicken which causes gastrointestinal illness, including nausea, vomiting,

abdominal cramps, diarrhea, and headaches (FDA, 1992). Salmonella

Typhimurium has the highest rate of hospitalization for any of the Salmonella
species (Fisker et al., 2003).

Listeria monocytogenes is a major threat to the food industry as a post-
processing contaminant. It is able to multiply on chill-stored poultry meat (Hart et
al., 1991; Bajard et al., 1996), and is well adapted to food and food processing
environments where the growth of other pathogens may be restricted. The
infection commonly results in fever, musoel aches, and sometimes gastrintestinal
symptoms such as nausea or diarrhea. Infections during pregnancy can cause
serious problems such as miscarriage, premature delivery, or infection of the
newborn (CDC, 2005).

Modified Atmosphere Packaging (MAP) is an effective way to suppress
normal aerobic spoilage microorganisms. Low level oxygen packaging combined
with a carbon dioxide and nitrogen flush inhibit Gram-negative bacteria. Jimenez
et al. (1997) showed that aerobic bacteria were suppressed by modified
atmosphere packaging. A combination of the antimicrobials (CIO; and AITC)
with MAP may increase antimicrobial activity through synergetic mechanisms.
Thus, the research hypothesis of this study is that gas phase CIO; or AITC plus
MAP can be successfully used to enhance microbial safety of fresh chicken.

In addition, CIO; is a highly oxidizing agent (Lenntech, 2006). Degradation
of packaging polymer properties by polymer oxidization has been reported in
several studies (Hara, 1970; Ozen et al., 2002). Use of CIO; in food packages
may involve higher concentrations than those used for package sterilization

alone (Ozen et al., 2002). At high CIO; concentration, the treatment could affect

physical properties of the packaging materials. In order to prove the hypothesis,
the effect of CIO; gas exposure at several different concentrations on common
packaging films (LDPE, PVC, and PS) was investigated. Thus, objectives of this
study are;

(1) To investigate antimicrobial activity and minimum concentration of
CIO; and AITC against Salmonella Typhimurium and Listeria
monocytogenes.

(2) To evaluate overall performance of CIO; or AITC plus MAP
combinations for Salmonella Typhimurium and Listeria monocytogenes
in inoculated fresh chicken.

(3) To evaluate the effect of CIO; on the mechanical properties of the

packaging films.

CHAPTER I LITERATURE REVIEW
1.1. Principle of antimicrobial packaging

Active packaging is a system which possesses attributes beyond basic
barrier properties, and which is achieved by adding active ingredients into the
packaging system (Ahvenainen, 2003). Antimicrobial packaging is a type of
active packaging which could have a significant impact on product shelf life
extension and food safety. Traditional methods used for microbial control are
canning, aseptic processing, and MAP. Canned food cannot be marketed as
fresh. Aseptic processing can be expensive and is limited to specific types of
foods. MAP can promote the growth of pathogenic anaerobic bacteria (Farber,
1991). Directly adding antimicrobial agents to the food has limited effectiveness
because the active substance can be neutralized on contact or absorbed into the
food surface to affect its original taste (Quintavalla and \ficini, 2002).

If packaging materials have controlled release ability, the packaging
system can be used to delay the lag phase or reduce the growth rate of
microorganisms by controlled release of small amounts of antimicrobial agents.
Han (Ahvenainen, 2003) explained the basic principle of antimicrobial packaging

as part of a hurdle concept (Figure 1).

 

 

 

 

 

 

 

 

_l
O
O . C
. O I O .
O

. . . . . . .. ‘

Moisture Oxygen Microbial

Barrier Barrier Barrier

 

 

 

Figure 1. An antimicrobial packaging system as part of a Hurdle technology

(Source: Ahvenainen, 2003)

The antimicrobial function of the packaging system is another hurdle designed to
help prevent product spoilage while the conventional layers satisfy the barrier
requirement and provide physical protection. Embedding the antimicrobial agent
into the packaging material provides active protection against microorganisms
which is not achievable by conventional barrier packaging.

“Antimicrobial agents can be categorized into two types; those
antimicrobial agents which intentionally migrate to the surface of the food, and
those that are effective against surface growth without intentional migration of the
active agent to the food (immobilization)” (Suppakul et al., 2003). The first type
includes antimicrobial agents impregnated into packaging materials or coated
onto their surface. The purpose is to migrate partially into or completely surround
the food, thus extending the lag phase of the target microorganisms, or possibly

inactivating the microorganisms. The antimicrobial agents can be in gas or solute

7

form. Solute agents have to be in direct contact with the food, while gaseous
agents can fill the package headspace and thus contact more of the food product,
whereas the solute agents can only control the direct contact area. Recently, a
new antimicrobial migration system was developed. Thijssen patented a
biologically triggered release system called the “BioSwitch” concept (Jong at al.,
2005). This system releases its agents only if bacterial growth occurs. The
advantage of this system is the lower active substance requirement because the
impregnated antimicrobial agents are released only when there is microbial
growth.

Immobilization systems do not release antimicrobial agents but inhibit
microorganisms in the direct contact areas. Some antimicrobial packaging
systems utilize covalently immobilized antimicrobial substances that suppress
microbial growth when the target microorganism comes into contact with the
antimicrobial surface (Kourai et al., 1994; Suppakul et al, 2003). This type of
antimicrobial packaging can be more effective with liquid food products than with

solids (Ahvenainen, 2003).

1.2 Types of antimicrobial packaging
1.2.1 Incorporation of antimicrobial agents into films

The antimicrobial packaging system is usually prepared by either
incorporating the antimicrobial agent into the packaging material or by coating
the active compound on the surface of the packaging film (Vermeiren at al.,

2002). The incorporation of an antimicrobial substance into a packaging material

can take several approaches. One approach is to add the active agents to the
resin mix in the extruder, and the film is then produced. However, this approach
is often limited because of the heat liability of the component during extrusion, or
incompatibility of the component in the packaging material (Quintavalla and Vicini,
2002). The process has poor cost effectiveness because the added active agents
that typically migrate less to the surface of food than the original input. To limit
heat loss, a master batch can be produced at low temperature. The antimicrobial
agents can be mixed, extruded, and pelletized to create the master batch at low
temperature (Brody, 2001). Han and Floros (1997) produced an LDPE master
batch containing 1% potassium sorbate to prevent heat decomposition. The film
extruded using the master batch inhibited the growth of Saccharomyces spp. on
agar plates. However, Weng and Hotchkiss (1992) failed to suppress mold
growth when they incorporated 1% sorbic acid into an LDPE film. Sorbic acid
was insufficiently released from the film because of different polarity between the
agent and polymer. While PE has been widely used as a heat seal layer in
packaging, the copolymer polytheylene-co-methacrylic acid (PEMA) has been
found to be preferable in some cases for incorporation of antimicrobials. Weng at
al. (1999) modified films with NaOH, and then incorporated benzoic acid and
sorbic acid. Tests then showed the antimicrobial films modified with NaOH
exhibited more antimicrobial effectiveness due to a high release from the film.
Some antimicrobial packaging systems are covalently immobilized with
active agents to suppress microbial growth. Appendini and Hotchkiss (1997)

produced cellulose triacetate (CTA) film containing Iysozyme. The presence of

the Micrococcus Iysodeikticus spp. in tryptic soy broth (T SB) was effectively

reduced by immobilized Iysozyme containing CTA films (0.01 cmzl ml TSB).

1.2.2. Edible films and coatings for antimicrobial packaging

Edible films with various antimicrobial compounds also have been recently
investigated for food packaging. Edible coatings and films are produced from
polysaccharides, proteins and lipids. Edible coatings have advantages such as
biodegradability, edibility, and good oxygen barrier properties. When using edible
films and coatings as carriers for antimicrobial agents, the safety and edibility of
the coating system is essential.

Rodrigues and Han (2000) investigated the effectiveness of whey protein
isolate (WPI) films containing nisin, lysozyme and ethylenediamine tetracetic acid
(EDTA). Both nisin and Iysozyme containing films effectively inhibited
Brochothirix thermosphacta but were not effective against Listeria
monocytogenes. Research was also done by Ouattara at al. (2000) who
prepared antimicrobial films with various organic acids and essential oils in a
chitosan matrix. They found that the growth of Enterobacteriaceae and Serratia
quuefaciens was reduced during 21 days of storage. Another study was done by
Cagri et al. (2002). They produced whey protein isolate films containing 0.5 to
1.0% p-aminobenzoic acid (PABA) and/or sorbic acid (SA). The PABA and SA
containing films were effective in decreasing L. monocytogenes, E. coli, and S.

Typhimurium populations on inoculated slices of bologna and summer sausage.

10

1.2.3. Photon excited polymer

Ultraviolet irradiation treatment can increase surface amine concentrations
which are surface active antimicrobial sites. Hagelstein treated nylon film with
UV irradiation in order to increase the concentration of surface amine, and
improved the film’s antimicrobial effectiveness (Brody 2001). The author reported
a 3 log reduction of Staphylococcus aureus by contact with the treated film.
Another antimicrobial film was developed using a UV excimer laser. The
polyamide film (nylon 6,6) was irradiated in air using a laser at 193 nm, and the
amide groups on the surface of the nylon films were converted to amines (Brody
2001). Irradiation at 248 nm did not convert the chemical functional group from

an amide to an amine. (Ozdemir et al., 1999)

1.2.4. Gas or volatile generating sachets

For volatile antimicrobial agents, a sachet type application is possible in
an antimicrobial packaging system. Many volatile food grade antimicrobials can
be encapsulated in a sachet. The release rate is then influenced by the
permeability of the sachet. Ethanol, carbon dioxide, chlorine dioxide, and allyl
isothiocyanate releasing sachets have been developed and commercialized for

food applications (Kruijf et al., 2002; Suppakul et al., 2003).

11

1.3. Commercially used antimicrobial agents for foods
1.3.1. Chlorine dioxide (CIO;)

Chlorine dioxide (CIO;) is a synthetic, yellowish - green gas which
possesses a similar odor to that of chlorine. It has been used for many years as
a powerful biocide which exhibits rapid kill over a wide range of organisms. It
works through oxidation, and penetrates bacterial cell walls and reacts with vital
amino acids in the cytoplasm of the cell to kill the organism (Clordisys, 2003).
Since its antimicrobial mechanism is oxidation (rather than chlorinating organic
material like chlorine), it does not form undesirable pollutants such as
trihalomethanes (T HMs) or dioxins. Thus, the use of chlorine dioxide has the
advantage that it produces less harmful byproducts than chlorine (Lenntech,
2006).

The first CIO; was produced in 1814 by Sir Humphrey Davy. He found that
CIO; gas can be generated by reaction between sulfurous acid (H;SO3) and
potassium chlorate (KCIO3). Nowadays, CIO; is generated by reaction of
hydrochloric acid (HCI) and sodium chlorate (NaC|03), which can be used to
produce large quantities inexpensively (Clordisys, 2003).

2NaCI03 + 4HCl -) ZCIO; + Cl; + 2NaCl +2H;O

Chlorine dioxide is currently used on food and medical equipment, counter
surfaces, and processing facilities. The compound is unstable, and easily
oxidized by sunlight. It is reduced to Cl‘, CIO;', and CIO3' as end products. Since
CIO; is an explosion hazard above 10% in air, it is difficult to store or transport as

a gas (EPA .1999). Therefore, it is often produced and used at the same location

12

(Lenntech, 2006). Thus, equipment for on-site chemical or electrochemical
generation is often put in place. However, the significant capital equipment and
operating costs have limited the use of chlorine dioxide to large-scale
applications only. The use of CIO; is also governed by strict hazardous material
handling regulations (Lenntech, 2006). Recently, CIO; sachets and films have
been developed to offer cost - effective alternatives to bulk CIO;. For example,
ICA Trinova and Engelhard manufacture CIO; release sachets. These utilize
sodium chlorite and zeolites with acidic minerals to generate small, precise
quantities of CIO; (Speronello, 2005). Bernard Technologies Inc. (Chicago,
Illinois) patented CIO; generating LDPE films (Microatmospherem) which consist
of hydrophobic materials containing an acid releasing agent and hydrophilic
materials containing anions that are able to react with hydronium ions to
generate a gas (US Patent, 1994). Since it is an irritating oxidizer, high
concentrations of CIO; gas may cause irritation of the eyes, skin, and lungs.
(OSHA, 2006) The use of chlorine dioxide for antimicrobial food packaging has
been recently permitted with “no objection” notification from FDA (2001 ). To
minimize any potential adverse effects associated with CIO;, it is important to
determine the minimum level of CIO; gas necessary to inhibit the target
organisms.

Several studies have examined the effect of CIO; gas treatment for food
packaging applications. Wellinghoff (1995) used CIO; to inhibit Escherichia coli in
a ground beef meat product, and attained a 2-6 log reduction, depending on the

dosage level. Knight (2001) reported a 90% microbial reduction using a high

13

concentration of CIO; (18 wt% of CIO;) on raw beef. On the down side, an
undesirable quality change was observed with the raw beef. The color of the beef
changed from red to dark green when treated with CIO;, Ellis et al. (2005) used a
combination of modified atmosphere packaging and CIO; (sachet) with fresh
chicken breast, and reduced Salmonella Typhimurium by 1 log. There was no off-

odor detected following CIO; treatment.

1.3.2. Allyl isothiocyanate (AITC)

Allyl isothiocyanate (AITC) is a major volatile pungent component of
mustard, horseradish and wasabi. AITC is present as a precursor in the form of
“sinigrin" in these natural plants. The sinigrin is hydrolyzed by the endogenous
enzyme myrosinase when the plant tissues were suffered by physical injury, and
yield ally isothiocyanate (Muthukumarasamy et al., 2003).

In the early 20th century, Hofmann discovered the effectiveness of AITC as
a preservative in mustard oil (Winther and Nielsen, 2006), Since then, the strong
antimicrobial effectiveness of both AITC liquid and vapor has been demonstrated
in many studies (Lin et al., 2000). AITC has properties similar to those of other
alkyl and aryl isothiocyanates which are used as a strong lachrymator or skin
vesicant. AITC can be used as a flavor fortifier for foods (ex. mustard and
horseradish) if used in the proper amount (Radomir et al., 1997).

The pungent AITC flavor is extremely reactive with nucleophiles such as
water and SH- and OH- groups commonly found in food (Drobnaica et al., 1977).

High concentrations of AITC are not typically useful for many food applications

14

because of its extremely pungent odor. Delaquis and Mazza (1995) mentioned
that the amount of isothiocyanate (AITC) that can be successfully be used for
food preservation is limited because of the possible negative sensory response it
can cause. Kim et al. (2002) also found that too much AITC (2000 pg) was not
acceptable to consumers due to its strong odor. Dunnick et al. (1982) observed
transitional cell papilloma in the urinary bladder (in 4 of 49 rats) at a high dosage
level (25 mg/kg/day). This indicates a possible safety concern at high
concentration.

AITC can be applied both as a liquid and gas type. Gas type AlTCs have
been shown to be more effective as antimicrobial agents. lsshiki et al. (1992)
presented data obtained from the Institute for Fermentation, Osaka (IF 0, Osaka),
the Japan Collection of Microorganisms (JCM, Saitama), and the American Type
Culture Collection (ATCC, Rockville, MD). The growth of bacteria and yeast were
suppressed at AITC concentrations of 31 to 420 ug/petri dish. These doses were
equivalent to 16 to 110 ng/ml. They suggested that AITC could be used as a
vapor for MAP. Nielsen and Rios (2000) used 3.5 jig/ml AITC in the gas phase
for hot dogs and rye bread. All fungi (Penicilium commune, P.roqueforti,
Aspergillus flavus, Endomyces filbuliger, etc) tested were eliminated, and it was
anticipated that the minimum inhibition dosage could be decreased when used
with MAP. Suhr and Nielsen (2003) reported that the minimum inhibitory
concentration was 250 times lower in the gas type than in liquid type.

The use of naturally extracted AITC in certain food packaging systems has

been allowed in Japan as a preservative (lsshiki et al., 1992). WasaOuro

15

(Mitsubish-Kagaku Foods Corporation, Japan) is an example of a
commercialized AITC preparation for food applications in Japan. The company
offers antimicrobial labels, and sheets to prevent mold or bacteria growth on
bakery, ready-to-eat foods, and vegetables. Recently, the use of AITC for food
packaging was granted a no objection status from FDA as GRAS (FDA, 2006).

1.3.3. Silver ions

The antimicrobial activity of metals is due to the ions formed by the metals.
Silver and copper ion, quaternary ammonium salts, and natural compounds such
as Hinokitiol are generally considered safe antimicrobial agents (Suppakul et al.,
2003). Silver is the most effective metallic antimicrobial at very low
concentrations (0.02 - 0.05 ppm). Silver does not release the ion easily,
compared to other metals such as copper. Thus, its antimicrobial activity is not as
strong in its metallic state, and its greatest potential appears to be as releasable
silver salts such as silver nitrate and silver zeolite. Silver substituted zeolite (Ag-
zeolite) is the most common metallic antimicrobial agent. It retards a range of
metabolic enzymes and has a uniquely broad microbial spectrum (Brody, 2001).
Its antimicrobial effectiveness has been demonstrated in many studies against a
wide range of bacteria, yeast, fungi and molds. Galeano et al. (2003) used
stainless steel surfaces coated with silver zeolite (Aglon) to inhibit vegetative
cells and Bacillus spores, and the silver zeolite coating inactived approximately 3

log of vegetative cells within a 5 to 24 h period.

16

AglONTM (Aglon Technology, Inc., Wakefield, MA) produces a
commercialized Ag-zeolite product. It is available in many food packaging
applications such as bulk food storage containers, paperboard cartons, plastic

and paper food wraps, and so on.

1.3.4. Ethanol

Ethanol (ethyl alcohol) is a flammable, colorless chemical compound with
a distinctive perfume-like odor. It is used conventionally in medical and
pharmaceutical packaging applications (Suppakul et al., 2003). Ethanol
generating films and sachets have been developed and marketed to prevent
mold growth of intermediate moisture foods, cheeses, and bakery products
(Smith et al., 1987; Suppakul et al., 2003). Labuza et al. (1989) introduced an
adhesive-backed film that could be taped on the inside of a package to provide
antimicrobial activity.

In Japan, several ethanol vapor - generating sachets have been
developed and commercialized, such as Ethicap (Freund Industrial Co. Ltd,
Japan) and Ageless type SE (Mitsubishi Gas Chemical 00,. Japan). Smith et al.
(1987) used a commercialized ethanol release sachet (Ethicap) to extend the
shelf life of apple pie, by reducing fungal growth. F ranke et al. (2002) also used
the sachet with pre-baked buns. Mold growth was delayed for 13 days in the
presence of the ethanol emitting sachet while that in the control package

normally occurred within 4-6 days. On the down side, though, the studies showed

17

that high ethanol release caused strong undesirable chemical odor in most food

products tested with the sachets (Franke et al., 2002; Smith et al, 1987).

1.3.5. Nisin

Nisin is a bacteriocin which is included in a group of microbial synthesized
proteinaceous antimicrobial agents. It is produced by the lactic acid bacteria,
Lactococcus lactis, particularly during the exponential phase of bacterial growth
(Buchman et al., 1988) and is effective against a broad range of Gram-positive
bacteria, including Listeria, Bacillus, Clostridium and other lactic acid bacteria
(Ahvenainen, 2003; Cleveland et al., 2001).

Nisin was the first bacteriocin to attain GRAS (General Recognized As
Safe) status. Siragusa et al. (1999) reported the potential of incorporating Nisin
directly into LDPE film, to enhance product safety. These peptide type
compounds can only inhibit bacteria on food closely in contact with the
microorganism. Nisaplin® (Danisco, Copenhagen, Denmark) is a commercialized
Nisin-based antimicrobial agent. It can be applied directly onto or to foods or be
incorporated into a packaging film. Cabo et al. (2001) applied Nisaplin® to fresh
pizza as a preservative, in combination with MAP. The results showed significant
increases in shelf life. Viskase Corp. (IL, USA) uses Nisaplin® in the casing of

frankfurters and other deli meat products (Charest, 2004).

18

1.3.6. Natural extract

. Antimicrobial extracts from plants are desirable for food packaging
applications because of their apparent safety and the consumer’s perception of
natural materials. Many plant extracts from grapefruit seeds, cinnamon, and
horseradish have been investigated as antimicrobials. CitrexTM (Citrex inc.,
Coconut Grove, FL) is a commercialized grapefruit seed extract (GFSE), that can
be incorporated at 1% w/w into LDPE film (Lee et al., 1998). Lee et al. observed
aerobic bacteria and yeast reduction on curled lettuce treated with GFSE. Ha et
al. (2001) extruded GFSE incorporated multilayer films, and found coated film
with 1% GFSE to have the highest effectiveness against E.coli, S.aureus, and

Bacillus subtilis.

1.3.7. Carbon dioxide (00;)

Carbon dioxide (CO;) is a well known antimicrobial gas which can
suppress Gram-negative psychrotrophic bacteria (Daniels et al., 1985). The use
of carbon dioxide to extend the lag phase and decrease the growth rate of
microorganisms on foods is well known (Kruijf et al., 2002). Therefore, this
antimicrobial has been commercially applied to many refrigerated products via
modified atmosphere packaging.

There are many commercialized CO; generating sachets, such as
FreshPax (Multisorb Tech Inc., USA), Ageless (Mitsubishi Gas Chemical Co.,

Japan), and Freshilizer (Toppan Printing 00., Japan). These sachets can also

19

be combined with an oxygen scavenging function. To enhance the antimicrobial
effectiveness and prevent package collapse as a result of oxygen absorption
(Smith et al., 1995), many commercialized CO; generating sachets are combined
with oxygen scavengers (Smith et al. 1995, Suppakul et al., 2003).

High CO; concentrations, however, can cause changes in product flavor
and the development of undesirable anaerobic glycolysis in fruits and vegetables.
For these reasons, CO; generators are more useful in applications such as fresh

meat, poultry, fish and cheese packaging (Suppakul et al., 2003).

1.4. Regulatory issues for antimicrobials in food packaging

Although antimicrobials have a promising potential, the current use of
antimicrobial agents in food packaging is limited, with one reason being the
regulations concerning their use. The use of antimicrobials in processed food and
food packaging must follow the guidelines and regulations of the FDA1 under
section 409 of the F FDCA2 (F DA/CSFAN, 1999). Agents used in antimicrobial
packaging are considered food additives. Antimicrobial packaging can only
contain agents which are approved by the authorization agency (FDA) or allowed
for use within the concentration limits for food safety enhancement or
preservation (Ahvenainen, 2003). Some organic acids, bacteriocins and volatile
compounds derived from plants have been approved by the FDA. Table 1 shows
the list of permitted antimicrobial agents which can be used for food packaging

materials.

 

‘ FDA: Food and Drug Administration
2 FFDCA: Federal Food, Drug and Cosmetic Act

20

Table 1. List of antimicrobial agents that are approved for food additive (food

packaging) materials in USA

 

 

AM agents Authority AM agents Authority
(US) (US)
Acetic acid GRAS Lysozyme FA
Allyl GRAS Malic acid GRAS
isothiocyanate
Adipic acid GRAS Natomycin FA
Benzoic acid GRAS Nisin GRAS
Calcium acetate GRAS Parabens FA
Carvarcol GRAS Phosphoric acid GRAS
Citral FA Polyphosphate GRAS
Citric acid GRAS Potassium sorbate GRAS
p-Cresol GRAS Propionic acid GRAS
Chlorine dioxide GRAS Propyl paraben GRAS
Ethanol FA Sodium acetate GRAS
EDTA FA Sodium GRAS
bicarbonate
Geraniol GRAS Sodium benzoate GRAS
Glucose oxidase GRAS Sorbic acid GRAS
Lactic acid GRAS Succinic acid GRAS
Lauric acid GRAS

 

1. Source: Suppakul et al. (2003), Davidson et al. (2005)

2. Classification in accordance with Food and Drug Administration (FDA) Title 21
of the Code of Federal Regulation (21 CFR) wherein substances intended for use
in the manufacture of food stuffs for human consumption are classified into 3
categories: food additives (FA), pre-sanctioned food ingredients and substances
generally recognized as safe (GRAS)

21

FDA safety reviews are sometimes in concert with other government
agencies such as EPA‘. EPA also regulates pesticide chemicals (antimicrobials)
with a tolerance or exemption from tolerance under section 408 of FIFRA2 (Misko
and Rothschild, 2001).

Historically, even if the antimicrobial is regulated as a food additive by the
FDA, an antimicrobial intended to have any ongoing effect on a permanent or
semi-pennanent food contact article needs to have its safety reissued as an
antimicrobial pesticide residue in or on the food packaging under section 408 of
FIFRA. Therefore, confusion can exist when antimicrobials are incorporated into
food packaging materials.

In 1998, the Antimicrobial Regulation Technical Corrections Act (ARTCA)
(21 CFR Part 173) was enacted in part to correct the regulatory authority
confusion between FDA and EPA. With this amendment, FDA now regulates all
antimicrobials incorporated in, or applied to, food packaging materials regardless
of whether the substance is intended to have an ongoing effect on any portion of
the packaging. However, the use of ethylene oxide and propylene oxide on
processed food is under the jurisdiction of EPA under section 408 of FIFRA
(Misko and Rothschild, 2001). EPA also regulates antimicrobials that are
incorporated in, or applied to, objects that have a semi-permanent or permanent
food-contact surface, other than food packaging, in order to provide a sanitizing

effect on surfaces such as a cutting board or a conveyor belt.

 

I EPA: Environmental Protection Agency
2 FIFRA: Federal Insecticide, Fungicide, and Rodenticide Act

22

In Europe, no specific legislation has yet been approved about active
packaging (including antimicrobial packaging). Antimicrobial compounds
released from packaging are still regarded as migration agents so that the
application has to follow the “Legislation for migration of components from the
packaging material to the product” (Quintavalla and Vicini, 2002). Van Beest
mentioned that the overall migration limit from packaging materials into a food is
set too small (60mg/kg) to be compatible with the aim of antimicrobial release
packaging (Suppakul et al., 2003). A new approach in food packaging
antimicrobial regulation is needed.

The use of natural plant extracts as antimicrobial agents in food packaging
goes through an easier regulating process when compared to chemical

antimicrobial agents (Ahvenainen, 2003).

1.5. Common pathogenic organisms in meat products
1.5.1. Salmonella Typhimurium
Salmonella spp. is Gram-negative, facultatively anaerobic bacteria of the
family Enterobacteriaceae, made up of nonspore-forming rods, usually motile
with peritrichous flagella. It is one of the major foodborne pathogens commonly
associated with raw poultry and poultry products (Mead, 2004). The usual
symptoms of Salmonellosis include diarrhea, abdominal pain, cramps and fever
12 to 72 hours after infection and may last for up to 7 days (FDA, 1992).
Salmonella causes an estimated 1.4 million illnesses in the USA annually.

The most common serotype was Typhimurium (CDC-b, 2006). Salmonella

23

Typhimurium DT 104 frequently causes serious complications within
Salmonellosis, and caused the highest rate of hospitalization within the
Salmonella species (Fisker et al., 2003). According to an epidemiological study
done in 1993, 34 of 83 Salmonella Typhimurium DT 104 gastroenteritis cases
required hospitalization and 10 people died (Bionewsonline, 2005). Another study
done in 2006 reported that 27 of 58 Salmonella Typhimurium DT 104 cases
resulted in hospitalization (CDC-b, 2006).

In July 1969, an outbreak of gastroenteritis due to Salmonella
Typhimurium occurred in the greater Spokane area of Atlanta. The outbreak was
associated with “ready to eat” barbecued chicken from a Spokane supermarket.
107 were known to have been ill, 29 were hospitalized, and two died (Wemer et
al., 1969). The Public Health Unit of the Dodge County Human Services and
Health Department (DCHSHD) in Wisconsin reported 107 confirmed and 51
probable cases of Salmonella serotype Typhimurium gastrointestinal illness in
1995, associated with eating contaminated raw ground beef during the winter
holiday season (CDC, 1995). One of the largest outbreaks of salmonella
gastroenterisitis was reported in Spain. As of 8 August 2005, a total of 2,138
cases were linked to processed chicken. The reported cases were linked to a
single brand of pre-cooked, vacuum packed roast chicken which was distributed
throughout Spain. All implicated chicken products were recalled from commercial
outlets (Nathnac, 2005). As of November, 2006, Centers for Disease Control and
Prevention reported a multistate outbreak of Salmonella Typhimurium linked to

contaminated tomato consumption at restaurant chains. The outbreak caused

24

183 cases in 21 states, most patients had fever and diarrhea, and 22 (12%) were

hospitalized; there have been no deaths reported (CDC, 2006c).

1.5.2. Listeria monocytogenes

Listeria monocytogenes is a psychrotrophic, Gram-positive, rod-shaped
bacterium that can grow under refrigeration, in anaerobic conditions and in low
oxygen atmosphere (Bonilauri et al., 2004). Listeria monocytogenes can persist
in cool, damp areas of raw meat or processed meat plants. Drains and
refrigeration freezing equipment can also harbor Listeria monocytogenes (Cox et
al., 1989). This is a common contaminant of raw poultry and up to 60% of
processed chicken carcasses may harbor low numbers of the organism (Cox et
al., 1999). Live birds are rarely found to be positive and contamination occurs
mainly during processing (Cox et al., 1989). Listeria monocytogenes is a major
concern to the food industry. Approximately 2,500 illnesses and 500 deaths were
caused by the pathogen, annually in the US (CDC, 2005b).

Consumption of Listeria monocytogenes contaminated foods causes the
human disease called Iisteriosis. Listeria monocytogenes is a particularly lethal
pathogen to the immunocompromised such as pregnant women and their fetuses,
AIDS patients, alcoholics, and the elderly. Dairy products can be particularly
susceptible to contamination by Listeria monocytogenes because cows can
shed the organism in the milk. Thus, contaminated raw milk could serve to
introduce the bacterium into dairy plants or foods made from raw milk (Donnelle,

1990). During August 1998 to February 1999, one listeriosis outbreak from hot

25

dogs resulted in 101 cases of listeriosis in 22 states, and created great concern
among ready-to-eat food manufacturers (CDC, 1999). From May to November,
2000, a multistate outbreak was caused by a Listeria monocytogenes
contamination with 29 illnesses, which were linked to eating deli turkey meat
(CDC, 2000). From July to September, 2002, another multistate outbreak was
reported of Listeria monocytogenes infections with 46 culture-confinned cases,
seven deaths, and three stillbirths or miscarriages in eight states. The outbreak
was also linked to sliceable turkey deli meat in Northeastern United States. The

meat was contaminated from a poultry processing plant (CDC, 2002).

1.5.3. Other foodborne pathogens

Thennophilic campylobacters, (especially Campylobacterjejuni), are
recognized as a major, worldwide cause of human enteritis (Mead, 2004). The
infection source is primarily poultry meat, contaminated water supplies, raw milk
and pets. Campylobacter human infections are often associated with
consumption of undercooked meat or handling of raw product since it is easily
transferred to kitchen tools such as cutting boards and plates. Most cases are
sporadic.

Eschericia coli include hundreds of bacterial strains which are commonly
found in the intestines of healthy cattle, deer, goats, and sheep. Although most
strains are harmless, one strain produces a powerful toxin that can cause severe
illness. Eco/i O-157:H7 is the most important serotype and has been the cause

of various food associated outbreaks (Mead, 2004). E. coli O157:H7 was first

26

recognized as a cause of illness in 1982 during an outbreak of severe bloody
diarrhea. People can become infected with E. coli 01 57:H7 in a variety of ways,
and most illness has been associated with eating undercooked, contaminated

ground beef (CDC, 2006a).

1.6. Chicken shelf life

Poultry meat is a highly perishable product because it provides an
excellent medium for microorganisms. Thus, the primary cause of spoilage in
fresh poultry is microbial growth, and shelf life can be extended by controlling
growth of the usual spoilage organisms. Mead (2004) said that “the meaning of
meat spoilage is that meat has become unfit for human consumption, due largely
to the growth and metabolic activities of particular microorganisms.” Thus, there
may be negative changes in the odor, flavor or appearance of meat making it
unacceptable. In the case of chilled stored chicken portions, Ayres et al (1950)
reported an ester-like odor and observed numerous small colonies on the surface
of the chicken, which developed eventually into a layer of slime. At this stage, the
odor becomes ammonium like and microbial populations exceed 1.0 x 10°
cfulcmz. Elliott and Michener (1961) found that off odors were associated with 1.6
x 105 to 1.0 x 108 cfu/cm2 organisms, and slime appeared between 3.2 x 107 to
1.0 x 109 cfu/cmz. At spoilage, the predominant organisms were pseudomonas
spp. (Cox et al, 1975). Most of the odor, color, and textural changes were
attributable to microbial metabolism. The main spoilage organisms of chilled

meat, in aerobic conditions, preferentially utilize glucose. During the logarithmic

27

phase of growth, glucose metabolism results in complex mixtures of short-chain
fatty acids, ketones and alcohols (Dainty, 1996). At microbial populations above
107 cfu/cmz, the source of glucose is soon depleted. Then, lactate and amino
acids begin to be utilized and malodorous compounds are formed, especially
from the sulphur - containing amino acids, cysteine, cystine and methionine.
These contribute to the putrid, sulphury off odors of spoilage (McMeekin and
Thomas, 1980). Once the pool of amino acids has been depleted and cells enter
the stationary growth phase, having reached maximum numbers, there may be
microbial proteolysis and lipolytic activity. The oxygen concentration in a meat
package is greatly diminished and the concentration of carbon dioxide increased,
resulting in a markedly reduced growth of Pseudomonas spp. This is mainly due
to the inhibitory effect of the high level of carbon dioxide. The slower growing
lactic acid bacteria tolerant to carbon dioxide then begin to dominate and ferment
glucose to products that include lactic, iso—butanoic, and acetic acids, which give

the meat a characteristic sour/cheesy odor (Mead, 2004).

1.7. Packaging of fresh meats

The oldest method of packaging and distributing fresh poultry meat is in a
“wet shipper". The wet shipper is a wax coated corrugated box in which whole
birds are placed with ice (Sams, 2001). Currently, poultry meat packaging is
including more small, consumer portions (known as “case ready”) processed and
packaged at central processing locations (Mead, 2004). Still, almost 90% of

these packaged chicken parts are packaged using highly oxygen permeable

28

polystyrene foam trays with a highly oxygen permeable PVC or other clear
stretch film overwrap. Whole carcasses are packaged in polymer bags, either
sealed or with clip closures (Sams, 2001).

Raw poultry meat is highly perishable even when stored in chilled
conditions because the growth of psychrotrophic bacteria will cause spoilage.
Normally, the shelf life of the conventional meat package is less than 5 days
(Ellis et al., 2005). Packaging is important to maintain the safety of perishable
products such as poultry and red meats. Modified atmosphere packaging (MAP)
is a useful method that can be used to extend the shelf life of fresh meat. MAP is
typically accomplished by flushing the package headspace with a desired gas
mixture to replace ambient air. Carbon dioxide (CO;) and nitrogen (N;) have
been used for this purpose. CO; particularly delays the lag phase of the microbial
growth and lengthens the generation times of the organisms. Ogilvy and Ayres
(1951) found that the shelf life of poultry meat can be expressed as a linear
function of CO; concentration in the air. However, too high CO; treatment
concentration can affect the color and texture of the meat product. For retail
packaging, the CO; concentration should be limited to 35% to minimize package
collapse and excessive purge (Sams, 2001). Many studies have been carried out
in order to study the effectiveness of different gas compositions on the
preservation of fresh meat. A minimum concentration of CO; is considered to be
at least 20% to obtain significant improvement in shelf life (Greengrass, 1993).
Jimenez et al. (1997) performed a study to determine the influence of gas

composition on the shelf life of chicken breasts at 4°C for 21 days. They found

29

that a 30%CO;/70%N; blend extended shelf life up to 14 days and
70%CO;/30%N; extended it to 21 days compared to 5 days for air-packaged
samples. Lawlis and Fuller (1990) reported that refrigerated meat shelf life is
typically 14 days with MAP, and the shelf life could be slightly longer if
accompanied by deep chilling. N; may be used as a filler gas to minimize purge
in the absence of oxygen. CO; with low levels of oxygen will inhibit many aerobic
spoilage organisms such as pseudomonas (the main spoilage organism in chilled
meat) (Mead, 2004). However, Sander and $00 (1978) reported that lactic acid

bacteria ultimately predominate in the presence of CO;

1.8. Poultry meat market segment & packaging

According to a “US poultry market research trend analysis“(Global
lnforrnation Inc., 2005), the total market growth from 1999 to 2004 has been 7%,
and reached $17.5 billion in 2004.

USDA’s foreign agricultural service reported that meat production
including beef, pork, and poultry has increased about 3% annually during the last
decade. These gains have been led by poultry. Poultry meat production has
expanded more than 5% each year on average, offsetting little or no growth in
beef production since 1988 (Greene, 1998).

There are several factors contributing to this growth. One of the main
factors is poultry’s good nutritional profile, versatility and low cost. For example,
poultry cuts like boneless, skinless breast have become popular because of their

healthy nutrients (low fat, low carbohydrate, and high protein content) and

30

convenience. Semi-prepared and pre-marinated varieties of poultry products are
also convenient and have been well-received. Cost is another reason; poultry
meat is a cheaper source of protein than beef, and it has been extremely well-
received in China, Russia, and Mexico (Greene, 1998). As a result of the strong,
growing demand for poultry meat, global exports advanced at a double digit pace
in the 19905. The US supplies about 53% of global poultry imports, and
promotion of poultry products in the fast food industry has also contributed to this

demand.

31

CHAPTER II. MATERIALS AND METHODS

2.1. Culture preparation

Three strains each of Listeria monocytogenes (1002, 1176, 1304) and
Salmonella Typhimurium (G10127, G10601, G10931) were obtained from Dr.
E.T. Ryser, Department of Food Science and Human Nutrition, Michigan State
University, East Lansing, MI. The cultures were stored at -70°C in trypticase soy
broth (T SB) (Difco Laboratories, Detroit, MI) containing 10% (v/v) glycerol (J.T.
Baker, Phillipsburg, NJ), and then subcultured twice in T88 containing 0.6%
(w/v) yeast extract (Difco) (TSBYE) at 35 °C/18~24 hours (1OQCFU/ml).The three
strains of Listeria monocytogenes and Salmonella Typhimurium culture were
then combined in equal volumes and agitated to obtain two three-strain cocktails.
Finally, the two cocktails were serially diluted in 9 ml of 0.1% peptone water to
produce different microbial densies (101-106). Figure 2 shows the Listeria

monocytogenes and Salmonella Typhimurium culture preparation procedure.

32

Frozen cultures stored at -70°C
Listeria monocytogenes (1 102,1 176,1304)

Salmonella Typhimurium (G10127,G10601,G10931)

l

lnoculate in 9 ml of trypticase soy broth with 0.6 % yeast extract (T SBYE)
35°C 122 hr

lnoculate in 9 ml of trypticase soy broth with 0.6 % yeast extract (TSBYE)
35°C 122 hr

Microbial culture (~109 CFU/ml)

Salmonella cocktail (3 strains) Listeria cocktail (3 strains)

Figure 2. Listeria I Salmonella culture preparation used for inoculation of the
chicken

33

2.2. Determine headspace concentration of chlorine dioxide (CIO;) and its
antimicrobial effectiveness

In order to investigate the antimicrobial effectiveness of CIO; against L.
monocytogene and S. Typhimurium, each culture was inoculated on a different
selective growth medium (agar), and incubated in the presence of different CIO;
concentrations. To perform this study, 1 quart (946 ml) Mason glass canning jars
were purchased from a local store (Meijer, Lansing). The metal lid of the glass jar
was modified by cutting with holes (0.25 in) into the lid and attaching, and flexible
Tygon tubes to the lid with 3l8 in diameter brass fittings. The tubes were closed
using laboratory pinch clamps. Figure 3 provides a schematic of the jar used for
CIO; analysis. A known amount of CIO; gas was injected into the jar headspace

to yield a range of headspace concentrations in the glass jar.

. clamp
. .1: ............... rubber septum

- -------------- tygon Tube

 

 

 

 

 

 

__ ............ mason glass jar

 

 

-----------

Figure 3. Schematic of canning jar used for CIO; concentration study

34

A CIO; sachet which is able to release CIO; gas in water was provided
from ICA Trinova. The sachet contains 29 of the company’s special formula, and
release of CIO; gas was begun by soaking the sachet in distilled water. One
sachet in 1 liter water provides approximately 40,000 ppm (107 mg/l) of dissolved
CIO; gas. The CIO; in the aqueous mixture was stable in the dark and
maintained a constant CIO; headspace concentration according to the vapor-
quuid equilibrium relationship of Henry’s law. (Montgomery, 1985)

X g = HPg

where,

X g = equilibrium mole of dissolved gas

H =Henry’s law constant (H of CIO; was 54 at 20°C, atm)

P8 = partial pressure of gas, atm

Figure 4 shows the glass container system used in the test. The CIO;
aqueous mixture was diluted with distilled water to produce 1000 ppm (2.76 mg/l)
of CIO; according to Henry’s law. The headspace concentration was then
measured using the ICA vapor equilibrated titration method (Appendix I). In order
to create desirable CIO; concentration in the canning jar (Figure 3), headspace
gas from the equilibrium CIO; generating container (Figure 4) was injected into
the canning jar. Injection volumes varied depending on desired headspace
concentration. For example, 1 ml of headspace gas containing 27.6 pg CIO; gas
was injected to produce approximately 27.6 pg CIO;/l (10 ppm) in the glass jar. In
the same way, 0.5 ml of headspace gas was removed from the equilibrium CIO;
gas generating container to produce 5 ppm.

35

------------- equilibrated
CIO; (1000ppm)

 

 

  

' H .. ~32: . - - -~— - - - aqueous CIO; mixture

Figure 4. Equilibrium CIO; gas generating system

To evaluate antimicrobial activity as a function of CIO; treatment, the glass
jar and lids were washed in hot soapy water, rinsed, and wiped with a 70%
ethanol solution to sterilize. After drying in a hood, an inoculated agar plate was
placed into the jar. 0.1 ml of each prepared cocktail (inoculum densities ranging
from 10° CF U/ml to 10°CFU/ml) was inoculated onto the agar plates (60 mm
diameter x 15 mm height). Modified Oxford agar (MOX) and Xylose Lysine
Deoxycholate (XLD) agar were used for Salmonella Typhimurium and Listeria
monocytogenes, respectively. After the metal lid was tightly closed, the prepared
CIO; gas was injected through the septum attached to the Tygon tubing (Figure
3). The closed jar was then placed in an incubator (37°C), and stored for 48 hr.
After the incubation period, the glass jars were removed, and evaluated to
determine the antimicrobial effectiveness with the amounts of injected CIO;. Lack
of growth on the plates was considered to be due to the inhibition of the inoculum

densities by CIO;. The amount of remaining CIO; in the jar was also monitored

36

during the incubation time. Headspace CIO; concentrations in the canning jar
were determined by using a chemical detector tube and Kwik—draw pump (MSA,
Pittsburgh, PA). The detector tube marks the concentration of CIO; by color
change (oxidation of aromatic amine by CIO;) after absorbing 120 cc of
headspace gas in the jar. To prevent development of a partial vacuum when the
headspace gas was pulled out of the jar, nitrogen gas was flowed into the jar

through another septum at an equal rate (approximately 150 cc/min).

 

Figure 5. Photo of the detector tube and pump used to determine CIO;
concentration

37

2.3. Headspace concentration of Allyl isothiocyanate (AITC) and its antimicrobial
effectiveness

1 quart (946 ml) glass canning jars (Mason, Lansing, MI) were purchased
from a local store (Menard, Lansing, MI). Their metal lid was equipped with a 3/8
in diameter septum adaptor. An orifice in the middle, sealed by silicon rubber,
served as a septum for sampling the headspace gas in the glass jar. Figure 5

shows the schematic of the jar.

; """""""""" adaptor

 

 

 

 

mason glass jar

petri dish

AL screen

 

 

 

AITC soaked filter paper

Figure 6. Schematic of canning jar used for AITC testing
AlT gas concentration was determined using a HP 6890 gas chromatograph
(Hewlett-Packard. Wilmington, DE), equipped with a crosslinked 5% PHME
siloxane column (30 m x 0.32 mm x 0.25 um film thickness). 1 ml of headspace
gas was withdrawn with a gas tight syringe from the glass jar, and injected into
the gas chromatograph. The flow rates of nitrogen carrier, hydrogen and air to

the FID detector were 30, 30, and 240 ml/min, respectively. The temperature of

38

the oven was programmed to hold at 45°C for 4.5 min and increase at 60°Clmin
to 230°C, and hold for 5 min. The temperatures of the injection port and detector
were set at 250°C and 290°C, respectively. To develop the standard curves, pure
AITC (95% Sigma-Aldrich, St Louis, MO) was mixed and diluted with hexane,
and a series of standard solution of known AITC amounts were injected into the
GC to prepare a standard calibration curve. The standard curve is shown in

Figure 7, and a copy of the actual chromatogram is shown in Figure 8.

3e+6

 

3e+6 j

26+6 -

2e+6 ~

Area

1e+6 -

5e+5 -

 

 

 

 

I

o 200 400 600 800 1000 1200
Concentration (ng/ml)

Figure 7. Standard curve of AITC concentration vs. area in a hexane solution

39

 

lemm-
Salm-

room-

vu- v-r-I '1‘u- ."'..q"r.‘l‘."‘ Irv‘liryv‘v-
lilo zilo 3.60 4.60 5m 6m rm 8]

 

 

 

 

Figure 8. Chromatogram of AITC in a Hexane- AITC solution

In order to determine the antimicrobial activity of AITC, pure allyl
isothiocyante (95%, Sigma Chemicals, St Louis, MO) was mixed in warmed
(30°C) corn oil, and AITC-oil mixtures of 0.5, 1, 2.5, and 5 ul/ml were added to
filter paper (4.24 cm diameter, Whatman Inc., New Jersey, USA).

Listeria monocytogene and Salmonella Typhimurium inoculated agar
plates were prepared as previously determined (section 2.2). To prevent direct
contact of the AITC soaked filter paper and the agar plate, the AITC treated filter
paper was positioned on an aluminum screen stand as is shown in Figure 5.
After 48 hr of incubation, the glass jars were opened and the agar plates
removed. Growth of the above microorganisms on the agar plate was then
determined. Lack of growth on each of the different densities of plates (101 - 10'5
CFU/ml) was considered to be due to the inhibition of the inoculum densities by

AITC.

40

2.4 Design of antimicrobial releasing system
2.4.1. CIO; release system

In order to produce a constant CIO; release, a specially designed
material providing CIO; release by mixing two dry solids, sodium chlorite
(NaClO;) and acid precursor in zeolite carrier, were received from ICA Trinova.
They were originally designed to release 3 pg/hr.g at ambient temperature (23°C)
for 31 days. CIO; is produced by a disproportion reaction as the two dry solids
are mixed:

4H+ 4' 5N80102 -> 4CI02 '1' NaCl + 4N8+ +2H20

2.4.2. AITC release system
2.4.2.1 AITC vapor pressure control

Since pure AITC solution is very volatile, it can quickly vaporize into the
atmosphere. In order to control AITC vapor pressure, Sekiyama et al. (1994) and
Lim and Tung (1997) used a triglyceride (ODO, oleic—capric-oleic) such as
vegetable oil to lower the vapor pressure of pure AITC. In this experiment, the
equilibrium AITC vapor pressures for various mole fractions in triglyceride
(equilibrium AITC vapor pressure in the liquid-liquid mixture) were investigated at
different storage temperatures.

5 ml of AITC liquid was introduced into a 10 ml vial with an orificed metal
cap. A rubber septum was inserted into the metal cap, and then the cap was
tightly closed using a metal clamp. The AITC containing vials were stored at

three temperatures (4, 22, and 37°C) for more than 30 min to produce a

41

saturated vapor in the vial. The equilibrium point was determined when the
headspace concentration did not increase during three serial injections. Then,
the saturated headspace gas was withdrawn and analyzed using gas
chromatography (GC). The AITC vapors were assumed to behave like an ideal
gas (pV = nRT). Partial vapor pressures were determined from the measured
concentrations, using the ideal gas equation. The experimentally determined
AITC vapor pressures for the triglyceride mixtures were than compared to the
results determined according to Raoult’s law

P1=P1° x x,

where

P1 = vapor pressure of the solvent with added solute

X1 = mole fraction of solvent

P1° = vapor pressure of the pure solvent
The relationship between vapor pressure and AITC mole fraction at different

temperatures was used to control the vapor pressure in the AITC canister.

2.4.2.2. AITC permeability in PE film
Figure 9 shows the schematic of the AITC canister used in this study. To
design a constant releasing canister system, the relationship between partial
vapor pressure and permeability was used to control the ATIC release rate.
The AITC vapor transmission rate through PE film was determined in
order to predict the organic vapor released from the canister. The Hatzidimitriu et

al. (1987) permeation cell method was used to determine the permeability

42

through the PE film. The cell consists of two stainless steel chambers tightened
around PE film. AITC liquid was placed in the lower part of the chamber, and the
permeated AITC vapor in the top cell was detected. The top part of the cell (total
volume 50 cc) contained an injection port with a rubber septum from which the
sample gas was extracted. An o-ring was used and vacuum sealing grease was
applied to assure a good seal between the film and surroundings. The
concentrations of the permeating vapors were monitored using gas
chromatography by removing 1 ml of headspace gas with a gas tight syringe. A
plot of the concentration versus sampling time was used to determine when
steady state was reached. Figure 10 shows a schematic of the stainless steel
permeability cell used in the test. Two cells were tightened together with a bolt
and nut, and a sample was withdrawn every 15 min until the perrneant flux

reached steady state.

43

 

....................... PP cap with 1.5/16 in hole

 

 

 

.m.—m.n.w.mmu~—m—.»~

PE film (1 mil)

  
  

------------------- glass vial (10ml)

“““““““““““““““““ AITC + triglyceride mixture

Figure 9. Schematic of the AITC release canister

Injection port

nnnnn

 

Figure 10. Quasi-isotatic cell used for measuring AITC permeability of the film
sample

2.5. Determination of the CIO; and AITC release rates needed to inhibit microbial
growth on fresh chicken

Both AITC and CIO; release systems were applied to fresh chicken
breasts and the microbial inhibition effectiveness of the two antimicrobial agents
investigated over a period of 8 days. Fresh boneless skinless chicken breasts
were obtained from a local grocery store (Goodridge ShopRite, East Lansing,
MI), transported to the microbiology laboratory (Department of Food Science and
Human Nutrition, Michigan State University, East Lansing, MI), and stored for no
longer than 1 day at 4°C before use. Each chicken breast was evenly cut with a
sterilized knife to get a standardized surface area. The average surface area and
weight were 286 cm2 and 200 g, respectively. Figure 11 shows how the chicken

samples were evenly cut before packaging.

 

Figure 11. Photo of chicken breast cut to provide a standardized surface area

The chicken samples were packaged in multilayer barrier trays, and heat

sealed with lid stock (CS 907, Cryovac lid 1050), which is typical packaging used

45

for fresh, modified atmosphere meat packaging. The total volume was 1010 ml
(7.5 in x 5.5 in x 1.5 in). A Multivac T-200 Traysealer (Multivac Inc, Kansas, MO)
was used to seal the MAP product in the package. After the chicken was placed
into the tray, it was inoculated evenly on the top surface of the chicken, and each
antimicrobial release agent (CIO; sachet or AITC canister) was inserted into the
package before the lid was sealed by the tray sealer. To prevent direct contact
between the chicken and antimicrobial, the antimicrobials were attached to the
walls of the package using pressure sensitive tape. Figure 12 shows a photo of

the tray after the antimicrobial sachets were applied.

 

Figure 12. Inoculated chicken sample with CIO; and AITC sachets

46

 

Figure 13. T-200 packaging machine used to pack the chicken breasts into the
trays

Each culture cocktail of Listeria monocytogenes and Salmonella.
Typhimurium (see 2.1) was inoculated onto separate chicken pieces to avoid any
interaction between them. 3, 4, 6, and 8 pg/hr CIO; releasing sachets were
prepared for the CIO; treatments, and 5 ml AITC-triglyceride mixtures able to
release 0.3, 0.6, 1,2, and 1.4 pg/hr of AITC continuously from the canister were
prepared for the AITC treatments. Each antimicrobial system was attached to the
inner wall of the tray, and sealed under a gas flush. A 70% N;/30% CO; gas
mixture was used for antimicrobial plus modified atmosphere packaging. An
ambient air flushed tray was also used to imitate conventional overwrapped
chicken packaging. All filled trays were stored at 7°C for 12 days.

Every two days, 10 packages (4 for CI02, 4 for AITC, 1 for MAP, and 1 for
ambient air) containing chicken were opened and the contents placed into a
stomacher filter bag. 200 ml of 0.1% peptone water was added to the stomacher
filter bag and homogenized for 2 minute. 1 ml of the homogenized sample was
diluted serially with 9 ml of 0.1% peptone water. The samples were plated on the

47

different selective growth media to quantify the number of Listeria
monocytogenes and Salmonella Typhimurium.

Modified Oxford agar (MOX) and Xylose Lysine Deoxycholate (XLD) agar
were used for each organism respectively. In order to recover the injured
pathogen organisms, the MOX and XLD were overlayed with Tryptic Soy agar
with 0.6% yeast extract (TSAYE) as shown in Figure 14, and the sample was
inoculated directly on the nonselective thin agar layer. Colony counts were
expressed as I091o colony forming units (cfu/g). Products from each packaging

treatment were sampled three times (n=3).

Nonselective agar (7 ml)

l

 

 

 

 

Petri dish

 

 

Selective differential media

Figure 14. Schematic illustration of the thin agar overlay method

48

2.6. Evaluation of the synergic effect of antimicrobials (CIO; and AITC) and MAP
combination.

The chicken samples were prepared and packaged as mentioned
previously (section 2.5). In order to confirm the MAP and antimicrobial synergic
effectiveness, half of the packages were packaged with air, and the other half
were packaged using a 70% N;/30% CO; gas flush. All packaged trays were
stored in an environmental chamber for 8 days with storage temperature of 4°C.
Then, the growth of Salmonella Typhimurium, and Listeria monocytogenes, and

total aerobic bacteria were enumerated on growth media (section 2.5).

2.7. Fresh chicken packaged with antimicrobial and stored for 21 day
2.7.1 Sample and packaging

To determine the effective minimum CIO; and AITC release necessary to
attain an antimicrobial impact, packages were prepared and packed with fresh
chicken breast and antimicrobials. Chicken was obtained from a local store (fresh,
boneless, skinless chicken breasts), cut, and packaged with CIO; or AITC after
gas flushing (30% CO;I70% N;). The CIO; amounts used were 8 and 16 g
sachets, with release rates of 4 pg/hr and 8 pg/hr, respectively at 4°C. For AITC
treatment, 1.0 and 1.5 ml of AITC were mixed in a 5 ml AITC-triglyceride mixture.
All of the antimicrobial packages were packed with a 30% CO;I70% N; gas
mixture. Samples were also packed without antimicrobial treatment as control
samples, to compare the microbial inhibition effectiveness. Half of the controls

were packaged in ambient atmosphere, and the other half were packaged with a

49

30% CO;I70% N; gas mixture. All sample trays were stored in an environmental

chamber for 21 days at 4°C.

2.7.2. Microbial analysis
Every three days, three trays from each treatment were removed and the amount
of microbial growth (0, 3, 6, 9, 12, 15, 18, and 21 day) determined. The microbial

analysis was discussed in a previous section (2.5).

2.7.3. Headspace gas analysis

In order to determine the internal atmosphere of the chicken packages,
gas headspace analysis was carried out throughout the storage period. Gas
composition of CO; and 0; within the package was monitored using a headspace
gas analyzer M-6600 (Illinois Instruments Inc, IL, US) at 0, 6, 12, 18, and 21 days.
Headspace samples from the tray packages were withdrawn at a rate of 40
ml/min using the internal pump of the Instrument, and passed by CO; and 0;

sensors.

2.7.4. pH measurement

During the storage period, pH was measured on the samples from storage
every 3 days. To determine pH, 10 g from each chicken piece was removed
before microbial analysis, diluted with 90 ml of distilled water (1 :10 dilution), and
homogenized in a blender for 2 min. Measurements were taken with a Corning

model 430 pH meter and electrode (Corning Inc, Canton, NY).

50

 

Figure 15. pH meter and assembly used to determine pH of chicken samples

2.7.5. Color measurement

From the consumer preference, chicken should not change color due to
antimicrobial treatment. Thus, the color of the breast meat treated with
antimicrobial was investigated. The surface colors from three random locations
on the chicken breast were monitored using a colorimeter (Hunter Color Machine
Hunter Associates Laboratory INC, Reston VA) during the storage period, and an
average value was then calculated from the three random locations for "L", "a",
and "b" values. Color measurements were made every three days. Three trays
from each treatment were removed and evaluated prior to the microbial test (0, 3,
6, 9, 12, 15, 18, and 21 days). Figure 16 shows a picture of the colorimeter used

for this test.

51

 

Figure 16. Colorimeter used to measure the color of antimicrobial treated chicken
breast
2.8. Physical property changes due to CIO; treatment

The same CIO; sachet from ICA (previously discussed in section 2.2) was
used for the CIO; treatment. A concentrated aqueous CIO; solution was diluted
with distilled water to make 100-2000 ppm of CIO; headspace concentration in 4
liter glass containers. The dilution amount of the concentrated aqueous CIO;
solution was varied depending on the vapor-liquid equilibrium relationship of
Henry’s law. Each headspace concentration in each prepared CIO; solution was
confirmed by titration (Appendix I). LDPE, PS, and PP films were pre—conditioned
at 23: 2 °C/50%:r 5% RH, and inserted into the container containing prepared
CIO; solution. Films were hung by an aluminum wire hook in the middle of a
metal lid, to prevent direct contact with the aqueous CIO; mixture. The lid
contained a 1/8 in hole, which was filled with silicone as a septum. For the
physical properties tests, the films were removed after 1, 12, 24, 48, and 72
hours, and mechanical, barrier, and optical property tests were performed on the
test films. An aluminum foil coated metal plate was placed immediately over the

mouth of the jar after the lid was opened, so that the loss of CIO; vapor was

52

minimized while sample films were being removed periodically. Figure 17 shows

a schematic of the CIO; treatment system used with these films.

silicon septum

Al wire hook

films

 

aqueous CIO; mixture

 

Figure 17. Schematic of CIO; film treatment system

2.8.1. Mechanical properties

Tensile strength (TS), elongation at break (EB), and toughness were measured
using an lnstron 4201 (lnstron Corp., Canton, MA) Universal Testing Machine
(Figure 18) according to ASTM D 882. Films were cut into strips (1 in width)
using a Precision Sample Cutter (Thawing Albert Instrument Co., Philadelphia,
PA), and conditioned according to ASTM D 618 at 23¢ 2 °C/50%i 5% RH. The
thickness of each film (PP, PS, and LDPE) was determined at three locations
with a TMI 549M micrometer (Testing Machines, |nc., Amityville, New York)

according to ASTM D 374-99.

53

 

Figure 18. lnstron 4201 used for mechanical properties evaluation of packaging
films

2.8.2. Measurement of oxygen transmission rate (OTR)

The oxygen transmission rate (OTR) was determined using an Oxtran Model
8001 unit (MOCON/Modern Controls, Inc., Minneapolis, MN) (Figure 19),
according to ASTM D 3985, at 0% RH and 23 °C. The area of the test film was 5

cm2. The test was continued until steady state was reached.

 

Figure 19. Oxtran 8001 unit used for oxygen transmission testing of film samples

54

2.9. Statistical analysis

Statistical evaluation of the data was performed using SPSS (SPSS Inc,
2004). Microbial growth depends on antimicrobial treatment and storage time
was analyzed using the ANOVA procedure. Significance levels were reported at
95% confidence (p=0.05) using the Tukey’s honestly significant difference (HSD)

multiple comparison. The results of statistical analysis are shown as mean values

:I: standard deviation.

55

CHAPTER III. RESULTS AND DISCUSSION

3.1. Inhibition performance of AITC and CIO;
3.1.1. Inhibition effectiveness of AITC against Listeria monocytogenes and
Salmonella Typhimurium

The inhibitory affect of AITC vapor on Listeria monocytogenes and
Salmonella Typhimurium was established in a series of microbial tests with
different inoculum densities ranging from 10° to 10° CFU/dish. Salmonella
Typhimurium and Listeria monocytogenes were stored in the presence of various
amounts of impregnated AITC (on filter paper). Storage times were 48 hr and 7
days at 37°C and 7°C respectively.

The antimicrobial activity of AITC depends on dosage and temperature,
as shown in Figures 20-21. Salmonella Typhimurium which had been inoculated
on the agar plates was completely inhibited by AITC concentrations of 160 pg/l or

more. At 7°C, it was completely inhibited with 70 119/I. Listeria monocytogenes
was more resistant to AITC vapor. At an AITC concentration of 160 pg, there was
a 3 log reduction at 37°C. However, the same concentration (160pg/l AITC)
reduced Listeria monocytogenes 6 log at 7°C. Complete inhibition was obtained

by 360pg/l AITC at both temperatures. Overall, AITC was more lethal to both

pathogens at 7°C. Delaquis and Sholberg (1997), Ward (1998), and lsshiki et al.

(1992) used similar model systems to determine the antimicrobial activity of AITC
vapor against various pathogenic bacteria. Delaquis and Sholberg (1997)

reported that Listeria monocytgoenes was strongly inhibited by an AITC

56

concentration of 1500 pg/l while Salmonella Typhimuirum and E.coli were barely
effected. Thus, the results of Delaquis and Sholberg are in conflict with these test
results. However, Ward (1998) showed that Salmonella Typhimuirum was more
susceptible to AITC vapor than Listeria monocytogenes. Salmonella
Typhimuirum was inhibited with concentrations of 4000 to 20000 jig/l. In Ward’s
experiment, the AITC vapor was distributed among 21 agar plates in a glass jar.
There was only one agar plate per glass jar, in this test. The range of minimum
inhibitory vapor concentrations in this research corresponded to the results from
lsshiki et al. (1992). They reported that 37 -110 ug/l inhibited bacteria growth on
agar. Figure 22 shows the gas headspace concentration profile during storage.
Decomposition of AITC in aqueous media is known to be suppressed in the
presence of dextrins and/or polysaccharides in the growth medium (Ohta et al.,
1995). The results indicate that AITC decomposed substantially during 7 days of

storage at 37°C. After 1 day, residual AITC concentration had decreased more
than 50% while AITC concentration at 7°C was down by 40%. On day 2, the

original 360 pg/l AITC was down by 90%. The difference in the reduction

between the two temperatures can therefore be ascribed to the difference in

decomposition rate. At 7°C, AITC concentration decomposition was substantial

but approximately 20% of the initial input remained until 6 days when the initial

level was 360 ug/l.

57

 

— S.Typhimurium
522:3 L.monocytogenes

 

 

 

Microbial reduction (log1o)

 

 

 

 

 

0 100 200 300 400
AH C concentrations (pg/l)

Figure 20. Reduction of Salmonella Typhimurium and Listeria monocytogenes on
agar due to AITC treatment (37°C for 2 days)

 

— S.Typhimurium
L.monocytogenes

 

 

NOD-bOTOJV

 

Microbial reduction (log1o)

 

 

 

 

 

 

 

0 100 200 300
AIT C concentrations (pg/l)

Figure 21. Reduction of Salmonella Typhimurium and Listeria monocytogenes on
agar due to AITC treatment (7°C for 7 days)

58

 

400 .-
e + 7°C
—t.B—- 37 °C

 

 

300

 

 

 

 

Headspace concentration (ug/I)

 

 

 

Storage time (day)

Figure 22. AITC headspace concentrations in glass jars (950 ml) with growth
media during storage.

3.1.2. Inhibition effectiveness of CIO; against Listeria monocytogenes and
Salmonella Typhimurium

For CIO; treatment, an experiment was performed with a known amount
of CIO; gas to yield desirable headspace concentrations. The inhibition
performance of CIO; on Listeria monocytogenes and Salmonella Typhimurium is
shown in Figures 23 and 24, respectively. CIO; treatment was shown to result in
a significant reduction of both pathogens. Listeria monocytogenes and

Salmonella Typhimurium were completely inhibited by 120—180 jig/l. At 120 ug/l

or more CIO; treatment, Listeria monocytogenes was inhibited completely at both

storage temperatures. At 37°C, 60 pg/I of CIO; reduced the Listeria

59

monocytogenes population by 4.0 log CPU. The strong lethality of CIO; against
Listeria monocytogenes has been also reported by other researches. Lee (2004)
used a gas sachet which generated 4.3-8.7 mg CIO; after 30 min, and reduced
the number of Listeria monocytogenes on lettuce leaves by 5.0 log, and Kaye et
al. (2005) reported a 5.8 log CFU/g maximum reduction of Listeria
monocytogenes on fresh vegetables with 4.1 mg/l CIO; gas. Salmonella
Typhimurium was more resistant to CIO; treatment than Listeria monocytogenes.
In order to eliminate Salmonella Typhimurium, 180 pg/l or more of CIO; was used,

and 2 log reductions were obtained using a 90 pg" treatment regardless of the
storage temperature.

At 37°C , the pathogens were more inhibited more than at 7°C. The
reductions in numbers of Listeria monocytogenes at 60 and 90 ug/I were 2 and 1
log higher at 37°C. Salmonella Typhimurium also suffered 1 log more destruction
at 120 119/I than at the lower temperature. The main reason is probably loss of

CIO; from the headspace. As is shown in Figure 23, the initial inputs were quickly
reduced at both temperatures. Regardless of storage temperature, all of the CIO;
in the headspace was reduced to zero within 1 day. The water solubility of CIO;
is 2.63 g/L at 40°C, and it is approximately 10 times more soluble in water than
chlorine (Lenntech, 2006). Both XLD and MOX agar have low density and are
water-based mediums. It is assumed that the heavy CIO; gas was absorbed into
the growth media, and then dissociated to Cl’, CIO', ClOg' as is shown in equation
3 (EPA, 1999). These CIO; byproducts are known to be weak antimicrobials
(Lenntech, 2006).

60

Equation 3: Decomposition reactions: CIO; + e' = CIO;'
ClO;‘+2H;O+4e‘=Cl‘+4OH'
Cl03'+H;O+2e'=ClO;'+20H'
ClO;‘+H;O+2e'=CIO;'+20H‘
Cl03'+2H+e'=CIO;+H;

At 7°C, the growth mediums (XLD and MOX) samples were held 5 days
longer than at 37°C. The pathogens would have had more time to recover from

CIO; treatment, particularly a psychrophilic bacteria such as Listeria
monocytogenes. A constant release of CIO; during storage may be more
effective than a onetime treatment. Ellis et al. (2005) reported that the low
release CIO; (2.5 mg for 22 day) from sachets was more effective in reducing

Salmonella Typhimurium on chicken than fast releasing sachets (6.6 mg for 12hr).

61

 

 

 

 

 

 

 

 

 

 

 

 

— S.Typhimurium
m L.monocytogenes
7
’5 cl
5
e 5,
S
a 4‘
3
E 3-
E
6 2'
.2
2 1 -
0
o 100 150 200
002 concentrations (pg/l)

Figure 23. Reduction of Salmonella Typhimurium and Listeria monocytogenes on
agar due to CIO; treatments (37°C for 2 days)

 

 

 

 

 

 

 

— S.Typhimurium
E2221 L. monocytogenes
7
’6 6 .
5
s s-
S
a 4‘
:3
E 3-
E
3 2‘
.9
2 1 -
0 . . _ ,
0 50 100 150 200
CI02 concentrations (pg/l)

Figure 24. Reduction of Salmonella Typhimurium and Listeria monocytogenes on
agar due to CIO; treatments (7°C for 7 days)

62

 

 

 

 

 

 

 

 

 

 

 

 

 

 

A 14 + 7°C

2 +37°c

e 12

8

“r:- 10E

8

3 8

C

. ll

9 6

8 E

(U

3 4:.

a \

f 2 3%“
0 t. w a r {B {3
0 2 4 6

Storage time (day)

Figure 25. CIO; headspace concentrations in glass jar (950 ml) containing growth
medium.
3.2. Development of AITC release system
3.2.1. AITC vapor pressure control

The objective of the test was to develop a model to predict the vapor-
liquid equilibrium (VLE) of a binary (AITC-triglyceride) mixture as a function of
temperature. In an ideal situation, the partial vapor pressure of a component in a
mixture is equal to the vapor pressure of the pure component multiplied by its
mole fraction in the mixture. Raoult’s law was used to develop the VLE model,
and an experimental study was conducted to verify the VLE (mixture of AITC and

triglyceride) model.

63

Firstly, P1° (vapor pressure of the pure solvent) as a function of
temperature was determined using the Clausius—Clapeyron equation. The
unknown temperature dependent vapor pressures were predicted using the

following equation.

wt%) = 5H7?

1 I
<—-——)
T2 T1
where,

P=pressure, AH =enthalpy of vaporization, T= temperature in Kelvin (K)

vap
R =ideal gas constant (8.3125 J mol'1 K")
P was determined using gas chromatography (GC) experiments. The

heat of vaporization (AHvap) was determined from the slope of the natural log of

the vapor pressure (In P) versus the inverse of temperature in Kelvin (1/T), The

slope of the plots was 47.53 KJ/mol with an R2 of 0.998. The AHW,p of AITC was

very close to the values determined by other researchers. Chickos and Acree
(2003) found it to be 47.6 KJ/mol and Lim and Tung (1997) obtained the same
values using a GC experiment. P1 and P; are the pressures at temperatures T1
and T; Figure 26 shows the predicted vapor pressure as a function of
temperature, and demonstrates that AITC vapor pressures are highly dependent

on temperature. The experimental data correspond well to the predicted values.

64

 

3000

 

 

 

0 Experimental values
2500

2000 /
1 500 /

1000 Z

500

 

 

 

 

vapor pressure, Pa

 

 

 

 

 

 

0 I I r I I
0 1 0 20 30 40 50 60

Temperature (°C)

Figure 26. Vapor pressure of AITC as a function of temperature

The AITC partial vapor pressure-temperature-mole fraction relationship

was obtained based on Raoult’s law. The vapor pressures of pure AITC (p2) at

specific temperatures and experimental validation are shown in Figure 27. The
vapor depression effects of the triglyceride diminished as temperature increased.
In order to achieve an equal vapor pressure at a lower temperature, a much

higher AITC mole fraction would be needed.

65

 

500

 

 

 

 

 

Prediction from Raoult's Law
------ Prediction from Margules equation
0 experiment at 4°C
0 experiment at 7°C
v experiment at 23°C
400 -
m 300 ..
o. ..
92' - v
3
in
m
2
9.
§
> 200 ~ ..
0'.
.0. v.
’ O
100 - ,.--°
O ..O ......O
O...
“,0 ......
’0'. v ....::3 000000 .
,.o ...:::6...0
..:001!"33O"
0 i .. l l I l
0.0 0.2 0.4 0.6 0.8
AITC mole fraction

Figure 27. Vapor pressures of AITC in a AITC-Triglyceride mixture at
temperatures 4, 7, and 23 °C
Results at 37 °C were out of scale so are not shown.

66

 

In order to verify the pressure-mole fraction model, the vapor pressure
was determined by GC analysis (described previously). The results showed that
the AITC-triglyceride mixtures deviated negatively from Raoult’s law. Specifically,
the deviation between model and experimental results was higher at lower
temperature. Lim and Tung (1997) reported similar results with an AITC-canola
oil mixture at 45, 35, 25, and 15°C. The author reported that there were larger
negative deviations of vapor pressure from Raoult’s Law at lower temperature.

Raoult’s law is strictly valid only under the assumption that the chemical
interactions between the two liquids are equal to the bonding within the liquids
such as with an ideal solution. Therefore, by comparing the actual measured
vapor pressures, it showed that the relative bonding strength between AITC and
triglyceride was greater than the bond strength within the individual liquids (AITC-
AITC). The deviations between Raoult’s ideal relationship and the real
experimental results were larger at lower temperature.

The non-ideal vapor-liquid equilibrium (VLE) relationship (p vs. x.) was

modeled using Margules equation (Metiu 2004). This is a widely used equation to

predict mixed component’s vapor pressures at a given temperature.
17 — p1 'x1( )n +1); -xz( >42

where, p = vapor pressure, x1(l),x2(l)=mole fraction of component x1

and x; in the solution, p10,p(2) =vapor pressure of pure liquid (x1 and x;),

r] (x1 (1)),r2(x2 (1)) =activity coefficients of component of x1 and x; in the solution,

"I =eXp[(A12 +2(A21-Alz)xl)-x22]. r2 =eXp[(A21+2(A12 -A21)X2)°XI2]

67

A12 and A21 are called Margules parameters. Reference for these parameters
was not available for AITC. Therefore, the activity coefficient (a ) was obtained
through experimental work (Figure 27). Since the vapor pressure of oil is
negligibly low, the vapor phase of the binary mixture can be taken to consist only

of pure AITC vapor, and x2 can be substituted for (1— x1). Therefore, the

equation yielded the following:
_ 0 _ O 2
p1 — p1-xl(l)-rl-p1~xt(l)-eXPla(1-XI) ]

The non-linear regression procedure of Sigmaplot (Systat software, 2004) was

used to determine a from the experimental data, and the a values varied
depending on temperature. pf was calculated from the previous vapor pressure-

temperature graph shown in Figure 26. The model had a much more satisfactory
fit than did Raoult’s Law in Figure 27. As is shown in Table 2, vapor pressure
predictions using Margules equation were shown to have R2 values greater than
0.98 at all temperatures.

As the temperature of the system increased, a values also increased.
The relationship between a and temperature was linear, as shown in Figure 28.
Therefore, by using the predicted a, a non-ideal relationship (AITC vapor
pressure vs. AITC mole fraction) may be able to be modeled at temperatures

between 4 and 37°C.

68

Table 2. Estimated values of a estimated using a non-linear regression
procedure and the coefficient of determination (R2)

 

 

Temperature (°C) a R2 value
4 -1.60 0.98
7 -1.57 0.99
22 -0.95 0.99
37 —0.57 0.99

 

a = — AH mix /RT (the activity coefficient from experimental data in Figure 27)
where, AH mix =heat of mixture, R =8.3143 m3-Pa°K'l -mol", T= Kelvin temperature

-02

 

-OA»a

-0.6 d

418 ~

-1l14

    

at value

-1.2 «

y=0.0328-1.74

R2=0.99

-1.4 ~
-1.6 (

6L8 -

 

 

'2.0 r I I
0 10 20 30 40

 

Temperature (°C)

Figure 28. Estimated a values as a function of temperature

69

3.2.2. AITC permeability of PE film and controlled release rate

The AITC permeability coefficient for PE film (1 mil) was determined, and
Figure 29 shows the permeation curve at ambient temperature (23 °C). The AITC
transmission rate through PE film was obtained from the slope of the straight line

once steady state was reached. The AITC permeability coefficient (P)

((ug ~I)/(in2 -hr - pa)) was calculated as follows:

_ Q°l
—A-t-Ap

 

where,

P = permeability, Q =the rate of permeation during the steady state

I = film thickness, A = area of the film exposed to permeant
t =time, Ap = partial vapor pressure

The equilibrated partial vapor pressure of AITC in the lower cell chamber
was 83 pa, and the permeability of AITC through PE film was determined to be
60.76 pg.mil/in2.day.pa. Despite the finite amount of AITC in the lower cell, the
change in the AITC mole fraction due to losses from permeation was very low
(less than 0.1%). Therefore, the AITC pressure drop within the lower permeation

cell was ignored.

70

 

 

 

 

 

a

a

E3 Y=0.05621-1.1479 .
g 8; R=0.9795

o.

3

.E

t; 6—

<

B

a 4*

a)

E

8 2;

“5

a

8 0e . . . . l . .
E o 20 4o 60 80 100120140160180

Time (min)

Figure 29. Quasi-isotatic permeation curves for AITC and PE film at 23 °C as a
function of time

The AITC release rate was designed using the vapor pressure (Pa) vs.
mole fraction (X3) relationship (Figure 28). Various AITC mole fractions in
triglyceride were prepared, and the AITC headspace concentrations in the glass
jar verified using GC analysis. Table 3 shows the controlled AITC release rate
from the canister system. The size of the orifice on the PE cap of the vial was
0.09 in. At a constant temperature, vapor pressure is the only factor needed to
control the AITC release rate. In the test, mole fractions (X3) varied from 0.3-0.8
to yield release rates from 0.3 to 1.8 pg AITC per hr. Figure 30 shows the AITC

concentration in the headspace of the empty container as a function of time.

71

Table 3. AITC vapor release system as related to AITC mole fractions in the

 

 

 

 

 

 

 

 

 

 

 

 

 

 

triglyceride mixture at 4°C
release rate release rate
AITC (ml) Xa Pa 1E9 lday) (pg/hr)
0.5 0.3 20 8 0.3
0.8 0.4 35 15 0.6
1.5 0.6 70 29 1.2
2 0.7 80 34 1.4
3 0.8 105 44 1.8
Total volume (AITC (ml) + Triglyceride (ml))=10 ml
1000
’5 O 0.3 pg/hr
E O 0.6 pg/hr /.
: 1‘in /
E - llQ i / /I / 1}
g 600 /' 7’
O I/ / / / 4
g / / A/ /
/ /‘
o . /
< ./ / _/
8 / /A /V
in I/ -/ /'/ / ’T
2 L x/ / 1 /
_ /% / / / O ’ ...—- ____ ____’
a 4% //AF ,,,aee '
I- d / : ,__, .._. a” “"
0 1Fé.’ I I I l
0 5 10 15 20
Time (day)

Figure 30. Theoretical vs. experimental AITC headspace concentrations of the

different release systems

The headspace AITC concentrations in the container consistently

increased as a function of time. The results indicate that the sachets continued to

emit AITC gas for more than 20 days. However, the concentration data from the

72

later tests (after 12~15 days) had lower concentrations than expected. Since the
empty container was flushed with 100% nitrogen gas, there was no
decomposition from oxidation or product interaction with AITC. It is suspected
that AITC suffered some amount of decomposition to the thiocyanate ion (SCN-).
Radomir (1997) found that 5% of AITC was converted to SCN- after 10 days of
storage. Another possible cause could have been sample loss during needle
penetration of the container (canning jar) during headspace sampling of AITC by
gas chromatography. The possibility of absorption into the package was
considered minor because the glass jar and metal lid provide almost perfect

barrier properties.

3.3. Development of CIO; release system
CIO; gas was generated from a powder mixture supplied by ICA. The two
different active compounds generated CIO; gas almost instantly with mixing, and

its release rate was designed to be 3 pg/hrg at room temperature (23°C) and 0.4
pg/hr.g at 4°C.

Therefore, the release rate was controlled by implementing the following simple

equation at constant temperature;

d—C- = K g x M
dt
Where,

C=gas phase concentration, t=time, Kg=gas generation rate

M=weight of sachet

73

Using this relationship, the CIO; release rate in the closed container was
controlled by the mass of powder placed into the sachet. The released CIO; gas
quickly decomposed during storage. Table 4 shows the concentration difference

between total released CIO; and remaining CIO; in the 1 qt glass jar.

Table 4. Total released CIO; amounts vs. experimental headspace concentration
as a function of time at 4°C

 

 

Time(hour) 4g 4g 8g 89
(calculation) (experiment) (calculation) (experiment)
5 3.6 0.2 7.2 1.0
10 7.2 0.8 14.5 2.5
24 (1 day) 17.4 0.8 34.8 2.5
48 (2 day) 34.8 1.0 69.6 3.0
72 (3 day) 52.2 0.8 104.3 2.5
96 (4 day) 69.6 0.8 139.1 2.5

 

1. The unit of all data are parts per million (ppm)
2. Calculated concentration was determined by CIO; conversion factor
(1 ppm= 2.76 mg/m3)

The CIO; concentration in both 4 and 8 9 treatments reached steady state in 10 -
24 hours. After the CIO; concentration reached steady state, it remained constant
during storage, though the concentration differed from the total amount released.
This shows that CIO; continuously decomposed during storage. There are many
possible reasons for this decomposition. The study was performed in an empty
container after nitrogen flushing. Thus, little or no oxygen remained. However,
some oxidation may still have occurred. In the gas state, as discussed in
equation 3 on page 56, CIO; is unstable, and free radicals are evenly formed and

dissociated. Thus, CIO; is converted to chlorite (CIO;), chlorate (CIO3') and

chloride (Cl‘) by oxidation. Another possible reason for the lower detected

74

amounts may be due to the instrument. When the detector tube absorbed
headspace gas, it needed to be vented using another Tygon tube opening to
avoid creating a partial vacuum. Thus, the sample gas might be mixed with air

and thus diluted during the extraction.

3.4. Determination of minimum release rate to inhibit target bacteria

The purpose of this study was to determine the AITC and CIO; release
rates which would inhibit Salmonella Typhimurium, Listeria monocytogenes, and
total aerobic bacteria growth on fresh chicken breast. The previous test results
showed that both AITC and CIO; either decomposed in the headspace or were
absorbed into the agar plates. Also, fast antimicrobial vapor release rates have
been known to cause adverse sensory problems. Muthukumarasamy et al.
(2003) mixed 0.7 ml of AITC with vegetable oil, and packaged it with ground beef.

The package system generated 1400 ug/l of AITC in the headspace immediately,

and reduced by 6 log the inoculated level of Listeria monocytogenes. The
packaged beef was observed to have a pungent flavor. In order to maintain the
antimicrobial inhibition, a constant slow release treatment was considered to be
an effective way to accomplish this with minimum sensory property changes.
Thus, several different release rates were selected and tested with chicken in this
study. The minimum effective release rates for both AITC and CIO; were selected
to be the lowest release rates at which microbial growth was significantly lower
compared to control samples. All antimicrobial treatments were combined with

modified atmosphere packaging (MAP) (30%CO;/70% N;). For control samples,

75

ambient air flushed packages and MAP were used. The tests were conducted at
a temperature of 7°C, a slightly abusive storage temperature for poultry products.
The pathogenic organisms grow faster at this temperature than at a more normal

storage temperature (3-4°C) for fresh chicken meat.

3.4.1. Target bacteria growth with AITC treatment

Four different constant AITC release systems (0.3, 0.6, 1.2 and 1.4 pglhr)
were developed and evaluated to determine their inhibition effectiveness. Growth
of Salmonella Typhimurium was lower in all AITC systems than for samples
which were packaged in modified atmosphere or ambient air packaging except
for the 0.3 pglhr release system (Figure 31). On day 12, the level of Salmonella
Typhimurium in the 0.6 pglhr release system was significantly lower than that
without AITC treatment (p<0.05) (Table 5). For the 1.2 pglhr and 1.4 pglhr AITC
release system, total counts of Salmonella Typhimurium were within 1.0 log on
day 12. No statistically significant difference between the 1.2 and 1.4 pglhr
release rate systems was observed.

The inhibition performance of AITC was also shown on chicken breast
meat inoculated with Listeria monocytogenes (Figure 32). The results show that
only the 1.2 and 1.4 pglhr release rate systems had a significant inhibition effect.
After 12 days, Listeria monocytogenes counts in the 1.2 and 1.4 pglhr AITC
release rate system were approximately 0.3 and 0.5 log lower than in the MA

package.

76

The antimicrobial effectiveness of AITC on total aerobic bacteria was
clear. Any treatment containing more than 0.6 pglhr AITC significantly inhibited
total aerobic bacteria compared to MA packages. On day 12, the reduction in
total bacteria with 0.6, 1.2, and 1.4 pglhrAITC treatments was 1.03, 1.41, and
1.73 logs compared to the MA package, respectively. The total bacteria counts of
the 0.3 pglhr AITC release samples were statistically lower than the MA
packaging on day 6. However, the effectiveness did not last until day 8.

Overall, 0.6 pglhr of AITC was the lowest dosage rate shown to
significantly inhibit Salmonella Typhimurium inoculated on the fresh chicken
breast. However, the minimum release rate which was effective against Listeria
monocytogenes was considerably higher. Listeria monocytogenes was inhibited

with 1.2 pglhr or higher AITC release.

77

ooh am 3559.? new “woos c9630 :0 36.385 E:=:E__._a>._. 36:92me :o 29 33.9 O._._< no fiotm ._.m 659“.

. :63 6E: 39on
N_. or m o v N o

 

 

l

 

 

5/l'I:lO 601

CD

o<2t£msi lul
c<2t£e33 l-I
$22385 lal
d<§i£m=md lbl
as). l0l

.2 lol

 

 

 

 

78

0..» “a c3909? new ammo... coono co 35.3005 mocomoiooeoE chose: :0 can. 636.2 ot< so sootw .mm 059”.

Q63 68: 69905
N? o_. m w v N o

p p — b n I— m

 

 

 

 

l

 

firnao 501

CO

93:59: lnT
o<2t£esme l-I
massaged lal
Q<E+L£mamd Ibl
as). l0l

.2. IT

in

 

 

 

 

79

Ooh um “32800.59 new 685 c9620 :0 3.083 058%.. .39 :0 99 38.2 3.2. so Beam .3 9:9...

 

 

 

 

 

Q63 66: 3905
e e v N o

. m
.. e
l s
a

$21,233.. lnT

. o<2t£e33 Ill
c<§t£e=ee ldl . a

o<2t£eamd I

as). IOI

._< Isl

 

 

 

 

or

firms 501

8O

Amodvav 299:5 $52.:ch 9m 9.828 8.3 E Bozo. EoLoEu 53> memos.
.cosmscp Emccmumncmes. .930 mo_n=c3

833$ 38.963 .5333 as .953 22551:

Begum: ateaove Devotee E 953 82.523

>863? >>>o a .983 Swansea :5 Bacon ot< .5966 Scams
engage Smashes are ammo 32.033 82 595.0 28.8 .98
33.033 caret: Sieve ”Fences c<s_ .

 

 

 

 

 

 

 

    
  

ocmogoocoE
steam:

 

E35829»...
m=ocoE_mw

 

Ooh “a Cmv—OEO cwmh. c0 mtg—own
O_QO._0m _mu0u UCN .WOCODOVAOOEOE mt6~w3 .E:t:EEQ>.—. G§OQOFSQW CO Ewgmmb O._._< m0 mmOCOZU—OOtw COEDEC. .m GEN-F

81

3.4.2. Target bacteria growth with CIO; treatment

The same microbial tests were performed on fresh chicken breast treated
with CIO;. The results obtained with Salmonella Typhimurium and Listeria
monocytogenes inoculated chicken are shown in Figures 34 to 36. The tested
organisms were significantly inhibited by CIO; treatment. However, a higher level
of dosage (4 pg to 8 pglhr) was required to get the same inhibition effectiveness
as with AITC.

The growth of Salmonella Typhimurium in the presence of 3, 4, and 6
pglhr release sachets did not shown any significant difference (p>0.05) from that
of the MA package (Table 6). 8 pglhr release was shown to have a significant
effect. The growth of Salmonella Typhimurium was 0.6 log lower during 12 days
storage. The inhibition effectiveness of CIO; was greater against Listeria
monocytogenes. At 4 pglhr or higher CIO;, the growth of Listeria
monocytogenes was significantly lower than in MA packaging. On day 21, the
reductions due to CIO;, 4.0, 6.0, and 8.0 pglhr were 0.5, 1.0, and 1.0 logs
compared to the MA package (Table 6). The effectiveness of 6 and 8 pglhr CIO;
treatments was statistically similar. The population counts in both release rate
systems became significantly lower than the other packages at 8 days.

The inhibition effectiveness of total aerobic bacteria in response to CIO;
treatment is shown in Figure 36. Even though a higher dosage was used, its
effectiveness was lower than that of the corresponding AITC treatment. Total
bacteria on chicken samples with 4 pglhr or higher release had significantly lower

(p<0.05) counts than those in MA packaging. At the end of storage, packages

82

with 4 pg/hr sachets had 0.7 log lower counts than that of the MA package. Total'
aerobic bacteria treated with 3 pglhr CIO; did not suffer any statistical increase in
inhibition during the storage period. Doubling the CIO; release rate did not double
the log reduction. Moreover, the effectiveness of 8 pglhr CIO; was statistically the
same as 6 pglhr CIO;. There may be a rate limiting effect other than
concentration. A similar trend was observed in microbial counts for AITC
treatment. Chicken subjected to more than 1.2 pglhr of AITC (1.2 -1.4 pglhr) had
statistically similar results. This may be due to mass transfer of the antimicrobials,
or higher microbial growth might have occurred on the bottom side of the chicken
in the glass jar. The contact area between the chicken and glass jar may have
been less accessible to the antimicrobials (either AITC and CIO;). A negative
side effect was observed at the highest (8.0 pglhr) CIO; release rate. The surface
color nearest to the sachet became brown-black. Details concerning color

change are discussed in the next section (3.6.6).

83

ooh E 639359 new 385 c.8620 9:0 33.305 EESEEEC. £265me :0 9.8 33.9 «0.0 so 80mm .3 9:9“.

:63 6E: 3965
3 up 9 m w v N o
P P F _ p p m.”

 

t of

 

 

 

 

1
II
“3.
v

firms 601

I
0.
IO

 

a<§t£maw IDI
d<§i£mam III
osztémsv ldl - we
d<§t£mzm Jrl
n25. IQI

.2 lol

 

 

 

 

0.0

84

UK. we 3.93:3. can “moms c9626 :0 85:85 32683822: @9qu :0 99 636.2 a0.0 ho fiotm .8 9:9“.

:83 65: cashew
3 NP 9 m o v N o
b b _ b — . m

 

IIUI

 

 

J.
Brndo 601

l
(O

 

m<§t£mzw IOI
d<2t£mao +
n_<s_t£m:v ldl .. A
n_<_2.:£m:m II
as). IOI

L_< IOI

 

 

 

 

85

Ooh F. 365999 pcm “amok. coono co p.583 0593 .99 .8 99 686.9 No.0 Co “norm. .8 p.59“.

CHE 6E: 399m
9 m o v N

- — P

 

 

 

n.<s.+L£m:m lnT
n.<.2+L£m:o +
o<2t£esv Idl
n_<.2+c£m:m Jrl
n.<.2 IOI

L_< IOI

 

 

 

 

0..

Brnzlo 501

86

ficdvav ESQEU >=cmoEc9m 05 ch3.8 9:8 c. 995. #:9ch 5.3 9.85.
60.33% Emucflwucmcé .930 moEED

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

so 333 negate 3333 at c cm a No 0 3.9.3
on ..oame.N SQONSN 32.95: at c cm H. No 0 3.9.3
2c. 333 9.952 33.62 at c cm a No 0 3.5.5.4 Becca
awesome steaSN sNNeaeN e s Roscoe N0.0 Essen 28.8 :28
5.033 sNSanN 38.38 e s Canoes as). .
xeoeasne thpaNNN >238N s Poaoee m2

Noe e Noe e see c as o ova
5.98... Became... Demons. .m a awesome ~05 3.9.3
3 N28... 32.953 amNeaNNe s ceases No_o Edge
Sign Stage 38.03 2. s a... 9 cc N 65 5.25... 88:88:.
2.8.033 accesses 35.03.”... a awesome 66 5.95...” Sou:
Erodes 258.35... 39.38... s as c. cm a o<s_
x9.oat.N xN Sage 333m... 3 Become m2.
38.38... 2... Sims smNeaBN s Page... 66 5.9.3
58.3%... >m SHE... 38.32... 3 Beam; 66 5.9.3
6933... >338... £332. 3 332+ 66 5.9.3 52.3%?
58.38... 5333 sedate s madam 3. N06 55qu 9685.8
>333... >30 at... 38.253 3 8.32.4 n_<_>_
25333 sNSamee 39.38... s mimosa; m2

Nan NF >3 w >9... v >3 0 0v.n. EmEMmLooEs.

 

0K 9m .5on0 coo: co
£563 059% .99 new .wocmuoiootoE 6:9»... .EESEEQPF 9.620296% :0 No.0 ho wmocozfioto 5.95.55 .m use...

87

3.5. Comparison of the antimicrobial treatment with MAP and antimicrobial
packaging

The purpose of the study was to estimate the antimicrobial effectiveness
of the antimicrobial in combination with MAP. Microbial enumeration was
performed on chicken packaged with an antimicrobial, with and without MAP. If
the effectiveness is not decreased, then the atmosphere for the chicken sample
would not have to be modified. N2 gas is substituted for ambient air to prevent
aerobic bacteria growth, and C02 gas is used to extend the shelf life of food by
hindering the growth of spoilage microorganisms (Jimenez et al., 1997) Thus, it
was hypothesized that an MAP-antimicrobial combination may be more effective,
than either alone.

Half the samples of each selected antimicrobial release system (0.6 pglhr
AITC, 1.2 pglhr AITC, 4 pglhr ClOz, and 8 pglhr CIO;) were combined with an
MAP gas mixture (30% 002/70% N2), and the other half were sealed in air. The
populations of Salmonella Typhiummium, Listeria monocytogenes, and total
aerobic bacteria were investigated. Figures 37-39 show the growth of these
organisms after 8 days. The results indicate that antimicrobial treated samples
with MAP had significantly more of an inhibition effect than those without MAP
(Table 7 and 8). Almost all antimicrobially treated samples packaged in ambient
atmosphere had more growth than samples in MAP. The difference in
effectiveness with/without MAP was more distinguishable for total aerobic
bacteria growth with slow release antimicrobials. At the end of storage, total

bacteria counts treated with 0.6 pglhr AITC with/without MAP were shown to be

88

statistically different. The bacteria growth was 8.1 log cfu/g in air and 4.6 log
cfu/g in MA packages. The total viable counts in 4 pglhr ClOz were 7.8 log cfu/g
in air and 5.5 log cuflg in MAP. The results indicate that the antimicrobial and
MAP combination is more effective than antimicrobial treatment alone. Without
MAP, a higher antimicrobial dosage is needed to get the same effectiveness as

the antimicrobial plus MAP combination.

 

- pkg with atmosphere
CI] pkg with MAP

 

Microbial reduction (logm)

  

 

  

  

 

   

 

 

 

 

   

 

 

 

Control 4~C|02 8-C|02 0.6-AITC 1 .2-AITC

Figure 37. Comparison of inhibition effectiveness of antimicrobial treatments with
MAP and without MAP on Salmonella Typhiummium on fresh chicken (after 8
days. at 4°C)

89

 

7
- pkg with atmosphere
pkg with MAP

 

Microbial reduction (log1o)

 

     

 

 

 

 

 

Control 4-C|02 8-C|02 0.6-AITC 1 .2-AITC

Figure 38. . Comparison of inhibition effectiveness of antimicrobial treatments
with MAP and without MAP on Listeria monocytogenes on fresh chicken (after 8

days, at 4°C)

90

 

10

 

 

 

 

 

 

 

— pkg with atmosphere
pkg with MAP
8 .
”5‘2
.5 6 l
‘8
3
'O
93
E 4 ‘ if:
D .33.}
g 3
.2 ,2:
2 " L:
2 ‘ <
0 _ l .J '

 

 

Control 4-C|02 8-C|02 0.6-AITC 1 .2-AITC

Figure 39. . Comparison of inhibition effectiveness of antimicrobial treatments
with MAP and without MAP on total aerobic bacteria on fresh chicken (after 8
days, at 4°C)

91

$06qu E00026 >_E00Ecm_0 0.0 26. 02.00 2_ 03200030 020 052200 02.00 c. 000.200.0030 E0..0t_u £25 02005.
.:o=0_>0_0 20020000502 .930 028...:

 

020.000
050.00 _00.o._.

0028268022
0.20203

EESEEQE.
020222.06

00v #0 c3030 :00... :0 0200000 05900 .002 0:0
602000300292 0.20203 .E:_.=E_.._Q>.r 02020228 co 32022002901028 00052000200 2055.22. 0o 50200250 N 0E0h

92

Amodvav 30.0.2.0 >..200....2m.0 0.0 .50. 02.00 2. 0.0200000 020 0223.00 02.00 2. 03.50.0000 30.0...0 2...... 0200.2
.20..0.>00 2002000200.). .930 00."..2...

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

000.008? 28.033... £305.? 60.38.03 0.32-6.0-0
00032.0: 08.3.0.0: £303.? 60.38.? 026.0...
08.0020. 0.0.38.3. (00.0020? 62 c «a 0.. 0326.0-.. 0.080..
000.382.: 02.380; 00.03:: 60.95. .0? 026.0... 0.8.8 .93
9. . 00.0.... 08.380; mm. 2o“. .0. (00.9.0. .0: 04...).
00200.00. 000.000.... 02.03 .0; «02 c i .0. 0....
m... 3.5:... <20: .05 5.0.38.0: 9.00.002 03 00.5.6.0-..
.5 3.50.3. (0200...? (00030.02 5.0.0000 0: 02.6.0...
020.9820; (5.00.80: (00.0080: 9.00.0000 0.. 0326.0... 8208.828.
02 . c. m... m. 83.38.? 203000.... (00.000003 026.0... 0.20.0:
00.00.20: 28.00205 28.0 30.0.; «m. c. m. v... n.<.>.
0.0... “RP 03.32.? 80.28.... «ml. 2 .2 v... 02
08.000009 28.33.? (8.0-4.8.? .02 c .00 .3 032.66...
63091.. 22.033: 3308...; (8.008.? 026.0...
$2002.03 202300.03 £0,320.05 9.8.38.0: 0.32-6.0-.. 2.252.223
£030.00; 08.0003..- .5330? 60.0080: 026.0... 0.03200
02.930... 3308...; 020080.. «8.0.8.0: 032
0030.00; 02300.... 88.880; 6.0.0080: 0.<

>00 0 >00 0 >00 0 >w00 C 0v... E0.20m.00.0..>.

 

0.... .0 2000.20 200.. 20 2.265 020.000 0.20.00 .03. 020
.002000.>.0020E 0.20.0... .E:..:E.20>... 020202200, 20 0.202200.. «0.0 3 000202.003 22.5.22. .0 200209200 .0 0.20.-

93

3.6. Fresh chicken packaged with antimicrobials for 21 days

The purpose of this test was to investigate the effect of antimicrobial
packaging on the growth of pathogen/spoilage bacteria, pH, and color of such
chicken breast samples stored at 4°C. The package systems used in this

research experiment are presented in Table 9.

Table 9. Gas mixture sample, and antimicrobial release rates used

 

 

Code Release rate Gas mixture

AIR none Ambient air
MAP none 30%002/70% N2
4-ClOz 4 pglhr CI02 30%002/70% N2
8-CIOz 8 pglhr ClOz 30%002/70% N2
0.6-AITC 0.6 pglhr AITC 30%C02/700/o N2
1.2-AITC 1.2pg/hr AITC 30%002/70% N2

 

Total package volume was 1010 ml

3.6.1. Listeria monocytogenes with antimicrobial treatments

Chicken samples were initially inoculated to contain 103 cfulg of Listeria
monocytogenes and examined during 21 days of storage. The population results
are shown in Table 10. The growth of Listeria monocytogenes at 4°C was slow.
The population in all packages studied was about the same until day 9 (except
for the samples in AIR). A significant effect was observed (p< 0.05) for Listeria
monocytogenes in the 4-CIOz, 8-ClOz, and 1.2-AITC samples from 15 to 21 days.
The most inhibition effectiveness was shown by the ClOz treatments. At day 15,
the population of Listeria monocytogenes with 8-Cl02 was 0.94 logs lower than
MAP (Figure 40). On day 21, the total counts on chicken in the 8-CIOz were 2.75

and 1.87 logs lower compared to AIR and MAP respectively, There was also a

94

significant difference as a function of the CIO;; release rates between 4-ClOz and
8-ClOz. From 12 to 21 days, the growth of Listeria monocytogenes in the 4-Cl02
was higher than in the 8-CIOz, On day 21, the microbial counts in 4-C|02 was
5.60 log cfulg while 8-C'02 had 4.84 log cfulg.

The growth of Listeria monocytogenes in the 1.2-AITC had a statistically
similar growth trend compared to 4-C|02 throughout the storage period. After 15
days, the growth of Listeria monocytogenes in 1.2-AITC was significantly lower
than in either AIR or MAP. On day 21, the counts in 1.2-AITC were approximately
1.65 and 0.77 logs lower than in AIR and MAP respectively. A statistical
difference between 0.6-AITC and MAP was not observed (p>0.05). However,
samples in 0.6-AITC had lower microbial counts than the controls on days 12, 15,

and 21.

95

mm

.000 .0 .02... 0005.0 .0 22.02:. 0 00 .0005 2000.20 200.. 20 20002000300202. 0.20.0.5 .0 2.30.0 .00 0.30.”.

cm

300. 02... 0m0.0.w

m.-

p

o.-

 

 

 

 

 

o.:<.~.. lnTr
02.2.0.0 Ill
No.00 ldl
No.90 lrl
n.<s_ IOI
m_< IOI

 

 

 

Brnao 501

96

Amodv0. .20.0...0 2.2005290 0.0 30. 02.00 2. 0.0200200 020 0225.00 02.00 2. 0.0..020000 .20.0t.0 2.25 0200.). 7
.20..0.>00 00020.0 0 200.2 9.00 00."..2... 9

000.0 800.0 000.0 m._00.0 000.0 (00.0 (00.0 (00.0 0 2.2.0..

 

 

 

”00¢.me Head .230 Hood “who...” “0N0. an F .m> awn-m

on .- .o 0mo¢> one. 02.. 00mm. 9.6m. (mad 9&0. 0.2.16.0
“3.0.025 HNQO> “no—mm?) Hm _. .0 0mm... Hmv.m> Hm .- .m> Hon...”

0 3.0 ooomd 000. o 09.6 ... 96m. (006 <0 .- .o 0.200. NO_Orm
«may. «end Hmmd “mod HNVd “um—.09 Hom.N> ”and:

mmmd 00.. F. 00m w. 00m P. 95m. $30.0 (mud (Rod ago-v

00.0.0x 00.0 000... 00.. .(s 000.0 000.0> 00.0., 000.00

m.000 000...; 80.. 0000 0 203.0, (00.0 (00.0 (00.0 0.52
000.0> 000.0> 000.0 000.0 000.3, 000.0> 000.0 000.0:

00.0 m0.0 002., 8.00. m.000 000.0 (00. (00.0 0.4.
000.0 000.0 000.0: 0.00.0 0000.. 00.0.? 000.0 000.0:
000 .0 ..00 0. 5%. .000 0t. 0000 >00 0 >00 0 000 0

0.0 .0 00.0.0.0 .0 0.00 .0 00.5.0 .020. 000 6.0. €02.02.
.0.20.0.2...20 0000.0. .20.0200 2...; .0005 2000.20 200.. 20 3.0.0 00.. 002000300202 0.20.0... .0 2.30.0 .o. 0.20..-

 

 

 

 

3.6.2. Salmonella Typhimurium with antimicrobial treatments

As seen in Table 11, the level of Salmonella Typhimurium was significantly
reduced (p<0.05) by the antimicrobial treatments. No matter the release rate, the
level of Salmonella Typhimurium present in the antimicrobial plus MAP was
significantly lower than the control, and the reduction difference increased as a
function of time. Inoculated Salmonella Typhimurium chicken breast delayed its
lag phase until 9 days while control packages were significantly higher at 6 days.
The greater inhibition effectiveness was found with the AITC treatment. On day
21, the microbial counts in 1.2-AITC were approximately 2.1 and 1.3 logs lower
than AIR and MAP, respectively. 0.6-AITC was shown to have a statistically
similar effect to 1.2-AITC. Except for day 18, the growth of Salmonella
Typhimurium on the chicken in both packages (0.6-AITC and 1.2-AITC) was not
significantly different. AITC treatment has been shown to have more antimicrobial
effectiveness on Gram-negative bacteria (such as Eschericia Coli, Samonella,
Peudomonas, etc) than Gram-positive bacteria (such as Listeria spp.,
Staphylococcus spp., etc) (lsshiki et al., 1992; Kim et al., 2002; Delaquis et al.,
1997). The results show that AITC had better antimicrobial performance against
Salmonella Typhimurium than Listeria monocytogenes.

CI02 sachet packages were also significant different (p<0.05) than the
control packages (AIR and MAP) against Salmonella Typhimurium as storage
time increased. However, overall inhibition performance was significantly lower
than with the AITC treatments. The amount of Salmonella Typhimurium in the 8-

ClOz package was approximately 1.40 and 0.70 logs lower than in the AIR and

98

MAP respectively. CI02 had shown strong antimicrobial performance against the
pathogens in the agar test. Both Salmonella Typhimurium and Listeria
monocytogenes treated with CIO;, were eliminated with 160 pg/I while 360 ug/l
AITC was required to eliminate these pathogens. With fresh chicken, however, a
much higher CIO; treatment was required to get any significant inhibition. The
released CIO; gas may be absorbed into the moist product mass due to its high

water solubility (2.63 g/l), where it will quickly decompose.

99

mm

.000 .0 .0E.. 000.90 .0 22.02:. 0 00 .0005 2020.20 200.. 20 2.:..:E.20>._. 0.020220% .0 2.30.0 .3. 0.30.“.

2.03 02... 000.05
00 0. 0.

- p F

L0

 

 

II
t

6/n;lo 60':

 

l
r.

0.:<.~.. IOI.
0.22.0.0 III
00.0.0 lql
«0.0.0 Irl
n_<.2 IOI
«=0. IOI

 

 

I
(D

 

 

 

100

$0.90. .20.0t.0 2.2005290 0.0 26. 02.00 2. 0.0200200 020 0520.00 02.00 2. 0.0200.0000 .20.0...0 2...... 0200s.
.20..0.>00 0502900200.... 9.00 00."..2:

 

.0..0
000.0>
0.0.0
000.0 x

00..
000.0005
000.0
000025
0......
000...,

000.0 000.0 000.0
.00 .0 .000. .00 .

000.0
000.0.
0m . .o
000.0x

.mm...

00....
00......
000.0,

0000.0
000.0VA
00.0
000.00;
000.0
00.0
00..
000.0>
000.0
00.0
000.

000.0
000.0
0000.0.
00.0..
000.0
000.0
0.0.
00.0>
.00....
00..0>

 

00... 0.0.0 00.0 000.0:
>00 0. x00 0 x00 0 N00 0

0000.0
000.0
0000....
000.0x
00.0

00.0
0.0....
000.0
.0...
0 .0
00...,

(00.0
00.0
0(0..

000.0

0..

. .
0......

000...,>
000.0>

00 .. .o

 

(0.0 (00.0 - .

0...... .0... ..

HBN.MD HNN.MD OP_<|®.O

(Omd ($0.0 «nu—01w

000.00 000.00

($0.0 ($0.0 ~O_01V

0.... .0...

0.0.00 000.00 02.2

(0.0 mmww 0....
0M0 0..

 

II!

0.0 .0 000.000 .0 0.00 .0 00.50 .05. 00.0 090. 0.00.5000
.0.2o.0.2...20 0000.0. .20.0200 2...... .0005 200.050 200.. 20 6.0.0 00.. 2.0..02..20>._. 0.520220% .0 2.30.0 2.. 0.20...

l

O

1

3.6.3. Total viable counts with antimicrobial treatments

The total plate counts of packaged chicken breasts, not inoculated with
any pathogenic organism, were also determined to investigate the effect of the
antimicrobials’ on broad spoilage bacteria. The results are shown in Table 12.
Antimicrobial treatments (CI02 and AITC) and their relative release rates had a
significant inhibition effect on total viable counts (p<0.05).

The highest inhibition effectiveness on chicken breast was obtained with
1.2-AITC. On day 3, the initial population decreased 0.7 logs with 1.2-AITC. The
largest difference between MAP and 1.2-AITC was 2.7 log cfulg on day 15. 0.6-
AITC was also shown to have a significant inhibition effect. On day 21, the total
viable counts on chicken breast were 9.3, 9.0, and 8.0 log cfulg in AIR, MAP, and
0.6—AITC. The results show that both AITC release rates inhibited total aerobic
bacteria growth during the storage period.

The 8-Cl02 had a statistically similar effect to 1.2-AITC, until day 15. Then,
the total aerobic bacteria increased rapidly. At the end of storage, there was 1.5
and 1.2 log cfulg less than in AIR and MAP respectively. No significant
effectiveness was observed in 4-ClOz until day 12. On day 15, the bacterial
counts in 4-Cl02 were 1.4 and 0.8 logs lower (p< 0.05) than samples in the
controls (AIR and MAP).

Overall, AITC performed better with smaller dosage than Cl02 treatment.
ClOz is able to kill a broad range of bacteria with a very small concentration (less
than 1 ppm) on a hard surface (Speronello, 2005). However, a much greater

ClOz dosage is required to get a significant antimicrobial effect on chicken breast.

102

CIO; is broadly effective against microorganisms at a wide range of pHs, but its
performance is reduced in high moisture food (Brody, 2006).

At a microbial population of more than 108 cfulg, the product developed
ester-like odors and numerous small translucent colonies on the surface of the
chicken. At this level of microorganisms, the product shelf life had ended (Mead,
2004). From a microbial standpoint, 0.6, 1.2-AITC, and 8-ClOz packages
extended the microbial critical point to more than 21 days. Many other factors for
shelf life evaluation (such as flavor, texture, color, and so on) were not
considered in this study. Further research will be required to clarify the time effect

of antimicrobial treatment on product shelf life of fresh chicken breast.

103

mm

ON

300. 0.2.. 000.05

9.

0..

P

 

 

 

 

 

 

 

 

0.0.4.0.. IOI
0..._<.o.o Ill
00.0.0 |¢|
No.0... lrl
0.<.2 IOI
m.< IOI

 

0..

 

Do? an 05.5 006.05 .0 c2853. 5 mm “wwwfi GOV—GEO p.30...— CO mtg—own OED—mm _mu0._. .NV 0.39“—

SITES 601

104

$0.90. .20.0...0 2.2005290 0.0 .50. 0.200 2. 0.0200200 .20 02.20.00 02.00 2. 0.0200.0000 .20.0...0 2.25 0200.2

.20..0.>00 2002300200.... .m..00 00."..2:

 

 

 

0.0.0 000.0 .0000 000.0 000.0 0000.0 0.0 00.0 0.0.0..
000... 000.0. 000.0x 000.0> 000.0 000.0 000.0x 000.00

000.0 00.0 000.0 000.0 0000. 00.0. 00.0 00.0 02.0.0.0
000.00,. 0000.2, 000.0 00.0x 0.0.0x 000.0x 000.0 000.00

0.0.0 000.0 000. 00.0 00.0 00.0 (0.. 00.0 00.0-0
000.0 x 000.0 x 000.00,. 000.00 000.0. 000.0x 000.0 000.00

000. 000.... 00.0 00.0 00.0 00.0 (0.. 00.0 006-0
000.0 000.0 000.0> 000.0 000.0 000.0 000.0 000.00

000. 0.00... 0.0.0 00.. 000.... 00.. 00.. (0.0 00.5.
000.0 000.0> 000.0 0.0.. 000.0> 000.0 000.0> 000.00

00.. 000.0 000.0, 00.. 000.0 000.0, 000.0 (0.0 m...
0000.. 00%: 00.0? 000.? “0.00.. 000.00 000.00 000.00

.0 L0. 07. m. 0 0r 0 0

 

 

0.0 .0 000.20 0.00.0.0 .0 00.50 .02....
020 00.0. 0.0.20.0.E..20 0000.0. .20.0200 2...... .0005 29.0.20 200.. 20 6.0.0 00.. 0.2000 020.002 0.20.00 .0.0... .N. 0.20 ..

5
0

1

3.6.4. pH of chicken breast during 21 days of storage

Many elements can affect the pH of a food during storage. Mead (2004)
said that the pool of amino acids (meat) may become depleted, by microbes or
spoilage bacteria. There may be microbial proteolysis and lipolytic activity. Under
these conditions, oxygen content in the package may be greatly diminished and
carbon dioxide increased, and thus, the bacteria population ultimately is
dominated by slow growing lactic acid bacteria which are tolerant to 002.
Therefore, pH value may increase at first, and then decrease due to the growth
of lactic acid bacteria. Dunnick et al. (1982) mentioned that increase in meat pH
during storage mainly comes from microbial growth. Thus, little change in the pH
of chicken meat may be an indication that little microbial growth has occurred in
storage. The average pH of the chicken breast in the different packages (AIR,
MAP, 4-Cl02, 8-ClOz, 0.6-AITC, and 1.2-AITC) and stored at 4°C for up to 21
days is shown in Table 13. Two sets of chicken breasts were obtained from
different sources, and each source had a different initial pH value (5.79 vs. 6.33).
Therefore, the results are reported separately in the table. Both had similar pH
profiles during storage.

As is shown in Table 13, the release rate of the CIO; treatments had a
significant effect on pH of the chicken samples. The pH values of chicken in 8—
CD; was significantly lower from day 6, and then decreased as a function of time.
The pH of the chicken taken from the last day of storage was 5.45 and 5.93 (for
the 18t and 2"d sets, respectively). 4 pglhr CIO; also had some effect on pH of the

chicken. On days 15-18, the chicken pH in the 4-Cl02 package was significantly

106

lower than initially. In contrast, no significant pH changes were observed in the
chicken treated with AITC. The pH profiles over 21 days were statistically the
same for both 0.6-AITC and 1.2-AITC. However, CIO; treatments significantly
decreased (p<0.05) the pH values of the fresh chicken. As shown in Table 13,
chicken breast exposed to the higher CIO; treatment had lower pH values. pH of
the chicken samples in 8-CIOz decreased from 5.79 to 5.45 in the first set, and
from 6.33 to 5.93 in the second after 21 days. No significant difference (p<0.05)
was observed with 4-ClOz in the first set, and a slight decease from 6.33 to 6.25
was noted in the second set. The pH drop may be due to CI02 gas absorption
into the product. CI02 can penetrate into the moist and tender chicken. The
absorbed ClOz may then be broken down, leaving chloride ion (OH on the
chicken (Ellis et al., 2005). This is likely the reason why the pH value of chicken
meat treated with ClOz was significantly lower than in other packages.

The average pH values of chicken in AIR significantly increased during
storage (p<0.05). The pH levels (initial 5.79 and 6.33) began to increase at 6
days, and rose to 6.31 and 6.54 (in the first and second sets, respectively) on
day 21. The pH in MAP was also affected by the storage time but the rate of
increase was significantly slower than in AIR. The pH values from each test were
statistically higher after day 12, and increased from 6.01 to 6.45. Total aerobic
bacteria in these samples (Table 12) in MAP and AIR became very high
(>108cfu/g) within 9 to 12 days. It is likely that there is a strong relationship
between microbial growth and pH as shown in the chicken breast samples. Aksu

(2006) also showed that the pH values of chicken breast and drumstick

107

significantly increased (p<0.01) during storage. Quio et al. (2002) showed that
the pH values of broiler breast were 5.82 to 6.23. The authors assumed the
reason to be the production of proteases by psychotropic bacteria during storage.
Allen et al. (1998) said that the ultimate pH of meat is highly dependent
upon the amount of glycogen present in the muscle. Chicken meat with high pH
is darker and redder than meat with low pH. Higher pH meat had a lower
lightness value (data will be shown in the next section). Phillips (1996) stated that
CO; from MAP is able to change the original pH values by subsequent ionization
of carbonic acid which can form on the surface of the food by 002 absorption.
However, it was a very small change, and did not cause any significant effect

compared to the pH of the samples in AIR.

108

699$ 280:6 >_EmoEcm_w 9m 39 9:3 :_ Eatomnsm ucm 3528 8:8 E 898885 Echoti 53> 850.2
:2 652m.- .nao whim Eo: 850w ”m .__2 6533 “mam. .9on 526000 So: 850m 3.

 

 

 

 

 

 

 

<86 <86 <86 <86 <86 <86 <86 <86 82.?

88> «8.8% «88% “88> “88> “8.06 «8.86 836

<86 <86 <86 <86 <86 <86 <86 <86 82.86

88> “8825 8.8 “88 «8.8 «88> 836 «8.86

086 88 6 n.88. 888. 88. 88.6 <86 <86 No.08 av
“8.8x «8.8x 868x «8.8 86.8 “8.8 836 “8.8: 83
<86 <86 <86 <8. <8. <8. <86 <86 No.08 N
8mm 8mm 83 83> 83> 83> H88: H88: 2
88.6) 88.65 88.65 m86 <86 <86 <86 <86 as).

88: «86 «8.8 «$8 6 “5.8 H88 836 «8.86

08.6 88. 88% 88. 886, 88% <8.6 <8.6 m_<

88 on «8 mp «88.. “we? «8.? «:89 H8 8.. «8 m:

<86 <86 <86 <86 <86 <86 <86 <86 82.?
83 “8.8 83 “88> 83> 83> £8.86 83.:

<8.6) <8. <8.6; <9 <6 <86 <86 <86 <86 0:86
#88 “8.8 83 «88> “88> 83> H88: 836

88. 88. 03%, 086 886 m.86 <86 <86 No.08 i
«88» «8.8.x «58x 88 88 «88 «886 836 88
<86 <86 <86 865 <8. <8. <86 <86 No.08 F
#36. 83 83 “88> 83> «88> so: 8%: ..
m.86 88.6,, 88. 886 <86 <86 <86 <86 n_<s_

£68) «8.8 «88 «88: 83> H88 8886 83:

N86 88.6, 08.6, U86 U86 88. <86 <86 «.2

in? «mg: “8.8.. «8.9. H88... «88.. £38 83..

a 48 mi? 8 m o m 6 9:

 

 

meow .80. Epcot? N Ho: .3808 B 98v 3 9.2.6 “woos coono 53.: 8o mo:_m> In 0...... .3 flow...

109

3.6.5. Headspace gas analysis of the chicken packages

The evolution of gas in the chicken packages is shown in Table 14.
Wolfe (1980) observed that dynamic changes in the composition of a package
atmosphere can be due to permeation, leakage and biochemical conversion due
to microbial respiratory activity. During microbial growth, oxygen concentration in
a meat package usually diminishes rapidly and carbon dioxide (C02) will be
increased. Without oxygen, anaerobic bacteria will grow in the package and also
produce 002, Therefore, the dynamic gas composition in a package can be used
to investigation the microbial growth during storage. The gas composition can be
affected by modified atmosphere or controlled atmosphere packaging systems.

Carbon dioxide (C02) concentration in antimicrobially treated packages
decreased with increasing time of storage until day 12, likely because the meat
absorbed the 002 gas via the surface of the fresh chicken. After 12 days, the
002 levels were increased. However, the increase was significantly lower than
that of the MAP control. 002 in AIR increased greatly during storage. The initial
concentration was 0.1%, and it rose to 18.55% on day 21. The results indicate
that there must have been a lot of microbial growth. Lipid oxidation could be

another reason for these results in the AIR.

110

Table 14. Change in gas composition in the package headspace during storage
of chicken breasts (n=3) at 4°C

 

 

0 6 12 18 21
AIR 02(0/0) 20.1 18.3 10.2 5.5 3.3
C02(°/o) 0.1 4.7 12.5 17.3 18.5
MAP 02(0/0) 0.5 0.0 0.0 0.0 0.0
COz(°/o) 29.3 24.1 25.0 31.0 29.70
4-Cl02 02(%) 0.5 0.0 0.0 0.0 0.0
C02(%) 29.0 22.1 22.0 23.2 25.0
80102 oz(%) 0.5 0.0 0.0 0.0 0.0
C02(°/o) 28.5 20.1 16.5 22.9 23.4
0.6-AITC 02(%) 0.5 0.0 0.0 0.0 0.0
COz(%) 29.9 19.0 21.0 23.8 24.0
1.2-AITC 02(%) 0.5 0.0 0.0 0.0 0.0
(Ev/o) 29.0 19.2 18.0 22.9 23.0

 

3.6.6. Surface color of fresh chicken during 21 days of storage

To the consumer, appearance is the major criterion leading to purchase
of meat, and is used to evaluate meat quality (Allen, 1998). Thus, meat color
plays a critical role in the consumer’s perception in deciding upon whether to
purchase a meat product. Other studies have shown that microbial growth did not
result in any significant color change (Allen et al., 1998; Quio et al., 2002; Sams,
2001 ). The purpose of this test was to investigate whether any discoloration
occurred due to ClOz or AITC treatment. The surface color of the chicken meat
was observed using an analytical instrument (calorimeter) as a function of
storage time, and the results are described using three parameters (L, a, and b),
as shown in Tables 15 -20.

The analytical color test was performed on two separate sets of chicken,
which had some initial color difference. The difference was too small to discern

with the naked eye, however, the difference was statistically significant. The

111

chicken in the second set also had a higher initial pH value. Thus, the results are
provided in separate tables for the first and second tests.

8-Cl02 and 1.2-AITC were shown to cause significant differences in
surface color of the chicken. The “L” values in 8-ClOz decreased by 4.88 (49.37
to 44.49) and 4.66 (44.14 to 39.49). At high CI02 treatment, the part of the
chicken nearest the ClOz sachet became brown black, and the discoloration
became more intense as time went on. Thus, the “L” value was reduced during
storage. The color changes in 1.2-AITC were nearly opposite to that of the CIO;)
treatment. The ”L” value was statistically higher at 18 days in both sets. On day
21, the overall “L” value increased to 5.3 (49.37 to 54.67) for the first set and 3.1
(44.15-->48.68) for the second. No significant difference was observed in 4-Cl02,
0.6- AITC, and MAP during storage. At the edge of the chicken breast nearest to
the 4pg/hr release CIO;: sachet, chicken meat samples were shown to be slightly
yellow black in color. However, overall test results did not show any statistical
difference.

The chicken breast in the second set was a little bit darker than the first.
Overall lightness (L) was lower than for the chicken of the first set. Even though
there was an initial color difference, the results were very similar in terms of color
change due to the antimicrobial treatments. Ngoka and Froning (1982) reported
that the pH of meat is highly dependent upon the amount of glycogen present in
the muscle. Yang and Chen (1993) confirmed that chicken meat with high pH
was darker, redder, and yellower in color than meat with lower pH. Thus, the

initial color difference may be due to the pH difference (table 13).

112

Both chicken samples (from the 1’t and 2"d batches) in AIR were shown
to have higher “L” values than MAP. On the last day, the “L" values in AIR were
slightly decreased, 3.22 (53.51 to 50.29) and 2.32 (46.14 to 43.23) for the 1“t and
2"d sets respectively, and the values were the same for 4-CIOz, 0.6- AITC, and
MAP. Lawrie (1998) stated that meat in an aerobic condition has a brighter red
color (high “L” and “a” value) than MAP due to the formation of oxymyoglobin,
and it is eventually oxidized to form metmyoglobin which is a dark color. Unlike
fresh red meat, however, fresh chicken breasts usually have a pinkish-white to
yellow color and color change by oxidation will not have a significant impact on
consumer perception. (Sams, 2001)

Table 17 - 18 show the “a” values recorded for chicken samples
packaged in the 13t and 2Ml sets. The “a” value from chicken breast treated with
8-CIOz was shown to be a positively correlated with the amount of dosage.
Chicken breast stored in 8-ClOz had a significantly increased “a” value (p<0.05)
from 9th day for the first test and 6"1 day for the second test. The values increased
as storage time increased. On day 21, The “a” in 8-C|02 increased by 1.54 (1.65
to 3.19) and 1.14 (3.54 to 4.68) for the 1" and 2“d tests, respectively. Oppositely,
“a” of chicken in 1.2-AITC was shown to have a negative correlation with amount
released and “a” value. After 18 days, “a” in 1.2-AITC was significantly lower
(p<0.05) than “a” on day 3. On day 21, the total decrease in “a” in 1.2-AITC was
1.35 (1.75 to 0.40) and 1.07 (3.62 to 2.63) for the first and second test,
respectively. No significant difference in “a” values were observed on chicken

breasts in 4-C102, 0.6-AITC, MAP, and AIR during storage.

113

There was a significant effect on the “b” values of chicken samples in 8-
CI02 and 1.2-AITC. As shown in Table 19 and 20, “b” values in these packages
increased significantly as a function of time. The chicken in 8—Cl02 had higher “b”
values after 6 days in both tests. On days 21, “b” values had increased 1.04
(2.39 to 3.43) in the 1" test and 2.43 (1.81 to 4.24) in the 2"d test. All chicken
samples in 8-ClOz exhibited the same discoloration. A black/greenish color
developed in areas close in proximity to the CIO; sachet, and the outside of the
main discolored portion had more yellow color. It is likely that the CIO; gas was
quickly absorbed into the chicken nearest the sachets. The reason for this
discoloration of meat has not yet been clearly investigated. Ellis et al. (2005) said
that the green/dark color (on chicken) could be due to choleglobin formation. The
iron atom in the center of a hematic nucleus of myoglobin may be reduced
through exposure to the reducing agent (CI02).

The “b” value in 1.2-AITC significantly increased after 18 days. On day 21,
“b” values in the 1m and 2Ml tests increased by 1.22 (2.23 to 3.50) and 1.16 (1.82
to 2.98), respectively. At the end of storage, the surface color of the chicken was
observed to be a bright and yellowish green over the whole surface. The color
change occurred over a wide area of the chicken in a slower manner while the
discoloration in 8-ClOz was localized in areas close to the sachet. It is likely that
the AITC was distributed more evenly on the surface of the chicken breast. The
color compounds in chicken treated with AITC were not identified in this
experiment. AITC breakdown reaction schemes involve amino acids or disulfides

in proteins. Under these reactions, byproducts such as diallyl disulfide which has

114

a pale yellow color, allyl thiourea (cobalt color), 2-thiohydrantione (orange-red),
and N-allyl thiocarbamoyl amino acid were produced (Ohta et al., 1995, OHSA,
2006, Pechacek et al., 1997, and Winther & Nielsen, 2006). These byproducts
may have accumulated on the chicken breasts, and caused the color change.
Chicken breast in 4-C102, 0.6-AITC, MAP, and AIR samples did not suffer any
significant change in “b” value throughout 21 days storage.

The overall color results clearly show that color changes occurred on
chicken breast treated with 8-ClOz and 1.2-AITC. Taking into consideration what
the consumer’s perception is of what the color should be, it is recommended that
the minimum effective concentration of AITC be used, and thus, it is noted that
0.6 pglhr of AITC did not cause any color change.

The C102 system was designed to release the gas from two sides to
reduce the effect on color. Figure 43 shows a color comparison between the one
side and two side release systems. The discoloration by the 4 pglhr sachet was
clearly reduced using the two side release system. The results indicate that
development of a 0102 releasing film may be able to provide microbial inhibition,

without causing a negative color change.

115

 

Figure 43. Appearance of chicken breast after 21 days at 4°C
A: 4pg/hr from 1 sachet, B: 8pg/hrfrom 1 sachet
C: 4ug/hr from 2 sachet, D: Bug/hr from 2 sachet

116

Godvav E8866 22.8588 8m 32 mEmm E 8983:“... new mcE:_oo mEmm E 865833 E90256 £5, mama—2 U

 
 
 

 

=2 .95ch 8mm. .8on notvooO Eo: venom 56:8 8.

 

 

 

 

886 886 826 1886 8:6 <86 <86 82.?
«8.8; «8.8x «8.8;: «8.8: «8.8: «8.8: «8.8:

<83 <86 <86 <86 <86 <88 <86 8286
«8.8> «863: «8.8;: «868; «8.68:: «8.8: «86¢:

886 83.6 886 886 863 <83 <83 No.08 26
«8.8: «8.8: «8.8: «86¢: «8.8: «36¢: «86¢: 8:
<8.? <86 <86 56 <86 <86 <85 «0.0.8 «
«88> «8s...> «8.8> «86¢: «8.8:> «8.8: «86¢: 8
<86 <86 <86 <2; <8.. <8.? <8.? n_<_2
«8.8> «88> «86.9 «8.8> «8.8:> «8.9.: «86¢:

886 8:6 886 <26 <8.? <86 <86 mi
«8.8 «8.8 «8.8 «8.8 «8.8 «8.8 «8.8

8 8 E l8 8 m «1

938 A86

“3de mm; cams am, o ‘89 00808 Co 966 Pm 9E2. .305 coono Emma E mm:_m> H... 8.00 .3 05m...

1

 

Amodvav «00.0.20 200005090 0.0 30. 00.00 5 30:00.03 0:0 0:60.00 0800 5 03:00.33 0:20.20 53> 0:005. m
:2 65000.. .020 whim E0: 0050w ”m

 

 

 

 

 

 

:36 686 086 0«666 6236 «<36 <36 0 «2&6
«66.3>> «66.3; «3.3:: «5.3; «5.3> «3.3: «9.3:

<63 <86 <86 <86 <36 <36 <36 0 «2.66
«3.3> «3.3> «8.3 «66.3; «:33 «3.3: «8.3:

:86 366 03.65: m$6 <-6 <26 <36 ”0.0.6 as
«366: «6666: «6666: «60.3: «3.3:: «3.3: «3.3: «mm:

<86 <86 <36 <36 <36 <86 <36 «0.0-3 N
«3.3> «53: «:63> «3.3:: «66.3: «3.3: «3.3: 2

<56 <26 <86 <86 <56 <36 <36 n_<s_
«66.3: «66.3: «3.3 «3.3 «06.3: «3.3: «3.3:

:36 £66 866% 866.6,: 56 <36 <36 3:.
«8.3. «2.3> «3.3 «3.3 «66.3 «2.3: «3.3:

N 2 E E 6 m m E
0.3 a 666

«and... 003 00.? L: 0 >09 00920 «0 $8.0 K 00:00 5005 000.020 :00... «0 000.? L: 00.00 .9 0.08.

l

 

 

 

 

 

Godvav 00000.00 200000.090 000 300 0.000 0. 03000000 000 0005.00 0000 0. 000.000.0000 “00.0.0.0 5.3 0000.2
:2 05000.. 000m. .0020 0000000 E0: 00500 000.020 2.

 

0....<.N. _.
0._._<.o.o
N0.0.0 A000
08:
N0.0.3 0F
n.<.2
m.<

 

<96 <26 8&6 8006 B6 36 36
«36x «006 «Re «5.; «34> «00.,> «0::
<0 «6 <8. <03: <36 <86 <86 <36
«.5 «000 «3.0 «,3 «6:> «0:: «E:
:60. 89.6, 83.6, 82.6, 826 <96 <8.6
«600: «006: «006: «62: «36: «05> «00.0>
<56 <86 <86 <86 <36 <36 <36
«00.? «0:: «30;; «3:: «0: «Ft: «6::
<86. <36 <36 <26 <80, <86 <36
«F: «.8:> «2.6 «65> «63 «NE: «6::
<3. <26 <06. <§6 <30, <36 <36
«000 «03 «:6 «02: «00.0: «26: «2.0:
a 2 2 E 0 0

 

 

00v “0 Gfiouwod 0.03 00.0> .0: 0 >09 000.90 ”.0 0>0

0 3 0000.0 «0005 0

FILE

00.0.00 000: 00 000.9 .0: 00.00 .2. 0.00...

 

9

1
1

Godve “00.0.0.0 200000.090 0.0 .50. 00.00 0. 0.0000000 000 0005.00 00.00 0. 000.000.0000 «00.0.0.0 5.2, 0000.2

:2 00.0000 00.0 0.60m E0... 0050w “m

 

 

 

 

 

 

0066 0006 800.6 <36 <26 <066« < 36.6 0 0.0.0.2

«00.0: «00.0: «02.0: «03.0: «00.0 30.0 «00.0:

<26 <86 <006 <26 <02. <00. <86 0 0<.06

«00.0: «00.0 «00.0: «00.0: «00.0: «00.0: «00.0

n.026 :06. 0066 0026 8066 mf6 < 2. 00.0-0 2.
«00.3x «60.3g «.03: «20.3: «30.0 «00.0 «30.0: .000

<26 <06 <26 <006 <02. <00. . <0.6 «063 060
«60.02,: «60.0: «00.0: «00.0: «03.0: «00.0: «60.0:

<006 <26 <06 <86 <006 <306 <006 as).
«00.0 «60.0 «20.0: «20.0: «00.0: «00.0: «00.0

<2. <006: <06 <06 <86 <006 56.. 02

«20.3 «00.3 «02.3: «00.3D «30.3 «00.3 03.3:

20 0L 0. 02 0 0 0 0E

 

 

 

0.3 .0 .326«20.3 00:: 0:.0: .0. 6 :00.

M00990 .0 0>m0 .0 00.50.0005 000.0?0 000.0 00 00:_0> ..0.. .200 0.. 0.00 ._.

20

l

 

l

2

Godvav 50.0.0.0 200000.090 0.0 30. 0.000 0. 000000000 000 0005.00 00.00 0. 000.80.00.00 50.0.0.0 5.3 0000.2 1

3‘ 659.54 «wmw .mhou—m 0.2.80.0 E0...— ghzom cmv—OEO H<

 

 

 

 

000.6 0.0036 U306 <26 436 <20.6 <36 0 22.0.2
«60.0: «00.0: «000:: «03.0 «3.0: «03.0: «00.0:
<26 <06 <06 <03. <006 <026 <006 0 00:06
«03.0: «60.0: «03.0: «03.0 «00.0 «63.0: «00.0:
00 26 0026 0006 800. 802. 0< 62.6 <0.6 660 2.
«03.0 «3.0: «30.0: «06.0: «06.0: «00.0: «00.0: .000
«00.0: «00.0: «0 2.0: «00.0 «0 2.0: «00.0: «60.0: 0
< o < o < o < a»: < o < o < o

00 6 03 6 03 6 20 3 6 03 6 32 6 as).
«00.0: «00.0: «00.0: «00.0 «02.0: «00.0: «00.0:
<006 <06 <36 <03: <36 <2 26 526 0.0.
«60.0: «02.0: «02.0: «06.0: «02.06 «2.0: «320:

20 02 02 02 0 0 0

 

0.3 .0 82630.0 00:: 020: .0. 6 :00. 0

00.000 .0 0>00 Fm 00.530005 09.0.00 000.. .0 00:.0> :0. .0.00 .00 0.00...

 

Godvav “00.0.20 200000.090 0.0 .50. 00.00 0. 0.0..00000 000 0005.00 00.00 0. 05.50.0000 000.0...0 5.3 0000.2
. :2 .00.000._ 00.0 0.0.0w 000.. 0050w ”m

 

 

 

 

I. . 1. < . < . < . <006 <026 - .
000. 0 02. 0 «$2.00 000202.”U «000.220 «00. 2: «00. P: 000. N 2
H00 N) “Fm .N) . > . D < .>D < . < N.O .
<26 <02 6 <02 6 <066 02 6 006 6. 053.06
. . « . «00.2 «00.2 «2.2 «00 2:
« 30 2 .
6206 0606 0306 060 6 02 02 . 00.0-0 2.
«30.3 «00.0 «00.0 «00.0: «20 .0: ”<20 .2: ¢0 .2: .000
<03»: <60. <2. <26 <0 2. 6 06. 6 02. 6 N0_0.3 060
«00.2 «30. 2: «20. 2: «20. 2: «00 .2: 0000 .2: «<60 .2:
< 06.... <26 <86 <26 <00. 6 00. 6 62. 6 00.2
. « . «0: «00.2 «00 2: «00 2: «00 .2:
H—R —. mm —. D 3 . .
: : . < . < <02 6 <06 6
<026 <006 <00 6 02. 6 00 6 «00.2 «20.2 0_<
«06.0p «06.2: «00 :6 «00 2: «36 0:: q: 0 :
20 E 02 02 0

 

0.3 00 60.00000; 00:: 0:.0> .0. 0 >09 000.000 .0 0.»

.00 N 00.50 0000.0 09.0.00 000.. 00 00:.0> .0. .0.00 .ow 0.00 ...

22

1

3.7. Physical property changes due to CI02 treatments

3.7.1. Mechanical properties

Changes in tensile strength (TS) and elongation at break (EB) of conventional
package films (PVC, LDPE, and PS) were determined after 1 day. The storage

temperature was 23°C and RH was 100% because these films were treated with

an equilibrium ClOz vapor from the aqueous ClOz solution. Since the selected
film samples have good water barrier properties, the effect of RH on the
mechanical properties was ignored. Figure 44 and 45 show the percentage
change in tensile strength and elongation at break, respectively. The tested films
did not show any significant mechanical property changes due to the CIO;
treatment except for PS. The percent EB of PS decreased by approximately 50%
at a 250 ppm or higher level of ClOz. Oppositely, an increase in treatment
concentration resulted in an increase in TS strength. PVC and LDPE films had no
statistically significant (p>0.05) changes in mechanical properties.

Razumovskii (1983) used ozone (03), another highly oxidizing agent, to
modify the mechanical properties of PS and PE films. Subjected to the 03
treatment, the films became brittle and opaque. The decrease in E8 might be
due to the degradation of the polymer chain. PS is considered more susceptible
to oxidation. Ozen et al. (2002) saw significant EB reduction in PE after ozone
treatment. The oxidation strength of 03 (2.07) is much higher than that of ClOz
(0.95) (Lenntech, 2006). 2000 ppm of ClOz did have a significant impact on
Cryovac C-1050, LDPE and PVC in this test. The TS of PS was significantly

increased by CIO; treatment. More than 500 ppm CIO; may cause degradation of

123

polymeric chains and strengthen the intermolecular forces as the result of the
formation of polar groups (Ozen et al., 2002). PS became more brittle because of
CIO; treatments. The functional group changes in PS by ClOz treatment should

be investigated in a further study.

40

 

 

 

 

 

 

 

 

 

 

 

2:0: 20 g ‘8
E g A
5 0
‘6
C
-.‘—3 -20
as
CD
8
E .40 + PVC
—0— LDPE
4 —v— PS
-60 . . . . .
0 500 1000 1500 2000 2500

ClOz concentration (ppm)

Figure 44. Percent (%) elongation at break with different CIO; concentrations for
films studied

124

 

 

 

 

 

Tensile strength(%)

 

+ PVC

—0— LDPE
+ PS

 

 

 

 

 

O 500 1000 1500 2000 2500
ClOz concentration (ppm)

Figure 45. Percent (%) tensile strength at different ClOz concentrations used for
the films studied

125

Table 21. Tensile strength and Elongation at break of packaging films (PVC,

LDPE, and PS)

 

 

 

 

0'02
concentration PVC LDPE PS
Oppm 2.98., 9.77.1 0.47..
EB 250ppm 2.95,, 9.71 a 0.22.,
500ppm 3.06,, 10.12., 0.25.,
1000ppm 3.20., 1 1 .35., 0.23.,
2000ppm 3.14,, 1 1 .23., 0.23.,

ClOz
concentration PVC LDPE PS
Oppm 15.76,, 4.44., 14.11.,
TS 250ppm 14.80,, 4.58.. 13.90,,
500ppm 15.05.. 4.59,, 15.16.,
1000ppm 15.00., 4.61,, 15.30.,
2000ppm 14.77,, 4.73,, 15.33.,

 

Means with different letters in the same column are significantly different (p<0.05)

3.7.2. Oxygen barrier properties

The oxygen transmission rate (OTR) of packaging films was determined
as a function of the different CI02 concentrations. Table 22 shows the
permeability results.

At more than 500 ppm ClOz, the permeability was significantly lower than
the control (p<0.05). Tsobkallo et al. (1988) found film (PE) oxidation impairs
mechanical properties but improves the molecular ordering of the PE. Further
exposure resulted in increased crystallinity and elastic modulus of the PE films.
Therefore, the PS barrier properties were enhanced by film oxidation. PVC and
LDPE did not show any significant permeability changes due to CIO; treatment
(p>0.05).

126

Table 22. Oxygen permeability of conventional packaging films (PS, LDPE, and

 

 

PVC) after CIO; treatment
W Oppm 500ppm 1000ppm 2000ppm
Films (Om/I) (1 .38mg/l) (2.76mg/l) (5.52mg/l)
PS(1 mil) 323, 292., 290., 271 ,
PVC(1 mil) 163, 159, 164, 154,
LDPE( 1 .25mil) 461 , 460, 463, 460,

 

Unit: cc.mil/100in2.day.atm
Means with different letters in same row are significantly different (p<0.05)

3.7.3. Color

PS, LDPE, and PVC were treated with different Cl02 concentrations and
stored for 24 hr. Change of color by CIO; treatment is shown in table 23. No
matter the treatment concentration, There were no significant changes observed

in LDPE and PVC. However, PS was shown to have a significant color change at

1000 ppm of CI02, At 1000 ppm, the L* value decreased by 4.55 and b*

increased by 2.86.

Table 23. Color changes of conventional packaging films (PS, LDPE, and PVC)

after CIO; treatment (n=3)

 

 

Films Conc. L* a* b* AE
PS 0 ppm Cl02 91.75, -0.92, -0.88, 91.76,
(1mil) 500 ppm ClOz 90.18, -0.90, -0.24, 90.18,
1000 ppm ClOz 87.20., -0.83, 1.98, 87.23.,
LDPE 0 ppm Cl02 91.88, -0.92, —0.91, 91.89,
(1 .25mil) 500 ppm ClOz 91.31, -0.92, -0.85, 91.32,
1000 ppm ClOz 90.45, -0.88, -0.93, 90.46,
PVC 0 ppm CIO; 92.53, -0.88, -0.84, 92.54,
(1mil) 500 ppm CI02 91.91, -0.89, -0.88, 91.92,
1000 ppm ClOz 91.87, -0.91, -0.90,I 91.88,

 

Means with different letters in the same column are significantlfiifferent (p<0.05)

 

127

CHAPTER IV. CONCLUSION AND FUTURE WORKS

Using a growth medium model system, chlorine dioxide (ClOz) and allyl
isothiocyanate (AITC) were shown to have strong antimicrobial activity against
Salmonella Typhimurium (G10127, G10601, G10931) and Listeria
monocytogenes (1002, 1176, 1304). At 7 and 37°C, 160 -360 ug/l of AITC
eliminated inoculated Salmonella Typhimurium and Listeria monocytogenes on
growth media. 120-180 ugll of CI02 also eliminated these organisms on growth
media.

From the tests (microbial growth, pH, gas composition) with fresh chicken
breast, the antimicrobials (C|02 or AITC) plus MAP showed potential to enhance
the microbial safety of a fresh poultry product. Compared to packaging in AIR,
the growth of Listeria monocytogenes were reduced by 1.15-2.75 log, and the
growth of Salmonella Typhimurium were reduced by 1.07-2.09 logs on fresh
chicken breast at the end of storage (21 days). The antimicrobials plus MAP
combination packages delayed the growth of total bacteria. On day 21, the
number of total aerobic bacteria on chicken breasts treated with 0.6-AITC, 1.2-
AlTC, and 8-Cl02 was less than 8 log cfu/g. The number of bacteria in AIR and
MAP packed chicken breast reached this level on the 9th and 15th days
respectively. The pH and gas headspace composition of MAP controls
significantly changed over 21 days, with antimicrobial (CI02 or AITC) plus MAP
combination packages suffered little or no significant change during storage,
except for a pH decrease in 8-Cl02. The results indicate that microbial growth on

the fresh chicken was suppressed by the antimicrobial plus MAP combination.

128

The fresh chicken color can be affected by high antimicrobial content.
Discoloration was observed in the 1.2-AITC, 4-ClOz, and 8-ClOz samples.
However, the results also show that discoloration due to Cl02 treatment can be
minimized or avoided by dispersed release. Chicken samples treated with 0.6-
AITC did not suffer any negative color change.

AITC or CI02 treatment with MAP could improve the stability and shelf life
of fresh poultry products. However, the use of high concentrations of AITC or
ClOz could result in adverse effects on the organoleptic properties of the fresh
chicken. Thus, in order to apply these compounds practically, the following
research is suggested:

1. Determine the maximum allowable treated concentration for consumer

acceptance at conventional fresh chicken storage temperatures

2. Investigate the use of other antimicrobial combinations for fresh

chicken to lead to lower dosages.

3. Identify and quantify the AITC or Cl02 byproducts in toxicological and

sensory aspect

4. Develop ClOz releasable film for dispersed treatment

5. Investigate a masking technique to minimize or prevent undesirable

flavor due to AlTC or CI02 treatment.

In addition, LDPE and PVC were observed to not suffer any significant
physical property changes due to CIO; treatment (up to 2000 ppm). These
polymers should not be affected by ClOz sanitization. However, CIO;. treatment

caused significant changes in mechanical, oxygen barrier properties, and optical

129

properties (color) of PS. These findings will be helpful in selecting appropriate
plastic packaging materials for a food process which uses CIO; as a food
preservative. Detailed structure and functional group analysis are needed to

determine the morphological effect of ClOz treatment on PS film.

130

APPENDIX A
Methods for analyzing chlorine dioxide and related components in gas
phases (ICA titration method»

131

GAS PHASE SAMPLING AND ANALYSIS

1. SCOPE & FIELD OF APPLICATIONS

This method describes the analytical procedure for withdrawing gas samples
from a CIO2 gas treatment container (such as a HDPE bucket), and analyzing
the gas.

2. APPARATUS

50 ml Beaker
50 ml Burette and Stand
Gas-Tight Syringe (60 cc) with Slip-Tip end

3. REAGENTS

Potassium Iodide Kl 10 wt% Solution

Sodium Thiosulfate Na28203 0.001 N from Certified Solution
Sulfuric Acid H2804 2 N — 4N (1 -2 M)

Starch indicator

4. PROCEDURE

4.1. Draw exactly 10 mL of 10 wt% KI solution into a syringe.

4.2. Hold the syringe upright and expel air from the syringe.

4.3. Draw syringe plunger back very slightly so that liquid meniscus is not right at
the tip.

4.4. Remove tape from the sampling port of the bucket (or other treatment
container).

4.5. Insert the tip of the syringe into the sampling port to form a gas tight seal.
4.6. Sample 50 mL of gas by slowly drawing back the plunger to the 60 mL mark.
4.7. Hold VERY slight backward pressure on plunger to prevent KI from entering
the container and remove from sampling port.

4.8. Place finger over tip of syringe and mix by shaking the contents for about 5
seconds. (C|02 will rapidly transfer to the liquid).

4.9. Repeat steps 2 through 8 until the desired cumulative volume of gas has
been sampled. Record this volume of gas as VS. (Typical volumes are 200 —
500 ml).

4.10. Transfer liquid from the syringe into a 50 mL beaker and add 5 to 25 mL of
distilled water.

4.11. Add 8-10 drops of starch indicator to the solution.

4.12. Titrate the solution to a colorless endpoint using 0.001 N sodium thiosulfate.
4.13. Record the volume of Thio used for the titration as VN (neutral titration).
4.14. Using the pH probe to stir the solution, add H2804 slowly to a pH of <20.
4.15. Allow the solution to stand for 5 minutes in the dark.

132

4.16. Titrate the solution to a colorless end point using 0.001 N sodium
thiosulfate.

4.17. Record the volume of Thio used for the titration as VA (acid titration).
Discard the solution.
5. CALCULATIONS

5.1. Calculate chlorine dioxide concentration in bucket:

5.2. To obtain results in mg per L of gas,

V,xNx67,500

C10 , /L =
2 (mg ) 4XVS

 

V
in --:-l
C12 ,(mg / L) = T XNX71,000

S

5.3. To obtain results in ppmv (parts per million by volume of gas), assuming
room temperature gas:

C102 , (ppmv) = 358xCIO2 , (mg / L)
Cl2 , (ppmv) = 34OxCIO2 , (mg / L)

where N = normality of sodium thiosulfate.

133

APPENDIX B
Growth model system raw data at 37°C and 7°C

134

Table 24. The reduction of Salmonella Typhimurium on agar plate (60 mm X 16
mm) in glass bottle (with soaked Allyl-isothiocyanate-oil mixture filter paper)
(37°C for 2 days)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Inoculation AITC 12ug/l 40ug/I 70ug/l 160ug/I 360ug/l
O X X X x
10‘ 0 x x x x
O X X X x
O X X X X
102 O X X x x
O X X x x
O X X X x
103 O X X X x
O X X X x
O X X X x
104 o o x x x
O O X X x
O O X X x
105 0 o x x x
O O X X x
O O X x x
106 O O X X x
O O O X x

 

 

 

 

 

 

 

1. O: microbial growth on the agar, X: No-microbial growth on the agar
2. The soaked mixture of AITC with corn oil was 1 ml
3. Atmosphere in the bottle was 100% N2

135

Table 25. The reduction of Listeria monocytogenes on agar plate (16 mm x 60
mm) in glass bottle (with soaked Allyl-isothiocyanate-oil mixture filter paper)
(37°C for 2 days)

 

AITC
Inoculation ”"9" 40W” 70W” 160%” 350U9/l

 

 

1o1

 

 

 

1o2

 

 

 

1o3

 

 

 

10‘

 

 

 

1o5

 

 

 

1o6

 

OOOOOOOOOOOOOOOOOO
OOOOOOOOOOOOOOOOOO
OOOOOOOOOOOOOOOXXX
OOOOOOOO)4><><><><><><><><><
O><><><><><><><><><><><><><><><><><

 

 

 

 

 

 

 

 

1. O: microbial growth on the agar, X: No-microbial growth on the agar
2. The soaked mixture of AITC with corn oil was 1 ml
3. Atmosphere in the bottle was 100% N2

136

Table 26. The reduction of Salmonella Typhimurium on agar plate (60 mm X 16
mm) in glass bottle (with soaked Allylisothiocyanate—oil mixture filter paper) (7°C
for 7 days)

 

AITC

Inoculation 12ug/l 40%" 70W" 160‘19” 360U9”

 

 

1o1

 

 

 

1o2

 

 

 

103

 

 

 

10‘

 

 

 

1o5

 

 

 

1o6

 

 

 

 

 

 

 

 

OOOOOOOOOOOOOOOXXX
OOOOOOOOOXXXXXXXXX
><><><><><><><><><><><><><><><><><><
><><><><><><><><><><><><><><><><><><
><><><><><><><><><><><><><><><><><><

 

l. O: microbial growth on the agar, X: No-microbial growth on the agar
2. The soaked mixture of AITC with corn oil was 1 ml
3. Atmosphere in the bottle was 100% N2

137

ill

Table 27. The reduction of Listeria monocytogenes on agar plate (60 mm X 16
mm) in glass bottle (with soaked Allylisothiocyanate—oil mixture filter paper) (7°C
for 7 days)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Inoculation AITC 12ugll 40ug/l 70ugll 160ugl| 360ug/l
O X X X X
101 O x X x x
O X X X x
O 0 x X x
102 o o x x x
O O x x x
O O X X x
103 O O X X x
O O X X x
O O x x x
104 0 o x x x
O O X X x
O O x x x
105 o o x x x
O O X x x
o o o x x
106 o o o x x
O O O X x

 

 

 

 

 

 

 

1. O: microbial growth on the agar, X: No-microbial growth on the agar
2. The soaked mixture of AITC with corn oil was 1 ml
3. Atmosphere in the bottle was 100% N2

138

Table 28. The reduction of Salmonella Typhimurium on agar plate (60 mm X 16
mm in lass bottle (with chlorine dioxide treatment) (37°C for 2 days)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

.noculaifhwg) 12ugll 30ugll 60ug/I 90ugll 120ugll 180ugll
O O O X X x
101 O O O X X x
O O O X x x
O O O X x x
102 0 o o x x x
0 O o x X x
0 O o O x x
103 o o o o x x
0 O O O X x
0 O O o x x
104 0 0 O o x x
0 O O O X x
0 0 o o o x
105 0 O O o o x
0 O 0 o o x
0 O O o o x
106 o o o o o x
0 O O o o x

 

 

 

 

 

 

 

 

1. O: microbial growth on the agar, X: No-microbial growth on the agar
2. Atmosphere in the bottle was 100% N2

139

Table 29. The reduction of Listeria monocytogene on agar plate (60 mm X 16

mm in lass bottle (with chlorine dioxide treatment) (37°C for 2 days)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Inocumlfzwg) 12ug/I 30ug/l 60ug/I 90ugll 120ugll 180ug/I
0 x x x x x
101 0 X X x x x
0 X X x x x
O X X X x x
102 0 X X x x x
0 X X x X x
0 0 x x x x
103 O 0 x x x x
0 O x x x x
0 O X x x x
104 0 0 X X x x
0 0 x x x x
0 0 o o x X
105 0 0 0 o x x
0 O 0 O X x
0 0 0 o x x
106 0 0 o o x x
0 O 0 o x X

 

1. O: microbial growth on the agar, X: No-microbial growth on the agar

2. Atmosphere in the bottle was 100% N2

140

 

Table 30. The reduction of Salmonella Typhimurium on agar plate (60 mm X 16
mm in lass bottle (with chlorine dioxide treatment) (7°C for 7 days)

 

|02(U9)
mom,“ 12ug/I 30ug/l 60ug/l 90ug/l 120ug/I 180ug/I

 

 

1o1

 

 

 

1o2

 

 

 

1o3

 

 

 

1o4

 

 

 

1o5

 

 

 

1o6

 

 

 

 

 

 

 

 

OOOOOOOOOOOOOOOOOO
OOOOOOOOOOOOOOOOOO
OOOOOOOOOOOOOOOOOO
OOOOOOOOOOOOXXXXXX
OOOOOOOOO><><><><><><><><><
><><><><><><><><><><><><><><><><><><

 

 

1. O: microbial growth on the agar, X: No-microbial growth on the agar
2. Atmosphere in the bottle was 100% N2

141

Table 31. The reduction of Listeria monocytogene on agar plate (60 mm X 16
mm in lass bottle (with chlorine dioxide treatment) (7°C for 7 days)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

InoculaLOXUg) 12“9’l 30U9/l 60ug/l 90ug/l 120ug/I 180ug/I
0 0 X X X x
10' O o x x x x
0 O X x x x
0 O X X X x
102 0 0 X X x x
0 O X x X x
0 O 0 x x x
103 0 0 0 X x x
0 o o o x x
0 0 o o x x
10‘ 0 0 o o x x
0 0 o o x X
0 O O O x x
105 0 O O O x x
0 0 o o x x
0 0 o o x x
106 0 O o o x x
0 O O 0 X x

 

1. O: microbial growth on the agar, X: No-microbial growth on the agar

2. Atmosphere in the bottle was 100% N2

142

 

Table 32. Headspace concentrations (ppm) of Chlorine dioxide (CIO2) with

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

growth medium (at 7 °C)
Wt 0 day 0.5 day 1 day 3 day 7 day
6.0 0.0 0.0 0.0 0.0
6°”9" 4.0 0.0 0.0 0.0 0.0
AVE 5.0 0.0 0.0 0.0 0.0
so 1.4 0.0 0.0 0.0 0.0
10.0 2.0 0.0 0.0 0.0
90"9" 10.0 1.0 0.0 0.0 0.0
AVE 10.0 1.5 0.0 0.0 0.0
so 0.0 0.7 0.0 0.0 0.0
15.0 5.0 0.0 0.0 0.0
120ug/l 15.0 1.0 0.0 0.0 0.0
AVE 15.0 3.0 0.0 0.0 0.0
so 0.0 2.8 0.0 0.0 0.0
15.0 7.0 0.0 0.0 0.0
180”“ 15.0 3.0 0.0 0.0 0.0
AVE 15.0 5.0 0.0 0.0 0.0
so 0.0 2.8 0.0 0.0 0.0
Table 33. Headspace concentrations (ppm) of Chlorine dioxide (CIO2) with
growth medium (at 37 °C)
Wt 0 day 0.5 day 1 day 3 day 7 day
6.0 0.0 0.0 0.0 0.0
6°”9" 4.0 0.0 0.0 0.0 0.0
AVE 5.0 0.0 0.0 0.0 0.0
so 1.4 0.0 0.0 0.0 0.0
10.0 2.0 0.0 0.0 0.0
9°”9” 10.0 1.0 0.0 0.0 0.0
AVE 10.0 1.5 0.0 0.0 0.0
so 0.0 0.7 0.0 0.0 0.0
15.0 1.0 0.0 0.0 0.0
120ug/I 15.0 1.0 0.0 0.0 0.0
AVE 15.0 1.0 0.0 0.0 0.0
so 0.0 0.0 0.0 0.0 0.0
15.0 1.5 0.0 0.0 0.0
180”" 15.0 1.5 0.0 0.0 0.0
AVE 15.0 1.5 0.0 0.0 0.0
so 0.0 0.0 0.0 0.0 0.0

 

143

 

 

Table 34. Headspace concentrations (ng/ml) of Allyl isothiocyanate (AITC) with

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

growth medium (at 7 °C)

Wt 0 dgL 1 day 2 daL 4 day 7 day

05” g /1 44.0 33.0 0.0 0.0 0.0

36.0 17.0 0.0 0.0 0.0

AVE 40.0 25.0 0.0 0.0 0.0

so 5.7 11.3 0.0 0.0 0.0

75.0 40.0 7.0 7.0 0.0

1'0““ 65.0 30.0 3.0 3.0 0.0

AVE 70.0 35.0 5.0 5.0 0.0

so 7.1 7.1 2.8 2.8 0.0

165.0 85.0 22.0 16.0 0.0

2'5“?" 155.0 83.0 8.0 14.0 0.0

AVE 160.0 84.0 15.0 15.0 0.0

so 7.1 1.4 9.9 1.4 0.0

50 g /1 385.0 170.0 55.0 25.0 0.0

335.0 150.0 45.0 15.0 0.0

AVE 360.0 160.0 50.0 20.0 0.0

so 35.4 14.1 7.1 7.1 0.0

 

 

 

 

 

 

 

 

 

wt(uglml): Soaked mg of AlTC with 1ml corn oil
Table 35. Headspace concentrations (ng/ml) of Allyl isothiocyanate (AITC) with
growth medium (at 37 °C)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Wt 0 day 1 day 2 day 4 day 7 day
44.0 25.0 6.0 0.0 0.0
0'5”9" 36.0 25.0 4.0 0.0 0.0
AVE 40.0 25.0 5.0 0.0 0.0
so 5.7 0.0 1.4 0.0 0.0
77.0 65.0 23.0 13.0 5.0
1'0”" 63.0 55.0 17.0 7.0 5.0
AVE 70.0 60.0 20.0 10.0 5.0
so 9.9 7.1 4.2 4.2 0.0
166.0 103.0 60.0 49.0 32.0
2‘5”" 154.0 97.0 60.0 39.0 28.0
AVE 160.0 100.0 60.0 44.0 30.0
so 8.5 4.2 0.0 7.1 2.8
5‘19” 385.0 230.0 125.0 85.0 55.0
335.0 210.0 115.0 75.0 45.0
AVE 360.0 220.0 120.0 80.0 50.0
so 35.4 14.1 7.1 7.1 7.1

 

wt(uglml): Soaked mg of AITC with 1mI corn oil

 

APPENDIX C
Summary tables of experimental AITC vapor pressures and model
vefificafion

145

Table 36. AITC vapor pressure based on mole faction at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

P A —
A'TC (x.) p (MaLgule’s equation) (yr " yi)2 (y -)’)z
0.0 0.0 0.0 0.0 3187.1
0.1 5.0 3.6 1.9 2792.5
0.2 9.0 9.5 0.2 2206.6
0.3 20.0 18.1 3.7 1472.6
0.4 35.0 29.7 28.3 716.9
0.5 42.0 44.2 5.0 149.2
0.6 70.0 61.3 75.5 23.6
0.7 80.0 80.0 0.0 554.9
0.8 105.0 99.1 35.4 1814.4
0.9 110.0 116.9 47.7 3654.9
1.0 145.0 132.0 169.0 5707.1
2 . - A. 2
R2 =1...(_yL__ii =0.93
(y - y)2
Table 37. AITC vapor pressure based on mole faction at 7°C
P A _
AITC (x.) p (Marglle’s equation) (Yr - ”)2 (y - )02
0.0 0.0 0.0 0.0 3187.1
0.1 7.0 4.6 5.9 2691.0
0.2 12.0 12.0 0.0 1978.9
0.3 22.0 22.7 0.5 1136.7
0.4 45.0 37.2 60.7 370.4
0.5 60.0 55.3 21.9 1.3
0.6 80.0 76.5 12.4 401.0
0.7 105.0 99.6 28.8 1864.1
0.8 135.0 123.2 139.5 4453.6
0.9 142.0 145.3 10.8 7891.7
1.0 164.0 164.0 0.0 11566.0
. _ A, 2
R2 = 1-91%!)— =o_gg
(y - y)2

146

Table 38. AITC vapor pressure based on mole faction at 22°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

P A —
AITC (x.) p (Margule’s equation) 0’.“ -y,-)2 (y -y)2
0.0 0.0 _ 0.0 0.0 3187.1
0.1 25.0 21.3 13.4 1233.0
0.2 40.0 50.2 104.9 38.6
0.3 - 87.0 - -
0.4 135.0 131.4 12.8 5619.8
0.5 185.0 182.6 6.0 15900.1
0.6 - 238.8 - -
0.7 280.0 297.9 321.8 58315.2
0.8 365.0 357.2 60.4 90465.9
0.9 410.0 413.6 13.0 1275600
1.0 464.0 464.0 0.0 1660933
R2 =1— M =1.00
(y - y)2
Table 39. AITC vapor pressure based on mole faction at 37°C
P A —
AITC (x.) p (Margule’s equation) (Yr “yi)2 (y -y)2
0.0 0.0 0.0 0.0 3187.1
0.1 - 74.6 - -
0.2 200.0 164.4 1270.2 11643.6
0.3 320.0 268.6 2639.9 45014.2
0.4 450.0 385.8 4121.6 1084684
0.5 570.0 513.6 3185.5 2089454
0.6 700.0 648.8 2618.4 3509087
0.7 820.0 787.9 1031.7 5349832
0.8 950.0 926.6 549.4 7570835
0.9 1065.0 1060.4 21.0 10079466
1.0 0.0 0.0 0.0 3187.1
R2 = p911"): =1 .00
(y - y)2

147

 

Table 40. The quantity of AITC penneant in the cell through PE film at 23°C as a
function of time

 

 

 

 

 

 

 

 

 

 

 

Time RT q

0 0 0.00
40 50589 0.86
60 155037 2.64
93 237948 4.05
1 17 323702 5.51
147 369219 6.28
170 525922 8.95

 

 

148

APPENDIX D
Raw data tables of microbial growth, headspace gas composition, pH, and
Color

149

Table 41. Raw data of Salmonella Typhimurium growth (log cfulg) on fresh
chicken based on AITC release rates at 7°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 4day 8day 12day
1 3.60 4.10 5.50 5.70
2 3.50 4.30 5.50 5.50
AIR 3 3.50 4.10 5.60 5.80
AVE 3.53 4.17 5.53 5.67
SD 0.06 0.12 0.06 0.15
1 3.60 3.90 4.50 5.10
2 3.50 3.50 4.75 5.15
MAP 3 3.50 3.65 4.55 5.10
AVE 3.53 3.68 4.60 5.12
SD 0.06 0.20 0.13 0.03
1 3.60 3.60 4.77 5.20
2 3.50 3.50 4.60 5.00
0.3ug/hr 3 3.50 3.70 4.65 5.15
AVE 3.53 3.60 4.67 5.12
SD 0.06 0.10 0.09 0.10
1 3.60 3.50 4.45 4.75
2 3.50 3.75 4.40 4.88
0.6ug/hr 3 3.50 3.60 4.60 4.90
AVE 3.53 3.62 4.48 4.84
SD 0.06 0.13 0.10 0.08
1 3.60 3.65 4.35 4.65
2 3.50 3.76 4.23 4.50
1.2ug/hr 3 3.50 3.80 4.25 4.52
AVE 3.53 3.74 4.28 4.56
SD 0.06 0.08 0.06 0.08
1 3.60 3.80 4.15 4.33
2 3.50 3.70 4.27 4.52
1.4ug/hr 3 3.50 3.60 4.10 4.45
AVE 3.53 3.70 4.17 4.43
SD 0.06 0.10 0.09 0.10

 

150

 

Table 42. Raw data of Listeria monocytogenes growth (log cfulg) on fresh
chicken based on AITC release rates at 7°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 4day 8day 1 ngy
1 4.50 4.60 6.00 7.20
2 4.10 4.50 5.80 7.00
AIR 3 3.80 4.70 5.80 7.30
AVE 4.13 4.60 5.87 7.17
SD 0.35 0.10 0.12 0.15
1 3.35 4.50 5.00 5.50
2 3.65 4.30 4.90 5.50
MAP 3 3.80 4.20 5.00 5.70
AVE 3.60 4.33 4.97 5.57
SD 0.23 0.15 0.06 0.12
1 3.35 4.40 5.00 5.70
2 3.65 4.10 5.10 5.30
0.3uglhr 3 3.80 4.40 5.00 5.50
AVE 3.60 4.30 5.03 5.50
SD 0.23 0.17 0.06 0.20
1 3.35 4.40 4.80 5.10
2 3.65 4.00 4.90 5.10
0.6ug/hr 3 3.80 4.00 4.70 5.00
AVE 3.60 4.13 4.80 5.07
SD 0.23 0.23 0.10 0.06
1 3.35 4.30 4.60 4.40
2 3.65 4.00 4.40 4.80
1.2ug/hr 3 3.80 4.50 4.50 4.70
AVE 3.60 4.27 4.50 4.63
SD 0.23 0.25 0.10 0.21
1 3.35 4.00 4.40 4.50
2 3.65 3.70 4.40 4.60
1.4ug/hr 3 3.80 4.20 4.50 4.80
AVE 3.60 3.97 4.43 4.63
SD 0.23 0.25 0.06 0.15

 

151

 

Table 43. Raw data of total aerobic bacteria growth (log cfulg) on fresh chicken
based on AITC release rates at 7°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 4day 6day 8day

1 5.50 7.70 9.10 9.50

2 5.50 7.50 9.50 9.60

AIR 3 5.80 7.90 9.30 9.80
AVE 5.60 7.70 9.30 9.63

SD 0.17 0.20 0.20 0.15

1 5.50 6.50 7.50 8.70

2 5.50 6.20 7.50 8.40

MAP 3 5.80 6.50 7.70 8.50
AVE 5.60 6.40 7.57 8.53

SD 0.17 0.17 0.12 0.15

1 5.50 6.30 7.40 8.70

2 5.50 6.20 7.50 9.00

0.3uglhr 3 5.80 6.50 6.90 8.80
AVE 5.60 6.33 7.27 8.83

SD 0.17 0.15 0.32 0.15

1 5.50 6.10 7.00 7.50

2 5.50 5.70 6.90 7.30

0.6uglhr 3 5.80 6.10 6.80 7.70
AVE 5.60 5.97 6.90 7.50

SD 0.17 0.23 0.10 0.20

1 5.50 5.40 6.20 7.10

2 5.50 5.50 6.50 7.25

1.2ug/hr 3 5.80 5.20 6.50 7.00
AVE 5.60 5.37 6.40 7.12

SD 0.17 0.15 0.17 0.13

1 5.50 5.20 6.10 6.60

2 5.50 5.10 6.20 6.80

1.4ug/hr 3 5.80 5.00 6.00 7.00
AVE 5.60 5.10 6.10 6.80

SD 0.17 0.10 0.10 0.20

 

152

 

Table 44. Raw data of Salmonella Typhimurium growth (log cfulg) on fresh
chicken based on CIO; release rates at 7°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 4day 8day 12day
1 4.50 4.30 5.50 5.60
2 4.10 4.20 5.30 5.70
AIR 3 3.80 4.40 5.50 5.80
AVE 4.13 4.30 5.43 5.70
SD 0.35 0.10 0.12 0.10
1 4.50 4.20 5.00 5.00
2 4.10 4.30 4.90 5.10
MAP 3 3.80 4.10 4.70 4.80
AVE 4.13 4.20 4.87 4.97
SD 0.35 0.10 0.15 0.15
1 4.50 4.10 5.00 5.10
2 4.10 4.30 5.10 4.90
3uglhr 3 3.80 4.10 4.90 4.70
AVE 4.13 4.17 5.00 4.90
SD 0.35 0.12 0.10 0.20
1 4.50 4.20 4.90 5.00
2 4.10 4.20 4.70 4.90
4ug/hr 3 3.80 4.00 4.80 5.00
AVE 4.13 4.13 4.80 4.97
SD 0.35 0.12 0.10 0.06
1 4.50 4.20 4.50 4.80
2 4.10 4.10 4.80 4.90
6ug/hr 3 3.80 4.10 4.70 4.95
AVE 4.13 4.13 4.67 4.88
SD 0.35 0.06 0.15 0.08
1 4.50 4.00 4.10 4.60
2 4.10 3.70 4.40 4.50
8ug/hr 3 3.80 4.20 4.30 4.65
AVE 4.13 3.97 4.27 4.58
SD 0.35 0.25 0.15 0.08

 

153

 

Table 45. Raw data of Listeria monocytogenes growth (log cfulg) on fresh
chicken depend on ClO2firelease rates at 7°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 4day 8day 12day
1 3.35 4.60 6.00 7.20
2 3.65 4.50 5.80 7.00
AIR 3 3.80 4.70 5.80 7.30
AVE 3.60 4.60 5.87 7.17
SD 0.23 0.10 0.12 0.15
1 4.50 4.50 5.00 5.50
2 4.10 4.30 4.90 5.50
MAP 3 3.80 4.20 5.00 5.70
AVE 4.13 4.33 4.97 5.57
SD 0.35 0.15 0.06 0.12
1 4.50 4.50 4.70 5.45
2 4.10 4.20 4.90 5.55
3uglhr 3 3.80 4.60 5.00 5.65
AVE 4.13 4.43 4.87 5.55
SD 0.35 0.21 0.15 0.10
1 4.50 4.50 5.00 5.53
2 4.10 4.30 4.70 5.60
4ug/hr 3 3.80 4.70 4.80 5.66
AVE 4.13 4.50 4.83 5.60
SD 0.35 0.20 0.15 0.07
1 4.50 4.50 4.80 5.23
2 4.10 4.30 4.70 5.15
6uglhr 3 3.80 4.20 4.50 5.30
AVE 4.13 4.33 4.67 5.23
SD 0.35 0.15 0.15 0.08
1 4.50 4.30 4.80 5.10
2 4.10 4.60 4.20 5.15
8ug/hr 3 3.80 4.10 4.50 5.10
AVE 4.13 4.33 4.50 5.12
SD 0.35 0.25 0.30 0.03

 

154

 

Table 46. Raw data of total aerobic bacteria growth (log cfulg) on fresh chicken
depend on ClO2 release rates at 7°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 4day 6day 8day

1 5.50 7.70 9.10 9.50

2 5.50 7.50 9.50 9.60

AIR 3 5.80 7.90 9.30 9.80
AVE 5.60 7.70 9.30 9.63

SD 0.17 0.20 0.20 0.15

1 5.50 6.50 7.50 8.70

2 5.50 6.20 7.50 8.40

MAP 3 5.80 6.50 7.70 8.50
AVE 5.60 6.40 7.57 8.53

SD 0.17 0.17 0.12 0.15

1 5.50 6.30 7.40 8.70

2 5.50 6.20 7.50 9.00

0.3uglhr 3 5.80 6.50 6.90 8.80
AVE 5.60 6.33 7.27 8.83

SD 0.17 0.15 0.32 0.15

1 5.50 6.10 7.00 7.50

2 5.50 5.70 6.90 7.30

0.6ug/hr 3 5.80 6.10 6.80 7.70
AVE 5.60 5.97 6.90 7.50

SD 0.17 0.23 0.10 0.20

1 5.50 5.40 6.20 7.10

2 5.50 5.50 6.50 7.25

1.2ug/hr 3 5.80 5.20 6.50 7.00
AVE 5.60 5.37 6.40 7.12

SD 0.17 0.15 0.17 0.13

1 5.50 5.20 6.10 6.60

2 5.50 5.10 6.20 6.80

1.4uglhr 3 5.80 5.00 6.00 7.00
AVE 5.60 5.10 6.10 6.80

SD 0.17 0.10 0.10 0.20

 

155

 

Table 47. Raw data of Salmonella Typhimurium growth on fresh chicken using

AITC treatment with MAP and without MAP at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 4day 8day 12day
1 3.60 4.80 5.20 5.80
2 3.60 5.00 5.30 5.60
AIR 3 3.70 5.20 5.50 5.30
AVE 3.63 5.00 5.33 5.57
SD 0.06 0.20 0.15 0.25
1 3.60 3.80 3.90 4.30
2 3.60 3.90 4.00 4.20
MAP 3 3.70 4.00 4.10 4.40
AVE 3.63 3.90 4.00 4.30
SD 0.06 0.10 0.10 0.10
1 3.60 4.00 4.89 5.00
2 3.60 3.80 4.98 5.20
4.0uglhr+AlR 3 3.70 3.90 5.00 5.50
AVE 3.63 3.90 4.96 5.23
SD 0.06 0.10 0.06 0.25
1 3.60 3.30 3.50 3.65
2 3.60 3.20 3.80 3.84
4.0uglhr+MAP 3 3.70 3.60 3.75 3.95
AVE 3.63 3.37 3.68 3.81
SD 0.06 0.21 0.16 0.15
1 3.60 4.10 4.00 4.50
2 3.60 3.30 4.20 4.40
8.0ug/hr+AlR 3 3.70 4.00 4.10 4.30
AVE 3.63 3.80 4.10 4.40
SD 0.06 0.44 0.10 0.10
1 3.60 3.20 3.50 3.75
2 3.60 3.00 3.30 3.55
8.0ug/hr+MAP 3 3.70 3.40 3.40 3.68
AVE 3.63 3.20 3.40 3.66
SD 0.06 0.20 0.10 0.10

 

156

 

Table 48. Raw data of Listeria monocytogenes growth on fresh chicken using

AITC treatment with MAP and without MAP at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

0day 4day 8day 12day
1 3.50 4.00 4.70 5.20
2 3.00 4.10 4.50 5.50
AIR 3 3.20 4.00 5.00 5.10
AVE 3.23 4.03 4.73 5.27
SD 0.25 0.06 0.25 0.21
1 3.50 3.50 3.50 3.90
2 3.00 3.10 3.40 3.80
MAP 3 3.20 3.30 3.50 3.60
AVE 3.23 3.30 3.47 3.77
SD 0.25 0.20 0.06 0.15
1 3.50 4.20 4.30 5.10
2 3.00 4.00 4.20 4.80
0.6ug/hr-I-AIR 3 3.20 4.20 4.40 5.00
AVE 3.23 4.13 4.30 4.97
SD 0.25 0.12 0.10 0.15
1 3.50 3.00 3.50 3.70
2 3.00 3.10 2.90 3.30
0.6ug/hr+MAP 3 3.20 2.90 3.20 3.60
AVE 3.23 3.00 3.20 3.53
SD 0.25 0.10 0.30 0.21
1 3.50 3.50 3.50 4.20
2 3.00 3.30 3.40 4.00
1.2ug/hr+AIR 3 3.20 3.50 3.30 4.00
AVE 3.23 3.43 3.40 4.07
SD 0.25 0.12 0.10 0.12
1 3.50 3.20 3.00 3.50
2 3.00 3.00 3.10 3.70
1.2ug/hr+MAP 3 3.20 3.00 3.00 3.20
AVE 3.23 3.07 3.03 3.47
SD 0.25 0.12 0.06 0.25

 

157

 

Table 49. Comparison of total aerobic bacteria growth on fresh chicken using

AITC treatment with MAP and without MAP at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

0day 4day 6day 8day

1 3.90 6.20 7.50 9.50

2 3.90 6.00 8.00 9.20

AIR 3 4.00 6.10 7.70 9.40
AVE 3.93 6.10 7.73 9.37

SD 0.06 0.10 0.25 0.15

1 3.90 5.20 6.00 6.50

2 3.90 5.00 6.10 6.70

MAP 3 4.00 5.30 5.90 6.50
AVE 3.93 5.17 6.00 6.57

SD 0.06 0.15 0.10 0.12

1 3.90 5.50 7.10 7.90

2 3.90 5.80 7.10 8.10

0.6ug/hr+AlR 3 4.00 5.90 6.90 8.20
AVE 3.93 5.73 7.03 8.07

SD 0.06 0.21 0.12 0.15

1 3.90 4.30 3.80 4.90

2 3.90 4.20 4.20 4.50

0.6ug/hr-I-MAP 3 4.00 4.30 4.50 4.50
AVE 3.93 4.27 4.17 4.63

SD 0.06 0.06 0.35 0.23

1 3.90 4.40 5.00 5.50

2 3.90 4.50 5.30 5.80

1.2uglhr+AlR 3 4.00 4.90 4.90 5.90
AVE 3.93 4.60 5.07 5.73

SD 0.06 0.26 0.21 0.21

1 3.90 4.00 4.10 4.20

2 3.90 4.50 4.20 4.50

1.2uglhr+MAP 3 4.00 4.20 4.50 4.40
AVE 3.93 4.23 4.27 4.37

SD 0.06 0.25 0.21 0.15

 

158

 

Table 50. Comparison of Salmonella Typhimurium growth on fresh chicken using

CIO2 treatment with MAP and without MAP at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

0day 4day 8day 12day
1 3.60 4.80 5.20 5.80
2 3.60 5.00 5.30 5.60
AIR 3 3.70 5.20 5.50 5.30
AVE 3.63 5.00 5.33 5.57
SD 0.06 0.20 0.15 0.25
1 3.60 3.80 3.90 4.30
2 3.60 3.90 4.00 4.20
MAP 3 3.70 4.00 4.10 4.40
AVE 3.63 3.90 4.00 4.30
SD 0.06 0.10 0.10 0.10
1 3.60 4.00 4.89 5.00
2 3.60 3.80 4.98 5.20
4.0uglhr+AlR 3 3.70 3.90 5.00 5.50
AVE 3.63 3.90 4.96 5.23
SD 0.06 0.10 0.06 0.25
1 3.60 3.30 3.50 3.65
2 3.60 3.20 3.80 3.84
4.0uglhr+MAP 3 3.70 3.60 3.75 3.95
AVE 3.63 3.37 3.68 3.81
SD 0.06 0.21 0.16 0.15
1 3.60 4.10 4.00 4.50
2 3.60 3.30 4.20 4.40
8.0ug/hr+AIR 3 3.70 4.00 4.10 4.30
AVE 3.63 3.80 4.10 4.40
SD 0.06 0.44 0.10 0.10
1 3.60 3.20 3.50 3.75
2 3.60 3.00 3.30 3.55
8.0ug/hr+MAP 3 3.70 3.40 3.40 3.68
AVE 3.63 3.20 3.40 3.66
SD 0.06 0.20 0.10 0.10

 

159

 

Table 51. Comparison of Listeria monocytogenes growth on fresh chicken using

Cl02 treatment with MAP and without MAP at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

0day 4day 8day 12day
1 3.50 4.00 4.70 5.20
2 3.00 4.10 4.50 5.50
AIR 3 3.20 4.00 5.00 5.10
AVE 3.23 4.03 4.73 5.27
SD 0.25 0.06 0.25 0.21
1 3.50 3.50 3.50 3.90
2 3.00 3.10 3.40 3.80
MAP 3 3.20 3.30 3.50 3.60
AVE 3.23 3.30 3.47 3.77
SD 0.25 0.20 0.06 0.15
1 3.50 3.35 4.80 6.00
2 3.00 4.80 4.40 5.60
4.0uglhr+AlR 3 3.20 4.00 4.60 5.00
AVE 3.23 4.05 4.60 5.53
SD 0.25 0.73 0.20 0.50
1 3.50 3.30 3.10 3.80
2 3.00 2.80 3.00 3.90
4.0uglhr+MAP 3 3.20 3.00 2.70 3.50
AVE 3.23 3.03 2.93 3.73
SD 0.25 0.25 0.21 0.21
1 3.50 3.70 3.70 4.20
2 3.00 3.30 3.60 4.30
8.0uglhr+AlR 3 3.20 3.80 3.90 4.40
AVE 3.23 3.60 3.73 4.30
SD 0.25 0.26 0.15 0.10
1 3.50 3.30 3.20 3.80
2 3.00 3.00 3.00 3.60
8.0ug/hr+MAP 3 3.20 2.80 3.10 3.70
AVE 3.23 3.03 3.10 3.70
SD 0.25 0.25 0.10 0.10

 

160

 

Table 52. Comparison of total aerobic bacteria growth on fresh chicken using

CIO2 treatment with MAP and without MAP at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

0day 4day 6day 8day

1 3.90 6.20 7.50 9.50

2 3.90 6.00 8.00 9.20

AIR 3 4.00 6.10 7.70 9.40
AVE 3.93 6.10 7.73 9.37

SD 0.06 0.10 0.25 0.15

1 3.90 5.20 6.00 6.50

2 3.90 5.00 6.10 6.70

MAP 3 4.00 5.30 5.90 6.50
AVE 3.93 5.17 6.00 6.57

SD 0.06 0.15 0.10 0.12

1 3.90 4.30 6.00 7.60

2 3.90 4.80 6.10 8.00

4.0uglhr+AlR 3 4.00 4.20 5.90 7.80
AVE 3.93 4.43 6.00 7.80

SD 0.06 0.32 0.10 0.20

1 3.90 4.00 5.00 5.50

2 3.90 4.50 5.10 5.40

4.0uglhr+MAP 3 4.00 4.30 4.70 5.50
AVE 3.93 4.27 4.93 5.47

SD 0.06 0.25 0.21 0.06

1 3.90 4.50 5.60 6.20

2 3.90 4.50 5.60 6.50

8.0uglhr+AlR 3 4.00 4.70 5.50 6.00
AVE 3.93 4.57 5.57 6.23

SD 0.06 0.12 0.06 0.25

1 3.90 4.00 4.30 4.70

2 3.90 4.20 4.10 4.30

8.0ug/hr+MAP 3 4.00 4.00 4.50 4.80
AVE 3.93 4.07 4.30 4.60

SD 0.06 0.12 0.20 0.26

 

161

 

Table 53. Raw data of Listeria monocytogenes (log cfulg) on fresh chicken breast
with constant release antimicrobial treatment during 21 days at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 3day 6day 9day 12day 15day 18day 21day
1 3.30 3.20 4.20 4.80 5.20 6.40 7.00 7.70
2 3.43 3.80 4.65 5.20 5.87 6.55 7.25 7.50
AIR 3 3.35 3.50 4.60 5.00 5.54 6.30 6.95 7.60
AVE 3.36 3.50 4.48 5.00 5.54 6.42 7.07 7.60
so 0.07 0.30 0.25 0.20 0.34 0.13 0.16 0.10
1 3.30 3.10 3.30 4.00 4.50 5.60 6.60 6.80
2 3.43 3.50 3.55 3.70 4.68 5.24 6.07 6.70
MAP 3 3.35 3.30 3.75 4.55 4.60 5.31 6.12 6.65
AVE 3.36 3.30 3.53 4.08 4.59 5.38 6.26 6.72
so 0.07 0.20 0.23 0.43 0.09 0.19 0.29 0.08
1 3.30 2.90 2.70 3.50 3.90 4.50 5.00 5.60
2 3.43 3.20 3.55 3.40 4.25 4.80 5.32 5.85
4-Cl02 3 3.35 3.45 3.35 3.95 4.15 4.65 5.25 5.36
AVE 3.36 3.18 3.20 3.62 4.10 4.65 5.19 5.60
so 0.07 0.28 0.44 0.29 0.18 0.15 0.17 0.25
1 3.30 2.80 2.60 3.10 3.50 4.10 4.50 5.20
2 3.43 3.00 3.46 3.70 3.80 3.80 4.10 4.40
8-Cl02 3 3.35 2.90 3.30 3.45 3.65 3.95 4.30 4.96
AVE 3.36 2.90 3.12 3.42 3.65 3.95 4.30 4.85
so 0.07 0.10 0.46 0.30 0.15 0.15 0.20 0.41
1 3.30 2.80 3.00 3.70 4.10 5.10 5.90 6.55
2 3.43 3.45 3.65 3.65 4.25 5.15 6.00 6.55
A1730 3 3.35 3.33 3.65 4.15 4.10 5.05 5.85 6.25
AVE 3.36 3.19 3.43 3.83 4.15 5.10 5.92 6.45
so 0.07 0.35 0.38 0.28 0.09 0.05 0.08 0.17
1 3.30 2.80 3.00 3.50 3.80 4.60 5.00 5.90
2 3.43 3.25 3.21 3.50 4.11 5.10 5.60 6.00
41120 3 3.35 3.45 3.50 4.00 4.25 4.85 5.30 5.95
AVE 3.36 3.17 3.24 3.67 4.05 4.85 5.30 5.95
so 0.07 0.33 0.25 0.29 0.23 0.25 0.30 0.05

 

 

162

 

Table 54. Raw data of Salmonella Typhimurium (log cfulg) on fresh chicken
breast with constant release antimicrobial treatment during 21 days at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 3day 6day 9day 12day 15day 18day 21day
1 3.20 3.60 5.00 5.80 7.00 7.10 7.50 7.50
2 3.37 3.80 5.07 6.07 7.13 7.00 7.20 7.50
AIR 3 3.25 3.75 5.24 6.05 7.26 7.55 7.85 8.00
AVE 3.27 3.72 5.10 5.97 7.13 7.22 7.52 7.67
so 0.09 0.10 0.12 0.15 0.13 0.29 0.33 0.29
1 3.20 3.20 3.60 5.20 5.15 6.00 6.50 6.90
2 3.37 3.21 3.99 4.71 5.22 6.15 6.50 6.85
MAP 3 3.25 3.71 4.05 4.98 5.35 6.40 6.60 7.00
AVE 3.27 3.37 3.88 4.96 5.24 6.18 6.53 6.92
so 0.09 0.29 0.24 0.25 0.10 0.20 0.06 0.08
1 3.20 3.50 3.65 4.50 5.10 5.70 6.15 6.55
2 3.37 3.00 3.90 4.36 5.00 6.00 6.25 6.60
4-croz 3 3.25 3.70 3.50 4.88 5.40 5.90 6.30 6.65
AVE 3.27 3.40 3.68 4.58 5.17 5.87 6.23 6.60
so 0.09 0.36 0.20 0.27 0.21 0.15 0.08 0.05
1 3.20 3.40 3.30 4.20 4.80 5.12 5.90 6.10
2 3.37 3.00 3.24 4.14 4.70 5.20 5.90 6.35
8-Cl02 3 3.25 3.70 3.35 3.95 4.75 5.15 5.75 6.25
AVE 3.27 3.37 3.30 4.10 4.75 5.16 5.85 6.23
so 0.09 0.35 0.06 0.13 0.05 0.04 0.09 0.13
1 3.20 3.40 3.50 3.90 4.15 4.90 5.60 5.90
2 3.37 2.80 3.30 3.65 4.22 4.60 5.30 5.80
.3436 3 3.25 3.60 3.50 4.28 4.10 4.60 5.45 6.20
AVE 3.27 3.27 3.43 3.94 4.16 4.70 5.45 5.97
so 0.09 0.42 0.12 0.32 0.06 0.17 0.15 0.21
1 3.20 2.90 2.90 3.70 4.10 4.70 5.00 5.70
2 3.37 2.80 3.10 3.75 3.78 4.15 4.50 5.50
£3,226; 3 3.25 3.10 3.30 3.60 3.94 4.45 4.80 5.55
AVE 3.27 2.93 3.10 3.68 3.94 4.43 4.77 5.58
so 0.09 0.15 0.20 0.08 0.16 0.28 0.25 0.10

 

163

 

Table 55. Raw data of total aerobic bacteria (log cfulg) on fresh chicken breast
with constant release antimicrobial treatment during 21 days at 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 3day 6day 9day 12day 15day 18day 21day
1 3.90 5.80 7.08 8.84 8.79 9.05 8.75 9.50
2 4.20 5.80 7.70 9.00 9.00 8.90 9.60 9.30
AIR 3 4.15 5.75 7.55 9.00 9.15 9.55 9.35 9.45
AVE 4.08 5.78 7.44 8.95 8.98 9.17 9.23 9.42
so 0.16 0.03 0.32 0.09 0.18 0.34 0.44 0.10
1 3.90 4.40 5.50 6.62 7.50 7.99 8.20 8.50
2 4.20 4.60 5.76 6.75 7.50 8.20 8.50 8.90
MAP 3 4.15 4.55 5.80 6.70 7.70 8.40 8.70 8.85
AVE 4.08 4.52 5.69 6.69 7.57 8.20 8.47 8.75
so 0.16 0.10 0.16 0.07 0.12 0.21 0.25 0.22
1 3.90 4.00 4.50 5.20 6.60 7.74 7.70 8.00
2 4.20 3.80 4.20 5.45 6.88 7.50 8.05 8.42
4-Cl02 3 4.15 4.00 4.40 5.48 6.80 7.75 7.95 8.55
AVE 4.08 3.93 4.37 5.38 6.76 7.66 7.90 8.32
so 0.16 0.12 0.15 0.15 0.14 0.14 0.18 0.29
1 3.90 3.80 3.85 4.10 5.50 6.00 7.00 7.15
2 4.20 4.00 3.90 4.40 5.80 6.40 7.25 7.45
8-Cl02 3 4.15 4.15 4.05 4.45 5.75 6.25 7.45 7.55
AVE 4.08 3.98 3.93 4.32 5.68 6.22 7.23 7.38
so 0.16 0.18 0.10 0.19 0.16 0.20 0.23 0.21
1 3.90 3.75 4.00 4.20 4.90 6.25 7.35 7.60
2 4.20 3.90 4.40 4.65 5.20 6.35 7.35 8.25
3%“: 3 4.15 3.95 4.60 4.85 5.45 6.40 7.55 8.15
AVE 4.08 3.87 4.33 4.57 5.18 6.33 7.42 8.00
so 0.16 0.10 0.31 0.33 0.28 0.08 0.12 0.35
1 3.90 3.20 3.60 3.90 4.40 5.00 6.50 7.05
2 4.20 3.50 4.00 4.30 4.80 5.80 6.80 7.35
A2263 3 4.15 3.55 4.15 4.40 4.75 5.75 6.88 7.45
AVE 4.08 3.42 3.92 4.20 4.65 5.52 6.73 7.28
so 0.16 0.19 0.28 0.26 0.22 0.45 0.20 0.21

 

 

 

 

 

 

 

 

 

 

164

 

Table 56. Raw data for pH values of fresh chicken breast for 21 days, 4°C (1“t

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

testset)
No. 0day 3day 6day 9day 12day 15day 18day 21day
AIR 1 5.79 5.82 6.00 5.99 6.05 6.20 6.30 6.32
2 5.77 5.79 6.03 6.01 6.11 6.25 6.25 6.31
3 5.80 5.80 5.99 5.89 6.14 6.23 6.20 6.31
4 5.79 5.82 6.02 5.92 6.03 6.18 6.25 6.30
AVE 5.79 5.81 6.01 5.95 6.08 6.22 6.25 6.31
SD 0.01 0.02 0.02 0.06 0.05 0.03 0.04 0.01
MAP 1 5.79 5.83 5.81 5.78 5.89 5.99 6.01 6.01
2 5.77 5.79 5.82 5.83 5.95 5.94 5.88 6.05
3 5.80 5.77 5.79 5.77 5.94 6.09 5.91 5.94
4 5.79 5.82 5.80 5.80 6.03 5.95 5.99 6.05
AVE 5.79 5.80 5.81 5.80 5.95 5.99 5.95 6.01
SD 0.01 0.03 0.01 0.03 0.06 0.07 0.06 0.05
4-CI02 1 5.79 5.79 5.77 5.80 5.81 5.81 5.72 5.75
2 5.77 5.80 5.76 5.82 5.79 5.77 5.72 5.75
3 5.80 5.71 5.80 5.77 5.84 5.93 5.80 5.75
4 5.79 5.77 5.79 5.78 5.79 5.64 5.77 5.71
AVE 5.79 5.77 5.78 5.79 5.81 5.79 5.75 5.74
SD 0.01 0.04 0.02 0.02 0.02 0.12 0.04 0.02
8-Cl02 1 5.79 5.77 5.66 5.45 5.42 5.45 5.50 5.40
2 5.77 5.66 5.60 5.42 5.49 5.24 5.55 5.45
3 5.80 5.82 5.54 5.50 5.40 5.58 5.50 5.40
4 5.79 5.79 5.57 5.48 5.42 5.35 5.49 5.55
AVE 5.79 5.76 5.59 5.46 5.43 5.41 5.51 5.45
SD 0.01 0.07 0.05 0.04 0.04 0.14 0.03 0.07
0.6- 1 5.79 5.81 5.84 5.82 5.82 5.89 5.82 5.88
AITC 2 5.77 5.84 5.68 5.79 5.79 5.72 5.81 5.83
3 5.80 5.80 5.71 5.88 5.88 5.71 5.85 5.79
4 5.79 5.79 5.77 5.78 5.63 5.85 5.83 5.85
AVE 5.79 5.81 5.75 5.82 5.78 5.79 5.83 5.84
SD 0.01 0.02 0.07 0.04 0.11 0.09 0.02 0.04
1.2- 1 5.79 5.79 5.88 5.80 5.84 5.88 5.82 5.88
AITC 2 5.77 5.81 5.72 5.72 5.82 5.72 5.82 5.83
3 5.80 5.83 5.81 5.79 5.77 5.69 5.80 5.85
4 5.79 5.82 5.72 5.80 5.79 5.75 5.81 5.82
AVE 5.79 5.81 5.78 5.78 5.81 5.76 5.81 5.85
SD 0.01 0.02 0.08 0.04 0.03 0.08 0.01 0.03

 

165

 

 

Table 57. Raw data for pH values of fresh chicken breast for 21 days, 4°C (2“

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

testset)
No. 0day 3day 6day 9day 12day 15day 18day 21day
1 6.32 6.43 6.34 6.44 6.45 6.48 6.51 6.52
2 6.32 6.32 6.39 6.45 6.44 6.42 6.43 6.55
AIR 3 6.36 6.35 6.43 6.42 6.47 6.51 6.55 6.55
4 6.33 6.40 6.46 6.45 6.45 6.49 6.45 6.53
AVE 6.33 6.38 6.41 6.44 6.45 6.48 6.49 6.54
SD 0.02 0.05 0.05 0.01 0.01 0.04 0.06 0.02
1 6.32 6.30 6.35 x 6.40 6.42 6.44 6.45
2 6.32 6.26 6.34 6.32 6.47 6.43 6.49 6.44
MAP 3 6.36 6.32 6.32 6.30 6.35 6.44 6.43 6.43
4 6.33 6.28 6.33 6.31 6.41 6.42 6.45 6.49
AVE 6.33 6.29 6.34 6.31 6.41 6.43 6.45 6.45
SD 0.02 0.03 0.01 0.01 0.05 0.01 0.03 0.03
1 6.32 6.40 6.27 6.30 6.28 6.37 6.34 6.24
2 6.32 6.34 6.29 6.31 6.22 6.18 6.27 6.18
4-CIOZ 3 6.36 6.34 6.29 6.27 6.30 6.22 6.18 6.31
4 6.33 6.32 6.30 6.31 6.22 6.25 6.20 6.25
AVE 6.33 6.35 6.29 6.30 6.26 6.26 6.25 6.25
SD 0.02 0.03 0.01 0.02 0.04 0.08 0.07 0.05
1 6.32 6.24 6.04 6.05 6.00 6.03 6.02 5.95
2 6.32 6.44 6.07 6.07 6.00 5.99 5.99 5.89
8-C|02 3 6.36 6.46 6.12 6.09 6.10 6.03 5.88 6.00
4 6.33 6.36 6.12 6.03 5.98 6.00 5.92 5.89
AVE 6.33 6.38 6.09 6.06 6.02 6.01 5.95 5.93
SD 0.02 0.10 0.04 0.03 0.05 0.02 0.06 0.05
1 6.32 6.39 6.28 6.30 6.25 6.28 6.24 6.34
2 6.32 6.36 6.29 6.32 6.30 6.15 6.25 6.40
0.6- 3 6.36 6.33 6.43 6.34 6.33 6.45 6.38 6.34
AITC 4 6.33 6.34 6.30 6.45 6.40 6.33 6.37 6.31
AVE 6.33 6.36 6.33 6.35 6.32 6.30 6.31 6.35
SD 0.02 0.03 0.07 0.07 0.06 0.12 0.08 0.04
1 6.32 6.32 6.45 6.34 6.34 6.35 6.34 6.33
2 6.32 6.36 6.32 6.42 6.32 6.37 6.35 6.35
1.2- 3 6.36 6.32 6.33 6.33 6.38 6.34 6.38 6.40
AITC 4 6.33 6.33 6.28 6.31 6.33 6.32 6.32 6.31
AVE 6.33 6.33 6.35 6.35 6.34 6.35 6.35 6.35
SD 0.02 0.02 0.07 0.05 0.03 0.02 0.02 0.04

 

 

 

 

 

 

 

 

 

 

 

166

 

Table 58. Raw data for color “L” values of fresh chicken breast for 21 days, 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

(1"testset)
No. 0day 3day 6day 9day 12day 15day 18day 21day
1 53.1 53.21 53.30 53.20 52.63 52.13 51.00 51.30
2 53.52 54.30 54.20 55.30 55.38 52.50 51.27 50.11
AIR 3 53.15 53.24 52.13 52.10 54.28 51.43 51.30 49.98
4 52.95 53.30 53.50 53.30 53.46 52.11 51.40 49.80
AVE 53.18 53.51 53.28 53.48 53.94 52.04 51.24 50.30
SD 0.24 0.53 0.86 1.33 1.17 0.45 0.17 0.68
1 53.1 49.00 48.27 47.25 49.00 50.00 48.00 47.50
2 53.52 47.00 49.38 49.38 48.27 49.25 48.29 48.25
MAP 3 53.15 51.00 50.27 50.25 47.25 48.28 49.30 47.25
4 52.95 50.68 51.38 49.25 51.28 50.28 48.25 49.56
AVE 53.18 49.42 49.83 49.03 48.95 49.45 48.46 48.14
SD 0.24 1.84 1.32 1.27 1.71 0.90 0.58 1.04
1 53.1 51.44 48.25 49.25 50.25 49.35 49.00 48.00
2 53.52 49.68 50.25 48.28 49.72 49.73 48.39 49.73
443.02 3 53.15 48.36 48.30 50.83 50.71 50.71 48.40 47.36
4 52.95 47.38 51.30 47.75 46.78 47.45 47.50 46.79
AVE 53.18 49.22 49.53 49.03 49.37 49.31 48.32 47.97
SD 0.24 1.76 1.51 1.35 1.77 1.37 0.62 1.27
1 53.1 50.36 49.25 48.54 45.13 45.50 44.00 44.30
2 53.52 49.36 51.50 48.56 47.22 44.50 43.25 43.22
8-Cl02 3 53.15 46.38 48.30 46.25 45.26 45.21 43.90 44.20
4 52.95 51.36 48.56 47.55 46.13 44.30 44.30 46.25
AVE 53.18 49.37 49.40 47.72 45.94 44.88 43.86 44.49
SD 0.24 2.15 1.45 1.09 0.96 0.57 0.44 1.27
1 53.1 51.26 50.35 51.28 50.57 50.25 49.50 48.50
2 53.52 50.26 47.25 49.83 51.37 51.29 51.25 49.25
0.6- 3 53.15 49.56 48.35 50.85 49.29 49.28 50.30 51.21
AITC 4 52.95 48.59 52.36 49.19 50.28 51.29 50.28 49.21
AVE 53.18 49.92 49.58 50.29 50.38 50.53 50.33 49.54
SD 0.24 1.13 2.25 0.95 0.86 0.97 0.72 1.16
1 53.1 49.25 51.00 52.00 53.00 52.46 53.10 56.40
2 53.52 50.20 50.40 51.26 50.40 50.12 53.30 53.50
1.2- 3 53.15 48.52 49.25 49.30 49.78 49.89 52.70 53.52
AITC 4 52.95 49.52 49.30 50.32 48.25 51.36 53.02 55.26
AVE 53.18 49.37 49.99 50.72 50.36 50.96 53.03 54.67
SD 0.24 0.69 0.86 1.17 1.98 1.19 0.25 1.42

 

 

 

 

 

 

 

 

 

 

167

 

Table 59. Raw data for color “L” values of fresh chicken breast for 21 days, 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2"d test set)
No. 0day 3day 6day 9day 12day 15day 18day 21day
1 45.50 45.70 46.00 46.10 45.60 45.10 44.20 43.50
2 46.40 46.30 45.70 46.20 44.40 43.20 44.30 44.10
AIR 3 46.54 46.20 46.50 45.80 46.10 44.40 43.90 43.00
4 47.90 46.30 46.20 46.10 46.50 45.10 44.40 42.30
AVE 46.59 46.13 46.10 46.05 45.65 44.45 44.20 43.23
SD 0.99 0.29 0.34 0.17 0.91 0.90 0.22 0.76
1 45.50 44.70 44.20 43.30 44.00 45.00 44.80 44.10
2 45.40 44.20 44.40 44.40 44.30 44.30 43.30 43.30
MAP 3 46.54 44.00 45.30 44.30 44.30 43.30 44.30 44.60
4 47.90 45.00 43.80 44.30 44.80 45.30 45.10 43.60
AVE 46.34 44.48 44.43 44.08 44.35 44.48 44.38 43.90
SD 1.16 0.46 0.63 0.52 0.33 0.89 0.79 0.57
1 45.50 44.40 44.50 44.30 44.60 44.40 43.00 43.00
2 45.40 44.70 43.70 43.50 44.70 44.70 43.40 43.70
4-CI02 3 46.54 44.80 44.30 44.80 44.70 43.70 44.40 42.40
4 47.90 43.90 45.30 43.80 43.80 44.50 43.50 43.50
AVE 46.34 44.45 44.45 44.10 44.45 44.33 43.58 43.15
SD 1.16 0.40 0.66 0.57 0.44 0.43 0.59 0.58
1 45.50 44.40 44.30 44.10 42.10 40.50 38.40 39.30
2 45.40 44.40 44.50 44.20 42.20 40.50 39.20 39.10
8-Cl02 3 46.54 43.40 44.40 44.20 41.30 39.50 40.10 39.20
4 47.90 44.40 44.60 44.60 41.10 39.30 38.30 40.30
AVE 46.34 44.15 44.45 44.28 41.68 39.95 39.00 39.48
SD 1.16 0.50 0.13 0.22 0.56 0.64 0.84 0.56
1 45.50 45.30 44.80 45.30 45.60 45.30 44.00 43.50
2 45.40 44.30 44.30 44.80 46.40 45.30 45.30 44.30
0.6- 3 46.54 44.60 44.40 44.50 44.30 44.30 44.30 46.20
AITC 4 47.90 44.60 44.70 44.20 45.30 45.30 44.60 44.20
AVE 46.34 44.70 44.55 44.70 45.40 45.05 44.55 44.55
SD 1.16 0.42 0.24 0.47 0.87 0.50 0.56 1.16
1 45.50 44.30 45.00 45.10 46.00 46.50 47.30 48.40
2 45.40 44.30 45.40 44.80 45.40 45.10 47.80 48.50
1.2- 3 46.54 43.50 44.30 44.80 44.80 45.90 48.40 48.50
AITC 4 47.90 44.50 44.40 45.30 45.30 46.40 48.50 49.30
AVE 46.34 44.15 44.78 45.00 45.38 45.98 48.00 48.68
SD 1.16 0.44 0.52 0.24 0.49 0.64 0.56 0.42

 

168

 

Table 60. Raw data for color “a” values of fresh chicken breast for 21 days, 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

1"testset)
No. 0day 3day 6day 9day 12day 15day 18day 21day
1 2.71 3.00 3.10 2.70 2.80 2.70 2.60 2.80
2 2.73 2.70 2.40 2.40 3.00 2.70 2.70 2.90
AIR 3 2.51 2.50 2.40 3.00 2.50 2.70 2.80 2.90
4 2.78 2.60 2.90 2.60 2.80 2.80 2.80 2.40
AVE 2.68 2.70 2.70 2.68 2.78 2.73 2.73 2.75
SD 0.12 0.22 0.36 0.25 0.21 0.05 0.10 0.24
1 2.71 1.70 1.80 1.90 1.90 1.50 1.20 1.70
2 2.73 1.50 1.70 1.50 0.90 1.90 1.90 1.70
MAP 3 2.51 1.80 1.50 1.70 2.60 1.70 1.50 1.70
4 2.78 1.80 1.90 1.30 1.40 1.80 1.90 1.80
AVE 2.68 1.70 1.73 1.60 1.70 1.73 1.63 1.73
SD 0.12 0.14 0.17 0.26 0.73 0.17 0.34 0.05
1 2.71 1.50 1.50 1.80 2.00 1.50 1.50 2.00
2 2.73 2.20 2.20 2.20 1.80 1.80 1.70 1.70
4-CIOZ 3 2.51 1.60 1.80 1.40 1.60 2.00 1.80 2.20
4 2.78 1.50 1.50 1.80 1.60 1.70 2.00 2.00
AVE 2.68 1.70 1.75 1.80 1.75 1.75 1.75 1.98
SD 0.12 0.34 0.33 0.33 0.19 0.21 0.21 0.21
1 2.71 1.80 1.70 2.60 2.50 2.70 3.10 2.90
2 2.73 2.00 1.60 2.30 2.60 2.60 2.90 3.20
8-Cl02 3 2.51 1.50 1.70 2.40 2.80 3.00 3.00 3.40
4 2.78 1.40 1.90 2.40 2.90 3.10 2.80 3.30
AVE 2.68 1.68 1.73 2.43 2.70 2.85 2.95 3.20
SD 0.12 0.28 0.13 0.13 0.18 0.24 0.13 0.22
1 2.71 1.80 1.30 1.60 2.30 2.40 1.80 1.70
2 2.73 1.50 2.20 2.00 1.50 2.10 1.50 1.80
0.6- 3 2.51 2.00 1.80 1.60 1.80 1.50 1.90 1.90
AITC 4 2.78 1.60 1.70 1.70 1.70 1.80 1.70 1.60
AVE 2.68 1.73 1.75 1.73 1.83 1.95 1.73 1.75
SD 0.12 0.22 0.37 0.19 0.34 0.39 0.17 0.13
1 2.71 2.00 1.50 1.50 1.70 1.00 0.60 0.50
2 2.73 1.50 2.20 1.40 1.70 1.10 0.80 0.30
1.2- 3 2.51 1.90 1.80 1.50 1.20 1.50 0.40 0.50
AITC 4 2.78 1.70 1.20 1.60 1.50 1.50 0.50 0.30
AVE 2.68 1.78 1.68 1.50 1.53 1.28 0.58 0.40
SD 0.12 0.22 0.43 0.08 0.24 0.26 0.17 0.12

169

Table 61. Raw data for color “a” values of fresh chicken breast for 21 days, 4°C
2"d test set)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 3day 6day 9day 12day 15day 18day '21day
1 4.50 4.50 4.10 4.30 4.15 4.35 4.55 4.22
2 4.22 4.26 4.70 4.15 4.15 4.06 4.22 4.10
AIR 3 4.60 4.35 4.15 4.30 4.56 4.22 4.10 4.15
4 3.90 4.56 4.22 4.20 4.12 4.10 4.05 4.35
AVE 4.31 4.42 4.29 4.24 4.25 4.18 4.23 4.21
SD 0.31 0.14 0.28 0.08 0.21 0.13 0.22 0.11
1 3.50 3.80 3.50 3.52 3.50 3.60 3.45 4.30
2 3.20 3.10 3.40 4.00 3.54 3.75 3.70 3.50
MAP 3 4.60 3.44 3.90 3.22 3.40 3.45 3.30 4.00
4 3.90 3.75 3.80 3.50 3.60 3.25 3.54 3.70
AVE 3.80 3.52 3.65 3.56 3.51 3.51 3.50 3.88
SD 0.61 0.32 0.24 0.32 0.08 0.21 0.17 0.35
1 3.50 3.50 3.44 3.64 4.00 3.74 3.85 4.02
2 3.20 3.70 3.80 3.50 3.66 3.33 3.64 3.88
4-CIOZ 3 4.60 3.30 3.75 3.30 3.50 3.45 3.55 3.75
4 3.90 3.50 3.35 3.34 3.54 3.54 3.34 3.95
AVE 3.80 3.50 3.59 3.45 3.68 3.52 3.60 3.90
SD 0.61 0.16 0.22 0.16 0.23 0.17 0.21 0.12
1 3.50 3.65 3.88 3.95 4.12 4.35 4.56 4.65
2 3.20 3.60 4.05 4.01 4.55 4.35 4.55 4.85
8-CIOZ 3 4.60 3.44 3.99 3.85 4.32 4.32 4.55 4.65
4 3.90 3.45 3.80 3.95 4.24 4.20 4.72 4.55
AVE 3.80 3.54 3.93 3.94 4.31 4.31 4.60 4.68
SD 0.61 0.11 0.11 0.07 0.18 0.07 0.08 0.13
1 3.50 3.50 3.80 3.65 3.45 3.75 3.54 3.30
2 3.20 3.56 3.40 3.64 3.33 3.40 3.64 3.64
0.6- 3 4.60 3.65 3.20 3.50 3.75 3.30 3.77 3.65
AITC 4 3.90 3.55 3.95 3.40 3.65 3.75 3.65 3.65
AVE 3.80 3.57 3.59 3.55 3.55 3.55 3.65 3.56
SD 0.61 0.06 0.35 0.12 0.19 0.23 0.09 0.17
1 3.50 3.60 3.55 3.50 3.35 3.45 2.55 2.50
2 3.20 3.65 3.77 3.65 3.45 3.25 3.10 2.65
12- 3 4.60 3.58 3.60 3.77 3.50 3.15 3.50 2.70
AITC 4 3.90 3.66 3.64 3.56 3.50 2.75 2.25 2.65
AVE 3.80 3.62 3.64 3.62 3.45 3.15 2.85 2.63
SD 0.61 0.04 0.09 0.12 0.07 0.29 0.56 0.09

 

170

 

1st test set)

Table 62. Raw data for color “b” values of fresh chicken breast for 21 days, 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

No. 0day 3day 6day 9day 12day 15day 18day 21day
1 2.15 2.15 2.22 2.15 1.90 2.30 2.13 1.98
2.22 2.22 2.30 2.32 1.95 2.10 2.35 2.00
AIR 3 2.35 1.95 2.12 2.00 2.21 1.99 2.00 2.32
4 2.10 2.25 2.05 2.25 2.15 2.22 2.10 2.50
AVE 2.21 2.14 2.17 2.18 2.05 2.15 2.15 2.20
SD 0.11 0.13 0.11 0.14 0.15 0.14 0.15 0.25
1 2.15 2.20 1.80 2.25 1.90 2.10 2.50 2.20
2 2.22 2.24 2.40 2.35 1.90 2.10 2.90 2.50
MAP 3 2.35 2.50 2.20 2.50 2.60 2.10 2.20 1.90
4 2.10 2.20 2.90 2.24 2.90 3.00 1.90 2.70
AVE 2.21 2.29 2.33 2.34 2.33 2.33 2.38 2.33
SD 0.11 0.14 0.46 0.12 0.51 0.45 0.43 0.35
1 2.15 2.30 2.40 2.13 2.40 2.50 2.90 2.60
2 2.22 2.20 2.00 2.20 2.30 2.30 2.20 2.50
4-C|02 3 2.35 2.20 2.20 2.34 2.00 2.00 2.85 2.75
4 2.10 2.50 2.30 2.33 2.60 1.80 2.25 2.77
AVE 2.21 2.30 2.23 2.25 2.33 2.15 2.55 2.66
SD 0.11 0.14 0.17 0.10 0.25 0.31 0.38 0.13
1 2.15 2.50 2.87 3.10 2.90 3.50 3.50 3.50
2 2.22 2.22 2.90 2.90 2.65 3.45 3.25 3.22
8-CI02 3 2.35 2.44 2.88 2.99 3.50 3.50 3.55 3.45
4 2.10 2.40 2.68 3.25 3.20 2.90 3.45 3.55
AVE 2.21 2.39 2.83 3.06 3.06 3.34 3.44 3.43
SD 0.11 0.12 0.10 0.15 0.37 0.29 0.13 0.15
1 2.15 2.50 2.35 2.40 2.50 3.30 2.80 2.30
2 2.22 2.80 2.55 2.15 3.00 2.30 2.65 2.40
0.6- 3 2.35 2.20 2.20 2.20 2.50 2.20 2.30 2.50
AITC 4 2.10 2.00 2.50 2.30 1.90 1.90 2.53 2.50
AVE 2.21 2.38 2.40 2.26 2.48 2.43 2.57 2.43
SD 0.11 0.35 0.16 0.11 0.45 0.61 0.21 0.10
1 2.15 2.70 2.50 2.45 2.25 2.90 3.00 3.00
2 2.22 1.70 2.20 2.50 2.60 2.46 3.50 4.00
1.2- 3 2.35 2.00 2.40 2.33 2.60 2.68 3.00 4.00
AITC 4 2.10 2.50 2.70 2.45 2.50 2.45 4.00 3.00
AVE 2.21 2.23 2.45 2.43 2.49 2.62 3.38 3.50
SD 0.11 0.46 0.21 0.07 0.17 0.21 0.48 0.58

 

171

 

APPENDIX E
Raw data tables of tensile strength (TS), elongation at break, oxygen
transmission rate, and color

172

Table 63. Raw data for color “b” values of fresh chicken breast for 21 days, 4°C

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

2"“testset)
No. 0day 3day 6day 9day 12day 15day 18day 21day
1 1.90 1.34 1.55 1.13 1.20 1.40 1.20 1.74
2 1.65 1.32 1.25 1.64 1.10 1.45 1.56 1.45
AIR 3 2.00 1.23 1.35 1.70 1.32 1.50 1.50 1.64
4 1.75 1.35 1.33 1.69 1.48 1.55 1.76 1.35
AVE 1.83 1.31 1.37 1.54 1.28 1.48 1.51 1.55
SD 0.16 0.05 0.13 0.27 0.16 0.06 0.23 0.18
1 1.90 1.96 2.30 1.70 1.84 1.69 1.60 1.70
2 1.65 1.95 1.70 2.40 1.85 1.77 1.75 1.77
MAP 3 2.00 1.80 1.79 1.74 1.84 1.65 1.87 1.73
4 1.75 1.77 1.82 1.76 1.67 1.76 1.76 1.65
AVE 1.83 1.87 1.90 1.90 1.80 1.72 1.75 1.71
SD 0.16 0.10 0.27 0.33 0.09 0.06 0.11 0.05
1 1.90 1.71 1.70 1.98 1.84 1.59 1.63 1.88
2 1.65 1.64 1.87 1.80 1.83 2.10 1.86 1.75
4-Cl02 3 2.00 1.88 1.80 1.87 1.86 1.99 1.65 2.04
4 1.75 1.97 1.84 1.76 1.67 1.84 1.9 1.88
AVE 1.83 1.80 1.80 1.85 1.80 1.88 1.76 1.89
SD 0.16 0.15 0.07 0.10 0.09 0.22 0.14 0.12
1 1.90 1.77 1.78 2.10 2.54 2.95 3.52 4.21
2 1.65 2.00 1.90 2.31 2.11 3.25 3.35 3.95
8-CIO2 3 2.00 1.80 2.07 2.31 2.22 3.18 3.89 4.44
4 1.75 1.66 1.89 2.11 2.46 3.54 3.98 4.35
AVE 1.83 1.81 1.91 2.21 2.33 3.23 3.69 4.24
SD 0.16 0.14 0.12 0.12 0.20 0.24 0.30 0.21
1 1.90 1.65 2.11 1.64 1.98 2.00 1.65 2.04
2 1.65 1.98 1.61 1.64 1.85 1.78 2.10 1.87
0.6- 3 2.00 2.10 1.71 1.88 1.75 1.69 1.89 1.84
AITC 4 1.75 1.77 1.65 1.95 1.87 1.88 1.87 1.65
AVE 1.83 1.88 1.77 1.78 1.86 1.84 1.88 1.85
SD 0.16 0.20 0.23 0.16 0.09 0.13 0.18 0.16
1 1.90 1.70 1.70 1.87 1.95 2.15 2.50 3.00
2 1.65 1.70 1.75 2.05 2.01 2.10 2.40 3.33
1.2- 3 2.00 2.00 2.20 1.75 1.84 2.33 2.80 2.75
AITC 4 1.75 1.89 1.8 1.86 2.1 2.25 2.75 2.85
AVE 1.83 1.82 1.86 1.88 1.98 2.21 2.61 2.98
SD 0.16 0.15 0.23 0.12 0.11 0.10 0.19 0.25

 

 

 

 

 

 

 

 

 

 

 

173

 

 

Table 64. Raw data for Tensile strength and Elongation at break of packaging
after CIO; treatment

films (PVC, LDPE, and PS)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

C|02(ppm) No. PVC LDPE PS
E.B T.S E.B T.S E.B T.S

1 2.97 15.50 10.17 4.18 0.61 14.3

2 2.43 13.40 8.50 4.12 0.36 14.3

3 3.16 16.90 11.38 5.20 0.75 14.2

Control 4 3.12 15.90 8.68 4.27 0.25 13.87
5 3.21 17.10 10.10 4.43 0.36 13.9

AVE 2.98 15.76 9.77 4.44 0.47 14.11

so 0.32 1.48 1.19 0.44 0.21 0.21

1 2.89 15.24 9.77 4.57 0.23 13.48

2 3.05 14.50 9.85 4.56 0.18 13.95

3 3.01 15.06 9.67 4.55 0.28 14

250 ppm 4 2.88 15.50 9.50 4.65 0.21 14.2
5 2.90 13.70 9.78 4.57 0.18 13.85

AVE 2.95 14.80 9.71 4.58 0.22 13.9

so 0.08 0.72 0.14 0.04 0.04 0.27

1 3.12 14.99 10.02 4.59 0.25 15.42

2 3.17 14.85 10.20 4.60 0.42 15.05

3 3.25 15.15 9.85 4.60 0.17 15.05

500 ppm 4 2.89 15.24 10.03 4.55 0.25 15.13
5 2.85 15.02 10.50 4.63 0.16 15.16

AVE 3.06 15.05 10.12 4.59 0.25 15.16

so 0.18 0.15 0.25 0.03 0.1 0.15

1 3.51 15.00 10.34 4.62 0.24 15.59

2 3.25 15.20 10.31 4.66 0.22 15.15

3 3.64 14.80 11.40 4.65 0.27 15.38

1000 ppm 4 2.75 15.08 11.35 4.56 0.18 15.26
5 2.85 14.90 9.45 4.57 0.25 15.14

AVE 3.20 15.00 10.57 4.61 0.23 15.3

so 0.39 0.16 0.82 0.05 0.03 0.19

1 2.85 15.10 9.90 4.43 0.2 15.7

2 2.64 14.50 10.08 4.70 0.14 15.05

3 3.33 15.06 10.06 4.94 0.24 15.38

2000 ppm 4 3.59 15.50 10.10 4.54 0.34 15.36
5 3.27 13.70 10.99 4.52 0.22 15.14

AVE 3.14 14.77 10.23 4.63 0.23 15.33

so 0.38 0.70 0.43 0.20 0.07 0.25

 

 

174

 

Table 65. Raw data for Oxygen permeability of packaging films (PVC, LDPE, and

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

PS) after CIO2 treatment

ClOz ([3me No. PS PVC LDPE
1 322.0 150.0 450.0
2 317.0 175.0 470.0
0 ppm 3 330.0 165.0 465.0
AVE 323.0 163.3 461.7

SD 6.6 12.6 10.4
1 292.0 155.0 455.0
2 302.0 162.0 456.0
500 ppm 3 282.0 160.0 470.0
AVE 292.0 159.0 460.3

SD 10.0 3.6 8.4
1 287.0 170.0 460.0
2 285.0 166.0 475.0
1000 ppm 3 300.0 158.0 455.0
AVE 290.7 164.7 463.3

SD 8.1 6.1 10.4
1 271.0 145.0 477.0
2 275.0 158.0 445.0
2000 ppm 3 267.0 160.0 458.0
AVE 271.0 154.3 460.0

SD 4.0 8.1 16.1

 

 

 

 

 

 

175

 

 

Table 66. Raw data for color changes of packaging films ((PVC, LDPE, and PS)

after CIO2 treatment

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cl02(ppm) No. L* a* b* Dele
1 91.48 -0.87 -0.95 91.49
2 91.48 -094 -0.84 91.49
0 ppm 3 92.29 -0.94 -O.84 92.30
AVE 91.75 -092 -0.88 91.76

so 0.47 0.04 0.06 0.47

1 92.51 -0.87 0.15 92.51
2 89.56 -095 -0.54 89.57
PS 500 ppm 3 88.46 -0.87 -0.32 88.46
AVE 90.18 -090 -0.24 90.18

so 2.09 0.05 0.35 2.09
1 86.08 —0.84 1.25 86.09
2 88.65 -0.85 2.35 88.69

1000 Ppm 3 86.87 -0.81 2.33 86.91
AVE 87.20 -0.83 1.98 87.23

so 1.32 0.02 0.63 1.33
1 92.54 -0.87 -0.78 92.55
2 93.52 -0.94 -0.88 93.53
0 ppm 3 91.54 -0.84 -0.86 91.55
AVE 92.53 -0.88 -0.84 92.54

so 0.99 0.05 0.05 0.99
1 91.48 -0.87 -0.87 91.49
2 92.29 -0.85 -092 92.30
PVC 500 ppm 3 91.96 -0.94 -0.86 91.97
AVE 91.91 -0.89 -0.88 91.92

so 0.41 0.05 0.03 0.41
1 91.46 -0.87 095 91.47
2 92.46 -0.91 -0.84 92.47
1000 ppm 3 91.68 -094 -092 91.69
AVE 91.87 -0.91 -090 91.88

so 0.53 0.04 0.06 0.52

 

176

 

 

Table 66 (continued)

 

 

LDPE

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

1 91.45 -0.87 -0.97 91.46

2 91.88 -0.94 -O.91 91.89

0 ppm 3 92.32 -0.94 -0.85 92.33
AVE 91.88 -0.92 -0.91 91.89

SD 0.44 0.04 0.06 0.43

1 91.15 -0.87 -0.94 91.16

2 92.45 -0.94 -0.64 92.46

500 ppm 3 90.34 -0.94 -O.97 90.35
AVE 91.31 -0.92 -0.85 91.32

SD 1.06 0.04 0.18 1.06

1 91.21 -0.88 -0.88 91.22

2 90.24 -0.94 -0.94 90.25

1000 ppm 3 89.89 -0.81 -0.97 89.90
AVE 90.45 —0.88 -0.93 90.46

SD 0.68 0.07 0.05 0.68

 

177

 

BIBLIOGRAPHY
Aberle ED, Forrest JC, Gerrard DE and Mills EW. 2001. Principles of meat
science. Kendall/Hunt Publishing Company

Ahvenainen R. 2003. Novel food packaging techniques. Woodhead Pubishing
Ltd.

Aksu Ml, Karaoglu M, Esenbuga N, Kaya M and Macit M. 2006. Effect of meat
piece. packaging and storage on pH thiobarbituric acid reactive substances and
microbial counts in broilers fed diets supplemented with ram horn hydrolysate.
Food Science and Technologies International. 12(2): 133-143.

Allen CD, Russell SM, and Fletcher DL. 1998. The relationship of broiler breast
color to meat quality and shelf life. Poultry Science. 77: 361-366.

Appendini P and Hotchkiss JH. 1997. Immobilization of Iysozyme on food contact
polymers as potential antimicrobial films. Packaging Technology and Science.
10(5): 271-279.

Ayres JC. Ogilvy WS and Stewart GF. 1950. Postmortem changes in stored
meats. Microorganisms associated with development of slime on eviscerated cut-
up poultry. Food Technology. 4: 199-205

Bajard S, Rosso L, Fardel G and F Iandrois JP. 1996. The particular behavior of
Listeria monocytogenes under sub-optimal condition. lntemational Journal of
Food Microbiology. 29: 201 ~21 1

Bionewsonline. 2005. What is Salmonella?. Automation in Microbiology and
Bioscience.
Available at: http://www.bionewsonline.coth/what_is_salmonella.htm.

Bonilauri P, Liuzzo G, Merialdi G and Bentley S. 2004. Growth of listeria
monocytogenes on vacuum-packaged horsemeat for human consumption. Meat
Science. 68: 671-674.

Brody A. 2001. Active packaging for food applications. Technomic Publishing Co..
Inc. Lancester. PA.

178

Brody A. 2005. The application of active packaging. IFT conference meeting.
Orlando. USA.

Brody A. 2006. State of the art of active l intelligent food packaging. IFT meeting.
Las Vegas. USA.

Buchman GW, Banteee S and Hansen JN. 1988. Structure, expression and
evolution of a gene encoding the precursor of nisin. a small protein antibiotic.
Journal of Biological Chemistry. 263: 16260-16266.

Cabo ML, Herrera JR, Sampedro G and Pastoriza L. 2001. Effectiveness of C02
and Nisaplin on increasing shelf life of fresh pizza. Food Microbiology. 18: 489-
498.

Cagri A, Ustunol Z and Ryser ET. 2002. Inhibition of Three Pathogens on
Bologna and Summer Sausage Using Antimicrobial Edible Films. Journal of Food
Science. 67(6): 2317-2324.

Center for Disease Control and Prevention (CDC). 1999. Multi-state outbreak of
Iisteriosis. MMWR. 49: 1 129-1 130.

Center for Disease Control and Prevention (CDC). 1995. Outbreak of Salmonella
Serotype Typhimurium Infection Associated with Eating Raw Ground Beef—
Wisconsin. Morbidity and Mortality Weekly Report (MMWR). 44(49): 905-909.

Center for Food Safety and Applied Nutritions (CFSAN). 1999. Antimicrobial food
additives-guidance. US Food and Drug Administration.
Available at: http://www.cfsan.fda.gov/~dmslopa-antg.html

Center for Disease Control and Prevention (CDC). 2000. Multistate Outbreak of
Listeriosis -- United States. 2000. Morbidity and Mortality Weekly Report
(MMWR). 49(50):1129-1130.

Center for Disease Control and Prevention (CDC). 2002. Public Health Dispatch:
Outbreak of Listeriosis --- Northeastern United States. 2002. Morbidity and
Mortality Weekly Report (MMWR). 51(42): 950-951.

179

Center for Disease Control and Prevention. 2005a. Listeriosis. Division of
Bacterial and Mycotic Diseases.

Center for Disease Control and Prevention. 2005b. Food Associated with Listeria
monocytogenes infections are common at long-term care facilities in eight states.
Foodborne Diseases Active Surveillance Network.

Center for Disease Control and Prevention. 2006a. Escherichia coli O157:H7.
Division of Bacterial and Mycotic Diseases.

Centers for Disease Control and Prevention. 2006b. Outbreak of multidrug-
resistant Salmonella enterica serotype Typhimurium definitive type 104 infection
linked to commerical ground beef. Northeastern United States. 2003-2004.
Clinical Infectious Diseases. 42: 742-752.

Centers for Disease Control and Prevention. 2006c. Salmonellosis-outbreak
investigation. October 2006. Division of Bacterial and Mycotic Diseases.

Chickos JS and Acree WE. 2003. Enthalpies of Vaporization of Organic and
Organometallic Compounds. Journal of Physical and Chemical Reference Data.
56: 1880—2002.

Cleveland J, Montville TJ, Nes IF and Chikindas ML. 2001. Bacteriocins: safe.
natural antimicrobials for food preservation. International Journal of Food
Microbiology. 71(1): 1-20.

Clordisys Co.. 2003. What is chlorine dioxide. where is it used. how does it work?.
Available: http:I/www.clordisys.comlwh§tiscg.html

Code of Federal Regulations (CFR). 1999. Synthetic flavoring substance and
adjuvant (Title 21. 3. Section 172.515). Washington. Federal Printing House.

Collins-Thompson D and Hwang CA. 2000. Packaging with antimicrobial
properties. encyclopedia of food microbiology (edited by Robinson. R. L.. al.).
Academic Press. London. 416-420.

180

Cox LJ, Kleiss T, Cordier C and Cordellana P. 1989. Listeria spp. in food
processing. non food and domestic environments. Food Microbiology. 6: 49-61.

Cox NA. Juven BJ. Thomson JE, Mercuri AJ and Chew V. 1975. Spoilage odors
in poultry meat produced by pigmented and non-pigmented Pseudomonas.
Poultry Science. 54: 2001-2006.

Cox NA, Bailey JS and Ryser ET. 1999. Poultry as a source of campylobacter
and related organisms. Journal of Applied Microbiology. 90: 96-114.

Davidson PM, Sofos JN and Branen AL. 2005. Antimicrobials in food. Taylor &
Francis.

David C. Business Manager. Danisco. Copenhagen. Denmark (2004. October).
Personal conmunication via e-mail

Daniels JA, Krishnamurthi, R and Rizvi, SSH. 1985. A review of the effects of
carbon dioxide on microbial growth and food quality. Journal of Food Protection.
6: 532-537.

Dainty RH. 1996. Chemical/biochemical detection of spoilage. lntemational
Journal of Food Microbiology. 33: 19-34.

Delaquis PJ and Mazza G. 1995. Antimicrobial properties of Isothiocyanates in
Food preservation. Food Technology. 43: 73-84.

Delaquis PJ and Sholberg PL. 1997. Antimicrobial activity of gaseous allyl
isothiocyanate. Journal of Food Protection. 60 (8): 943-947.

Donnelle CW. 1990. Concern of microbial pathogens in associated with dairy
foods. Journal of Dairy Science. 73: 1656-1661.

Drobnaica L, Kristian P and Augusin J. 1977. The chemistry of the NSC group. in
the chemistry of cyanates and their thio derivates. John Wiley & Sons Inc.. New
York. 1003-1221.

181

 

Du J, Han Y and Linton RH. 2002. Inactivation by chlorine dioxide gas of Listeria
monocytogenes spotted onto different apple surfaces. Food Microbiology. 19:
481~490.

Dunnick JK, Prejean JD, Haseman J and Thompson RB. 1982. Carcinogenesis
bioassay of allyisothiocyanate. Fundamental and Applied Toxicology. 2: 114-120.

Elliott RP and Michener HD. 1961. microbiological standards and handling codes
for chilled and frozen foods- A review. Applied Microbiology. 9: 452-468.

Ellis M, Cooksey K, Dawson P, Han | and Vergano P. 2005 Quality of Fresh
Chicken Breasts Using a Combination of Modified Atmosphere Packaging and
Chlorine Dioxide Sachets. Journal of Food Protection. 69(8): 1991—1996.

Environmental Protection Agency (EPA). 1999. Section 4. Chlorine dioxide in
Alternative disinfectants and oxidant. EPA Guidance Manual.

Farber JM. 1991. Microbial aspects of modified atmosphere packaging
technology-A review. Journal of Food Protection. 54(1): 58~70.

Fisker N, Vinding K, Molbak K and Hornstrup MK. 2003. Clinical Review of
Nontyphoid Salmonella Infections from 1991 to 1999 in a Danish County. Clinical
Infectious Diseases. 37: 47—52.

Food and Drug Administration (FDA). 1992. Bad bug book. Salmonella spp..
Available at: www.dfsamlfda/~mow/chap1.html

Food and Drug Administration (FDA). Center for Food Safety & Applied Nutrition
(CSFAN). 1999. antimicrobial food additives-guidance.
Available at: http:llwww.cfsan.fda.gov/~dmslopa-antg.html

Food and Drug Administration (FDA). 2001. Agency Response Letter GRAS
Notice GRN 000062.
Available at: http:llwww.cfsan.fda.gov/~rdb/opa-gO62.html

182

Food and Drug Administration (FDA). 2004. Agency response letter GRAS notice
GRN 000144.
Available at: http:llwww.cfsan.fda.gov/~rdb/opa-g144.htm|

Food and Drug Administration (FDA). 2006. Agency response letter. GRAS
Notice GRN 0001333.
Available at: http:llwww.cfsan.fda.gov/~rdb/opa-g133.htm|

Franke I, Wijma E and Bouma K. 2002. shelf life extension of pre-baked buns by
an ACTIVE PACKAGING ethanol emitter. Food Additives and Contaminants.
19(3): 314-322.

Galeano B, Korff E and Nicholson WL. 2003. Inactivation of vegetative cells. but
not spores. of Bacillus anthracis. B. cereus. and B. subtilis on stainless steel .
surfaces coated with an antimicrobial silver and zinc containing zeolite i
formulation. Applied and Environmental microbiology. 69(7): 4329-4331.

 

Global lnforrnation Inc. 2005. US poultry market research. trend. analysis. Mintel
lntemational Group Inc.

Greengrass J. 1993. Films for MAP foods. in principles and application of
modified Atmosphere Packaging of foods (ed. Parry R T). Blackie Academic and
Professional. 63-100.

Greene J. 1998. US. Red Meat 8. Poultry Markets in a Global Setting.
Agricultural Outlook. USDA. 10-12.

Ha JU, Kim YM and Lee DS. 2001. Multilayered antimicrobial polyethylene films
applied to the packaged ground beef. Packaging Technology and Science. 15:
55-62.

Han JH and Floros JD. 1997. Casting antimicrobial packaging films and
measuring their physical properties and antimicrobial activity. Journal of Plastic
Film 8. Sheeting. 13: 287-298.

Han JH. 2000. Antimicrobial food packaging. Food Technology. 54(3): 56~65.

183

Han Y, Linton RH, Nielsen SS and Nelson PE. 2001. Reduction of Listeria
monocytogenes on green peppers by gaseous and aqueous chlorine dioxide and
water washing and its growth at 7°C. Journal of Food Protection. 64: 1730~1738.

Hara K. 1970. Introduction of polar groups by liquid phase oxidation to
polyethylene. Studies on Adhesion of Polyethylene. 229(1): 4-7.

Hart CD, Mead GC and Norris AP. 1991. Effects of gaseous environment and
temperature on the storage behaviour of Iisteria monocytogenes on chicken
breast meat. Journal of Applied Bacteriology. 70: 40-46.

Hatzidimitriu S, Gilbert G and Loukakis G. 1987. Odor Barrier Properties of Multi-
Layer Packaging Films at Different Relative Humidities. Journal of Food Science.
52(2): 472—474.

lsshiki K, Tokuoka K, Rori R and Chiba S. 1992. Preliminary examination of allyl
isothiocyanate vapor for food preservation. Bioscience. Biotechnology and
Biochemistry. 56:1476-1477.

Jimenez SM, Salsi MS, Tiburzi MC and Rafaghelli RC. 1997. Spoilage microflora
in fresh chicken breast stored at 4C: influence of packaging methods. Journal of
Applied Microbiology. 83: 613-618.

Jong AR, Boumans H, Slaghek T, Van Veen J and Rijk R. 2005. Active and
intelligent packaging for food: is it the future?. Food Additives and Contaminants.
22(10): 975-979.

Kaye VS, Murray MB, Harrison D and Beuchat LR. 2005. Evaluation of gaseous
chlorine dioxide as a sanitizer for killing salmonella. Escherichia coli O157:H7.
Listeria monocytogenes. and yeasts and molds on fresh and fresh-cut produce.
Journal of Food Protection. 68(6): 1176-1187.

Kim YS, Ahn ES and Shin DH. 2002. Extension of shelf life by treatment with
Allyl isothiocyanate in combination with acetic acid on cooked rice. Journal of
Food Science. 67(1): 274~279.

184

 

Kourai H, Manabe Y and Yamada Y. 1994. Mode of bactericidal action of
zirconium phosphate ceramics containing silver ions in the crystal structure.
Journal of Antibacterial and Antifungal Agents. 22: 595-601.

Knight TD, Murano PS and Murano EA. 2001. Effect of chlorine dioxide releasing
packaging film on the objective. microbial. and sensory quality of beef. IFT
Annual Meeting. New Orleans. Louisiana.

Kruijf N, Van Beest M, Rijk R, Paseiro P, Sipilainen-Malm T and Meulenaer D.
2002. Active and intelligent packaging: applications and regulatory aspects. Food
Additives and Contaminations. 19: 144-162.

Lawlis TL and Fuller SL. 1990. Modified atmosphere packaging incorporating an
oxygen barrier shrink film. Food Technology. 44(6): 124-125.

Labuza TP and Breene WM. 1989. Application of active packaging for
improvement of shelf life and nutritional quality of fresh and extended shelf life
foods. Journal of Food Processing and Preservatives. 13(1): 1-69.

Lawlis TL and Fuller SL. 1990. Modified atmosphere packaging incorporating an
oxygen barrier shrink film. Food Technology. 44(6): 123-125.

Lawrie RA. 1985. Meat Science. Pergamon Press. 200—216.

Lenntech Co. 2006. Chlorine dioxide as a disinfectant.
Available : http:llwww.Ienntech.com/water-disinfection/disinfectants-chlorine-
dioxide.htm#Wat%202ijn%20de%20eigenschappen%20van%20chIoordioxide ?

Levland J. 2001. Bacteriocins: safe. natural antimicrobials for food preservation.
lntemational Journal of Food Microbiology. 71(1): 1-20.

Lee DS, Hwang Y1 and Cho SH. 1998. Developing antimicrobial packaging film
for curled lettuce and soy bean sprouts. Food Science and Biotechnology. 7(2):
1 17-121.

185

 

Lim LT and Tung MA. 1997. Vapor pressure of Allyl isothiocyanate and its
transport in PVDC/PVC copolymer packaging film. Journal of Food Science.
62(5): 1061-1066.

Lin CM, Kim J, Du WX and Wei CL. 2000. Bactericidal activity of isothiocyanate
against pathogens on fresh produce. Journal of Food Protection. 63(1): 35~30.

McMeekin TA and Thomas CJ. 1980. Microbiological problems associated with
refrigerated poultry. CSIRO Food Research Quarterly 40: 141-149.

Mead GC. 2004. Poultry meat processing and quality. Woodhead Publishing
Limitted.

Metiu H. 2004. Physical chemistry-Therrnodynamics. Taylor & Francis. 519-546.

Misko G and Rothschild KL. 2001. The regulation of the use of antimicrobials in
food packaging: a new ball game. an article of packaging law magazine.
Available at: http:llwww.packaginglaw.comfIndex_mf.cfrn?id=41

Montgomery JM. 1985. Water treatment principle and design. Consulting
engineers INC.. JMM.

Muthukumarasamy P, Han JH and Holley RA. 2003. Bactericidal effects of
Lactobacillus reuteri and allyl isothiocyanate on Escherichia coli O157:H7 in
refrigerated ground beef. Journal of Food Protection. 66 (11): 2038-2044.

Nathnac. 2005. Salmonella outbreak in Spain.
Available at: http:llwww.nathnac.org/travel/news/salmonella_spain_110805.htm

Ngoka DA and Froning GW. 1982. Effect of free struggle and pre-slaughter
excitement on color of turkey breast muscles. Poultry Science. 61: 2291-2293.

Nielsen PV and Rios R. 2000. Inhibition of fungal growth on bad by volatile
components from spices and herbs. and the possible application in active
packaging. with special emphasis on mustard essential oil. lntemational Journal
of Food Microbiology. 60: 219-229.

186

 

Occupation safety & health administration (OSHA). 2006. sampling & analytical
method.

Available at: Mtg/[WWWoshggomts/sltc/methods/partial/t-Qv2086-01-8305-ch/t-
Q2086-01-8305-chhtml

Ogilvy WS and Ayres JC. 1951. The effect of atmospheres containing carbon
dioxide in prolonging the storage life of cut-up chicken. Food Technology. 5: 97.

Ohta Y, Takatani K and Kawakishi S. 1995. Decompostion rate of allyl
isothiocyanate in aqueous solution. Bioscience Biotechnology and Biochemistry.
59(1): 102-103.

Ouattara B, Simard RE, Piette G and Begin A. 2000. Diffusion of acetic and
propionic acids from chitosan-based antimicrobial packaging films. Journal of
Food Science. 65(5): 768-773.

 

Ozdemir M, Yurteri CU and Sadikoglu H. 1999. Physical polymer surface
modification methods and applications in food packaging polymers. Food
Science and Nutrition. 39(5): 457-477.

Ozen BF, Mauer LJ and Floros JD. 2002. Effects of ozone exposure on the
structural. mechanical and barrier properties of select plastic packaging films.
Packaging Technology and Science. 15: 301-311.

Perchacek R, Velisek J and Hrabcova H. 1997. Decomposition products of allyl
isothiocyanate in aqueous solutions. Journal of Agricultural Food Chemistry. 45:
4584-4588.

Phillips C. 1996. Review: Modified atmosphere packaging and its effects on the
microbiological quality and safety of produce. International Journal of Food
Science and Technology. 31: 463-479.

Quintavalla S and Vicini L. 2002. Antimicrobial food packaging in meat industry.
Meat Science. 62: 373~380.

Quio M, Fletcher DL, Northcutt JK and Smith DP. 2002.The relationship between
raw broiler breast meat color and composition. Poultry Science. 81: 422-427.

187

Radomir P, Velisek J and Hrabcova H. 1997. Decomposition products of allyl
isthiocyanate in aqueous solutions. Journal of Food Chemistry. 45: 4584-4588.

Razumovskii SD. 1983. Degradation and protection of polymeric materials in
ozone. Developments in polymer stabilization. 6: 239-293.

Rodrigues ET and Han JH. 2000. Antimicrobial whey protein films against

spoilage and pathogenic bacteria. proceedings of the IFT annual meeting; Dallas.

Tex. Institute of Food Technologists.
Sams AR. 2001. Poultry meat procession. CRC Press. New York.

Sander EH and $00 HM. 1978. Increasing shelf life by carbon dioxide treatment
and low temperature storage of bulk pack fresh chickens packaged in nylon
Surlyn film. Journal of Food Science. 43: 1519-1534.

Sekiyama Y, Mizukamin Y, Takada A and Numata S. 1994. Vapor pressure and
stability of allyl isothiocyanate. Journal of Food Hygiene and Society of Japan.
35(4): 365-370.

Sekiyama Y, Mizukami Y, Takada A and Oosono M. 1996. Effect of mustard
extract vapor on fungi and spore-forming bacteria. Journal of Antibacteria and
Antifungus Agents. 24(3): 171-178.

Singh N, Singh RK, Bhunia AK and Stroshine RL. 2002. Efficacy of Chlorine
Dioxide. Ozone. and Thyme Essential Oil or a Sequential Washing in Killing
Escherichia coli O157:H7 on Lettuce and Baby Carrots. Lebensmittel-
WIssenschaft und-Technologie. 35: 720~729. '

Siragusa GR, Cutter CN and WiIIett JL. 1999. Incorporation of bacteriocin in
plastic retains activity and inhibits surface growth of bacteria on meat. Food
Microbiology. 16(3): 229-239.

Smith JP, Ooraikul B, Koersen WJ and Van De Voort FR. 1987. Shelf life
extension of a bakery product using ethanol vapor. Food Microbiology. 4(4): 329-
337.

188

 

 

Speronello B. Associate Researcher. Engelhard. Iselin. New Jersey. (2005.
Feburary). Personal communication via e-mail.

Suhr and Nielsen. 2003. Antifungal activity of essential oils evaluated by two
different application techniques against rye bread spoilage fungi. Journal of
applied microbiology. 94: 665-674.

Suppakul P, Miltz J, Sonneveld K and Bigger SW. 2003. Active packaging
technologies with an emphasis on antimicrobial packaging and its applications.
Journal of food science. 68(2): 408-420.

Tsobkallo ES, Petrova LN and Khagen V. 1988. Influence of Ozone on the ,
Structure and Mechanical Properties of PE Film. International Polymer Science
and Technology. 15: 42-44.

US. Patent and Trademark Office. 1994. Chlorine dioxide generating polymer
packaging films. Patent 5360609.

Verrneiren L, Devlieghere F and Debevere J. 2002. Effectiveness of some recent
antimicrobial packaging concepts. Food Additives and Contaminants. 19: 163-
171.

Ward SM. 1998. Inhibition of spoilage and pathogenic bacteria on agar and pre-
cooked roast beef by volatile horseradish distillates. Food Research lntemational.
31(1): 19-26.

Wellinghoff ST. 1995. Keeping food fresh longer. Technology Today Magazine.
Available at: http:llwww.swri.edu/3pubslttoday/summer95/keepfood.htm

Weng YM and Hotchkiss JH. 1992. Inhibition of surface molds on cheese by
polyethylene film containing the antimycotic imazalil. Journal of Food Protection.
55(5): 367-369.

Weng YM, Chen MJ and Chen W. 1999. Antimicrobial food packaging materials
from poly(ethy|ene-co-methacrylic acid. Lebensmittel-Wissenschaft und-
Technologie. 32(4): 191-195.

189

 

Werner SB, Allad J and Ager EA. 1969. Salmonellosis from chickens prepared in
commercial rotisseries: report of an outbreak. American Journal of Epidemiology .
90(5): 429-437.

VVInther M and Nielsen PV. 2006. Active packaging of cheese with allyl
isothiocyanate. an alternative to modified atmosphere packaging. Journal of
Food Protection. 69(10): 2430-2435.

Wolfe SK. 1980. Use of CO- and CO2- enriched atmospheres for meats. fish. and
produce. Food Technology. 24: 55-58.

Yang CC and Chen TC. 1993. Effect of refrigerated storage. pH adjustment. and

marinade on color of raw and microwave cooked chicken meat. Poultry Science.
72: 355-362.

190

   

111111111rill