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This is to certify that the

thesis entitled

FRUIT RESPIRATION AND DETERMINATION OF LOW OXYGEN LIMITS

FOR APPLE (MALUS DOMESTICA, BORKH.) FRUIT
presented by

Christopher David Gran

has been accepted towards fulfillment
of the requirements for

M.S. d . Horticulture
egree 1n

 

 

K536) professor ’7

Date gig/‘5

0-7639 MS U is an Affirmative Action/Equal Opportunity Institution

 

 

PLACE IN RETURN BOX to remove this checkout from your record.
TO AVOID FINES return on or before due due.

  

DATE DUE DATE DUE DATE DUE

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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MSU to An Affirmative Action/Equal Opportmity InetItution
cmmwt

 

 

FRUIT RESPIRATION AND DETERMINATION OF LOW OXYGEN LIMITS
FOR APPLE (MALES DQMESJICA, BORKH.) FRUIT

By

Christopher David Gran

A THESIS

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

MASTER OF SCIENCE

Department of Horticulture

1993

ABSTRACT

FRUIT RESPIRATION AND DETERMINATION OF LOW OXYGEN LIMITS
FOR APPLE (W5 DQMESDQA. BORKH.) FRUIT

By

Christopher David Gran

Low oxygen (02) storage of apple (Malus domestica, Borkh.) fruit is an
important practice which slows respiration rates and maintains fruit quality longer
than air storage. The experiments conducted employed modified atmosphere
packaging (MAP) techniques to determine low 02 limits of commercially stored
apple cultivars over a range of temperatures. The low 0; limit was determined
from fruit respiratory quotient (RQ) breakpoint and level of ethanol (EtOH) in
package headspace. Generally, as temperature increased, the low 0; limit
increased, however the extent of the effect was cultivar-dependent. C02 removal
from package headspaces did not alter low 02 limits. RQ breakpoint and
elevated headspace EtOH were found to be useful tools for determining low 02
limits. Low 02 limits determined by elevated EtOH levels are in agreement with
to recommended storage atmospheres, while values of low 02 limits determined

by the RQ breakpoint were slightly lower.

Capyright by
CHRISTOPHER DAVID GRAN

1993

For Victor L. Leidel

1919-1988

If I had a choice
I’d break it
bury my memory
bones
wander off into the sea
and cast my lonely face
towards home
no crashing screams
no faded shore
I know too well the
nature balanced score

iv

ACKNOWLEDGMENTS

The author cares to express his appreciation for the opportunities and
assistance provided by Dr. R. M. Beaudry throughout the course of the work
presented within this volume. I also care to thank Dr. D.R. Dilley and Dr. K.L.
Poff for encouragement in addition to thesis review and service on my guidance
committee.

I would like to personally thank those who have assisted my academic and
scientific efforts throughout my tenure in the Postharvest Lab. Your assistance
with experiments and studies, critical review of various manuscripts, and
friendship was invaluable. I can only hope that I have given as much as I have
received.

To Sandra K. Ruthsatz, for selfless assistance. To Mikki for critical thesis
review in addition to personal encouragement, support, and friendship. To

Mother, for everything. Thank you all.

Guidance Committee:

The journal paper format was chosen for this thesis in accordance with
departmental and university regulations. The thesis is divided into 3 chapters in
which the first has been accepted for publication in Postharvest Biology and
Technology. The second and third chapters have been prepared according to
format requirements for Postharvest Biology and Technology but have not been

submitted.

TABLE OF CONTENTS

Page
LIST OF TABLES ......................................... ix-x
LIST OF FIGURES ........................................ xi-xv
LIST OF SYMBOLS AND ABBREVIATIONS ................... xvi-xvii
LIST OF BOTANICAL NAMES ............................... xviii
LITERATURE REVIEW .................................... 1-21
REFERENCES ............................................ 22-25
CHAPTER 1:
Determination of the Low Oxygen Limit for Several Commercial Apple Oiltivars
by Respiratory Quotient Breakpoint ............................. 26
ABSTRACT ............................................ 27
INTRODUCTION ....................................... 27-29
MATERIALS AND METHODS ............................. 29-31
RESULTS AND DISCUSSION ............................. 31-39
REFERENCES ......................................... 40-41
CHAPTER 2:

Modified Atmosphere Packaging of Apple: The Effect of Temperature on the
Lower Oxygen Limit as Determined by Respiratory Quotient Breakpoint . . 42

ABSTRACT ............................................ 43
INTRODUCTION ....................................... 43-45
MATERIALS AND METHODS ............................ 45-47
RESULTS AND DISCUSSION ............................. 47-65
REFERENCES ......................................... 66-67

vii

TABLE OF CONTENTS (cont.)

Page
CHAPTER 3:
Modified Atmosphere Packaging of Apple: The Effect of Temperature on the
Lower Oxygen limit as Determined by Headspace Ethanol Accumulation . 68

ABSTRACT ............................................ 69
INTRODUCTION ....................................... 69-72
MATERIALS AND METHODS ............................ 72-74
RESULTS AND DISCUSSION ............................. 74-88
REFERENCES ......................................... 89-90
Summary and Conclusions .................................... 91-97
REFERENCES ......................................... 98
APPENDIX A ............................................. 99
C02 Production for 8 Cultivars of Apple Fruit at O.1°C. ............. 1.00-101
APPENDIX B ............................................. 102

The Effects of Removing Carbon Dioxide from Headspaces of Modified
Atmosphere Packages on Lower Oxygen Limits .................... 103-116

viii

LIST OF TABLES

Table Page

Chapter 1

. General equations and values of constants describing the relationship between

steady state 0; partial pressures and 02 uptake for apple cultivars stored at

0.1 °C. ................................................. 37
Chapter 2

. Estimated low 02 limit ranges for apple fruit of 3 cultivars based upon RQ
breakpoint. ............................................. 52

. General equations and values of constants describing the relationship between
steady state 02 partial pressures and 02 uptake for 3 apple cultivars in

modified atmosphere packages. .............................. 61
Chapter 3
. Estimated low 02 limits for apple fruit of 3 cultivars indicated by elevated
headspace EtOH levels. ................................... 80
APPENDIX B

. General equations and values of constants describing the relationship between
steady-state and 0, partial pressures and 02 uptake for 3 apple cultivars stored
at a range of temperatures. ................................. 110

. General equations and values of constants describing the relationship between
steady state 02 partial pressures and 02 uptake for ‘Golden Delicious’ apple
fruit over a range of temperatures with CO2 removed from package
headspaces. ............................................. 112

. Estimated low 02 limits, as indicated by elevated headspace EtOH, of ‘Golden
Delicious’ apple fruit over a range of temperatures with C02 removed from

packages. ............................................... 112

LIST OF TABLES (cont.)

Table Page

APPENDIX B (cont.)

4. General equations and values of constants describing the relationship between
steady state 0, partial pressure and 02 uptake for ‘Marshall McIntosh’ apple
fruit over a range of temperatures with CO, removed from package
headspaces. ............................................. 115

5. Estimated low 02 limits, as indicated by elevated headspace EtOH, of
‘Marshall McIntosh’ apple fruit over a range of temperatures with C02
removed from package headspaces. ........................... 115

LIST OF FIGURES

Figure Page
Literature Review
. Transport models for lipid membranes. ........................ 4
. Structure of apple skin modeling the difference in cell structure and
intercellular air spaces with changes in relative humidity. ........... 9
Chapter 1
. Effect of steady state 02 partial pressures on the respiratory quotient (R0) of
9 cultivars of apple fruit held at O.1°C. ......................... 33
. Effect of steady state 02 partial pressures on the 02 uptake of 9 cultivars of
apple fruit held at O.1°C. .................................. 35
. Effect of steady state 02 partial pressures on the respiratory quotient (R0)
and headspace EtOH of ‘Law Rome’ apple fruit held at O.1°C. ...... 36
Chapter 2

. Effect of temperature on the respiratory quotient (R0) and the RQ
breakpoint of ‘Marshall McIntosh’ apple fruit. ................... 48

. Effect of temperature on the respiratory quotient (R0) and the RQ
breakpoint of ‘Redmax McIntosh’ apple fruit. ................... 49

. Effect of temperature on the respiratory quotient (R0) and the RQ
breakpoint of ‘Golden Delicious’ apple fruit. .................... 50

. Effect of temperature on the respiratory quotient (R0) and the RQ
breakpoint of ‘Early Red One Red Delicious’ apple fruit. .......... 51

. Low 0, limit boundaries for 3 apple cultivars over a range of temperatures as
determined by RQ breakpoint. .............................. 52

LIST OF FIGURES (cont.)
Figure Page
Chapter 2 (cont.)

6. Effect of temperature and [02],... on RRcoz of ‘Marshall McIntosh’ apple
fruit. .................................................. 55

7. Effect of temperature and [02118: on RRcoz of ‘Redmax McIntosh’ apple
fruit. .................................................. 56

8. Effect of temperature and [02],.“ on RRCOZ of ‘Golden Delicious’ apple
fruit. ................................................. 57

9. Effect of temperature and [02],,“ on RR€02 of ‘Early Red One Red Delicious’
apple fruit. ............................................. 58

10. Comparison of 02 uptake curves for ‘Marshall McIntosh’ apple fruit over a
range of temperatures. ..................................... 59

11. Comparison of 02 uptake curves for ‘Redmax McIntosh’ apple fruit over a
range of temperatures. ..................................... 59

12. Comparison of 02 uptake curves for ‘Golden Delicious’ apple fruit over a
range of temperatures. ..................................... 60

13. Comparison of 02 uptake curves for ‘Early Red One Red Delicious’ apple
fruit over a range of temperatures. ........................... 6O

14. Apparent whole fruit K. for 02 of ‘Marshall McIntosh’ over a range of
temperatures, with and without scrubbing CO; from package headspaces. 63

15. Apparent whole fruit K. for O; of ‘Golden Delicious’ over a range of
temperatures, with and without scrubbing CO, from package headspaces. 63

xii

LIST OF FIGURES (cont.)

Figure Page

Chapter 3

. Effect of temperature on headspace EtOH levels as an indication of the low

0, limit of ‘Marshall McIntosh’ apple fruit. ..................... 76
. Effect of temperature on headspace EtOH levels as an indication of the low
0, limit of ‘Redmax McIntosh’ apple fruit. ..................... 77
. Effect of temperature on headspace EtOH levels as an indication of the low
0, limit of ‘Golden Delicious’ apple fruit. ...................... 78
. Effect of temperature on headspace EtOH levels as an indication of the low
0; limit of ‘Early Red One Red Delicious’ apple fruit. ............ 79
. Low 0; limit boundaries for 3 apple cultivars over a range of temperatures as
determined by elevated [EtOH]m. ........................... 80
. Correlation of headspace EtOH levels with macerated tissue EtOH
levels. ................................................. 81
. The accumulation of EtOH with time in the headspace of packages at steady-
state respiration over a range of 02 partial pressures. ............. 82
APPENDIX A
. Effect of steady state 0; partial pressure on C0, production of 8 cultivars of
apple fruit at O.1°C. ....................................... 101
APPENDIX B

. O, uptake and predicted 02 uptake for ‘Golden Delicious’ apple fruit with C02
removed from package headspaces, over a range of temperatures. . . . . 104

xiii

LIST OF FIGURES (cont.)

Figure Page

APPENDIX B (cont.)

. 02 uptake and predicted 02 uptake for ‘Golden Delicious’ apple fruit over a
range of temperatures. .................................... 104

. O, uptake and predicted O, uptake for ‘Marshall McIntosh’ apple fruit with
C02 removed from package headspaces, over a range of temperatures. 105

. O, uptake and predicted 02 uptake for ‘Marshall McIntosh’ apple fruit over a
range of temperatures. .................................... 105

. Effect of temperature and 02 partial pressure on RRom of ‘Marshall McIntosh’
apple fruit. ............................................. 106

. Effect of temperature and 0, partial pressure on RRcoz of ‘Redmax McIntosh’
apple fruit. ............................................. 107

. Effect of temperature and 0; partial pressure on RRC02 of ‘Golden Delicious’
apple fruit. ............................................. 108

. Effect of temperature and 0; partial pressure on RRcoz of ‘Early Red One
Red Delicious’ apple fruit. .................................. 109

. Effect of steady state 0; partial pressure on the 02 uptake of ‘Golden
Delicious’ apple fruit over a range of temperatures with CO, removed from
package headspaces. ...................................... 111

10. Effect of steady state 0; partial pressure on the low 0, limit of ‘Golden

Delicious’ apple fruit over a range of temperatures with C02 removed from
package headspaces. ...................................... 113

11. Effect of steady state 0; partial pressure on the O, uptake of ‘Marshall

McIntosh’ apple fruit over a range of temperatures with C02 removed from
package headspaces. ...................................... 114

xiv

LIST OF FIGURES (cont.)
Figure Page
APPENDIX B (cont.)
12. Effect of steady state 02 partial pressure on the low 0, limit of ‘Marshall

McIntosh’ apple fruit over a range of temperatures with C02 removed from
package headspaces. ...................................... 116

atm

CA

02H.
Ca(0H)2

[C02]...
Icozim
EtOH
[E10H],,,

LIST OF SYMBOLS AND ABBREVIATIONS

surface area

atmosphere

centimeter

celsius

controlled-atmosphere

ethylene (ethene)

hydrated lime

carbon dioxide

atmospheric carbon dioxide partial pressure
package carbon dioxide partial pressure
ethanol (ethyl alcohol)

package ethanol level

gram

hour

water

kilogram

kilopascal

low density polyethylene
modified-atmosphere

modified-atmosphere packaging

minute

millimole

oxygen

atmospheric oxygen partial pressure
package oxygen partial pressure

parts per million (pl/l)

partial pressure of carbon dioxide

partial pressure of oxygen

carbon dioxide permeability coefficient
oxygen permeability coefficient

water vapor permeability coefficient
relative humidity

respiratory quotient

resistance (5 - cm") of tissue to gas transport
resistance (5 - cm“) of tissue to H20 vapor transport
resistance (5 - cm") of tissue to CO, transport

LIST OF SYMBOLS AND ABBREVIATIONS (cont.)

resistance (s - cm") of tissue to 0; transport
resistance (s - cm") of tissue to CJ-I. transport
rate of carbon dioxide production

rate of oxygen uptake

temperature (°C)

second

fruit weight

thickness of low density polyethylene
microliter

LIST OF BOTANICAL NAMES

Common name

Apple
Avocado

Banana
Blueberry
Cantaloupe
Citrus
Grapefruit
Mango
Onion
Orange
Peach
Pear
Pumpkin
Strawberry

Tomato

Botanical name

Malus domestica, Borkh.
Persea americana, Mill.
Musa, L. sp.

Vaccinium corymbosum sp.
Cucumis melo, L.

Citrus, L. sp.

Citrus paradisi, Macf.
Magtfera indica, L.

Allium sp.

Citrus sinensis, L. Osbeck
Prunus pension, L.

Pyms communicus, L.
Cucwbita pepo, L.
Fragaria x ananassa, Duch.

Lycopersicon esculentum, Mill.

LITERATURE REVIEW

2

Fruit respiration, fruit storage, the internal atmospheres of fruit, and gas
movement through bulky tissues have interested plant scientists for over 100 years.
The movement of water vapor (H20), oxygen (02), carbon dioxide (C02), and
ethylene (C211,) through fruit cuticles has significant implications for the
respiratory activity, ripening, and ultimately, the storability of bulky plant tissues.
The purpose of this literature review is to investigate specific paths of gas
movement in fruits and the physiology of low 0, storage of fruits.

'h'anspirational vapor movement.

The characteristics of H20 vapor movement through bulky plant tissues varies
from the other biologically active gases (ie. 02, C02, and C,H.). Resistance to
H20 transport (r,) in ‘Valencia’ oranges (Citrus sinensis, L. Osbeck) has been
measured at 110 s ~cm", compared with transport resistance values of 5700, 6000,
and 6900 s ~cm" for CO, (rem), O2 (rm), and QIL (ram), respectively (Ben-
Yehoshua, et al., 1985).

Pieniazek (1944), examining apple fruit transpiration, found no correlation
between cuticle thickness and rate of transpiration. Blocking of lenticels with
paraffin reduced transpirational weight loss by 8 to 25%, while the removal of the
wax coating from the cuticle using cheesecloth resulted in a 7% to 88% increase
in transpiration rate. Increases in transpiration rates as low as 7% indicate
incomplete removal of the wax coating.

Horrocks (1964), in his study of the waxes of apple fruits, concluded "...the

cuticular wax is a prime factor determining the impedance to water vapour of an

3

apple cuticle. It is possible that the density or geometry of the lenticels is
important or it may be that the distribution or type of wax plays the dominant
part". In addition, H20 vapor permeability (P',) of apple tissue was found to be
similar to that for isolated peels.

Ben-Yehoshua et al., (1985), using a scanning electron microscope (SEM),
showed waxing to partially or completely plug open stomatal pores of ‘Valencia’
oranges, increasing the resistance to the transport of C02, 02, and 02H. by 140,
250, and 100%, respectively, while only increasing the rv by 25%. Conversely,
sealing individual fruits within high density polyethylene (HDPE) increased r, by
1400%, while increasing r002, rm, and ram by 72, 230, and 25 %, respectively.
Results indicate H20 vapor moves through the cuticle by a pathway different from
that of C02, 02, and QIL.

Schénherr (1976), examining the influence of cuticular waxes on P2,, found
diffusion for H20 through the cuticle membrane to be completely determined by
the permeability coefficient of the cuticular waxes for citrus, pear (Pynts
communicus, L.) and onion (Allium, sp.). Schénherr and Schmidt (1979) found P',
to be independent of cuticle thickness. Further studies which consisted of the
extraction of cuticular waxes increased P', by 300- to 500-fold without cuticle
thickness alterations of equal magnitude. Cuticular resistance is not a function of
thickness, and therefore for isolated cuticles H20 vapor movement does not obey
Fick’s Law of Diffusion, but is a function of the diffusional matrix and the

diffusion coefficient resulting from cuticular composition.

4
Schénherr and Schmidt (1979) next tested the hypothesis that vapor pressure

deficits alter cuticular permeability coefficients. Using isolated cuticles of citrus
they determined P',, transpiration (Ju), and r,, and found that each varied with
both pH and H20 vapor activity (am), where aw=relative humidity (RH)/ 100.
Based upon the observations of both citrus and artificial membranes, two models
were developed for membrane transport (Sch6nherr and Schmidt, 1979).

Model I is a porous membrane, which consists of a lipid matrix containing
HZO-filled pores (Figure 1). These pores are such that pore length exceeds
membrane thickness, and they form as a result of hydration of polar functional
groups within the membrane matrix. Varying a“, will both increase the driving

force of diffusion and the flux of H20 vapor, reducing the H20 content of the

 

  

a): a widget):

  

Model 1' Model 11

 

 

 

 

Figure 1. Transport models for lipid membranes. Model I. Porous membrane.
Model 11. Solubility membrane. Adapted from Schénherr and Schmidt, 1979.

5

membrane and, therefore, the P', for the membrane. The effect of aw, upon the

diffusion coefficient can be described by:

I": D-n-n-r (1)

 

where P', is directly proportional to D, the diffusion coefficient (SchOnherr and
Schmidt, 1979). The relationship of the number of pores (n), and the square of
the pore radius (r), are inversely proportional to diffusional path length (9). The
driving force for diffusion is the sum of the vapor pressure gradients for each
given gas. Experimental results indicated that for cuticles, pore number increased
with increasing pH, while pore diameter was independent of pH (Schénherr,
1976). This model is believed to represent gas phase movement in apples of 02,
C02, and 0,11,, but not H20 vapor, with C02, 02, and QIL gas movement
facilitated by residually open lenticels.

Model II represents a solubility membrane, in which the molecules of the
membrane are tightly packed and no pores develop which traverse the membrane.
H20 molecules move through the cuticle independently, and this movement is
only accomplished with the breaking of all H bonds between neighboring H20
molecules (Figure 2). Molecular movement through the membrane occurs in the
gas phase, with transition to the gas phase occurring at the membrane / liquid
phase interface, or the interior surface of the membrane structure. Because of

the semi-closed nature of this system, diffusion and the permeability coefficient

6

are constant and independent of a“, the H20 vapor activity. The driving force for
movement across the membrane is the partial pressure gradient for H20 across
the membrane.

The driving forces for H20 vapor movement across the membrane in the two
models differ in that Model I, the porous membrane, the driving force is the
difference in H20 potential between the inner and outer surface of the
membrane, while in Model 11 the driving force is the partial pressure of H20
vapor differences across the membrane. Data indicate that Model 11 is
representative of H20 vapor diffusion and transpirational resistance (Burg, 1990).
This conclusion is based upon determinations of transpirational resistance for
fruits ranging from 33 to 200 s - cm“, compared with cuticular resistance to water
of 30 to 290 s -cm'1 for the adaxial surfaces of hypostomatous leaves (Ben-
Yehoshua et al., 1985). Because
r, is not a function of cuticle thickness for
isolated cuticles (Schénherr and Schmidt, 1979), a change in the resistance to gas
exchange must occur with the isolation process.

In studies of transpirational H20 loss and the openness of lenticels, apple
(Malus domestica, Borkh,) peels stored under increased RH had less resistance to
H20 vapor loss (Anon., 1953). Models developed by Fockens and Meffert (1972)
show loss of mass due to evaporation depends upon the cuticular resistance of the
fruit, with cell shape and intercellular space affecting r,. High RH results in H20

uptake by the cell, circular shaped cells, increased intercellular spaces, and

7

decreased diffusional resistance (r) of skin tissue, and visa versa. SchOnherr and
Schmidt (1979) have shown that transpirational H20 loss, as well as gas phase
diffusion, depended upon the barrier matrix and phase of transfer inherent for
each tissue system.

The barriers to gas movement.

Maintaining 02 levels sufficient for aerobic respiration in tissues at the center
of fruit tissues, especially large fruits, poses an interesting problem. What are the
driving forces for gas movement into and out of fruit tissues? What is the
behavior of gas movement through fruit tissues, and what factors determine gas
movement behavior? Are gas exchange characteristics static through the stages of
maturation?

When examining a fruit, the most obvious barrier to gas exchange is the
epidermis, or skin, of the fruit. Kidd and West (1934) hypothesized that cuticle
permeability influenced fruit respiration during controlled-atmosphere (CA)
storage. Pcm and P02 values were determined ”...by dividing at corresponding
times the rate of carbon dioxide-production by the concentration of carbon
dioxide and by the oxygen deficit in the internal atmosphere respectively."

(Kidd and West, 1949). Examination of the influence of apple skin permeability
upon respiration determined low PC02 decreased respiration rates. Apple cultivars
varied in storage life, internal gas compositions, and susceptibility to storage
damage. Time course data indicated cuticle P002 and P02 did not change

significantly through the climacteric, although after 60 days storage at 12° C

8

cuticular r02 increased more rapidly and to a greater extent than rem. Similar
increases in ro2 in comparison with r002 were noted by Trout et al., (1942). For all
cultivars examined, permeability to P'02 was less than P'Coz. Differences in F02
and P'cm within the fruit may directly influence storage disorders in CA storage.

Another factor thought to influence tissue permeabilities is tissue cell
structure. Fockens and Meffert ( 1972), demonstrated that changes in cell
structure were correlated with changes in RH. This underscores the importance
of cell structure in relation to diffusional resistance. Increased RH results in H20
uptake by the cell, swollen cells, and larger intercellular airspaces, and vise versa
(Figure 2). Diffusional resistance is high when intercellular space is reduced and
cells have a reduced cross-sectional width. Park (1990) has also shown a change
in the gas exchange characteristics of fruit through development with variations
thought to be directly influenced by the collapse of stomatal guard cells and the
blocking of skin pores.

Trout et al., (1942) tested the hypothesis that diffusional resistance of the
fleshy tissues is negligible compared to cuticular resistance. Removal of the apple
fruit’s skin, 0, levels increased from 6% to 20% while CO, levels decreased from
7.2% to 2.4%. Similar changes in Oz and C02 concentrations were seen when
small sections of the peel were removed. Based upon these results, Trout et al.,
concluded that, for apples, determination of the cuticular resistance to diffusion is
an accurate measure of a fruit’s total diffusional resistance.

Biochemical analyses of cuticles have determined their structure to be a

 

 

 

 

 

 

 

 

High Relative Humidity Low Relative Humidity
Low Diffusional Resistance Diffusional Resistance High

 

 

 

 

Figure 2. Structure of apple skin modeling the differences in cell structure and
intercellular air spaces with changes in relative humidity. Adapted from Fockens
and Meffert, 1972, J. Fd. Sci. Agric.

polymer membrane consisting of a cutin polymer matrix. This matrix contains
imbedded lipids which are often referred to as waxes (Schénherr and Bukovac,
1973). These layers of lipids are oriented in a fashion parallel to the surface. P’m,
P’om, and P'am may also change when the composition of the cuticle, or waxy
layer, changes.

Gas movement through bulky fruit tissues.

Burg and Burg (1965) investigated gas movement and asked whether diffusion

of gases through fruit tissues obeys Fick’s First Law, which may be represented as:

10

’x'A'Dt'ww‘Cmm) (2)
T

fli =

dt
where ds/dt is the rate of transport, D is the diffusion coefficient, Chm“ and
Cm the concentrations of gas within and outside the fruit, respectively, T the
effective thickness of the barrier to diffusion, ‘A the surface area of the fruit, and
x the fraction of the surface area through which gas exchange occurs (value from
0-1). This relationship will hold when the gases are at steady state. From this
formula, a value r, the resistance coefficient for the barrier to a specific gas, can
be generated. For a gas such a C02, which has an approximate Cm,“ value of 0,

Fick’s Law can be reduced to:

 

CW“ = ____'T - r (3)

dud: x-AOD-

Note that the resistance factor, r, when Cw has a value of 0, divided by the
rate of transport (ds/dt), is the inverse of the permeability factor used by Kidd
and West (1934) in their determination of gas exchange rates, analogous to the
relationship of resistance and conductance for the flow of current.

Diffusion of C02 and (32H4 for preclimacteric tissues of cantaloupe (Cucumis
melo, L.) fruit (Lyons et al., 1962), bananas and mangoes (Magifera indica, L.)
were found to obey F ick’s First Law (Burg and Burg, 1962). Climacteric avocado

(Persea americana, Mill.) fruit (Ben-Yehoshua et al., 1963) stored apples (Trout et

11
al., 1942), and post-climacteric bananas (Leonard and Wardlaw, 1941), however,

do not. This indicates that a changed diffusional pathway or the barrier matrix to
diffusion alters diffusional resistance during the course of the climacteric or aging.
Cellular breakdown and the loss of cell turgor with cell senescence and the loss of
water by transpiration, which then results in reduced intercellular airspace for the
diffusional movement of gases into and out of the fruit, may be the cause of the
deviation in results.

Studies using avocado and peach (Prunus persica, L.) by Rodriguez et al.,
(1989) show the correlation between loss of flesh firmness and increased r02, rm,
and ram. The drastic changes in flesh texture and associated losses in
unobstructed intercellular pathways have direct influences on gas exchange
characteristics with fruit ripening. The dynamic nature of fruit maturation and
ripening further complicate the mechanisms of fruit gas exchange and respiration.

Increases in the matrix barrier resistance, which would occur with cell wall
softening, cellular membrane ruptures, and subsequent flooding of the
intercellular airspaces of tissues and the pathways of gas diffusion, would result in
an alteration in gas exchange properties. During flooding of intercellular air
spaces, gas movement through this liquid matrix is substantially slower for 02 and
(3H, when compared with CO, rates of diffusion. More importantly, flooding
would reduce levels of 02 available to internal cells because the diffusivity of O,
in water is 10‘ less than in air (Burg and Burg, 1965). Examinations of the

intercellular air space of apples have shown 30-35% of the total fruit volume

12

consists of intercellular gas (Smith, 1947). Although a significant percent of the
fruit’s volume is open space, minor losses of intercellular space will reduce the
continuous pathways for the unrestricted movement of gaseous Oz and CO,
through fruit tissues.

The rate limiting step in the outward passage of CO, and Czl-I4 gas occurs in a
media where both possess nearly identical diffusivities (Burg and Burg, 1965; Ben-
Yehoshua et al., 1985). On the basis of solubilities of C02, QIL, Oz, and water
vapor in lipids, the rates of diffusion should be H20 > (12H, > CO2 > 02.

In an 0, atmosphere enriched with N2 or He diffusion of QIL through an
apple peel mimics diffusion of (121-14 through pores in apple peels and paper of a
known diameter. The dependance of C02 and QH. gas exchange upon
atmospheric pressure and gas composition supports the theory that movement of
CO, and QIL through apple peels occurs through air-filled pores, with open
lenticels an obvious candidate for this passageway (Burg and Burg, 1965).

The driving force for gas movement of 02, C02, QIL, and N2 would be partial
pressure gradients, although the specific paths for gas movement differ.
Mathematical models, based upon Fick’s Law of Gas Diffusion, as well as other
models related to gas phase movement of 02, C02, H20, and 02H, have indicated
that water vapor moves preferentially through the cuticle of the fruit in the liquid
state, while other gases move preferentially through available pores of the tissue.
Partial pressure gradients for 02, C0,, and QIL gas allow for mass transfer of

these molecules through the tissue.

13
Tissue gas gradients.

Rajapaske et al., (1989) have found that 02 concentration gradients exist from
fruit centers to epidermal tissue. Gas gradients varied with cultivars and were
dependant upon the respiration rate and volume of intercellular airspace.
Solomos (1987) has also reported C02 gradients within apple tissues. The
existence of 02 and C02 gradients between tissue areas indicates that in certain
cases tissue resistance to diffusion may not be negligible, especially under
conditions of reduced atmospheric 02 levels, when the existence of 0; partial
pressures 0.1% below that maintained in the atmosphere may result in
fermentation and skin discoloration (Lau, 1989).

Pathways for gas movement.

Devaux (1891), after conducting experiments involving forcing gas through
submerged tissues, postulated that gases pass through both pore openings and
through the cuticle, with movement through the cuticle possible in both the free
and dissolved state. Devaux suggested movement of gases depended on both the
permeability and the porosity of the peridermic membrane, and that 02 enters
pumpkin (Cucurbita pepo, L.) fruit primarily through pores and C02 exits
primarily through the membrane. Although these experiments were conducted
well before determination of cuticle chemistry and the advent of gas
chromatography, it is very interesting that data from recent experiments examining
gas exchange support Devaux’s conclusions.

Burg and Burg (1965) concluded that gas exchange in apples and other fruits

14
was governed by Fick’s law of diffusion, with the peel of apple fruit providing the

primary resistance to gas exchange. CO, and QH. had similar diffusivities, with
exchange dependent upon atmospheric pressure and gas composition. The
diffusion barrier for C02 and QIL exchange was considered to consist of air filled
pores, with indirect measurements of pore size in agreement with the expected
size and number of lenticels present on the apple cuticle. Calculations of barrier
thickness using Fick’s Law generated estimated cuticle thicknesses similar with
actual measurements.

Examinations of stomatal density and responsiveness for banana (Musa sp.)
fruit have shown fruit stomatal density to be 30 times less than that for leaf
tissues, averaging approximately 450 stomates/cm’, with residual opening being
influenced positively by high RH conditions and light exposure treatments
(Johnson and Brun, 1966). Other anatomical studies indicate 1.8 lenticels/cm2 of
apple fruit cuticle (Clements, 1935), compared with 40,000 stomates/cm2 for apple
leaf tissue (Curtis and Clark, 1950). Consideration of these figures point to the
reduced avenues of gas phase diffusion in fruits relative to leaves. For citrus
(Citrus, L.) fruits, when diffusion coefficients are considered, residual stomatal
opening of less than 0.4% initial open pore area is sufficient to account for the
needed residual gas exchange through these means (Ben-Yehoshua et al., 1985).
Therefore, in certain fruits, gas exchange does not occur solely in the dissolved
state through the cuticle. Some gases move through pores in the cuticle.

Ben-Yehoshua et al., using oranges and grapefruit (Citrus paradisi, Macf.),

15

examined the effect of waxing and sealing of fi'uits with high density polyethylene
(HDPE) films and indicated mass transport of H20 vapor and other gases occurs
by different mechanisms. Values of ram, rm, and ram are similar for untreated
fruits, while the r, is 60-100 times less (Ben-Yehoshua et al., 1985). Waxing of
fruits inhibits C02, 02, and QIL transport but not H20 vapor, while wrapping
fruits with HDPE film mainly restricted H20 transport. Waxing is thought to
restrict non-H20 vapor transport by blocking of open stomates and lenticels, while
not effecting H20 vapor transport due to the tendency of the wax layer to pit and
crack. An increase of 1,400% in r, was seen with HDPE wrapping compared with
increases of 72, 230, and 25% for raw, rm, and ram, respectively. These results
arise from differences in the selective permeability of the fruit cuticle and the
barrier to transpirational H20 loss created by the film.

Cameron and Yang (1982) determined coefficients of diffusion for bulky plant
tissues by means of loading of ethane (C,H.) gas, allowing this system to come to
steady-state, and then measure gas efflux levels in order to determine specific
paths of gas movement and each pathway's diffusion coefficient. Cameron and
Yang determined 94, 81 and 67% of QIL, C02, and H20 vapor gas exchange in
tomato (Lycopersicon esculentum Mill., Cv. ‘Ace’) fruit occurred through the stem
scar, and that cuticular r, was 1,000 times less than r002, rm, and ram. The calyx
end of ‘Golden Delicious’ apple fruit account for 29, 24 and 2% of QIL, C02,
and H20 vapor exchange, respectively. On this basis Cameron and Yang

concluded that lenticular gas exchange is insignificant for total fruit gas exchange.

16
It should be noted, however, that the physical structure of ‘Golden Delicious’

apple fruit, used by Cameron and Yang, vary significantly when compared with
‘McIntosh’ apples, which were used in the studies of Burg and Burg (1965).
While the calyx end of ‘Golden Delicious’ is usually quite open and free to gas
exchange, this is often not the case, especially with ‘McIntosh’. It should also be
noted that for ‘Golden Delicious’ lenticels are numerous and raised, and the fruit
lack an apparent waxy bloom, which is quite opposite ‘McIntosh’ fruit.

Recent experiments by Park (1990), involving blocking of the lenticels of
’McIntosh’ apples also support Burg and Burg’s conclusion that for apple fruit,
lenticels are the primary pathway for gas exchange, excluding water vapor
exchange. Park also indicated that resistance to gas exchange is a factor of
lenticel structure, not strictly related to lenticel density.

Respiration and low oxygen tolerance of apple fruits.

Rate of respiration affects storage life, value, and quality of stored fruits. The
rate at which stored carbohydrates are converted by respiration to CO; may
influence the length the stored fruits maintain marketable quality. In the study of
low 0, storage and the low 0, limits of fruits, the influence of temperature, RH,
and the influence of elevated CO, levels upon respiration are very important
because of their abilities to alter respiratory activity and gas movement.
Knowledge of these factors is vital to understanding those factors which affect
respiration and the internal gas composition of the fruit present for respiration.

The fruit respiration studies initiated by Kidd and West, and continued by

17
Fidler and North, at the Low Temperature Research Station at Cambridge,

England established many of the fundamental characteristics of climacteric fruit
respiration, testing the influence of temperature, RH, and atmosphere
composition on respiratory rates.

Decreased storage temperatures are known to lower respiration rates, slow
metabolic processes in tissues, and maintain marketable value and edible
qualities. With low temperature or high CO, damage (Fidler and North, 1964)
respiration rates increase in comparison with apples at the same temperature.
Respiratory quotients (RQ) increased with decreasing temperatures, from 1.3-1.4
at 38°C to 1.6-1.7 at 32°C (Fidler and North, 1966). Lower temperatures (0°-
3.5°C) alter the respiratory substrate from the expected 1:1 ratio of carbon (C)
lost to C evolved (Fidler and North, 1968).

Kidd and West (1933) have shown respiration rates of apple tissue can be
increased with elevated 0, partial pressures (P02) (SO-100%) or decreased with
lowered P02 (5%). Elevated C02 partial pressures (Pm)(5-10%) delayed the
respiratory climacteric. With further PC02 increases, continued decreases in
respiration rates were observed.

Hardy (1949) found internal P002 concentrations in apple fruit vary with
temperature, from 1.7% to 13.2% for 0° to 25°C respectively, and respiration
rates varied as well. This reflects an increase in respiration rates relative to
increased tissue permeability with increased temperatures.

The R0 for apple tissue is unaffected by fluctuations of P0, from 2-21%, but is

18

lowered with progressive increases in PM (Fidler and North, 1964). When stored
in COz-free atmospheres, the RQ can range from 1.5 to 1.8, with peaks of 3.0 at
0-2°C. Results using modified atmosphere packaging (MAP) follow these
patterns, with RQ’s ranging from 1.3 to 1.5 for the 9 cultivars tested at 01°C
(Gran and Beaudry, 1993).

For apple fruit metabolism, malic acid synthesis occurs with exposure to high
Pm concentrations, which may occur with climacteric respiration (Anon., 1950).
Addition of malic acid to post-climacteric fruits will increase CO, production
without an increase in O, uptake, but only in the presence of 02, thus increasing
the RQ (Anon., 1955). Pyruvic acid also has this effect in both pre- and post-
climacteric fruit, when 02 is present or absent. A saturation of enzymes
converting pyruvate and malic acids into the Krebs cycle is indicated. Hulme
(1961) hypothesized that the C02 climacteric results from increased activity of
malic enzyme and pyruvic carboxylase, which should not result in an equivalent
rise in 02 uptake, thusly creating the observed increase in R0 during the
climacteric. The R0 for apples through the climacteric, however, has been
observed to remain rather constant at 1.1 to 1.3 at 7 to 12°C (Fidler and North,
1965).

Storage of fruits at P02 levels below the low 0; limit will lead to low 02
damage, characterized by glycolytic conversion of pyruvate to acetaldehyde and
ethanol, which can lead to the production of off-flavor compounds and visible

tissue damage. Fruits with low 02 damage have noticeable buildups of EtOH in

19

the damaged areas, along with tissue browning.

Park (1990) has shown respiration is negatively correlated with resistance to
gas exchange at picking. It may be the case that respirational gas exchange is
restricted within those tissue areas highly susceptible to low 0, damage, creating
anaerobic conditions which lead to fermentation of tissues. We have observed
that low 02 damage of fruits generally occurs in areas of the fruit having smaller
cell size and decreased intercellular air spaces. With extended storage under
anaerobic conditions, the products of fermentation build up. to levels toxic for
normal cellular functions, with cell death and tissue browning the result.

Gas exchange characteristic are intricately related to physical characteristics of
the tissue cell mass. Changes in physical characters have been seen to influence
physiological processes during ripening. Maximum respiration of bananas is
achieved only when RH levels exceed 80% (Grierson and Wardowski, 1978).
Lowered RH is also known to decrease storage breakdown of ‘Jonathan’ apples,
as well as decrease volatiles and flavor components of bananas and mangos and
increase strawberry (Fragan’a x ananassa, Duch.) and pear flavor development
(Grierson and Wardowski, 1978).

The anaerobic extinction point.

The examination of low 02 limits and the effects of limiting P02 on 02 uptake,
CO2 production, and the various indicators of increased tissue EtOH integrate
those properties defined as the anaerobic extinction point (Blackman and Parija,

1928; Parija, 1928; Blackman, 1928). The anaerobic extinction point was defined

20

as the theoretical "point such that just enough oxygen enters the cells to convert
the whole product of D (terminal substrate of aerobic carbohydrate metabolism)
to OA + OR (the final oxidation products) and there is no longer any NR (the
product of nitrogen respiration) production. This marks a definite physiological
state...(and we are conducting experiments in order to locate the) external
concentration of oxygen that, for a given tissue, coincides with this ‘extinction
point’ of NR.” Blackman and Parija further define the anaerobic extinction point
by stating "One physiological index of it, in a living apple, may be that this point
there is a minimum production of C02. Incidentally it has the significance that at
this value there is presumably no longer any accumulation of alcohol in the
tissues." (Blackman, 1928). It is by the physiological indicators of reduced 02
uptake, minimal CO, production, and tissue EtOH production that we may
determine the anaerobic extinction point. Although Blackman and Parija could
not specify the substrates and products of aerobic and anaerobic respiration, the
definition is useful for a better understanding the physiological considerations of

CA and MA storage.

21

The processes of fruit respiration, ripening, and senescence in storage are
influenced by a complex interaction of physical properties and physiological
processes. Low 02 limits and the physiology of CA and MA stored fruits depend
upon the interaction of tissue gas exchange characteristics, metabolic and cellular
changes with maturation and ripening, and changes in tissue properties with
maturation and ripening. The following studies examine low 02 limits,
considering the factors of gas exchange and physiological processes in MA storage,

in order to better understand the requirements of low 0; storage.

REFERENCES

22

REFERENCES

Anon. 1950. Rep. Fd. Invest. Bd. for 1950. 32.
Anon. 1953. Rep. Fd. Invest. Bd. for 1953. 38.
Anon. 1955. Rep. Fd. Invest. Bd. for 1955. 44.

Ben-Yehoshua, S., Robertson, R.N., and Biale, J .B. 1963. Respiration and
internal atmospheres of avocado fruit. Plant Physiol. 38:194-201.

Ben-Yehoshua, S., Burg, S.P., and Young, R. 1985. Resistance of citrus fruit to
mass transport of water vapor and other gases. Plant Physiol. 79:1048-1053.

Blackman, RF. 1928. Analytic studies in plant respiration. III. Formulation of a

catalytic system for the respiration of apples and its relation to oxygen. Proc.
Roy. Soc. Lond. (Ser. B) 103:491-523.

Blackman, RF. and P. Parija. 1928. Analytic studies in plant respiration. I. The

respiration of a population of senescent ripening apples. Proc. Roy. Soc.
Lond. (Ser. B) 103:412-445.

Burg, S.P., and Burg, EA. 1962. Role of ethylene in fruit ripening. Plant
Physiol. 37:179-189.

Burg, S.P., and Burg, EA. 1965. Gas exchange in fruits. Physiol. Plant. 18:870-
884.

Burg, SP. 1990. Theory and practice of hypobaric storage. Food Preservation by
Modified Atmospheres. CRC Press, Boca Raton, FL 353-372.

Clements, HF. 1935. Morphology and physiology of the pome lenticels in Pynts
malus. Bot. Gaz. 97:101.

Curtis, OE and Clark, D.G. 1950. An Introduction to Plant Physiology.
McGraw-Hill, New York. 212.

23

Cameron, AC. and Yang, SF. 1982. A simple method for the determination of
resistance to gas diffusion in plant organs. Plant Physiol. 70:21-23.

Devaux, H. 1891. Etude experimental sur l’aeration des tissues massifs
(experimental study on the aeration of solid tissues: Introduction to the study
of the mechanisms of gaseous exchange in aerial plants). Annales Des
Sciences Naturalles Botanique. Ser. 7, Vol. 14:297-395.

Fidler, J .C., and North, CJ. 1964. Respiration of apples in controlled
atmosphere conditions. Rep. Fd. Invest. Bd. for 1963-1964. 23-24.

Fidler, J.C., and North, CJ. 1965. The respiration of apples. Rep. Fd. Invest.
Bd. for 1964-65. 33-35.

Fidler, J.C., and North, CJ. 1966. The effect of varying temperature on the

respiration of apples, and on the incidence of low temperature injury. 1966.
Rep. Fd. Invest. Bd. for 1965-1966. 21-23.

Fidler, J .C., and North, CJ. 1968. The carbon balance in the respiration of
apples. Rep. Fd. Invest. Bd. for 1967-1968. 22.

Fockens, SH, and Meffert, H.F. Th. 1972. Biophysical properties of
horticultural products as related to loss of moisture during cooling down. J.
Sci. Fd. Agric. 23:285-298.

Gran, CD, and Beaudry, RM. 1993. Determination of the low oxygen limit for
several commercial apple cultivars by respiratory quotient breakpoint.
Postharvest Biol. Techno]. (in press).

Grierson, W., and Wardowski, W.F. 1978. Relative humidity effects on the
postharvest life of fruits and vegetables. HortScience. 13(5):570-574.

Hardy, J .K. 1949. Diffusion of gases in fruit: The solubility of carbon dioxide
and other constants for ‘Cox’s Orange Pippin’ apples. Rep. F (1. Invest. Bd. for
1939. 105-109.

Horrocks, R.L. 1964. Wax and the water vapour permeability of apple cuticle.
Nature. 4944:547.

Hulme, AC. 1961. Advances in Horticulture Science and their Application.
Pergamon Press, London. 77.

Johnson, BE. and Brun, W.A. 1966. Stomatal density and responsiveness of
banana stomates. Plant Physiol. 41:99-101.

24

Kidd, F., and West, C. 1933. The influence of the composition of the atmosphere

upon the incidence of the climacteric in apples. Rep. Fd. Invest. Bd. for 1933.
51-57.

Kidd, F. and West, C. 1934. Rep. Fd. Invest. Bd. for 1934. 110.

Kidd, F. and West, C. 1949. Resistance of the skin of the apple fruit to gaseous
exchange. Rep. Fd. Invest. Bd. for 1939. 59-64.

Leonard, E.R., and Warlaw, CW. 1941. Studies in tropical fruits. XII. the
respiration of bananas during storage at 53°F and ripening at controlled
temperatures. Ann. Bot. 5:379-391.

Lyons, J .M., McGlasson, W.B., and Pratt, H.K. 1962. Ethylene production,
respiration, and internal gas concentrations in cantaloupe fruits at various
stages of maturity. Plant Physiol. 37:31-36.

Lau, 0.1.. 1989. Control of storage scald in ‘Delicious’ apples by diphenylamine,
low oxygen atmosphere, and ethylene scrubbing. Proc. 5th Int. C.A. Res. Conf.
(Wenatchee). 169-176.

Parija, P. 1928. Analytic studies in plant respiration. II. The respiration of apples
in nitrogen and its relation to respiration in air. Proc. Roy. Soc. Lond. (Ser.
B) 103:446-490.

Park, Y.M. 1990. Gas exchange in apples: Pathway for gas exchange, changes in
resistance to gas diffusion during fruit development and storage, and the
factors affecting the change. PhD. Thesis, Cornell University. pp. 116.

Pieniazek, SA. 1944. Physical characteristics of the skin in relation to apple fruit
transpiration. Plant Physiol. 18:529-536.

Rajapakse, N.C., Hewett, E.W., Banks, NH, and Cleland, DJ. 1989. Oxygen
diffusion in apple fruit flesh. 5th Proc. Int. C.A. Res. Conf. (Wenatchee). 13-
21.

Rodriguez, L., Zagory, D., and Kader, AA. 1989. Relation between gas diffusion
resistance and ripening in fruits. 5th Proc. Int. C.A. Res. Conf. (Wenatchee).
(2):1-7.

Schénherr, J .S. 1976. Water permeability of isolated cuticular membranes: The
effect of cuticular waxes on diffusion of water. Planta. 131:159-164.

25

Schénherr, J.S. and Bukovac, MJ. 1973. Ion exchange properties of isolated
tomato fruit cuticular membrane: Exchange capacity, nature of fixed charges
and cation selectivity. Planta. 109:73-93.

Schénherr, J .S. and Schmidt, H.W. 1979. Water permeability of plant cuticles:
Dependance of permeability coefficients of cuticular transpiration on vapor
pressure saturation deficit. Planta. 144:391—400.

Smith, H.W. 1947. A new method of determining the composition of the internal
atmosphere of fleshy plant organs. Ann. Bot. 11:363-368.

Solomos, T. 1987. Principles of gas exchange in bulky plant tissues. HortScience.
22(5):766-771.

Trout, S.A., Hall, E.G., Robertson, R.N., Hackney, F.M.V., and Sykes, SM. 1942.
Studies in the metabolism of apples. Austr. J. Exptl. Biol. Med. Sci. 20:219-
231.

Chapter 1

Determination of the Low Oxygen Limit for Several Commercial Apple Cultivars

by Respiratory Quotient Breakpoint

26

27
ABSTRACT:

Low oxygen (02) limits for apple (Malus domestica, Borkh.) fruit were
determined using modified-atmosphere packaging (MAP) techniques. Fruits were
sealed in low density polyethylene (LDPE) packages and placed at 01°C until
steady-state respiration was reached. Steady state 02 and CO; partial pressures
were varied by altering package thickness, package surface area, and the total
fruit weight within the package. Based on measured permeabilities of the LDPE
packages to 02 and C02, package partial pressures were used to determine the
gas flux for 02 and CO, for each package, with gas flux rates representing
respiratory rates. The effect of O2 partial pressure on O, uptake, C02 production,
and the respiratory quotient (R0) was then determined. As the 0, partial
pressure decreased to approximately 2 kPa, RQ remained relatively constant. A
marked increase in the R0 (the RQ breakpoint) occurred below 2 kPa 0,, which
was associated with elevated EtOH concentrations. The low 0, tolerance limit
was then estimated for each cultivar as the 02 level at the RQ breakpoint; these
values ranged from 0.7 kPa O, for cultivars ‘Red Delicious’ and 0.8 kPa for ‘Law
Rome’ to approximately 1.9 kPa for ‘McIntosh’.

INTRODUCTION:

Low oxygen storage of apple fruit is an important commercial practice that
slows respiration rates and maintains fruit quality longer than air storage. The
storage experiments of Kidd and West (1927, 1933) established that both reduced

02 (5 kPa) and elevated CO, concentrations (5-10 kPa) delayed and reduced the

28
magnitude of the climacteric rise in respiration (1 %= 1.0135 kPa @ 1 atm).

Fidler and North (1966, 1967 and 1968) have shown that the respiratory quotient
(R0) for apple tissue is unaffected by variations in 0, partial pressures ranging
from 21 kPa down to 2 kPa, while being lowered with progressive increases in
CO, partial pressures.

If 02 levels fall below those supporting aerobic respiration, glycolytic
conversion of pyruvate to acetaldehyde and EtOH occurs. The 02 level at which
tissue fermentation is induced may be taken as the lower 0, limit. Although
tissue tolerance to anaerobic conditions is variable, extended exposure to these
conditions leads to tissue fermentation, browning, and a loss of economic value.

Commercial storage at 02 levels below approximately 3 kPa was formerly
limited by the technological limitations of controlled atmosphere facilities.
Recent improvements in room construction, atmosphere generation and the
control and sensing of Oz and CO, partial pressures facilitates the regulation of
gas levels to approximately 10.1 kPa. As a result of these improvements, storage
at partial pressures as low as 0.7 kPa for ‘Red Delicious’ for 7 or more months is
being conducted with minimal fruit loss (Lau, 1989). Minimizing the 0; partial
pressures at which fruit are stored increases the necessity for an accurate
determination of the lower 0, limit.

Establishment of the lower 0, limit for stored fruits has previously been
accomplished empirically via a gradual decrease in the storage 02 partial pressure

until intolerable storage damage occurred. Each commodity and new cultivar

29

required a large investment in time, equipment and materials to establish the
lower 0; limit for successful long term storage. We describe a method for
determining the lower 0; limit that is rapid, relatively simple, requires a minimal
number of fruit and is based on the measurement of physiological responses,
rather than empirical observations. This method measures the commodity’s R0
and the dependence of R0 upon 02 partial pressure. This information may then
be used to establish the lower 02 limit, as defined by the 0; level at the upswing
in the R0 (R0 breakpoint), as 02 levels become limiting to aerobic respiration.
This article reports upon (the determination of low 0, limits using this method for
9 commercially stored apple cultivars at 0°C.

MATERIALS AND METHODS:

Apple fruits of the cultivars ‘Empire’, ‘Red Fuji’, ‘Golden Delicious’, ‘Ida Red’,
‘Jonathan’, ‘Law Rome’, ‘McIntosh’ (strain MacSpur), ‘Northern Spy’, and ‘Red
Delicious’ were harvested at the preclimacteric stage as monitored by internal
ethylene levels and stored at 1°C under 1.5 kPa Oz and 3.0 kPa C02 and
packaged 1 to 2 weeks later.

As described by Cameron et al., (1989), a range of package headspace
atmospheres with steady-state O, and C02 partial pressures was produced by
varying film thickness, surface area and total fruit weight. Four thicknesses of low
density polyethylene (LDPE) (DOW Chemical Co., Midland, MI) (0.00254,
0.00508, 0.00762, and 0.0116 cm) and two surface areas (450 and 1125 cm’) were

used. Total fruit weight ranged from approximately 100-250 g and 450-800 g for

30
the 450 and 1125 cm2 packages, respectively.

Fruits were removed from storage, weighed, and heat-sealed into LDPE
packages. A septum consisting of cured silicone rubber (General Electric Clear
Silicone II) dolloped on vinyl plastic electrical tape (Scotch, 3M Super 88) was
attached to each package for gas sampling (Boylan-Pett, 1986). Packaged fruits
were left at room temperature for approximately 1 day to maintain high
respiratory rates and rapidly decrease 02 levels towards the range of expected
steady-state partial pressures. Fruits were then placed at 01°C and gas partial
pressures were monitored until steady-state respiration was reached 50-70 days
later. Gas samples (100 pl) were drawn from each package through the self-
sealing silicone septum using a 0.5 ml insulin syringe. Gas samples were analyzed
for O, (Servomex Paramagnetic Oz Transducer, Series 1100, Servomex Co.,
Sussex, England) and CO2 (ADC analytical infra red CO, Analyzer, 225 -MK3,
Analytical Development Co., Hoddesdon, England) in series, with N2 as the
carrier gas (flow rate= 100 ml ~min"). Gas samples were drawn from 4 or less
packages at a time and immediately analyzed, in order that samples not be
contaminated by atmospheric gases. This was especially important for those
packages with very low 02 levels.

Respiratory rates were determined according to the following equations:

p0, - A
x - (Iozlm-loszh) (1)

02 W

 

 

31

P A

1'3;— - ([cogk-[cogag (2)

‘02— W

 

where RRO2 and RRc02 are the rates of O, uptake and CO, production (mmol-kg‘
1-h"); P02 and Poo, are 0; and C02 permeability coefficients measured for the
LDPE at 0°C (mmol-cm-cm"-h" ~kPa"); A is the surface area (cm’); X is the
thickness of LDPE (cm); [02],... and [02],“ are the external atmosphere and
package 0, partial pressures (kPa); [COdm and [C02]..., are the package and
external atmosphere C02 partial pressures; and W is fruit weight (kg).
Permeabilities of the LDPE film packages to 02 and CO, were determined as
described previously (Beaudry et al., 1992). The effect of 0, partial pressure on
02 uptake, CO2 production, and the R0 was then determined using equations 1
and 2. Rates of C02 production as calculated using MAP techniques have been
verified in a flow through system using carrot tissue (Lennington and Beaudry,
unpublished data).

EtOH headspace concentrations were determined by gas chromatography
(Carle GC with 45.7 cm column, .32 cm bore, Haysep N packing, at 120° C with
gas flow rates of 40, 40 and 200 ml ~min‘1 for H2, He and air, respectively).
RESULTS AND DISCUSSION:

Steady state headspace O, and CO, partial pressures ranging from 16.0 to 0.2
kPa 02 (Figure 1) and 1.0 to 18.0 kPa CO, (data not shown) were generated. As

0, partial pressure decreased to approximately 2 kPa, the RQ remained

_‘4f.

2...

32
practically constant. Rapid increases in R0 were seen below 2.0 kPa 02. Low 02

limits were estimated for each cultivar at the RQ breakpoint, and ranged from 0.8
kPa for ‘Northern Spy’ to approximately 2.0 kPa for ‘McIntosh’. C02 partial
pressures ranged from 6.0 to 18.0 kPa for those packages with elevated RQ
values.

Whereas the RQ remained rather constant from 16 to 2 kPa O, for all
cultivars, the rate of 02 uptake varied among cultivars as steady-state 02 levels
decreased (Figure 2). For instance, while 02 uptake for ‘Northern Spy’ decreased
slightly from 16 to 2 kPa Oz, 02 uptake for ‘Law Rome’ decreased steadily.

Measurement of EtOH headspace levels for ‘Law Rome’ showed fruit with 02
partial pressures below the RQ breakpoint exhibited elevated headspace EtOH
concentrations (Figure 3), as well as detectable levels of acetaldehyde (data not
shown). Packages with O2 partial pressures above the RQ breakpoint had no
detectable EtOH or acetaldehyde in the package headspace.

As 0, partial pressures decreased to approximately 2 kPa, the RQ remained
practically constant in agreement with the observations of Fidler and North
(1967). Assuming the RQ breakpoint to indicate the lower 0, limit, there is
clearly variation between cultivars for the lower 0, limit (Figure 1). R0
breakpoints for the 9 cultivars tested ranged from 0.7 kPa to 2.0 kPa. Assuming
the cuticle provides the primary resistance to gas exchange (Burg and Burg, 1965;
Cameron and Yang, 1982), we hypothesize that variation in the RQ breakpoint

between cultivars largely reflects differences in cuticular resistance to diffusion,

33

 

 

 

 

 

 

 

 

 

 

 

 

 

 

   
   
  

 

 

 

 

 

 

 

 

  
   

  

 

 

 

 

 

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44 3 T 343 1
C 1 10
m .1 2 ‘1 2c: -1
’1: 3°“ . VV '° .. ° smart“
g' 121604812160481216
O 41 Red Fuji 1 41 Jonothon~ 4 Northern1
o 1 1 1 Sp 1
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0 1° 1 1
+’ 2 o 21 - 2
O . o 3% °1 1'. gq ’0 o ‘ 00¢ co
.5 1 M 1 8 o 1Q%%@O °
0. o 11 8 1'21'6 o «'1 8 1216 o 4 8 1216
m "'I"'I"'r'*'l
(31:) 41 Golden 1 4 4 Red .
1 Delucuous Delucnous
3.? . 3 3 .
‘ 1
2.1% . 2 .1 2
1 “00 00° . o? 06 «yea» «0° °
Irr'r'r'rrrrr'r'l I 1
0 4 8 12 16 O 4 8 12 16 0 4 8 12 16

Steady State 02 (kPa)

Figure 1. Effect of steady state 0; partial pressures on the respiratory quotient
(R0) of 9 cultivars of apple fruit sealed in low density polyethylene and held at
O.1°C. Estimated lower 02 limits as follows: Empire, 0.8; Red Fuji, 1.4; G.
Del.,1.3; Ida Red, 1.1; Jonathan, <2.0 (limited data); Law Rome, 0.8; McIntosh,
1.9; Northern Spy, 0.9, and R. Del., 0.7. All values given in kPa.

34

and that with increasing resistance to diffusion, internal 0, levels are more readily
reduced to levels below the lower 02 limit. Results from various studies indicate
that, in fact, permeability coefficients for 02, C02, and QIL gas varies between
apple cultivars (Kidd and West, 1949; Burg and Burg, 1962; Cameron, 1982; and
Rajapakse et al., 1990). The RQ breakpoint at 0° C for ‘McIntosh’, a fruit with a
notable heavy wax sheen, closed calyx end, and few visible lenticels, of 1.9 kPa 0,
compared with a RQ breakpoint of approximately 1.3 kPa O, for ‘Golden
Delicious’, a fruit with less sheen, an often open calyx end, and noticeable
lenticels, supports the hypothesis that the lower 02 limits for fruits are in part a
reflection of cuticular permeability.

Banks (1984), has observed increasing variation in internal 02 concentrations
with factors that increase 02 gradients by reducing cuticular permeability. It
would follow that fruit with lower cuticular permeabilities would tend to have
more variable internal 0, partial pressures relative to fruit with highly permeable
skin. Increased variability in the internal atmospheres of fruits
under low 0; storage would likely result in an increased percentage of fruits with
internal 02 partial pressures below the lower 0; limit, and a greater incidence of
damage. For those varieties having relatively high skin permeabilities and,
therefore, more homogeneous internal atmospheres, one may expect smaller
numbers of fruits to exhibit damage symptoms at the lower 0, limit. The
influence of temperature upon the low 02 limits for apple fruit has yet to be

examined. At increased temperatures and correspondingly increased

Oz Uptake (mmole-kg-1.hr-1)

 

 

 

 

 

 

 
  
 

QOlden
DeIICIOus

 

0" o.
0 4 81216

35

 

   

  

8’
Ida Red

 

 

 

 

 

o.
0 4 81216

 

  
   
 
    

 

 

 

 

 

 

 

81
Red .
Delucuous

0481216

 

Steady State 02 (kPa)

Figure 2. Effect of steady state 0, partial pressures on the 02 uptake of 9
cultivars of apple fruits sealed in low density polyethylene and held at O.1°C. See
Table 1 for equations describing curves.

36

 

 

 

 

5 El 300

4—4 - -
C ‘@ 0 Law Rome RQ @ 0°C _
(l)
L: 4-1 1:] Law Rome CszOH F
O - _
3 j —200
O 3- ”

- 1
Ex - _
O ' O '
E. 1 Idbo © C19 W O O O O O '
m it] '
<1) , 1
a: o . CED 11:11 1:11:11 maria] 1:1 13 1:1 1:1 1:1 1—0

I I I I l I I I I I I T I I
O 2 4 6 8

Steady State 02 (kPa)

Headspace CZHSOH (ppm)

Figure 3. Effect of steady state 02 partial pressure on the respiratory quotient
(R0) and headspace EtOH of ‘Law Rome’ apple fruit sealed in low density

polyethylene and held at 01°C

37

Table 1. General equations (Eq. [1]) and values of constants describing the
relationship between steady state 02 partial pressures (kPa) and 02 uptake
(mmol okg'1 oh") for the given apple cultivars sealed in LDPE and stored at O.1°C.

511- [1]: RRoz = b1'{1 - expl-b2°(02)ml}”3

 

 

Cultivar b1 b2 b, r2

Empire 0.04074 0.30732 0.19208 0.408
Red Fuji 0.05048 0.05688 0.35304 0.721
Golden Delicious 0.07960 0.07300 0.48863 0.886
Ida Red 0.03972 0.07076 0.21003 0.655
Jonathan 0.04074 0.30732 0.19208 0.408
Law Rome 0.05637 0.09455 0.29418 0.794
McIntosh, (strain MacSpur) 0.04442 0.30241 0.39690 0.871
Northern Spy 0.03873 0.09112 0.13650 0.658
Red Delicious 0.04064 0.07697 0.18953 0.560

 

respiration rates, the influence of tissue permeability upon low 02 damage would
likely be accentuated. Beaudry et al., (1992), have shown the 02 level at the RQ
breakpoint increased with increasing temperatures in-blueberry (Vaccinium
corymbosum) fruits. The authors suggest respiration rates increase at elevated
temperatures more rapidly than skin permeability, creating an increased 0,
gradient across the skin.

Park (1990), using the ethane efflux method of Cameron and Yang (1982) to
determine cuticular resistance to gas diffusion, showed considerable (i1 x 10‘
s - cm‘1 at ripe) variability of cuticular resistance coefficients within apple cultivars
from year to year. Variations in cuticular resistance also occurred through
development, and these changes were largely correlated with stomatal and

lenticular structure. 02 gradients in the cortex of air-stored apple fruit have been

38
reported recently by Dadzie et al., (1990). In examination of ’Golden Delicious’

apple fruit 0, levels were consistently lower in the calyx end. This variation in 02
levels within an individual fruit was hypothesized to result from tissue-specific
differences in cortex resistance due to variation in intercellular airspace and cell
size.

Headspace EtOH data show active fermentation within fruit tissues at 02
levels at or below the RQ breakpoint, indicating anaerobic conditions. This does
not, however, directly reflect tissue damage and the loss of value which may be
expected when fruit are stored below the lower 0; limit. Tolerance to anaerobic
conditions appears to be quite variable (Lau, 1989), with ‘Red Delicious’ apples
showing only 8% skin discoloration after 7 months at 0.5% 02 versus 61%
discoloration in ‘Spartan’ apples. The effects of long-term exposures to 02 levels,
at or below the RQ breakpoint, and the cause of cultivar variations are not clearly
defined. Tolerance of ‘Valencia’ oranges (Citrus sinensis, L. Osbeck) to short-term
anaerobic conditions for insect control has been shown (Kc and Kader, 1990).
Numerous reports indicate a greater tolerance of low 0, exposures by non-
climacteric fruits versus climacteric fruits, although differentiation between fruit
tolerance of the metabolic products of anaerobic respiration and the tolerance of
off-flavor compounds by consumers is required. Further studies relating factors of
gas movement through tissue to low 02 damage may clarify cultivar tolerance
variations and the effect of increased respiratory demands during the climacteric.

Rates of 02 uptake with variations in 0; partial pressure (Figure 2) show the

39

reduction of respiration rates with decreased 0, partial pressures. The decrease
in 02 uptake may also reflect the effects of elevated CO, levels, as C02 was not
removed from the headspace of packages. Cultivar variations in the effect of
decreased 02 levels upon 02 uptake, or respiration rates, shows reducing
atmosphere 02 levels does not always affect respiration to an equal extent.
Whereas some cultivars (McIntosh, Red Fuji) showed decreases in rates of 02
uptake below 4 kPa, other cultivars (Northern Spy) did not show a decrease in
rate of 02 uptake until 02 levels fell below 1 to 1.5 kPa 02. This may be
influenced by both skin and tissue resistances to Oz movement.

Additional examination of the effects of temperature and atmosphere
composition on fruit respiration is required. Apple fruit respiration rates in initial
experiments did not generate the full range of headspace atmospheres for each
cultivar tested as hoped, and additional data will more definitively pinpoint the
effect of 0, partial pressure on 02 uptake, C02 production and the R0 for apple
fruits. Further studies of the physiology of modified-atmosphere fruit respiration,
and the mechanisms of gas movement through bulky plant tissues should provide

valuable information for optimizing low 0, storage of apples and other fruits.

REFERENCES

40

REFERENCES

Banks, NH. 1984. Internal atmosphere modification in Pro-Long coated apples.
Acta Hort. 157:105-112.

Beaudry, R.M., Cameron, A.C., Shirazi, A., and Dostal-Lange, D.L. 1992.
Modified-atmosphere packaging of blueberry fruit: Effect of temperature on
package oxygen and carbon dioxide. J. Amer. Soc. Hort. Sci. 117(3):436-441.

Boylan-Pett, W. 1986. Design and function of a modified-atmosphere package
for tomato fruit. MS Thesis, Michigan State University, East Lansing, MI.

Burg, S.P., and Burg, EA. 1962. Role of ethylene in fruit ripening. Plant
Physiol. 37:179-189.

Burg, S.P., and Burg, EA 1965. Gas exchange in fruits. Physiol. Plant. 18:870-
884.

Cameron, AC. 1982. Gas diffusion in bulky plant organs. PhD Thesis,
University of California, Davis. pp. 117.

Cameron, AC, and Yang, SF. 1982. A simple method for the determination of
resistance to gas diffusion in plant organs. Plant Physiol. 70:21-23.

Cameron, A.C., Boylan-Pett, W., Lee, J. 1989. Design of modified atmosphere
packaging systems: Modeling oxygen concentrations within sealed packages of
tomato fruits. Inst. Food Tech. 54:1413-1416, 1421.

Dadzie, B.K., Banks, N.H., Hewett, E.W., and Cleland, DJ. 1990. Variation in
internal atmosphere composition within single apples. XXIII International
Horticulture Congress Abstracts. (Firenze, Italy). 659.

Fidler, J .C., and CJ. North. 1966. The effect of varying temperature on the

respiration of apples, and on the incidence of low temperature injury. Rep.
Fd. Invest. Bd. for 1965-1966. 21-22.

41

Fidler, J.C., and North, CJ. 1967. The effect of conditions of storage on the
respiration of apples. 1. The effects of temperature and concentration of
carbon dioxide and oxygen on the production of carbon dioxide and uptake of
oxygen. J. Hort. Sci. 42:189-206.

Fidler, J .C., and North, CJ. 1968. The carbon balance in the respiration of
apples. Rep. Fd. Invest. Bd. for 1967-1968. 22.

Ke, D., and Kader, AA. 1990. Tolerance of ’Valencia’ oranges to controlled
atmospheres as determined by physiological responses and quality attributes.
J. Amer. Soc. Hort. Sci. 115:779-783.

Kidd, F., and West, C. 1927. A relation between the concentration of oxygen and

carbon dioxide in the atmosphere, rate of respiration, and the length of
storage life of apples. Rep. Fd. Invest. Bd. for 1927. 41-42.

Kidd, F., and West, C. 1933. The influence of the composition of the atmosphere

upon the incidence of the climacteric in apples. Rep. Fd. Invest. Bd. for 1933.
51-57.

Kidd, F., and West, C. 1949. Resistance of the skin of the apple fruit to gaseous
exchange. Rep. Fd. Invest. Bd. for 1939. 59-64.

Lau, 0.1.. 1989. Control of storage scald in ’Delicious’ apples by diphenylamine,
low oxygen atmosphere, and ethylene scrubbing. Int. Contr. Atmos. Res. Conf.
(Wenatchee) 5:169-176.

Park, Y.M. 1990. Gas Exchange in Apples: Pathway for gas exchange, changes
in resistance to gas diffusion during development and storage, and factors
affecting the changes. PhD Thesis, Cornell University, pp. 116.

Rajapakse, N.C., Banks, N.H., Hewett, E.W., and Cleland, DJ. 1990.
Development of oxygen concentration gradients in flesh tissues of bulky plant
organs. J. Amer. Soc. Hort. Sci. 155:793-797.

_W ‘ 3‘5!

Chapter 2

Modified Atmosphere Packaging of Apple: The Effect of Temperature on the

Low Oxygen limit as Determined by the Respiratory Quotient Breakpoint

42

43
ABSTRACT:

This study examines the interaction of temperature and cultivar on the low
oxygen (02) limit for apple (Malus domestica, Borkh.) fruit. Low 02 limits were
determined by measuring the 0, partial pressure at which the respiratory quotient
(RQ) increased (RQ breakpoint), using modified atmosphere packaging (MAP)
techniques. ‘Golden Delicious’, ‘Early Red One Red Delicious’, ‘Redmax
McIntosh’, and ‘Marshall McIntosh’ apple fruits were sealed in low-density
polyethylene (LDPE) packages and placed at 0.1, 5, 10, 15, 20, and 25°C, with
3°C substituted for 01°C for ‘McIntosh’ strains, until steady-state respiration was
reached. For all apple cultivars tested, the package 0, partial pressure ([Oflm) at
which the RQ breakpoint occurred increased with increased temperature. From
3°C to 25°C, the RQ breakpoint increased from 0.75 to 2.5 and 1.2 to
approximately 4.0 kPa (1%= 1.0135 kPa @ 1 atm) O; for ‘Redmax McIntosh’ and
‘Marshall McIntosh’, respectively. From 01° to 25°C, the RQ breakpoint for
‘Red Delicious’ and ‘Golden Delicious’ increased from 0.5 to 0.8 and 0.6 to 1.0
kPa 02, respectively. 0, uptake (RROQ decreased with decreasing temperatures
and decreasing [02],“. CO2 production (RRCOz) rate decreased with decreasing
temperatures; RR002 also decreased as [02],“ fell below 6 to 8 kPa, and then
increased at [02],“ below the RQ breakpoint as anaerobic respiration began.
INTRODUCTION

Decreased 02 partial pressures (P02), elevated CO, partial pressures (P002),

and low storage temperature extend the storage life of a wide range of

44
horticultural commodities, including apple fruit (Kidd and West, 1927). Each

factor may significantly inhibit rates of respiration. The minimum P02 at which
fruits may be stored without inducing anaerobic respiration may be influenced by
both P00, and temperature. Normal storage strategies alter temperature and P02
while maintaining PM below levels that would limit aerobic 02 uptake for fruit
metabolism. Aerobic respiration decreases as Pm decreases.

In addition to the influences of temperature and Pam on low 0, limits, defined
here as the minimal Pm at which controlled-atmosphere (CA) or modified-
atmosphere (MA) storage may be conducted without inducing anaerobic
respiration, commodity-specific atmosphere requirements exist (Meheruik, 1989).
Safe atmosphere compositions for CA or MA storage vary greatly among
individual commodities as the result of inherent differences in organ respiration
rates, ripening characteristics, physiological state, and gas exchange properties
related to tissue and/ or organ morphology and composition. In addition to
specific atmospheric requirements, chilling-sensitive, tropical fruits such as mango
(Mangifera indica, L) and banana (Musa, L. sp.) require that storage temperatures
not fall below critical temperatures, at which chilling injury may occur. Critical
temperatures also vary by commodity and length of exposure. In the case of
apple fruit, optimal storage temperatures and CA atmospheres vary with cultivar
and strain (Meheruik, 1989).

Fruit quality must be retained and optimized to warrant the added expenses of

CA and MA storage. Previous experiments have shown that the low 02 limit

45
determined by RQ breakpoint varies with temperature (Fidler and North, 1967).

A modified-atmosphere packaging (MAP) approach and use of R0 breakpoint
appear to be useful in determining the low 02 limit for apple and blueberry
(Vaccinium corymbosum) fruit (Gran and Beaudry, 1993; Beaudry et al., 1992). In
these studies the [02],.“ at which the RQ breakpoint occurred was taken to
represent the low 0, limit. Hypothesizing that low 02 limits increase with
temperature, and that increases in low 0, limit vary with cultivar, experiments
were conducted in order to determine the effect of temperature on RR” RRCOZ,
and low 0, limits determined by RQ breakpoint for three cultivars of apple fruits.
MATERIALS AND METHODS:

Apple fruit of the cultivars ‘McIntosh’ (strains ‘Redmax’ and ‘Marshall’),
‘Golden Delicious’, and ‘Red Delicious’ (strain ‘Early Red One’) were harvested
at the preclimacteric stage as monitored by internal ethylene (QIL) levels
(520% of fruit $0.2 ul/l) and stored at 3°C in air until packaged 2 to 6 days
later.

As described by Cameron et al., (1989), a range of steady-state [02],.“ and
package C02 partial pressure ([COz],.,,) were produced by varying film thickness
and total fruit weight in the package. Four thicknesses (0.00254, 0.00508, 0.00762,
and 0.0116 cm) of low density polyethylene (LDPE) (Dow Chemical Company,
Midland, M1) were used. Package surface area was 960 cm’; fruit numbered from
2 to 5 fruit and total fruit weights were approximately 250 to 800 g per package.

Fruits were packaged as previously described (Gran and Beaudry, 1993) and

46

30 or more packages of differing combinations of fruit weight and film thickness
were placed at 0.1, 5, 10, 15, 20, and 25°C for ‘Golden Delicious’ and ‘Red
Delicious’ cultivars, with 3°C substituted as the low temperature for ‘McIntosh’
strains. The lowest treatment temperatures used are those normally
recommended for commercial CA storage (Dilley, 1989; Meheriuk, 1989). Over
30 additional packages for ‘Marshall McIntosh’ and ‘Golden Delicious’ fruits were
prepared with 9 x 6 cm Tyvek" pouches containing approximately 40 ml of
hydrated lime [Ca(OH)2], in order to reduce [COflm to below ambient levels.
Packages containing [Ca(OH)2] pouches were stored over the same range of
temperatures according to cultivar. [02],“ and [C02]m were monitored until
steady-state respiration was reached.

Duplicate gas samples (25 [1.1) were drawn from each package through a self-
sealing silicone septum using a 50 n1 glass syringe (Hamilton Co., Reno, NV).
Samples were analyzed for O2 (Servomex Paramagnetic O2 Transducer, Series
1100, Servomex Co., Sussex, England) and CO, (ADC analytical infra red C02
Analyzer, 225-MK3, Analytical Development Co., Hoddesdon, England) in series,
with N2 as the carrier gas (flow rate= 100 ml-min").

RRO, and RR», were determined using measured permeabilities for O; (P'm)
and C02 (Fem) previously described (Beaudry et al., 1992; Gran and Beaudry,
1993). The effect of [02],“ on RRO” RR“), and the R0 was determined at each
storage temperature, and the effect of temperature on the low 02 limit was

determined.

47

R0 breakpoints and low 02 limit boundaries were determined by marked
increases (commonly 0.1-0.2 units) in R0 values above the basal aerobic RQ
value which occurred between packages with contiguous [02],“. Low 0; limit
boundaries were defined by the [02],.“ of the packages between which a change in
R0 was observed.

Prediction of RR.)2 over the range of temperature treatments was done using .1
SAS' nonlinear regressions analysis. Data were fit to the Michaelis-Menten 11-
kinetic equation [Eq. (1)] given as:

V" ' [02]"!— (1)

02 ' 1gp, + 10,1”

 

where V.“ is the maximum initial velocity and K_Oz is the Michaelis-Menten
constant for 02 for whole fruit tissue. RRm curves were generated for ‘Golden
Delicious’ and ‘Marshall McIntosh’ cultivars, for packages both with elevated
[COAN and packages scrubbed for C02.

RESULTS AND DISCUSSION:

A range of steady-state [02],“, both aerobic and anaerobic, were generated for
each apple cultivar at each treatment temperature. Steady-state [COflm ranged
from approximately 3 kPa for aerobic headspace atmosphere up to 15 kPa for
anaerobic package atmospheres. For all cultivars examined, the [02],“ at which
the RQ breakpoint occurred increased with increased temperature. Low 02 limits

of ‘Golden Delicious’ and ‘Red Delicious’ fruits were affected to a lesser

Respiratory Quotient (R0)

48

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Figure 1. Effect of temperature (3, 5, 10, 15, 20, and 25°C) on the respiratory
quotient (R0) and the RQ breakpoint of ‘Marshall McIntosh’ apple fruit.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Steady State 02 (kPa)

Figure 2. Effect of temperature (3, 5, 10, 15, 20, and 25°C) on the respiratory
quotient (R0) and R0 breakpoint of ‘Redmax McIntosh’ apple fruit.

50

 

 

 

 

 

 

 

 

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Figure 3. Effect of temperature (0.1, 5, 10, 15, 20, and 25°C) on the respiratory
quotient (R0) and R0 breakpoint of ‘Golden Delicious’ apple fruit.

 

 

51

 

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Figure 4. Effect of temperature (0.1, 5, 10, 15, 20, and 25°C) on the respiratory
quotient (R0) and R0 breakpoint or ‘Early Red One Red Delicious’ apple fruit.

52

Table 1. Estimated low 02 limit ranges for apple fruit of 3 cultivars based upon
RQ breakpoint (Figures 1-4).

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Low 02 limit
Cultivar
Temperature Golden Red Redmax Marshall
(°C) Delicious Delicious McIntosh McIntosh
0.1 / 3 0.55-0.60 0.45-0.60 0.75 1.20-1.25
5 0.50-0.70 0.35-0.45 0.85-0.95 1.25-1.60
10 0.45-0.65 OAS-0.50 1.15 1.85-2.00
15 0.40-0.60 0.55-0.70 1.75-1.85 2.50-3.00
20 0.70-0.95 0.60-0.75 1.90-2.15 4.00-4.10
25 0.80-1.00 0.55-0.85 2.45-2.50 3.80-4.90
5
o GDU (R0)
8 4 0 R0 (RD)
“E 4 MAX (R0)
0 4'4 o MRU (R0) 8 <>
U
C -1
8
a3 3"
.é -‘
:1 2‘
o" 1 ‘
3
3 A u a
O I I I I
O 5 10 15 20 25

Temperature (°C)

Figure 5. Low 0, limit boundaries for 3 apple cultivars over a range of
temperatures as determined by RQ breakpoint. (GDU, ‘Golden Delicious’
unscrubbed; RD, ‘Red Delicious’; MAX, ‘Redmax McIntosh’; MRU, ‘Marshall
McIntosh’ unscrubbed)

53
extent by temperature than either strain of ‘McIntosh’ (Figures 14). Low 0, limit

boundaries increased from 0.6 to 1.0 kPa and 0.5 to 0.8 kPa 02 for ‘Golden
Delicious’ and ‘Red Delicious’ as temperature increased from 01°C to 25°C,
while for ‘Redmax McIntosh’ and ‘Marshall McIntosh’ the low 02 limit increased
from 0.75 to 2.5 kPa and 1.2 to 4.0 kPa 02, respectively, as temperature increased
from 3 to 25°C. Estimated low 02 limits and the specific [02],,“ at which the RQ
breakpoint occurred for treatment temperatures are given for each cultivar in
Table 1. Distinct differences in the low 02 limits of different cultivars and strains
of cultivars were observed. Changes in low 02 limits occurred most markedly
above 10°C for all cultivars.

The minimum P02 determined to be safe for storage, at the lowest
experimental temperature, was greater for both ‘McIntosh’ strains than either
‘Golden Delicious’ and ‘Red Delicious’ (Figure 5). Results reflect commercial CA
storage practices, with 02 atmosphere recommendations for CA storage of
‘Marshall McIntosh’ fruits being 1.0 kPa higher than either ‘Red Delicious’ or
‘Golden Delicious’.

RQ levels for fruits under aerobic conditions decreased in most cases from 1.4
at 15 and 20°C to 1.0 at 0.1 or 3°C (Figures 1-4). This may indicate a loss of
organic acids with time. CA storage, at a P02 of 1 %, maintains organic acid
content of stored ‘Bartlett’ pears (Pyrus communis, L.) at levels higher than for air
stored fruits (Yoshida et al., 1986).

For all cultivars examined, fruit RQ values were calculated between 1.0 to 1.4

54

at [021m above the anaerobic extinction point (Figures 1-4). An exception was for
the fruit stored at higher temperatures (20 and 25°C), where aerobic RQ levels
ranged up to 1.8. The aerobic R0 for apple fruit has been determined to range
from 1.5 to 1.8, with peaks of 3.0 at 0°C in C02-free atmospheres (Fidler and
North, 1964). Time to steady-state respiration varied from 5 to 7 days at 25°C to
75 days or longer at O.1°C. The occurrence of mold and general tissue
breakdown at higher treatment temperatures required quick sampling and did not
allow data collection beyond 15 days.

Differences in aerobic RQ values may be the result of chilling-induced
ripening and an associated increased respiratory rate (Purvis, 1985) as observed
with chilling of avocado (Persea americana, Mill.) fruit (Cooper et al., 1969). Fruit
ripening within packages may have occurred at lower treatment temperatures due
to slow headspace [02],“ reduction or large fluctuations in [02],“ before steady-
state respiration was established. In the case of ‘Marshall McIntosh’ at 3°C,
[02],“ fluctuated by 3.5 kPa between the 12th and 17th day following packaging,
with [02],.“ traversing the subsequently determined low 0; limit. Such
fluctuations in [02],.“ would alter lRRo2 and fruit ripening in storage.

A concurrent increase in the rate of respiration, as determined by RRcm
(Figure 6-9) and predicted RR02 (Figures 10-13), occurred above 10°C. RRm
increased with increased temperatures for all cultivars examined. The effects of
temperature on RRO2 uptake varied with cultivar (Figures 10-13). In the case of

both ‘McIntosh’ strains, particularly ‘Marshall McIntosh’, respiration and the effect

55

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Steady State 02 (kPa)

Figure 6. Eflect of temperature (3, 5, 10, 15, 20, and 25°C) and [021.11. on RRom
~ of ‘Marshall McIntosh’ apple fruit.

56

 

 

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CO2 Production (mmol-kg"-h")

 

 

 

 

 

 

T T r I I I I r I I I I I I I I I I I I I

0 3 6 9 12 15 18 0 5 g 9 12 15 18
0'” T I I I I I I I I I T I I I—FI I 0.x I I I I I I I I I I at I I r I I I

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Dim I I T I ‘r I I I I I I I I I I I com I I III I I r l I I I fir ' I f

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Steady State 02 (kPa)

Figure 7. Effect of temperature (3, 5, 10, 15, 20, and 25°C) and [02],.“ on RRcm
of ‘Redmax McIntosh’ apple fruit.

‘_ .
m ‘b..‘l_

57

 

 

 

 

 

 

 

 

 

 

 

 

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Steady State 02 (kPa)

Figure 8. Effect of temperature (0.1, 5, 10, 15, 20, and 25°C) and [02],“ on RRcm

of ‘Golden Delicious’ apple fruit.

.?*WW

58

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Steady State 02 (kPa)

Figure 9. Effect of temperature (0.1, 5, 10, 15, 20, and 25°C) and [021m on RRcm
of ‘Early Red One Red Delicious’ apple fruit.

bf‘f‘.’ " 'l

 

 

 

 

 

 

 

 

0.25 1 I r r I 1 I 1 I l 1 fl
25°C
0 0.20- -
*6 A zo'c
a 7
3 f 0.15— 15'c
o" 3
U -'— .
o — 1o
3 E 0.10- C
o 1
'6 s
:5 51°C I
0.05“ ‘J'C .
l
Marshall McIntosh E
0.001 I I I I I l l l I I I I
O 8 12 16
P02 (kPa)

Figure 10. Comparison of 02 uptake curves for ‘Marshall McIntosh’ apple fruit
over a range of temperatures. See Table 2 for equations describing curves.

0.20 , .

 

0.15-

0.10~

Predicted O2 Uptake
(mmol kg" h")

0.05-

 

 

 

 

 

 

 

T l I I I T I I I I 25.C
- 20°C
#- 15°C
10'c
_ 5°C
3°C
Marshall McIntosh (-co,)
I l I r 1 l I I I F
8 12 16
F302 (kPa)

Figure 1 1. Comparison of 02 uptake curves for ‘Redmax McIntosh’ apple fruit
over a range of temperatures, with C02 removed from package headspaces. See
Table 2 for equations describing curves.

 

 

 

 

 

 

 

 

 

 

0.25 I I I I I I I I I I
25'0
0 0.20— -
.x
3 C‘
Q I
D ‘7: 015- 20°C
0 3'
U L6 0
#3 E 0.10- ‘5‘:
'6 E ,
33 V 100
CL
0.05 '5’c
0.1'c
Golden Delicious
000' I I l l l l l I I l
O 8 12 16

P02 (kPa)

Figure 12. Comparison of 02 uptake curves for ‘Golden Delicious’ apple fruit
over a range of temperatures. See Table 2 for equations describing curves.

 

 

 

 

 

 

 

 

 

 

0.20 I I T I 1 I I I I I
zs'c
3 0.15- _
U A
4—1 T .
g .g: 200
N r
O a? 0.10~ 15'C
'0 _
‘3 E 10'c
" E
8 v .
i 0.05 ‘50
0.1'c
Golden Delicious (-co,)
0'00 I I F I I I I I I I r
O 8 12 16
F302 (kPa)

Figure 13. Comparison of 02 uptake curves for ‘Early Red One Red Delicious’
apple fruit over a range of temperatures. See Table 2 for equations describing

curves.

61

Table 2. General equations (Eq. (1) and Eq.(2)) and values of constants
describing the relationship between steady state [02],“ (kPa), RRm (mmol - kg'
‘- hr"), and storage temperature (°C) for 3 apple cultivars in modified atmosphere

packages.

Eq.(l) = {(a“ exp[b'temperature]c) ‘ {021*} / {(d + e‘temperature) + [02]le

Eq.(2) = { (a‘ exp[b'temperature]c) " {021”} / {(d‘exp[e‘temperature] + f) + [02],.,}

 

Cultivar

Marshall
McIntosh

a

-.7187

b
-.0167

C

.7325

d
1.1463

 

1-2

.7889

 

 

Marshall
McIntosh
(-C02)"

.0329

.0757

-9. 176

10.284

.8218

 

Golden
Delicious

.0367

.0741

-.0079

.1514

.0071

.7893

 

Golden
Delicious
(~C02)

 

.2721

 

.0158

 

-.2450

 

-.2128

 

-.9451

 

.2038

 

.7528

 

‘ EQ- (1)
" Eq.(2)

""1

62
of temperature on RR02 and RRCO, appears variable with [02],“, indicating

variation in gas exchange characteristics and subsequently individual fruit
respiration.

Respiration, as indicated by RRCOZ, was decreased with decreased
temperatures similar to RROT At [02],.“ below the anaerobic extinction point,

where rapid decreases in RRO2 were seen, rates of RR», increased rapidly

""" n"‘9‘"

(Figures 6-9), and RRCOZ curves often exhibited a characteristic ‘checkmark’ I,
shape with decreasing [02],“. This increase in RRcm is an indication of
anaerobic respiration.

In addition to reducing rates of respiration, treatment temperature also
affected the [02],“ at which RRO2 was decreased, indicating a change in apparent
K. for 02. Apparent K,n for 02 for ‘Marshall McIntosh’ increased linearly with
temperature, from approximately 1.0 kPa 02 to 3.5 kPa 02 from 3° to 25°C
(Figure 14), while apparent K... for 02 for ‘Golden Delicious’ cultivars increased
nonlinearly with temperature, increasing sharply from 0.05 kPa 02 at 01° to
approximately 0.25 kPa 02 at 10°C were values remained relatively constant up to
25°C (Figure 15). Kill for 02 values for ‘Golden Delicious’ and ‘Red Delicious’
cultivars ranged from 0.1 to 0.4 kPa 02. Removal of CO, from package
headspaces may have reduced apparent KIll for 02. Previous MAP studies using
blueberry fruits show reduction of RR” by decreased [02],“ varies with
temperature (Beaudry et al., 1992). Whereas RRO, decreased if [02],“ fell below

approximately 6 kPa at 01°C, RRoz decreased continually up to 15 kPa 02 at

63

 

 

 

 

4.0
you 3.0.“ ’
éxe ‘
LL-O-J
q) C 2.0"
6 8
_C 8 ..
3 C).
< 1.0—
4 H MRU
H MRS
0.0 I I I I I I I I I
O 5 10 15 20 25

Temperature (°C)

Figure 14. Apparent whole fruit K... for 02 of ‘Marshall McIntosh’ apple fruit
stored over a range of temperatures, with and without scrubbing of C02 from
package headspaces (MRU, ‘Marshall McIntosh’ unscrubbed; MRS, ‘Marshall
McIntosh’ scrubbed).

 

 

      

 

 

 

 

0.4
N —— 4 I
#0 0.3
'éxe _
LL“ 02 ‘ ‘ A
(DC . —‘ ' ' v
68
_CO
g&
< 0.1—
HGDU
0.0I l I I I l I IHIGDSI
O 5 10 15 20 25

Temperature (°C)

Figure 15. Apparent whole fruit KIn for 02 of ‘Golden Delicious’ apple fruit
stored over a range of temperatures, with and without scrubbing of C02 from
package headspaces (GDU, ‘Golden Delicious’ unscrubbed; GDS ‘Golden
Delicious’ scrubbed).

64

25°C. This indicates that at higher temperatures peel and tissue resistances to 02
diffusion (r02) may limit RRoz at a higher [021m than at lower temperatures.
Removal of CO; from package headspaces may have produced noticeable
differences in RRo, for ‘Marshall McIntosh’ (Figures 1 and 2, APPENDIX B) or
‘Golden Delicious’ fruits (Figures 4 and 5, APPENDIX B) when compared with
RRm curves for packages with [C02],., up to 12 kPa, although the removal of
[C02],.,' which ranged up to 15 kPa unexpectantly did not increase respiration
rates as determined by RROZ, but rather decreased measured respiration rates.
Calculations of whole fruit apparent KIII for 02 (Figures 14 and 15), using non-
linear regressions of RROZ, temperature, and [02],“, allowed further indications of
differences in gas exchange properties for the cultivars examined. Whereas Km for
02 for ‘Marshall McIntosh’ changed linearly with changes in temperature,
indicating r02 is limiting respiration, ‘Golden Delicious’ changed non-linearly,
increasing quickly from 0.1 to 10°C and then remaining unchanged at
temperatures above 10°C, indicating r02 is not limiting respiration for this cultivar.
Determination of the low 0, limit of ’Marshall McIntosh’ was difficult due to
variable respiration rates of fruits, with variability of RR“ and RR“), increased
with temperature. The RQ breakpoint at higher temperatures was poorly defined,
which may indicate variable gas exchange characteristics between individual fruits.
Variations in low 02 limits with individual fruit and changes from year to year
(Chapters 1 and 2) may result in storage problems, particularly in the case of

storage at very low 02 levels. Differences in low 02 limits for different strains of

65
apple cultivars, as indicated by comparison of ‘Marshall McIntosh’ and ‘Redmax

McIntosh’ data, underscores the difficulties associated with CA and MA storage at
low P02 when cultivar-specific requirements of storage are not known. Problems
in optimizing storage conditions also arise with variable low 02 limits of strains
when facilities or other limitations require storage of multiple strains of fruits with
variable low 02 limits in the same CA or MA storage unit.

The results show temperature control for low 0, storage is important,
especially in the case of cultivars with a propensity for low 02 damage.
Temperature control is particularly important for commercial MAP applications,

with atmosphere maintenance dependent upon rates of respiration.

REFERENCES

66

REFERENCES

I.”

-11 as

Beaudry, R.M., Cameron, A.C., Shirazi, A., and Dostal-Lange, D.L. 1992.
Modified-atmosphere packaging of blueberry fruit: Effect of temperature on
package oxygen and carbon dioxide. J. Amer. Soc. Hort. Sci. 117(3):436-441.

F
. .
A I A

Cameron, A.C., Boylan-Pett, W., and Lee, J. 1989. Design of modified-
atmosphere packaging systems: Modeling oxygen concentrations within sealed
packages of tomato fruits. Inst. Food Tech. 54:1413-1416, 1421.

Cooper, W.C., Rasmussen, GK, and Waldon, ES. 1969. Ethylene evolution
simulated by chilling in citrus and Persea sp. Plant Physiol. 44:1194-1196.

Dilley, DR. 1989. 1989 recommended storage conditions for Michigan apples.
1989 Michigan State University Controlled Atmosphere Storage Clinic
Handbook. 51.

Fidler, J .C., and North, CJ. 1964. Respiration of apples in controlled
atmosphere conditions. Rep. Fd. Invest. Bd. for 1963-1964. 23-24.

Fidler, J.C., and North, CJ. 1967. The effects of conditions of storage on the
respiration of apples. 1. The effects of temperature and concentrations of

carbon dioxide and oxygen on the production of carbon dioxide and the uptake
of oxygen. J. Hort. Sci. 42:189-206.

Gran, CD, and Beaudry, RM. 1993. Determination of the low oxygen limit for
several commercial apple cultivars by respiratory quotient breakpoint.
Postharvest Biol. Technol. (in press).

Kidd, F., and West, C. 1927. A relation between the concentration of oxygen and
carbon dioxide in the atmosphere, rate of respiration, and the length of
storage life of apples. Rep. Fd. Invest. Bd. for 1926. 41—42.

67

Meheriuk, M. 1989. CA storage of apples. Proc. 5th Int. C.A. Res. Conf.
(Wenatchee). (2):257-284.

Purvis, AC. 1985. Low temperature induced azide insensitive oxygen uptake in
grapefruit flavedo tissue. J. Amer. Soc. Hort. Sci. 110(6):782-785.

Yoshida, T., Borgic, D.M., Chen, P.M., and Mielke, E.A. 1986. Changes in
ethylene, acids, and brown-core development of ‘Bartlett’ pears in low-oxygen
storage. HortScience 21(3):472-474.

Chapter 3
Modified Atmosphere Packaging of Apple: The Effect of Temperature on the

Low Oxygen Limit as Determined by Headspace Ethanol Accumulation

68

69
ABSTRACT:

The purpose of this study was to determine the extent to which temperature
alters the low 02 limit for apple (Malus domestica, Borkh.) fruit of three cultivars,
by measuring the accumulation of ethanol (EtOH) in the headspace of modified
atmosphere packages. For the cultivars tested, the highest package 02 partial
pressure ([02]ka inducing ethanol (EtOH) accumulation in package headspaces,
termed here as the ‘fermentation threshold’, increased with increasing
temperature. From 3° to 25°C the [02],.“ at which headspace EtOH levels
[EtOH],.,, accumulated increased from 0.75 to 4.0 kPa and 2.0 to 6.6 kPa
(1%=1.0135 kPa @ 1 atm) for ‘Redmax McIntosh’ and ‘Marshall McIntosh’,
respectively, while increasing from 0.5 to greater than 1.0 kPa and 0.6 to 1.2 kPa
from 0.1 to 25°C for ‘Early Red One Red Delicious’ and ‘Golden Delicious’,
respectively. The accumulation of EtOH in the headspace of modified
atmosphere packages at steady-state respiration appears to be a useful tool for
quick, accurate determination of low 02 limits of apple fruit.

INTRODUCTION:

Successful long term storage of pome fruits depends on optimized temperature
and gas atmosphere compositions. Maintenance of the proper storage
temperature is required in order to maintain reduced respiration rates and to
maximize the retention of firmness and other quality characteristics, especially for
those fruits to be stored greater than 6 months. The storage temperature must

also be adjusted for various storage related disorders which may be attenuated or

E===t <"1

70
aggravated by temperature effects (Wills et al., 1989). Needed adjustments of

storage temperature may be related to cultural conditions and practices,
environmental factors, mineral nutrition of the fruit, time from harvest to storage
atmosphere establishment, and the predisposal of various varieties to develop
storage disorders. A number of storage disorders can be characterized as chilling
disorders, including superficial scald, core flush, Jonathan spot, and low I
temperature breakdown (Fidler et al., 1973). Higher than optimal storage H
temperatures may also lead to the development of disorders such as senescent
breakdown, especially in the case of fruit stored at an over-mature stage.

Storage 02 and CO; partial pressures (P02 and P002) are known affect
temperature-related disorders, as well as induce specific 02- and COz-related
storage disorders. Superficial scald was reduced or eliminated by decreased P02
(Lau, 1989) or increased PCO2 (Tomkins, 1966). The effectiveness of elevated PC02
to reduce the incidence of superficial scald may vary with storage ventilation, and
was especially ineffective when elevated P002 is achieved by reduced venting
(Meigh, 1970).

The induction of tissue fermentation by limiting P02 or toxic Poo, results, in
part, in the production of EtOH and acetaldehyde. The buildup of these
fermentation products may result in tissue browning, cell death, intercellular
flooding, and production of off-flavor compounds. Tissue EtOH production is an
indicator or cause of many controlled atmosphere storage disorders (Fidler et al.,

1973).

71

A primary deterrent to commercial applications of modified atmosphere
packaging (MAP) technologies for storage and transit of fruits and vegetables is
the likelihood that packages and the commodities they contain will be exposed to
a wide range of temperatures throughout the transport chain. An increase in
temperature will increase respiration rates to a greater extent relative to changes
in film 02 permeabilities (P'oz). With headspace atmosphere dependent upon the
temperature effect on tissue respiration rates, temperature shifts will alter package
atmosphere. In the case of blueberry (Vaccinium corymbosum) fruit, increased
temperature lowered [02],.“ (Beaudry et al., 1992). Temperature effects on
packaged fruit physiology, with the disparity of changes in P}; and RR” can be
marked when the low Oz limit increases with temperature. Current research
efforts center on the development of packages which would be able to alter
atmospheres in response to deleterious storage atmospheres and temperatures.

It is generally accepted that respiration rates vary among individual fruits and
throughout the various physiological states of maturation. Ripening and
senescence processes, variation in physiological state, environmental conditions for
growth, and environment after harvest affect respiration rates. Previous
experiments have shown the low 02 limit of a variety of commodities varies with
temperature (Fidler and North, 1967). Studies in our laboratory have shown
MAP systems well suited for determination of the low O, limit for apple fruit at
01°C (Gran and Beaudry, 1993) and blueberry fruit (Beaudry et al., 1992). For

apple fruit at 01°C, the [02],“ at which a large increase in [EtOH]m occurred

72
was taken to represent the low 0; limit, and coincided with the RQ breakpoint

(Gran and Beaudry, 1993).

Transport temperature shifts would change tissue respiration, potentially
causing headspace atmosphere fluctuations, fermentation, and reducing tissue
preservation with MAP transport and storage of apples. Hypothesizing that low
O, limits increase with increasing temperatures, and that the extent of the
increase varies with cultivar, experiments were conducted to determine the effects
of temperature on low 02 limits for three cultivars of apple fruit using [EtOHlm
accumulation as the indicator.

MATERIALS AND METHODS:

Apple fruit of the cultivars ‘McIntosh’ (strains ‘Redmax’ and ‘Marshall’),
‘Golden Delicious’, and ‘Red Delicious’ (strain ‘Earli Red One’) were harvested at
the preclimacteric stage as determined by internal ethylene (OJ-L) levels and
stored at 3°C in air until packaged 2 to 6 days later.

As described by Cameron et al., (1989), packages with a range of steady-state
[02],“ and C02 partial pressure ([COzlm) were produced by varying film thickness
and total fruit weight in the package. Four thicknesses (0.00254, 0.00508, 0.00762,
and 0.0116 cm) of low density polyethylene (LDPE) (Dow Chemical Company,
Midland M1) were used. Package surface area was 960 cm’; fruit number from
with a range of 2 to 5 fruit to generate fruit weights of approximately 250 to 800 g
per package.

Fruits were packaged as previously described (Gran and Beaudry, 1993) and

73
30 or more packages of differing combinations of fruit weight and film thickness

were placed at 0.1, 5, 10, 15, 20, and 25°C for ‘Golden Delicious’ and ‘Early Red
One Red Delicious’, with 3°C substituted for the low temperature for ‘McIntosh’
strains. Over 30 additional packages of ‘Marshall McIntosh’ and ‘Golden
Delicious’ fruits were prepared with 9 x 6 cm Tyvek' pouches containing
approximately 40 ml of hydrated lime [Ca(OH)2], in order to reduce [COZLn to
below ambient levels. Packages with Ca(OH)2 were placed at the same range of
temperatures according to cultivar. [02],“ and [COJN were monitored
periodically for each cultivar stored at each treatment temperature until it was
determined that fruits were at steady-state respiration.

Duplicate gas samples (25 p1) were drawn from each package through a self-
sealing silicone septum using a 50 pl glass syringe (Hamilton Co., Reno, NV), and
analyzed for O, (Servomex Paramagnetic Oz Transducer, Series 1100, Servomex
Co., Sussex, England) and CO2 (ADC analytical infra red C02 Analyzer, 225-
MK3, Analytical Development Co., Hoddesdon, England) in series, with N2 as the
carrier gas (flow rate= 100 ml ~min").

Gas samples (250 pl) were drawn from packages using a 500 pl gastight glass
syringe (Hamilton Co., Reno, NV) and [EtOH]m were determined by gas
chromatography (Carle GC with 45.7 cm column, .32 cm bore, Haysep N packing,
at 120°C with gas flow rates of 40, 40, and 200 ml omin" for H2, He and air,
respectively). EtOH levels were determined by comparison with standard

solutions consisting of 100 pl - 100 ml" deO placed at treatment temperatures,

74

with adjustments for standard vapor pressure changes with temperature (Harger et
al., 1950).

[EtOH],,, and tissue EtOH levels were measured and compared using tissue
samples from packages having previously been sampled for [EtOH]m. Tissue
samples were taken from core and cortical tissues, pooling 2-3 g of tissue sliced
from multiple fruit removed from packages. Samples of approximately 10 g were
macerated, placed in a 25 m1 flask, capped with a rubber septum, and placed at
original treatment temperatures. Gas samples (250 pl) from the sealed flasks
were analyzed after 2 hours (Pesis and Avissar, 1990). [EtOHLn were determined
over a range of [021m each storage temperature, and the effect of temperature on
the low 0, limit was determined.

EtOH analysis was limited to 10- 15 packages at each temperature, with the
sample population consisting primarily of packages with a range of [021m 13 kPa
Po, from the low O, limit determined by the RQ breakpoint (Chapter 2).

Low O, limit boundaries, indicated by elevated [EtOH]m, were determined by
marked increases (commonly 1050 ppm) in [EtOHlm above the basal aerobic
[EtOH],., values which occurred between packages with contiguous [02]” Low
O, limit boundaries were defined by the [O,],,, for the packages between which an
increase in [EtOH]m was observed.

RESULTS AND DISCUSSION:
A range of steady state [O,],,, were generated for each apple cultivar at each

treatment temperature. [0,1],“ ranged from 0.1 to 6 kPa 0,. Time until steady-

75

state respiration varied from 5 to 7 days at 25°C to more than 75 days at O.1°C.
Aerobic [EtOH],.‘ measured 510 to 50 ppm at 0.1 or 3 to 25° C, respectively,
until reaching the anaerobic extinction point (Figures 1-4).

For all cultivars examined, the [021m at which elevated [EtOH]m were
observed increased with temperature (Figures 1-5). Low 0, limits are expressed
as a range (Table 1), representing [O,],,,, between which an increase in [EtOHLh
increased above aerobic levels was observed. In some instances the range is
small, while for others the increment of [021m was separated by larger amounts,
which did not allow accurate determination of specific low O, limit values.
Estimated low O, limits at lower treatment temperatures are comparable with
recommended atmospheres for controlled atmosphere (CA) storage of apple fruits
(Dilley, 1989; Lau, 1989; Meheriuk, 1989).

Low O, limits increased from approximately 2.0 to 6.6 kPa and 0.75 to 4.0 kPa
for ‘Marshall McIntosh’ and ‘Redmax McIntosh’, respectively, as temperature
increased from 3 to 25°C (Table 1). Low 0, limits for ‘Golden Delicious’ and
‘Red Delicious’ increased from approximately 0.6 to 1.0 kPa and 0.5 to 1.2 kPa
from 0.1 to 25°C. Low O, limits increased most dramatically from 15 to 25°C,
while increasing only slightly from 0.1 or 3 to 10°C (Figure 5).

[EtOHLh was highly correlated with EtOH levels of flasks containing
macerated tissue (Figure 6). Internal and external tissue samples exhibited EtOH
levels which did not differ significantly with area of fruit sampled. Packages with

[EtOHlm above aerobic [EtOH]m showed increased [EtOHlm with time,

d

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Figure 1. Effect of temperature (3, 5, 10, 15, 20 and 25°C) on headspace EtOH
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77

 

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78

 

 

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Figure 3. Effect of temperature (0.1, 5, 10, 15, 20, and 25°C) on headspace EtOH
levels as an indication of the low 02 limit of ‘Golden Delicious’ apple fruit.

79

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Figure 4. Effect of temperature (0.1, 5, 10, 15, 20, and 25°C) on headspace EtOH
levels as an indication of the low 02 limit of ‘Early Red One Red Delicious’ apple
fruit.

80

Table 1. Estimated low 0, limit ranges for 3 cultivars of apple fruit indicated by
elevated headspace EtOH.

 

   

 

 

 

 

 

 

 

 

Temperature Golden Red Redmax Marshall
(°C) Delicious Delicious McIntosh McIntosh
0.1/3 0.55-1.25 0.45-0.60 0.75-0.80 1.80-2.25
5 0.70-0.75 0.55-0.85 1.00-1.10 1.75-2.05
10 0.65 0.90-1.20 1.15 2.15-2.70
15 O.55-1.45 1.15-2.20 1.60-2.50 2.35-3.00
20 OAS-0.95 1.70-2.00 2.00-2.35 4.95-5.20
25 1.15-1.65 0.90-3.35 3.85-4.25 6.60-6.85
7

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Temperature (°C)

 

Figure 5. Low 0, limit boundaries for 3 apple cultivars over a range of
temperatures as determined by elevated [EtOH]m. Treatments include packages
with C02 removed from headspaces. All low 02 limits determined by [EtOH],.,,

accumulation.

81

 

 

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levels. r2 = 0.948.

82

 

   

  

 

 

 

 

 

 

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steady-state respiration (Day 1), with a range of headspace 02 partial pressures
(‘Redmax McIntosh’, at 20° C, with a low 0; limit of 2.00-2.35 kPa).

83

indicating continued fermentation of tissues (Figure 7), while packages with an
aerobic [EtOH],.,, of approximately 5 10 and 550 ppm, at 0.1/3 and 25°C,
respectively, did not show increased [EtOH]m during the same period.

[EtOHLh increased with the accumulation of EtOH in fermenting tissues.
The usefulness of an observed breakpoint or increase in [EtOH1m to be an
accurate indicator of the low 02 limit was examined. Packages with anaerobic
[02],“ should exhibit elevated [EtOHLh with levels of EtOH increasing with time.
[EtOHLh in packages possessing [02],,“ above the low 0, limit would theoretically
remain at a basal aerobic level. Observation of low level EtOH production by
apples in storage is common (T. Solomos, personal communication). Packages
were sampled at or shortly after it was determined that they had reached steady-
state respiration. Although there were increases with time in [EtOH]m for fruits
determined to have elevated [EtOH]m and assumed to be under anaerobic
conditions (Figure 6), those fruits at higher [02],“ did not exhibit increased
[EtOH]m. N o indication of EtOH production as a result of tissue breakdown was
observed for those packages at higher [02],“ 6 days after steady-state respiration.

For ‘Marshall McIntosh’ fruit with C02 removed from headspaces, the [02],“
at which elevated [EtOHLh were observed increased with increased temperatures
(Figure 6, APPENDIX B). For ‘Golden Delicious’, the low 02 limits did not
increase noticeably from 0.1 to 20°C, increasing from 20 to 25°C (Figure 3,
APPENDIX B). Low 02 limits for fruit stored in packages with CO2 removed

from headspaces are given in Tables 2 and 4 (APPENDIX B), and did not vary

84
from those with [C021m ranging up to 12 kPa (data not shown).

Recent studies of Dadzie et al., (1992) support the observation that the lowest
P02 at which fruits may be stored is not constant, but is temperature dependent,
and the lowest P02 safe for storage varies to the extent of changes in respiration
rate, temperature quotient (Qw) values (defined as the ratio of change in
respiratory rate with a temperature change of 10°C and with a common value of
approximately 2) and differences in surface resistance to gas diffusion.
Differences in low 02 limit for apple fruit with temperature may be directly
related in part to variable tissue permeabilities and Q“, values, although tissue
resistance to gas exchange has been shown to change significantly with ripening
and aging (Rodriguez et al., 1989), and 0,0 values are not constant and may be as
high as 7 with a change of temperature from O to 10°C (Wills et al., 1989).
Changes in Q“, values for respiration-linked reactions may vary among the
cultivars examined.

The three cultivars examined possess different gas exchange characteristics,
storage characteristics, and vary in susceptibility to storage disorders. ‘McIntosh’
cultivars are generally less permeable to gases and are especially prone to storage
disorders, including low 02 damage (Fidler et al., 1973; Meheriuk, 1989).
‘Marshall McIntosh’ are predisposed to storage damage and are particularly
problematic for storage. ‘Golden Delicious’ and ‘Red Delicious’ cultivars, in
contrast, tend to be more permeable to gas exchange and less susceptible to

atmosphere-related storage disorders. An interdependence of fruit morphology,

85

gas exchange characteristics, lowest P02 safe for storage, respiration rates, and
susceptibility to low 02 and high CO, storage disorders is evident with
comparisons of low 02 limits and fruit characteristics.

The determination of the low 02 limit of ‘Marshall McIntosh’ was difficult due
to variations in [02],“ at which EtOH accumulated for individual fruits and
packages. Variable [02]," for EtOH accumulation were accompanied by variable
RQ breakpoints (Chapter 2), indicating fruit-to-fruit differences in gas exchange
characteristics. Burg and Burg (1965) concluded that unblocked, open lenticels
serve as a primary route for gas exchange of apple fruit. The differences in gas
exchange characteristics of individual fruits may be the result of differences in the
amount of cuticular waxes or the number of open lenticels present on individual
fruit. It would also be expected that variations in gas exchange characteristics and
low 02 limits may be seen with variable growth conditions. Climate and
environmental factors such as growing season moisture levels, humidity, and air
pollution may alter the makeup and amount of cuticular waxes present, affecting
the number of open lenticels closed by natural waxing. Observations of unwaxed
orange (Citrus sinensis L. Osbeck) fruit and grapefruit (Citrus paradisi Macf.) fruit
peels revealed lenticels were often clogged by natural waxing or foreign debris
(Ben-Yehoshua et al., 1985).

In the case of apple fruit with cuticles possessing few lenticels and heavy
waxing, as is the case with ‘Marshall McIntosh’, the lenticels appear less likely to

be the sole route of gas exchange. We cannot assume, for all cultivars of apple

86

fruit, that the number of open lenticels, pore size, and the diffusive resistance of
the cuticle serve as the sole determinants of gas exchange, thereby determining
low 0, limits for storage. The flooding of intercellular air space, measured at 30-
35% of the total volume of apple fruit (Smith, 1947), would alter the diffusive
resistance to gases, especially gases with low solubility in H20 such as 02.

Watercore, a disorder seen especially in over-mature fruits, would alter gas

 

exchange properties, increasing tissue resistance, and increase the low 02 limit. 11.,

While accumulation of EtOH in the headspace of modified atmosphere
packages appears to be a quick and accurate means to determine the low 0; limit
of apple fruit using a limited number of fruit, it does not provide a definite
indication of actual fruit damage and propensity for the development of storage
disorders. The studies of Lau (1989), examining fruit tolerance to anaerobic
conditions, showed that while 61% of ‘Spartan’ apple fruit showed discoloration
after 7 months at 0.5% 02, 8% of ‘Red Delicious’ apples discolored. The
development of storage disorders, including low 02 damage, appears to be cultivar
dependant with variations in tolerance to intermittent or continuous anaerobic
conditions. While EtOH has not been determined to be the cause of any storage
disorder, it may indicate tissue fermentation, which is a commonly associated
symptom of many storage disorders.

The results of this study underscore the possible problems in developing and
implementing commercial MAP of fruits and vegetables through the marketing

chain, given the current packaging films available and their permeability

87

characteristics. Data such as that generated herein may be used to calculate
proper combinations of produce weight, package size, and film thickness to
generate low 02 and elevated CO, atmospheres, which in turn may extend storage
life and maintain high fruit quality. For commodities exposed to a range of
temperatures in transit and holding, rates of respiration would fluctuate, causing
package [02],.“ to decline to P02 below the anaerobic extinction point.
Temperature increases would thereby induce tissue fermentation, while
temperature decreases would elevate P02, such that shelflife would not be
extended. Recent review articles by those working with the storage and transit of
horticultural commodities reiterate these concerns for commercial applicability of
MAP technology given the temperature exposures from harvest to consumer
(Sharp et al., 1993), the properties of films available for MAP, and the inability to
control atmosphere compositions with changes in temperature (M.E. Saltveit,
personal communication).

The determination of low 02 limits by EtOH accumulation, compared with
calculations of R0 values and R0 breakpoints (Chapter 2), another indicator of
low 0, limits, establish higher low 02 limits, with disparity in the determined low
02 limits increasing with increasing temperatures for both strains of ‘McIntosh’.
Previous experiments examining low 02 limits of ‘I.aw Rome’ apple fruit indicated
RQ breakpoints and [EtOHLfl accumulation occurred at the same [02],“ (Gran
and Beaudry, 1993). The previous experiment, however, was conducted at 0.1° C

and it appears that differences between the techniques in determining low 0;

88

limits increased with increased temperatures. Law Oz limits determined by
[EtOH],., accumulation are very close to the atmospheres recommended for
commercial storage, while comparison of RQ-determined low 02 values suggest
slightly lower than recommended storage atmospheres at 0.1 and 3° C.

Using MAP to determine the low 0, limits by means of [EtOHLh '
accumulation is a simple and accurate technique. Results are similar to previous r
large scale, commercial storage studies. The procedures do not require extensive
facilities or the large quantity of fruit as is the case for CA storage experiments.
While technical problems may plague the commercial application of MAP
technologies, the system may prove a useful tool for applied studies of ripening
and stored fruit physiology.

Differences in gas exchange characteristics must be considered for storage
atmosphere recommendations and are likely to be helpful in establishing storage
atmosphere regimes for new varieties. Gas exchange characteristics should not be
overlooked as a factor affecting successful storage on a year to year basis.
Commercial storage recommendations for P02 continue to decrease, with ultra low
0, storage (1 kPa or less P02) atmospheres increasingly used. Further reductions
of P02 for storage will accentuate the consequence of a variation in gas exchange
characteristics. Qfltivar-specific data relating fruit-to-fruit variability of gas

exchange characteristics may prove important for successful storage in the future.

 

REFERENCES

89

REFERENCES

Beaudry, R.M., Cameron, AC, Shirazi, A., and Dostal-Iange, D.L. 1992.
Modified-atmosphere packaging of blueberry fruit: Effect of temperature on
package oxygen and carbon dioxide. J. Amer. Soc. Hort. Sci. 117(3):436-441.

Ben-Yehoshua, S., Burg, SP, and Young, R. 1985. Resistance of citrus fruit mass
transport of water vapor and other gases. Plant Physiol. 79:1048-1053.

Burg, SP, and Burg, EA. 1965. Gas exchange in fruits. Physiol. Plant. 18:870-
884.

Cameron, A.C., Boylan-Pett, W., and Lee, J. 1989. Design of modified-
atmosphere packaging systems: Modeling oxygen concentrations within sealed
packages of tomato fruits. Inst. Food Tech. 54:1413-1416, 1421.

Dadzie, B.K., Banks, N.H., Cleland, D.J., and Hewett, E.W. 1992. Role of skin
resistance in the response of fruits to modified atmospheres. In Abstracts of
Postharvest 1992 International Symposium, University of California, Davis. 13.

Dilley, DR. 1989. 1989 recommended storage conditions for Michigan apples.
Michigan State University Controlled Atmosphere Clinic Handbook. 51.

Fidler, J .C., and North, CJ. 1967. The effects of conditions of storage on the
respiration of apples. 1. The effects of temperature and concentrations of

carbon dioxide and oxygen on the production of carbon dioxide and the uptake
of oxygen. J. Hort. Sci. 42:189-206.

Fidler, J .C., Wilkinson, B.G., Edney, KL, and Sharples, RD. 1973. Physiological
Disorders. In The Biology of Apple and Pear Storage. Commonwealth
Agricultural Bureaux. pp. 235.

Gran, CD, and Beaudry, RM. 1993. Determination of the low oxygen limit for
several commercial apple cultivars by respiratory quotient breakpoint.
Postharvest Biol. Technol. (in press).

90

Harger, R.N., Raney, B.B., Bridwell, E.G., and Kitchell, M.F. 1950. The partition
ratio of alcohol between air and water, urine, and blood; Estimation and

identification of alcohol in these liquids from analysis of air equilibrated with
them. J. Biol. Chem. (183):197-213.

Lau, 0.1.. 1989. Control of storage scald in ‘Delicious’ apples by diphenylamine,
low oxygen atmosphere, and ethylene scrubbing. Proc. 5th Int. C.A. Res. Conf.
(Wenatchee). 5:169-176.

Meheriuk, M. 1989. CA storage of apples. Proc. 5th Int. C.A. Res. Conf.
(Wenatchee). (2):257-284.

Meigh, DP. 1970. Apple Scald. The Biochemistry of Fruits and Their Products.
A.C. Hulme, Ed. Acad. Press. 555-569.

Pesis, E., and Avissar, I. 1990. The effect of postharvest applications of
acetaldehyde vapour on strawberry decay, taste and certain volatiles. J. Sci.
Food Agric. 52:377-385.

Rodriguez, L., Devon, 2., and Kader, AA. 1989. Relation between gas diffusion
resistance and ripening in fruits. 5th Proc. Int. C.A. Res. Conf. (Wenatchee).
(2):1-7.

Sharp, AK, Irving, AR., and Morris, SC. 1993. Does temperature variation limit
the use of MA packaging during shipment in freight containers? Proc. 6th Int.
C.A. Res. Conf. (Ithaca). (in press).

Smith, H.W. 1947. A new method of determining the composition of the internal
atmosphere of fleshy plant organs. Ann. Bot. 11:363-368.

Tomkins, KG. 1966. The storage of fruits and vegetables. Small scale gas
storage experiments. Rep. Fd. Invest Bd. for 1965-1966. 8-10.

Wills, R.B.H., McGlasson, W.B., Graham, D., Lee, TH, and Hall, E.G. 1989.
Postharvest. An Introduction to the Physiology and Handling of Fruit and
Vegetables. New South Wales University Press, Australia. pp. 174.

Summary and Conclusions

91

92
Low 0, Limits and Storage Studies

Results from two years of experimentation show low 0, limits vary with
temperature, cultivar, and slightly from year to year. Low 02 limits increased with
increasing temperature for all cultivars examined, using both RQ breakpoint and
elevated EtOH levels as indicators. The extent to which increasing temperatures
affected low 0, limits varied highly between cultivars, with low 0, limits for
‘McIntosh’ fruits, particularly the ‘Marshall’ strain, changing an 4 kPa 0, from 3 to
25 °C.

Low 0, limit values determined by RQ breakpoint were slightly lower than
those indicated by elevated [EtOH]m. Low 02 limits indicated by elevated
[EtOH]m compared closely with commercially recommended storage 02
atmospheres. Differences in low 02 limits among the various cultivars examined
may be related to tissue gas exchange characteristics.

Variations in Low 0, Limits Determined by RQ Breakpoint and Elevated
Headspace EtOH Levels

The extensive storage studies initiated by Kidd and West and continued by
Fidler and North at Ditton Laboratory in England established general
physiological characteristics of fruit ripening and climacteric respiration. In these
studies, RQ values associated with fruits in aerobic atmospheres were found to
rise with increased temperature, be reduced with increased Pm, and normally
ranged from 1.5 to 1.8, but up to 3.0 under certain conditions. The results of this

study confirm many previous results and thereby demonstrate the usefulness of

93

MAP as a tool to examine the physiology of fruits in MA storage.

An interesting note from the study, particularly when examining the effects of
temperature on low 02 limits, is the apparent difference in low 02 limit
determined by RQ breakpoint and the low 02 limit, or anaerobic extinction point,
determined by headspace EtOH accumulation. In almost every instance, RQ
breakpoint values determined and taken to represent low 0, limits were slightly
lower than those determined by elevated headspace EtOH levels.

Why is there an apparent difference in the determined low 0; limit with
alternate methods? Both methods rely upon physiological characteristics rather
than empirical determination of threshold levels below which excessive damage
occurs. One primary difference is the dependence of R0 breakpoints upon
calculated flux levels, while headspace EtOH levels are measured directly.
Differences in actual and measured fihn P'Cm would result in miscalculated RQ
values. This error would primarily affect basal RQ values, while affecting RQ
breakpoints to a lesser extent. Errors in calculated [02],“, however, could effect
both aerobic RQ levels and the determination of the [02],,“ at which the RQ
breakpoint occurs.

A possible cause of variation in low 0, limit determined by the two methods
may be the differences in tissue permeability for the ‘McIntosh’ strains compared
with ‘Golden Delicious’ and ‘Red Delicious’. Comparisons of apparent whole
fruit K,II for 02 (Figures 14 and 15, Chapter 2) indicate relatively high and low

tissue gas permeabilities for ‘Golden Delicious’ and ‘Marshall McIntosh’,

94

respectively, over a range of temperatures. While the apparent K, for O, of
‘Marshall McIntosh’ increased linearly with increasing temperature, the apparent
K. for 02 for ‘Golden Delicious’ apple fruit increased rapidly from 0 to 10° C and
remained constant at temperatures up to 25° C.

An additional indication of respiration being limited by tissue permeability for
‘Marshall McIntosh’ is the shape of predicted 02 uptake curves (Figures 10 and
11, Chapter 2). With increased temperatures, predicted 02 uptake curves
approach linearity, indicating limited 02 uptake with any decrease in [02],“ at 25°
C. Predicted 02 uptake curves for ‘Golden Delicious’ (Figure 12 and 13, Chapter
2), in comparison, did not change markedly with increased temperature.

Variations in determined low 02 limits may result from the indicated tissue
permeability differences. Limited cuticle gas permeability may cause 02 and CO;
gradients within the fruit tissue. If the principal route of Oz and CO, gas
exchange into ‘Marshall McIntosh’ is through the calyx pore rather than lenticels,
thought to be the principal route for gas exchange into apple fruits (Burg and
Burg, 1965), those areas with the lowest level of P02 would be directly below the
cuticle rather than at the center of the fruit tissue, as is normally though to be the
case for apple fruit and other bulky fruit tissues. Localized anaerobic areas
created under the cuticle in an aerobic atmosphere for the whole fruit tissue may
induce fermentation and EtOH production in portions of the ‘Marshall McIntosh’

fruit tissue at aerobic O, atmospheres, indicating low O, limits at higher a [02],“.

95
Variations in O, Uptake Between Cultivars

Another issue for examination relates to variations in RROZ between cultivars.
Presentation of this data has brought differing opinions from fellow scientists,
some believing this to be an actual phenomenon related to limiting RRoz with
variation in flesh permeability, others believing differences to be artifacts of the
experimental methods. Further examination of this is required. Arguments that
the apparent differences are actually artifacts include the heritable enzymatic
systems involved in fruit respiration. Any differences in RRoz indicate possible
differences in K,n values for 02. Apparent KIll for O, for whole fruit is not a
function of cytochrome oxidase alone, but involves other oxidases, as well as
whole tissue gas permeability to Oz. Differences in K“, for 0, may be indicative
of the extent to which tissue resistance to gas diffusion limits respiration of
individual fruit or fruit cultivars.

Variations in tissue P‘m, P'cm, P2,, P',32m (Burg and Burg, 1965; Cameron,
1982; Ben-Yehoshua et al., 1985; Rodriguez et al., 1989) have been shown for a
number of fruit tissues throughout the ripening process. Variations for these
tissue permeabilities have also been shown between individual cultivars of apple
fruit (Park, 1990).

The internal atmospheres of fruits, both stored in air and under CA
conditions, have been examined from the advent of CA storage (Kidd and West,
1949), with 02 and CO, gradients found for various biological gases. The tissue

gradients for various physiologic gases appear to be transient (Kidd and West,

96

1949), vary from year to year and between individual fruits and regions of each
fruit (Dadzie et al., 1992), and change significantly with maturation and ripening
(Kidd and West, 1949). MAP techniques may be useful in the examination of gas
exchange and internal gas gradients of fruits.

Problems of Modified Atmosphere Packaging Examination of Fruit Storage

One problem associated with the use of MAP techniques for examination of
fruit physiology is the dependance of atmosphere establishment upon respiration,
the long length of time for atmosphere pulldown at low temperatures, and
changes in respiration with time. With delayed atmosphere establishment,
ripening would progress in packages, allowing changes in carbohydrate and
organic acid content. Such changes are detrimental for stored fruits, and would
not be representative of those conditions desirable for commercial storage studies
and applications.

The influence of CO2 on fruit ripening, and the means by which elevated CO;
partial pressures act to extend CA and MA storage life, are still unresolved.
Further work must determine the specific mechanisms by which the pleiomorphic
effects of C02 are induced. Use of MAP techniques to study these factors may be
limited by the mechanical constraints of the methodology. While it is possible to
completely remove CO, from package headspaces, establishment of precise CO;
levels in combination with specific 0, levels is more difficult, incorporating MAP
and the methods of small scale CA studies. In this system, headspace CO;

atmospheres depend upon film permeability, and with variation in film

97

permeabilities of 10% on any given lot of film, additional experimental error may
be introduced. The increased complexity of experiments and calculations with a
possible decrease in accuracy using MAP techniques to study CO, effects are not
warranted given the availability of CA techniques. Finally, the physiological
responses of whole fruit to C0, are well studied and documented, with studies
using MAP techniques unlikely to increase the information on specific C02
effects.

Further Applications of Modified Atmosphere Packaging Techniques

The experiments conducted demonstrate the usefulness of MAP techniques for
quick and relatively simple determination of low 02 limits. These techniques may
be particularly useful when CA storage of newly introduced cultivars is desired.
When CA storage facilities are not available for storage research, MAP
techniques may serve as a useful substitute.

In addition to the study of MA storage of commodities, MAP techniques may
prove useful for the study of changes in protein content and syntheses induced by
CA and MA storage. Using MAP techniques it is possible to generate a range of
Oz atmospheres in small increments from which samples may be extracted and
analyzed. Use of this tool may allow for quick, accurate generation of samples for
the study of specific enzymes, proteins, or genes which are modified in

transcription, translation, or activation under modified atmospheres.

REFERENCES

98

REFERENCES

Ben-Yehoshua, S., Burg, S.P., and Young, R. 1985. Resistance of citrus fruits to
mass transport of water vapor and other gases. Plant. Physiol. 79:1048-1053.

Burg, S.P., and Burg, EA 1965. Gas exchange in fruits. Physiol. Plant. 18:870-
884.

Cameron, AC. 1982. Gas diffusion in bulky plant organs. PhD thesis, University
of California, Davis. pp. 117.

Dadzie, B.K., Banks, N.H., Cleland, DJ., and Hewett, E.W. 1992. Role of skin
resistance in the response of fruits to modified atmospheres. In Abstracts of
Postharvest 1992 International Symposium, University of California, Davis. 13.

Kidd, F. and West, C. 1949. Resistance of the skin of the apple fruit to gaseous
exchange. Rep. Fd. Invest. Bd. for 1939. 59-64.

Park, Y.M. 1990. Gas exchange in apples: Pathway for gas exchange, changes in
resistance to gas diffusion during fruit development and storage, and the
factors affecting the change. PhD. Thesis, Cornell University. pp.116.

Rodriguez, L., Devon, 2., and Kader, AA. 1989. Relation between gas diffusion
resistance and ripening in fruits. 5th Proc. Int. C.A. Res. Conf. (Wenatchee).
(2):1-7.

APPENDIX A

99

100

The following data constitute the C02 production rates for 8 cultivars of apple
fruit packaged at 0.1° C for the first years experiments (1991). This data is

presented as a supplement to data included in Chapter 1.

101

 

 

0'1: rVVV‘VVVjVVVVTVVVI’VfV' 00" VVVrVV V I VV V'V V V' V VV

   
 
   
    
   
 
     

 

 

 

 

 

-1 0 J 0 ‘
1 0.101 1 0.10.
0.101 1 o.1o- -
AA 4
A A 996‘ ‘
A -
A ‘ AAA“ 4
Red Fuji.
'IV" "T1"‘1*"
4 5 12 16 20

 

 

 

 

 

 

 

 

 

CO2 Production (mmol-kg"-h“)

1
fi
1 c1 4
A
.1 . A
28? § 3 1‘ A an : ° 4
A g A
0.06 .. 0,05- A26 A A ..
A . . A” .
Law Rome, . Marshall McIntosh
00m rVVV‘lV—VV 'V V V ' r1 V ' VVV cum I V VI VVV V l V V V I V V VI V V V
0 4 8 12 10 20 0 4 0 12 10
06‘s 00‘: V V V l V V V I T V V l V V V l V V V
q o n
It 0'1 C q
0.10 0.10- -

1 .1

q a

4 A A A
AA A -1 1 A %4
A A _, 0.05 A w A gab & A q

 

0.05 3‘ ‘ “AAA

Red Delicious.

0 UN V V T V T V V V V V V V V V V
(’1 ' 8 1'2 1'6 20

Steady State 02 (kPa)

 

 

 

12 16 20

Figure 1. Effect of steady state 0; partial pressure on C0; production of 8
cultivars of apple fruit at O.1°C.

APPENDIX B

102

103

The following data are derived from the second year of experiments (1992).
This material consists of raw data for 02 production rates for the 3 cultivars of
apple fruit examined with best fit curves and equations describing those curves,
the raw data for SAS' predicted 02 uptake analysis for ‘Marshall McIntosh’ and
‘Golden Delicious’ cultivars, the headspace [EtOHLu for packages of ‘Marshall
McIntosh’ and ‘Golden Delicious’ with C02 removed by scrubbing with hydrated
lime [Ca(OH)2], and the low 02 limits estimated for these varieties using

headspace EtOH as the indicator.

104

 

 

 

 

0.3
(D _
x
B 7,5 ‘
g E 0.2—«
6 :3 ‘ ° 0 889%0 0 ° <2;
E) E d 00 ° 00
U ——+
‘1’ V ‘ %&i
0. _
(3.0—fie I I I l Golden DeliICious(— C302)I
0.00 0.10 0.20 0. 30

O2 Uptake (kPa)

Figure 1. 02 uptake and predicted 02 uptake for ‘Golden Delicious’ apple fruit,
with CO2 removed from package headspaces, over a range of temperatures, using
SAS nonlinear regression fit.

 

 

 

 

0.3
Q) —4
x A
*3 T 0 0 Q.) o o o
L 0 2— o o
3 -C 4 o
N '- 00
o :9 _ o q)
(l) 0 (£0 08 O
73 E o o
'6 E 00
0) v
L
Q
Golden Delicious
0.0 I I I T I I T I I I I

 

0.00 0.10 0.20 0.30
02 Uptake (kPa)

Figure 2. 02 uptake and predicted 02 uptake for ‘Golden Delicious’ apple fruit
over a range of temperatures, using SAS' nonlinear regression fit.

Predicted O2 Uptake

(mmol-kg"-hr“)

105

 

 

 

 

0.3

I
0.2-1

fl ° ° °°9

00 090m
4 o8 0%09’ 0 0‘9 0
0 0° ° 0

01‘ gaoggooo O

1 fl" °°°

d Marshall McIntosh (—C02)
0.0 I I I I I I I I I

l
0.10

0.20 0.30

02 Uptake (kPa)

Figure 3. 02 uptake and predicted 02 uptake for ‘Marshall McIntosh’ apple fruit,
with CO2 removed from package headspaces, over at a range of temperatures,
using SAS' nonlinear regression fit.

Predicted O2 Uptake

(mmol-kg“-hr“)

 

0.3

—<
.1

'1

 

 

 

Marshall McIntosh

 

I I I I

0.10

I I I I I I
0.20 0.30

02 Uptake (kPa)

Figure 4. 02 uptake and predicted 02 uptake for ‘Marshall McIntosh’ apple fruit
over a range of temperatures using SAS' nonlinear regression fit.

106

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

0150 V l V I V I V V I T V l V V1 0.40‘ V V I V I I V I V I V V d
o ‘ " o '1
0.40.1 25 C I. 0.32.“ 20 C J
'1 'l -1
c1 4 1 cl
0.301 1 0 J
1 I 1
:1 O O A ——t\ J I‘
0.20: a) U0 - - 1 %
0.101 O" 1 1
C. ‘ 1 l I 1
| 0.00 v v I 1% I fl I 1 V I v v I t 7 0.00 Vfi I V V I V T I 1 V I V v I V v
.C o 3 6 9 12 15 18 o 3 6 9 12 15 18
T 0020 V V ' bV ' V—r I V V I V V t V V 00‘: V V I V V ' fr ' V V 1' V V I V V ‘
'l o
3‘ 0.151 ° 15°C 10c: i
— I o 1
O 0.121 n 1
1 o
E 1 o 0 I
V 0.041 1
Q) i I
xo.oc..,..,..,..,..,.- ..,.
C o 3 6 9 12 15 15 18
4.;
Q 0.10d V V Ifi' V—I V V I V V I Y‘ 1 r V T 4 001c‘—r V—I V_V_ r V ‘V I V I V l V V J
3 : . 1 : . :
0.081 o 5 C - 0.08- 3 C 1
N 1 o 1 1 1
O 0.061 10.061 0 1
. . 3 O O 1
0.04- 1‘ 0.041 J
. i I o 1
. o . .
0.021 1 0.021 0° ° 1
0.00 V V I VVV l V V I rf' er V V 0.00 V V V V V1 V VjVV V I V l V V
o 3 6 9 12 15 18 o 3 6 9 12 15 18

Steady State 02 (kPa)

Figure 5. Effect of temperature (3, S, 10, 15, 20, and 25°C) on RRo, of ‘Marshall
McIntosh’ apple fruit. See Table 1 for equations describing curves.

107

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

0050‘ V V ' V V I VV I VV I V V I V V Oow‘ V V I V V I V V ' V V T V V l V V
d o " o
0.40: 25 C 20 C
0.301 ':
0.201 ° ° 1
. o 0 o 1
0.10: 1 .1
A i ‘ :
| 0.00 V V V V I V r I V V I TV I V V 00% V V ' V V ' VV 1 V V r V V I V V
1: 0 3 6 12 15 18 0 3 6 9 12 15 18
T 0020+ V V T V V I V V I V V T VVr I V V 4 0.154 V V r V V l V V I V V I V V I V V ‘
O 1 ° 1 1 ° 1
.x 0.161 15 C 1 0.121 10 C 1
_._ 1 O 1 1 .1
1 I d 1
CE) 0.121 Q) 1 0.091 1
: o I I ‘
E 0.08: O O -1 0.06: ‘1
q 1 q d
V 0041 o 1 0.031 1
(D : 1 i
x 0.00 VI V V l V V I VV‘I V V I V V 0.00 V V I V V l V V l V V l V V l' V V
O 0 3 6 9 12 15 18 0 3 6 12 15 18
4—1
(10101., l..,..T..Tfi10.1c..,..,., ,. , 1
D . : : . :
0.081 o 5 C 1 0.081 3 C 1
ON : : :o :
0.061 o 1 0.06- 0 1
1 o 1 1 II
1 1 I o d
J a 1 .
0.04.: 1 0.04: 0 j
: o ‘ : 0° ‘
0.021 1 0.021 0 0° 1
0.00 V V I V V I V V I V V I V V I VV . 00°C V V l V V I V V l V V l V V I V V
0 3 6 12 15 18 0 3 6 12 15 18

Steady State 02 (kPa)

Figure 6. Effect of temperature (3, 5, 10, 15, 20, and 25°C) on RRo, of ‘Redmax
McIntosh’ apple fruit. See Table 1 for equations describing curves.

108

 

 

 

 

0.50 V T V V 1 V V I V V ‘ o.“ 1
0 . .

0.40 25 C 1 0.32 1
0.30 1 0.24 1
O " 1

on u «1

0 0° : 1

GAO

 

 

 

 

 

'8

TV'VVVIVV'VV‘VV OIOCIV

0'3 6 9121518

 

 

    

0.20 .,..,+.,..,..,..10.15 1
0.16 150C 1012 1
0.12 10.09 1
0.08 ° 10.06 1

fi

1 0.03

llllll

 

 

 

 

d
IrTrI'ITIr1II 0.00lvv

O 3 6 9 12 15 18

F’
c:
<3

 

d
00

 

V'VV'VV.OO1C
5C 10.08

 

O2 Uptake (mmol°kg"-h“)

(106

AIL].

(L04

llellllJlllJl

I'LL

1 0.02

 

14111

 

 

V V I VV' VV‘OIOC' V V l V V l V V V
12 15 18 0 3 6 9 12 15

Steady State 02 (kPa)

 

 

 

.3
m

Figure 7. Effect of temperature (0.1, S, 10, 15, 20, and 25°C) on RRm of ‘Golden
Delicious’ apple fruit. See Table 1 for equations describing curves.

109

 

 

0050 V V ' V V r f V r T V I V V I V r 004°

0.40 o 25 C

1114‘!

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

0 I1
0.32 20 C 1
0.30 10.24 1
0.20 10.16 1
0.10 10.08 1
A ; -
| 0.00 V V OOOCI T V I V V I V V I V V V l V V
.C 18 0 3 6 9 12 15 18
T 0020 r V l V V I V V l V V I V V I V V ‘ 0": Vi ‘ V V [j V I V V ' V V I V V.
O . : . :
x 0.16 150 10.12 10C 1
.1:— 0 j j
(E) 0.12 10.09 1
E 008 o 10.06 1
1 1
V 0.04 1 0.03 1
Q) I
x0001 Iv't"1"t"rfi O'OCI‘TI"I"I"I"I"
O 0 3 6 9 12 1 18 o 3 6 9 12 15 18
4.;
Q0112 .....r..j-.,..,..10.101...fi,-.,.-r..,fir1
D . : : . :
0.08 5 C .. 0,03- 0.1 C 2
N : : :
O 0.06 10.061 1
1 I o 2
0.04 -1 0.04-1 0° 0 00 -1
i 1 o :
1 1 o :1
0.02 10.02.:W0 .1
1 ‘ 1
0900 l V f‘ V V 1 V V l V V I T V I V V 0000 V V V V V I VV' V V

 

 

 

o.—
{D—

0 3 6 91215180 3'

Steady State 02 (kPa)

12 15

.5
0

Figure 8. Effect of temperature (0.1, 5, 10, 15, 20, and 25°C) on RRm of ‘Early
Red One Red Delicious’ apple fruit. See Table 1 for equations describing curves.

110

Table 1. General equations (Eq. (1)) and values of constants describing the
relationship between steady state 0; partial pressure (kPa) and 02 uptake
(mmol-kg‘1 oh") for 3 apple cultivars in modified atmosphere packages over a
range of temperatures. Eq.(l): RR.» = bl -{1-exp[—b2-(Oz)m]}"3

 

 

Cllltivar T b1 b2 b3 1'2
(°C)
‘Marshall McIntosh’ 3 0.03858 1.04802 1.07847 0.357

5 0.09628 0.23805 0.82028 0.787
10 0.08918 0.36431 0.91880 0.652
15 0.11381 1.16961 7.14997 0.469
20 0.22397 0.32543 1.45 153 0.465
25 0.20351 0.20891 0.87544 0.531

 

‘Redmax McIntosh’ 5 0.04790 1.21530 1.19230 0.465
10 0.07677 1.40427 1.84727 0.445
15 0.10944 1.05992 1.56692 0.551
20 0.18612 0.31237 0.69226 0.663
25 0.18529 0.60942 1.34822 0.798

 

‘Golden Delicious’ 0.1 0.03927 0.32535 0.3136 0.259
5 0.05529 0.51940 0.42338 0.629
10 0.13979 0.07294 0.29358 0.436
15 0.13571 0.58672 0.52379 0.521
20 0.18988 0.48562 0.48583 0.545
25 0.20109 0.69397 0.34256 0.461

 

‘Early Red One Red Delicious’ 0.1 0.03818 0.09332 0.22647 0.306
5 0.06949 0.18928 0.35375 0.876
10 0.06669 2.34255 10.3644 0.607
15 0.11496 0.86286 0.69101 0.616
20 0.38478 0.07736 0.48758 0.850
25 0.38988 0.07546 0.48661 0.85 1

 

 

 

 

 

 

111

 

   
   

 

 

 

 

 

 

 

 

 

0.25 *

 

 

 

 

 

 

 

 

 

 

 

 

04' I I I I I I I I I I I I I I r I T q I I I I I r I I I I I BI I I I I q
0 ° 2 0 ° 2
0.28 25 C 1 0.20 20 C 1
1 ° 3
0.21 - 0.15 0 0 —
o“ 0 o :
0.14 0 ° -' 0.10 ° 0° 0 -‘
o 1 1
0.07 4 0.05 1
A 3 j
7 00m I I I I I I I I I I I I I I I I I I 0.00 I I I I I I Ij I I I I I I II 1 I I
.C 0 3 6 9 12 15 18 0 3 6 9 12 15 18
T 0.15 I I I I I I I I I I I I I I L I 001: 1 I I I I I I I I I I I I Ifi' 1 I I ‘
3 ° ° 15 c 10°C ° 1
CE) ° 0 15 Q
o o oo _‘
é a
Q) 1
x r I I I I I I T IOCOO lfi' r I I I I I I I 1 I I rT I I I
O 12 15 18 3 6 9 12 15 18
4—0
Q 0010 7 I I I I I I I I I III I I I I I I q 0010 . I I I I I I I I I I I I I I I I I d
D o 2 0 2
0.08 5 C - 0.08 0-1 C -
N 0 : :
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Steady State 02 (kPa)

Figure 9. Effect of steady state 02 partial pressure on the 02 uptake of ‘Golden
Delicious’ apple fruit over a range of temperatures with C02 removed from
package headspaces. See Table 2 for equations describing 02 uptake curves.

112

Table 2. General equations (Eq. (1)) and values of constants describing the
relationship between steady state 02 partial pressure (kPa) and 02 uptake
(mmol -kg" oh") for ‘Golden Delicious’ apple fruit over a range of temperatures
with C02 removed from package headspaces.

 

 

 

 

T (°C) b, 62 b, r2
0.1 0.03499 0.11569 0.10414 0.070
5 0.06087 1.04650 0.61064 0.495
10 0.08503 0.43285 0.41455 0.588
15 0.11967 0.39001 0.45600 0.459
20 0.15026 0.74490 0.32932 0.398
25 0.52207 0.00292 0.25203 0.354

 

 

 

 

 

Table 3. Estimated low 02 limits, as indicated by elevated headspace EtOH, of
‘Golden Delicious’ apple fruit sealed in packages with C02 removed from the
headspaces, over a range of temperatures.

 

 

 

 

T (°C) Estimated low 0; limit (kPa)
0.1 ~ 0.90
5 070-075
10 0.55-0.70
15 0.70-1.00
20 GAO-0.90
25 . 0.95-2.40

 

 

113

 

 

 

 

 

 

 

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Steady State 02 (kPa)

Figure 10. Effect of steady state 0; partial pressure on the low 02 limit, as
indicated by elevated headspace EtOH levels, of ‘Golden Delicious’ apple fruit
over a range of temperatures with C02 removed from the package headspaces.
See Table 3 for estimated low 02 limits.

114

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Steady State 02 (kPa)

Figure 11. Effect of steady state 0; partial pressures on 02 uptake of ‘Marshall
McIntosh’ apple fruit over a range of temperatures with C02 removed from
package headspaces. See Table 4 for equations describing curves.

115

Table 4. General equations (Eq.(1)) and values of constants describing the
relationship between steady state 0; partial pressures (kPa) and 02 uptake

(mmol ~kg‘l -h") for ‘Marshall McIntosh’ apple fruit over a range of temperatures

with CO, removed from package headspaces.

 

 

T (°C) b, b, b3 :2
3 0.06152 0.18805 0.71192 0.730
5 0.05156 0.74187 1.41744 0.609
10 0.13914 0.15775 1.14515 0.712
15 0.08079 0.89873 2.46464 0.291
20 0.12236 0.42082 0.93988 0.104
25 0.17133 0.87569 6.08627 0.280

 

 

 

 

Table 5. Estimated low 02 limits, as indicated by elevated headspace EtOH, for

 

 

 

 

 

‘Marshall McIntosh’ apple fruit sealed in packages with C02 removed from the
package headspaces, stored over a range of temperatures.

 

 

 

 

1.65-2.20
1.45-1.65
2.75-3.30
2.60-4.00
3.45-3.50

116

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Steady State 02 (kPa)

Figure 12. Effect of steady state 02 partial pressure on the low 0; limit, as
indicated by elevated headspace EtOH levels, of ‘Marshall McIntosh’ apple fruit
over a range of temperatures with CO2 removed from package headspaces. See
Table 5 for estimated low 02 limits.

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