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Development, Evaluation and Application
Casein-Based Edible Films

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DEVELOPMENT, EVALUATION, AND APPLICATION OF CASEIN-BASED
EDIBLE FILMS

By

Jay Lyle Chick

A THESIS
Submitted to
Michigan State University
in partial ﬁilﬁllment of the requirements
for the degree of

MASTER OF SCIENCE

Department of Food Science and Human Nutrition

1 996

ABSTRACT

DEVELOPMENT, EVALUATION, AND APPLICATION OF CASEIN-BASED
EDIBLE FILMS

By
Jay Lyle Chick

This study was conducted to determine the effect of protein type, plasticizer type,
and protein to plasticizer concentration on properties of casein-based ﬁlms, and then
applying ﬁlms to a food system. Films were produced from lactic acid or rennet
precipitated casein, and either sorbitol or glycerol, mixed in distilled water. A lactic acid
casein and sorbitol ﬁlm and a lactic acid casein and glycerol ﬁlm were used to wrap
processed cheese slices, with low density polyethylene (LDPE) wrapped and unwrapped
samples used as controls. Films were tested for barrier properties, water vapor and
oxygen permeability, and mechanical properties, elongation and tensile strength. The
cheese slices and ﬁlms were tested for moisture content and color change over a 30 day
storage period (22°C, 88% relative humidity).

Films produced displayed good oxygen barrier properties, but poor water barrier
properties compared to synthetic ﬁlms. Films made with sorbitol exhibited signiﬁcantly
better (p<0.05) water barrier and tensile properties than those made with glycerol. Higher
protein concentrations also produced stronger ﬁlms. Processed cheese slices wrapped in
casein-based ﬁlms lost a signiﬁcant amount of moisture (p<0.05) as compared to the
LDPE wrapped control slices, which was the cause of a signiﬁcant color change (p<0.05)
of the cheese slices. However, moisture lost by the cheese slices was retained in the

casein-based ﬁlms, shown by a large increase in moisture content of these ﬁlms.

Dedicated to my Parents, Lyle and Lois Ann Chick,
for all their love, support, patience, and

raising me to be the person I am today

iii

ACKNOWLEDGMENTS

I would like to give a special thanks to my advisor Dr. Zeynep Ustunol, ﬁrst for
giving me the opportunity to continue my education and also for the tremendous amount
of help and guidance she gave me throughout graduate program. Her friendship and
guidance helped push me to expand myself educationally and professionally.

I would like to thank my committee members Dr. J.P. Partridge, and Dr. B. Harte
for their guidance throughout my graduate program. I would also like to thank the
Michigan State Agricultural Experiment Station and the State of Michigan Research
Excellence Funds via the Crop and Food Bioprocessing Center for their partial support of
this research. New Zealand Milk Products Inc. is also acknowledged for providing me
with the casein used in this study.

A special thanks goes to my brothers and sisters Jeff and Lisa, Jackie and Russ,
Jerry, and Jon. I also want to thank Dr. Luis Rayas, Gineth Trank, and Dr. Virginia Vega-
Wamer for their much appreciated help with lab techniques, and my lab mates Chris,
Renee, Xuemei, Han, Seong-Joo Kim, Dr. Jong Hwa Lee, Dr. H.K. Jeong, Dr. Choi Lan
Ha, Julie, Heather, and Manee for making it fun to be in the lab. A big thanks to Eric
Cole, Jamie Merritt, Alicia Orta-Ramirez, Chris Daubert, Anne Smyth, Sheau-shya Wu,
and all my other good ﬁiends I have made during graduate school for their friendship and

making my decision to go back to school one of the best ones I have ever made.

iv

TABLE OF CONTENTS

page
LIST OF TABLES .......................................................................... viii
LIST OF FIGURES ........................................................................ x
INTRODUCTION ......................................................................... 1
REVIEW OF LITERATURE ........................................................... 4
Formation of Edible Films and Coatings .................................... 4
Components of edible ﬁlms ............................................... 4
Processes of edible ﬁlm and coating formation ........................ 6
Properties of Films Important for Food Applications ...................... 7
Barrier ....................................................................... 7
Mechanical .................................................................. 13
Properties of Protein-Based Films .............................................. 14
Water vapor permeability ................................................. 14
Oxygen permeability ...................................................... 15
Tensile strength and elongation .......................................... 16
Properties of Milk Protein-Based Films ....................................... 17
Water vapor permeability .................................................. 17
Oxygen permeability ...................................................... 19
Tensile strength and elongation .......................................... l9

Milk protein-based edible ﬁlm and coating applications .............. 20

Properties of Plasticizers ........................................................ 21
Properties of Milk Proteins ...................................................... 22
Casein ...................................................................... 22

The casein micelle .......................................................... 26

Casein manufacturing ...................................................... 28
MATERIALS AND METHODS ........................................................ 30
Film Components and Formation .............................................. 3O
Thickness ............................................................................ 35
Water Vapor Permeability ...................................................... 35
Oxygen Permeability ............................................................. 36
Mechanical Properties ........................................................... 37
Storage Study ...................................................................... 38
Statistical Analysis ................................................................ 40
RESULTS AND DISCUSSION .......................................................... 41
Film Development ................................................................. 41
Barrier Properties ................................................................. 41
Water vapor permeability ................................................. 41
Oxygen permeability ...................................................... 46
Mechanical Properties ........................................................... SO
Tensile strength ............................................................. 50
Elongation .................................................................. 54

vi

Comparison to Other Films ..................................................... 58

Storage Study ...................................................................... 61
Moisture content ........................................................... 61

Color ........................................................................ 65
CONCLUSIONS ........................................................................... 71
RECOMMENDATIONS ................................................................. 72
LIST OF REFERENCES ................................................................. 73

vii

LIST OF TABLES

Table 1. Compositions of Caseins Used .................................................. 31
Table 2. Compositions of Casein-Based Edible Films Produced ..................... 33

Table 3. Formulations of Film Forming Solutions for
Casein-Based Edible Films ................................................ 34

Table 4. Eﬁ‘ect of Plasticizer Type on Water Vapor Permeability (WVP)
of Casein-Based Edible Films (3 78°C, 90% RH.) ................... 43

Table 5. Effect of Casein Type on Water Vapor Permeability (WVP)
of Casein-Based Edible Films (378°C, 90% RH.) ................... 44

Table 6. Effect of Plasticizer Type on Oxygen Permeability (OP)
of Casein-Based Edible Films (23°C, 0% RH.) ....................... 47

Table 7. Effect of Casein Type on Oxygen Permeability (OP)
of Casein-Based Edible Films (23°C, 0% RH.) ....................... 48

Table 8. Effect of Plasticizer Type on Tensile Strength (TS) of
Casein-Based Edible Films (23°C, 50% RH.) ........................ 51

Table 9. Eﬁ‘ect of Casein Type on Tensile Strength (TS) of
Casein-Based Edible Films (23°C, 50% RH.) ......................... 52

Table 10. Effect of Plasticizer Type on Elongation (E%) of
Casein-Based Edible Films (23°C, 50% RH.) ......................... 55

Table 11. Effect of Casein Type on Elongation (E%) of
Casein-Based Edible Films (23°C, 50% RH.) ......................... 55

Table 12. Comparison of Selected Casein-Based Films and Synthetic Polymers 59

Table 13. Comparisons of Various Protein-Based Edible Films ...................... 6O

viii

Table 14.

Table 15.

Table 16.

Table 17.

Table 18.

Table 19.

Effect of Storage Time on Moisture Content of Processed Cheese
Slices Packaged in Various Wraps °
(Stored at 22°C, and 88% RH.) ........................................ 62

Effect of Wrap Type on Moisture Content of Processed Cheese
Slices (Stored at 22°C, and 88% RH.) ................................ 63

Eﬁ‘ect of Storage Time on Moisture Content of Edible Films Used
as a Processed Cheese Wrap (Stored at 22°C, and 88% RH.) ..... 64

Weight Gain of Processed Cheese in Different Wraps During
Storage (Stored at 22°C, and 88% RH.) ............................. 66

Effect of Wrap Type on Color Changes in Processed Cheese
During Storage (Stored at 22°C, and 88% RH.) .................... 67

Comparisons of Color Changes of Wraps After Storage on
Processed Cheese (Stored at 22°C, and 88% RH.) ................. 69

LIST OF FIGURES

Figure 1. The Diffusion Cell for Permeability Testing ................................. 10
Figure 2. Mass Transport Proﬁle Curves ................................................ 11
Figure 3. The Casein Micelle (Slattery and Evard Model) ............................. 27
Figure 4. Schematic Diagram of the Film Forming Process ........................... 32

Figure 5. Packaging of Processed Cheese Slices Using
Casein-Based Edible Films ............................................... 39

Figure 6. Water Vapor Permeability (WVP) of Casein-Based
Edible Films (378°C, 90% RH.) ....................................... 45

Figure 7. Oxygen Permeability (OP) of Casein-Based Edible Films
(23°C, 0% RH.) ........................................................... 49

Figure 8. Tensile Strength (TS) of Casein-Based Edible Films
(23°C, 50% RH.) ......................................................... 53

Figure 9. Elongation (E%) of Casein-Based Edible Films
(23°C, 50% RH.) ......................................................... 57

INTRODUCTION

In 1909 Dr. Leo Hendrick Baekland reacted phenol and formaldehyde together to
form a polymer which was called Bakelite, and this is known as the ﬁrst synthetic plastic.
Soon to follow was the development of many other new synthetic polymers, like cellulose
acetate and polyvinyl chloride in 1927. Polyethylene, the major food packaging plastic,
was developed in 1935 by Imperial Chemical Industries in England. The ability to extrude
plastic into ﬁlms was developed in the mid 1940’s (Hanlon, 1992). As these synthetic
polymers have been further developed to perform useful, packaging and other, functions
their use has become widespread. One possible alternative to synthetic polymers that has
gained increased attention, due to their value added properties they can impart, is the use
of edible ﬁlms.

Edible ﬁlms have actually been in use for centuries, for example, the coating of
ﬁrm with wax or coating food with lard for longer preservation. There are a number of
advantages to be gained through the use of edible ﬁlms. First, they can enhance ﬁrnctional
and nutritional properties of a food. They can be used to protect small pieces or portions
of food, or can be used inside heterogeneous foods to separate components. Finally, since
they are edible there is little to no waste generated (Guilbert, 1986).

Edible ﬁlms will probably never be able to replace the qualities that synthetic

materials possess, but they still can perform many of the same functions to a lesser extent.

2

The ﬁmctions desirable in these ﬁlms consist of lowering the migration of moisture, fats,
and oils. They also decrease the transport of gases like oxygen and carbon dioxide, while
forming a barrier against the contamination from outside microorganisms (Kester and
Fennema, 1986; Donhowe and Fennema, 1994).

The formation of edible ﬁlms has been accomplished using high molecular weight
polymers, which are necessary in order to form a polymer matrix with enough cohesive
strength. The types of high molecular weight polymers used in the making of edible ﬁlms
fall into three categories, being hydrocolloids, lipids, and a composite of both a
hydorcolloid and lipid. Lipids used are fatty acids, fatty alcohols, or a combination of
both, common types being acetoglycerides, surfactants, and waxes. Hydrocolloids used
can be either proteins or polysaccharides, common ones being corn, soy, wheat, and milk
proteins or pectin, starch, and cellulose derivatives (Kester and Fennema, 1986).

In this research we will develop edible ﬁlms using casein from milk. Casein was
chosen because it is abundant, inexpensive, and are extensively used in the production of
adhesives and coatings. Their ﬁrnctional properties make them very suitable for ﬁlm
production, of which they have not been as extensively studied as whey proteins from
milk. The objectives of this research will be to develop optimal formulations and
processes for the formation of these casein edible ﬁlms. These ﬁlms will be comprised of
either rennet or lactic acid precipitated casein, and either sorbitol or glycerol as a
plasticizer. Once the ﬁlms are developed they will be tested for their mechanical
properties (tensile strength and elongation) and barrier properties (oxygen and water
permeability). Comparisons will be made to determine the effect of protein type,

plasticizer type, protein to plasticizer concentration. Then compare these properties with

3

those of other protein-based ﬁlms and synthetic polymers whose properties are known.
The casein-based ﬁlms with the best overall properties will then be selected for further
evaluation in an actual food system to determine their effectiveness as an alternative

packaging material.

LITERATURE REVIEW

Formation of Edible Films and Coatings

There are three basic steps usually followed in the formation of a protein edible
ﬁlm or coating. The mixing of the ﬁlm forming constituents in the solvent must be done
to obtain a dispersion of the high molecular weight polymer. This is followed by casting a
thin layer of the ﬁlm forming solution onto a smooth level surface or the food item in the
case of coatings. This then undergoes a drying process to allow the solvent to evaporate,
allowing the protein to ﬁ'om a matrix and subsequently the coating or ﬁee standing ﬁlm

(Cuq et al., 1995).

Components of edible ﬁlms

A high molecular weight polymer is the one basic requirement for the formation of
an edible ﬁlm or coating. This is needed because ﬁlms with enough cohesive strength
require long chain polymeric structures (Banker, 1966). Two types of high molecular
weight polymers used in the formation of edible ﬁlms and coatings, which are
hydrocolloids and lipids. There are two categories of hydrocolloids used, polysaccharides
and proteins. The polysaccharides consist of starches, gums, and modiﬁed starches.
These include alginate, carrageenan, amylose, and cellulose derivatives. Proteins used in
edible ﬁlms and coatings include com, soy, wheat, collagen, peanut, and milk protein
among others (Donhowe and Fennema, 1994; Kester and Fennema, 1986). The lipids fall
into two categories, neutral lipids of glycerides that are esters of glycerol and fatty acids,
and waxes which are esters of long chain monohydric alcohols and fatty acids (Hemandez,

4

5

1994). Acetylated monoglycerides, natural waxes, and surfactants are the common lipids
used in the manufacturing of edible ﬁlms and coatings (Kester and F ennema, 1986;
Hernandez, 1994). These have included lauric, oleic, and stearic acid, carnauba,
candelilla, and beeswax, and corn, soybean, and palm oil to name a few.

A solvent system is often used in the formation of edible ﬁlms and coatings, usually
when a hydrocolloid or a composite ﬁlm is being produced. This makes it possible to
solubilize and spread the high molecular weight polymer into a thin layer. The two
primary solvents used for these are water and ethanol (Kester and Fennema, 1986).

There are a number of additives that can also be incorporated into the ﬁlm or ﬁlm
forming solution that alter the properties of the edible ﬁlm or coating. There purpose is to
impart more desirable properties to the ﬁlm or coating, or to give an added value to the
food system. These additives can include plasticizers, crosslinkers, vitamins, antioxidants,
ﬂavors, colors, and antimicrobials (Donhowe and Fennema, 1994; Guilbert, 1986).

Plasticizers are widely used in hydrocolloid and composite ﬁlms. These reduce
brittleness and increase ﬂexibility by interfering with intermolecular bonding between
adjacent polymer chains (Koelsch, 1994; Guilbert, 1986; Kester and Fennema, 1986).
Common plasticizers used in edible ﬁlms and coatings are glycerol, polyethylene glycol,
sorbitol, and sucrose. Crosslinkers have been used to impart an increase in cohesive
strength by enhancing intermolecular bonding. These have included such things as
transglutarninase, tannic acid, and formaldehyde. Formaldehyde, not being edible imposes
some limits on it’s use for edible packaging. Antioxidants are incorporated to prolong the
food degradation by way of oxidation. Ascorbic acid, citric acid, butylated hydroxyanisole

(BHA), and butylated hydroxytoluene (BHT) are commonly used food antioxidants.

6

Antimicrobials are used to hinder the growth of microbes, that can cause spoilage of the

food product. These include sorbic acid and potassium sorbate among others.

Processes of edible ﬁlm and coating formation

One of several processes can be used to form an edible ﬁlm or coating. These
include solidiﬁcation of melt, coacervation, and solvent removal. Solidiﬁcation of melt is
the common process by which lipid ﬁlms and coatings are produced. This involves the
melting of the lipid followed by it’s subsequent cooling to resolidify (Donhowe and
F ennema, 1994). Coacervation is the separation of the ﬁlm forming material from solution
by heating, changing pH, adding solvents, or changing the charge. This can be simple,
where only one high molecular weight polymer is involved, or complex, where two
oppositely charged high molecular weight polymersare used (Donhowe and Fennema,
1994', Kester and Fennema, 1986). The most common process used to form hydrocolloid
edible ﬁlms is by solvent removal. In this process the ﬁlm forming constituents are
dispersed in an aqueous phase, which then undergoes a drying process to remove the
solvent (Donhowe and F ennema, 1994).

Casting involves spreading the ﬁlm forming solution in a thin layer, so evaporation
of the solvent and formation of the ﬁlm can occur. Numerous surface types have been
used to cast edible ﬁlms. The requirements for these surfaces is that it be smooth and
level, it is able to contain the ﬁlm forming solution during drying, and that the ﬁlm is able
to be peeled intact ﬁom it’s surface aﬁer drying. Materials used for casting protein ﬁlms
have included glass, teﬂon (polytetraﬂuoroethylene), polystyrene, plexiglass

(polymethacrylate), polyethylene (PE), and polyvinyl chloride (PVC).

7
A glass surface has been successful for the production of cereal protein ﬁlms

(Rayas, 1996; Gennadios eta1., 1993a). PE, plexiglass, and PVC have also been used for
wheat gluten ﬁlms, while PE and teﬂon have been used with soy ﬁlms (Herald et al., 1995;
Gontard et al., 1992; Redl et a1. , 1996; Stuchell and Krochta, 1994; Brandenburg et al.,
1993). Teﬂon has been used in the production of caseinate, whey protein, and non-fat dry
milk ﬁlms. Polystyrene has also been successful with the caseinate and the whey protein
ﬁlms (Banerjee and Chen, 1995). Whey protein ﬁlms have also been cast on plexiglass
and non fat dry milk ﬁlms have been cast on high density polyethylene and teﬂon surfaces

(McHugh et al., 1993; Maynes and Krochta, 1994).

Properties of Films Important for Food Applications
m I

The quality of many food products is dependent on their loss of or exposure to
vapors and gases. These include water (H20), oxygen (02), carbon dioxide (C02), or
volatiles (ﬂavors, antioxidants, etc.). One of the roles of polymers used in packaging is to
control the migration of these gases and vapors (mass transport) into or out of the
package.

The rate at which these vapors or gases pass through a polymer is known as
permeation. There are three steps involved in the action of permeation, adsorption of the
vapor or gas into the polymers surface, followed by it’s diﬁ'hsion through the polymer, and
ﬁnally by it’s desorption through the opposite surface (Sperling, 1992; Birley er al., 1992).

This is expressed by the equation:

P=DxS
where P is the permeability coefficient, D is the diffusion coefﬁcient, and S is the solubility
constant. Solubility is based on the fact that like dissolves like, so gases and vapors with
similar solubility parameters to the particular polymer will dissolve more easily into the
polymer. Henry’s law expresses this action as:

CD = Sp
where CD is the dissolved equilibrium concentration, and Sp is the gas solubility constant.
This expression holds true if Sp is a linear function of the volumetric proportion of the
amorphous phase, but a temperature dependence and the presence of polymer crystallinity
can affect sorption (Sperling, 1992; Birley er al., 1992). To compensate for this a dual
sorption model has been developed, based on Henry’s law. This expression states:

C' = CD + CH
where C. is the total eﬁ‘ective gas concentration, and Cu is the gas concentration assumed
to be adsorbed into the holes of the polymer (Birley et al., 1992).

Diffusion is the transport of the gas or vapor molecules through the polymer. This
occurs in a direction from high concentration to that of low. This process is expressed by
Fick’s laws, F ick’s ﬁrst law of steady state transfer states:

J = -D (SC/5x)
where J is the ﬂux, the rate of transfer per unit area, D the diffusion coefficient, and 5C/6x
is the concentration gradient of the perrneant in the x-direction. The diﬂ‘usion coeﬂicient
is very temperature dependent. For unsteady state diffusion Fick’s second law states:

5cm: = D(62C/6x2)

9

where the change in rate is proportional to the change in concentration gradient with
permeant penetration depth (SIC/65x2 = O at steady state) (Sperling, 1992; Birley etal.,
1992)

There are two basic processes for determining the permeation of the gas or vapor,
this is by either the isostatic or quasi-isostatic method. In both cases you have the
permeant ﬂowing over one side of the ﬁlm, which will then permeate through the ﬁlm and
collected. The difference in the two methods is that with the isostatic method the
permeant is constantly swept out of a diffusion cell (Figure 1) and carried to a sensor,
while with the quasi-isostatic method the permeant is allowed to collect in the diffusion
cell and is sampled at certain time periods. Typical proﬁle curves are shown in Figure 2.
The test is generally run until steady state is reached.

Protein used alone as the high molecular weight polymer in edible ﬁlms have
generally not displayed good water vapor barrier properties as compared to many
synthetic polymers, this is due primarily to their being hydrophilic in nature. Water vapor
transmission rate (WVTR) is generally tested using one of two established methods under
the ASTM guidelines (ASTM, 1990). The ﬁrst, ASTM standard E 96-80, is a cup
method, while the second, ASTM standard F 1249-90, is a method using an infrared
sensor.

In the cup method WVTR is determined by the amount of water weight gained or
lost depending if the cup is ﬁlled with dessicant or water salt solution respectively. In this
method you place a layer of ﬁlm over a non-corroding water impermeable cup and observe

weight change over a period of time. This is continued until a steady rate of weight

10

 

Carrier gas in

+ Upper cell half:

collection of permeant

 

Carrier gas out

 

 

 

Permeation through material

 

 

 

 

 

 

 

Lower cell half: permeant source

 

Figure 1. The Diffusion Cell for Permeability Testing

11

a) Isostatic Method

 

Quantity Perrneated

 

 

Time

b) Quasi-Isostatic Method

Quantity Permeated

 

 

Time

Figure 2. Mass Transport Proﬁle Curves

12
change is observed. McHugh et al.(1993) modiﬁed this procedure for use with

hydrophilic edible ﬁlms. This was developed to account for the water vapor partial
pressure gradient that is present in the stagnant airspace of the test cup between the salt
or water solution and the ﬁlm. This is expressed by the equation:

WVTR = slope / A
where slope = slope of line of weight loss vs. Time, A = area of test ﬁlm.

This is then used to calculate the corrected water vapor partial pressure of the ﬁlms inner
surface in the cup (p2) by:

WVTR=PxDan[(P-p2)/(P-p1)]/(RxTxAz)
where P = total pressure, D = diffusivity of water through air at the testing temperature, R
= the gas law constant, A2 = mean stagnant air gap height (this should be < 14 mm), p1 =
water vapor partial pressure at the solution surface in the cup. You can then use p; to
determine the true water vapor permeance by:

Permeance = WVTR / (p2 - p3)
where p2 = water vapor partial pressure at the ﬁlms outer surface.

The method using an infrared sensor, ASTM standard F 1249-90, does not take
into account weight gain or loss, but rather directly by the amount of a water vapor
present in a sample of air. For example, in the isostatic method you would have a carrier
gas constantly sweeping out the upper chamber of the diffusion cell, carrying that gas to a
sensor which determines the amount of water in it. With the quasi-isostatic method you
would inject a certain quantity of an air sample into a sensor which would then determine

the amount of water in it.

13
To obtain the water vapor permeability (WVP) you take into account the WVTR

thickness of the ﬁlm, and the partial pressure. This is shown by the equation:

WVP = (WVTR x l) / Ap
where l = thickness, and Ap = partial pressure of water at the test conditions.

Oxygen permeability (OP) is tested in basically the same way that as the infrared
method for determining WVP, established under the ASTM standard D 3985-81. First, by
obtaining the oxygen transmission rate (OTR) accomplished again by continuously passing
a known amount of oxygen containing gas through the lower portion of a diﬂirsion cell
and either collecting and sampling, at certain time intervals, the permeant in the upper
portion of the diffusion cell, or continuously sweeping out the permeant in the upper
portion of the cell, with carrier gas, into a sensor. This is performed until, as in WVTR
steady state is reached. Again, the thickness of the ﬁlm and the partial pressure is taken
into account shown by the equation:

OP=(OTRxl)/Ap

where l = thickness, and Ap = partial pressure of oxygen (2 1% 02 being 1 atmosphere).

Mechanical

The main mechanical properties most commonly tested for in edible ﬁlms are
tensile strength (TS) and elongation (E%). Tensile strength is a measure of the force per
unit area required to pull apart the ﬁlm (F / A), and is an indicator of how strong a ﬁlm is.
Elongation is the length of displacement per original length when a force acts to pull a ﬁlm
apart (Al / I) reported as a percentage, and is an indicator of the ﬁlms toughness and

ﬂexibility. Speed at which the force is applied to the sample will affect it’s mechanical

14
properties. The higher the speed at which the force is applied the sample will act more

brittle and stiff (Birley et al., 1992). These same results will occur as temperature and
RH. decreases (Birley et al., 1992). The established method for the testing of tensile

strength and elongation is ASTM standard D-882-3.

Properties of Protein-Based Films
Water Vgor PerrneaLilig

Proteins display hydrophilic tendencies so on their own they have not shown to be
very good water vapor barriers. These properties can be affected by different parameters
which include protein type and concentration, plasticizer type and concentration, pH of the
ﬁlm and ﬁlm forming solutions, environmental conditions, and crosslinking agents.

Corn zein ﬁlms are typically made using ethanol as the solvent. Park and Chinnan
(1990) made 83.1% zein protein, 16.9% glycerol ﬁlms (all ﬁlm percentages based on dry
weight unless otherwise speciﬁed) and reported their WVP to range from 7.69-11.49
g mm/m2 day kPa (tested at 21°C and 85% RH.) Aydt et a1. (1991) observed corn zein
ﬁlms made with glycerol as a plasticizer and ethanol as the solvent to display a WVP of
35.15 g mrn/m2 day kPa (tested at 26°C and 100% RH. inside the test cup and 50% RH.
outside the test cup). 9

Soy protein ﬁlms are typically made using distilled water as the solvent.
Gennadios et al. (1993a) tested the WVP of soy protein isolate (SP1) ﬁlms as they are
eﬁ'ected by pH. They reported that WVP decreased as pH increased above the isoelectric
point (pI) of soy protein (pl = 4.5). Stuchell and Krochta ( 1994) tested the effect of

varying amounts of plasticizer (glycerol) on WVP and observed that as plasticizer

15

concentration increased WVP increased. They also studied the effect of pH change and
had the same results as Gennadios et al. (1993 a).

Wheat gluten ﬁlms use both ethanol and distilled water as a solvent, because it
contains both water and alcohol soluble proteins. Park and Chinnan (1990) tested a wheat
gluten ﬁlm, 75.6% wheat gluten and 24.4% glycerol, and reported it to have a WVP range
of 52. 1-54.4 g rum/m2 day kPa (tested at 21°C and 85% RH.) Aydt et al. (1991)
observed a WVP of 108.4 g mm/m2 day kPa (tested at 378°C and 100% RH. inside the
cup and 50% RH. outside) using a ﬁlm consisting of 71 .4% wheat gluten and 28.6%
glycerol. It was concluded that wheat gluten ﬁlms give the lowest WVP’s at the extreme
pH’s, again away from the isoelectric point (p1 = 7.5, average for wheat glutens)
(Gennadios et al., 1993a). 60an et a1. (1992) demonstrated that the amount of ethanol
in the ﬁlm forming solution and it’s pH had an important role in WVP. They found that a

neutral pH and a low ethanol content gave the lowest WVP’s.

@1an Permeability

Protein ﬁlms have generally possessed good oxygen barrier properties as compared
to synthetic ﬁlms. These properties haven’t been as widely studied as WVP or mechanical
properties. Again, as with WVP factors that can effect these values are protein type,
plasticizer, and environmental conditions.

Park and Chinnan (1990) with ﬁlms comprising of 83.1% corn zein and 16.9%
glycerol observed OP’s that ranged from 13.0-44.9 cc urn / m2 day kPa (tested at 30°C
and 0% RH.) Aydt er a1. (1991) reported an OP of 76.63 cc um / m2 day kPa (tested at

378°C and 0% RH.)

l6
Brandenburg et a1. (1993) with a 62.96% SP1 and 37.04% glycerol ﬁlm observed

an OP of 4.75 cc um / m2 day kPa (tested at 25°C and 0% RH.) They also concluded
that as pH increased ﬁom 6-12 OP decreased.

Gennadios et al. (1993b) obtained an OP of 3.82 cc um / m2 day kPa with a ﬁlm
consisting of 71 .4% wheat gluten and 28.6% glycerol (tested at 23°C and 0% RH), and
concluded that OP increased as temperature increased. This was veriﬁed by Aydt et a1.
(1991) using a ﬁlm of the same composition, but tested at 378°C and 0% RH, where the
OP was observed to be 7.78 cc um / m2 day kPa. This was also demonstrated by Rayas
(1996) with commercial bread ﬂour, who also showed that the use of crosslinkers
(cysteine, formaldehyde, and glutaraldehyde) increased OP, and an increase in pH, from 4

to 11, also increased OP.

Tensile Strength and Elongation
Gennadios et al. (1993c) showed that changes conditioning environments (RH.

and temperature) had an effect on tensile strength. They concluded that as RH. increased,
with no change in temperature TS decreased. As temperature increased, with no change
in RH., TS increased. They relate this to the moisture content in the ﬁlms, with more
moisture at higher RH. ’s, acting as a plasticizer, and less moisture present in ﬁlms at
higher temperatures. They observed these ﬁlm to have a very low E%, ranging ﬁ'om 3-
7%.

Gennadios er al. (1993a) showed T8 for SP1 ﬁlms (62.5% SP1 and 37.5%
glycerol) to be 3.0-3.6 MPa at a pH of 6-11 and ﬁom 1.9-2.3 at a pH in the range of 13

MPa. E% peaked for these ﬁlms between pH 7 and 11, which ranged from 130-

17
190%. Stuchell and Krochta (1994) showed in SPI ﬁlms that TS decreased as plasticizer

% increased. They found that as plasticizer increased E% increased, ﬁlms with 20%
glycerol had an E% of 16.8%, while ﬁlms with 23% glycerol had an E% of 23.8%.

Gennadios er al. (1993a) observed that as pH of the ﬁlm increased TS increased,
0.5, 1.9, and 4.4 MPa at pH’s of 4, 9, and 13 respectively for ﬁlms containing 73.2%
wheat gluten and 26.8% glycerol. No signiﬁcant diﬁ’erences were detected in E% at
different pH’s, ranging from 156.7 to 259.6%. It was demonstrated that as temperature
increased, so did TS for wheat protein ﬁlms. Rayas (1996) showed that the addition of
the formaldehyde, a crosslinker, caused and increase in TS, ﬁom 2.35 MPa to 4.33 MPa,
while cysteine and glutaraldehyde (crosslinkers) had no effect. The addition of the

crosslinker decreased the observed E%.

Properties of Milk Protein-Based Films
Wator Vapor Permeabilig

Distilled water is typically used as the solvent for the formation of edible ﬁlms
from milk proteins. Edible ﬁlms made from non-fat dry milk (NFDM) incorporate all the
milk proteins into the ﬁlm. Maynes and Krochta (1994) produced edible ﬁlms from
various NFDM types. These included commercial blends varying in protein content ﬁ'om
85%-87%, a lactose extracted blend (63.8% protein), and an ultraﬁltered (UF) NFDM
blend (82.5% protein). The ﬁlms were consisted of 75% protein mix and 25% glycerol.
They observed that the UF-NFDM ﬁlms gave the best WVP’s, at 70.3 g mm/m2 day kPa,
while the rest were not signiﬁcantly different, ranging from 80.1-86.3 g mm/m2 day kPa

(tested at 30°C with 100% RH. in the cup, and 0% RH. outside the cup).

18
Several groups have studied the properties of whey protein-based edible ﬁlms.

Banjeree and Chen (1995) compared properties of whey protein concentrate (WPC),
76.6% protein, and whey protein isolate (WPI), 93.6% protein. The ﬁlms consisted of
66.7% protein and 33.3% glycerol. They reported that the WPC ﬁlms gave a lower WVP
then WPI, 10.64 g mm/m2 day kPa and 12.12 g mm/m2 day kPa respectively (tested at
23°C with 100% RH. in the cup, and 55% RH. outside the cup). McHugh er al.
(1994a) concluded that both plasticizer type and plasticizer concentration effect the WVP
of a ﬁlm. They demonstrated that sorbitol, as a plasticizer, displayed better water barrier
properties than either polyethylene glycol (PEG) or glycerol at the same concentration,
with glycerol having the worst barrier properties. Values of 50% WPI, 50% plasticizer
being 3.53, 5.61, and 6.44 g mm/m2 day kPa for sorbitol, PEG, and glycerol respectively
(tested at 25°C and z 77% RH. inside the cup and 0% outside). They also showed that
as the concentration of plasticizer increased WVP of the ﬁlms increased. Films with
62.5% protein an 37.5% sorbitol had a WVP of 2.58 g mm/m2 day kPa, while ﬁlms with
50% wpr and 50% sorbitol had a WVP of3.53 g rum/m2 day kPa.

Various caseinates have been studied in the production of ﬁlms (including sodium,
calcium, potassium, and magnesium caseinates). Banjeree and Chen (1995) reported that
calcium caseinate (CC) ﬁlms gave a lower WVP than sodium caseinate (SC) and
potassium caseinate (PC) ﬁlms, values being 7.91, 12.90, and 12.12 g mm/m2 day kPa
respectively (tested at 23°C and 100% RH. inside the cup and 55% outside). Avena-
Bustillos and Krochta (1993 a) also obtained the same results, with CC ﬁlms providing a
better water barrier than SC ﬁlms. They also tested the effect of calcium crosslinking and

pH adjustment on WVP properties, and showed that soaking SC ﬁlms in a calcium

l9

chloride solution (calcium crosslinker) or in buffers to lower pH to 4.6 (pl of casein)
decreased the WVP of the ﬁlm. Ho (1992) produced ﬁlms out of 80% magnesium
caseinate and 20% glycerol, and 80% rennet casein and 20% glycerol, and obtained

_‘ WVP’s of 43.9 and 56.0 g mm/m2 day kPa respectively (tested at 25°C and a corrected

RH. of 77%).

04¢an Permeability

McHugh and Krochta (1994b) studied the eﬁ’ect of plasticizer type, plasticizer
concentration, and relative humidity on OP of WPI-based ﬁlms. They concluded that ﬁlms ,
containing sorbitol will give lower OP’s than ﬁlms containing glycerol, at equal
concentrations. At 70% WPI and 30% plasticizer the OP of the ﬁlm using sorbitol had an
OP of 4.3 cc um / m2 day kPa, compared to 76.1 cc um / m2 day kPa for glycerol (tested
at 23°C and 50% RH.) They demonstrated that as plasticizer concentration increased

OP increased, and as relative humidity increased OP increased.

Tgsilo Strength and Elongation

Maynes and Krochta (1994) observed TS’s up to 9.1 MPa with 75% protein and
25% glycerol NFDM ﬁlms. 'UF-NFDM ﬁlms displayed the lowest 15% at 5.2%, the
lactose extracted NFDM ﬁlm had an 13% of 12.2%, while the commercial brands ranged
from 22.1-38.5%, with E% increasing as protein content decreased. McHugh and
Krochta (1994b) reported TS increased as plasticizer concentration decreased in WPI and
glycerol ﬁlms. 85% WPI and 15% glycerol ﬁlms had a TS of 29.1 MPa, while 70% WPI

and 30% glycerol ﬁlms had a TS of 13.9 MPa. As plasticizer concentration increased E%

20
increased. Banjeree and Chen (1995) demonstrated that WPI ﬁlms produced stronger

ﬁlms than WPC, with TS’s of 5.94 and 3.36 MPa respectively (66.7% protein and 33.3%
glycerol). E% for the WPC ﬁlm was 20.84% and 22.74% for the WP1 ﬁlm. Banerjee and
Chen (1995) observed that CC ﬁlms produced stronger ﬁlms than SC or PC, TS’s of 4.25,
2.98, and 2.97 MPa respectively. The PC ﬁlm had the largest E% followed by SC and
CC, elongations being 42.80, 29.89, and 1.45%, respectively. Motoki et a]. (1987) used
transglutarninase as a crosslinker, which increased TS of out-casein ﬁlms from 4.1 to 10.6

MPa.

Milk protein-based ediolo ﬁlm and ooating applications

The ultimate goal for developing these protein edible ﬁlms is for their possible
application into a food system. There has been numerous studies using these protein
solutions as a coating on either meat, seafood, nuts, ﬁ'uits, and vegetables (Baker etal.,
1994). These studies have shown the ability to reduce spoilage problems, like degradation
from oxidative rancidity and browning, in these foods. These coatings are either brushed
on, sprayed on, or the food item is dipped into the coating. ‘

Stuchell and Krochta (1995) used a WPI and acetylated monoglyceride coating on
frozen king salmon and found it to delay lipid oxidation, decrease peak peroxide value,
and reduce the amount of moisture loss. Lerdthanangkul and Krochta (1996) observed
that SC coatings caused an increase in internal C02 and a decrease in internal 0; in green
bell peppers, showing their good gas barrier properties. Avena-Bustillos et al. (1993b)
showed that SC-lipid and CC-lipid coatings reduced white blush on minimally processed

carrots, and also reduced water vapor transmission. Most of this research dealt with

21

composite (protein/lipid) coatings, because the protein coatings on their own do not
provide good water barrier properties. There has been little reported research with free-
standing protein edible ﬁlms, this is due largely to the fact that a way to seal these ﬁlms

has not yet been reported.

Properties of Plasticizers

Glycerol and sorbitol are naturally occurring carbon backboned polyhydric
alcohols. Besides use as plasticizers they are often used as sweeteners, humectants, and
pharmaceutic aids.

Glycerol originates from oils and fats, usually as a by-product in the manufacturing
of soaps and fatty acids. It has a molecular weight of 92.09 daltons and is a three carbon
molecule with one hydroxyl group (C3HsOg) (Merck, 1989). It is a liquid at room
temperature, possessing a melting point of 178°C. It is miscible in water and alcohol and
have the ability to absorb moisture from the air. It’s viscosity at 20°C is 1.143, 2.095,
6.050, and 22.94 centipoise for solutions of 5, 25, 50, and 70% glycerol respectively
(Merck, 1989).

Sorbitol was discovered in 1872 in the berries of mountain ash (Sorbus aucuparia
L.) and now are produced by high pressure hydrogenation or electrolytic reduction of D-
glucose, or by catalytic hydrogenation of dextrose (Merck, 1989). It has a molecular
weight of 182.17 daltons, consisting of a 6 carbon chain with 4 hydroxyl groups
(C5111406) (Merck, 1989). It is mainly found in the stable crystalline (y-) form, with a
melting point of 96°C. It is highly soluble in water (solubility at 25°C being 234g/100g

water), but insoluble in most other organic substances (Sicard and Leroy, 1983; Sicard,

22

1982). It is stable and chemically unreactive. It is more viscous than lower weight
polyhydric alcohols, with viscosities at 20°C being 1.230, 2.689, 11.09, and 185 centipoise
for solutions of 5, 25,50, and 70% sorbitol respectively (Merck, 1989). At equal RH.’s
the water content of sorbitol will be lower than that of glycerol. Sorbitol is also more
resistant to changes in water content as relative humidity changes (Sicard and Leroy,

1983)

Properties of Milk Proteins
Milk is comprised of 3.3-3.9% protein, of which 20% are whey proteins and 80%
are casein proteins (Swaisgood, 1985). Caseins possess many desirable characteristics

that make them suitable for the production of edible ﬁlms.

mo

On average, 38-45% of the caseins are comprised of oat-casein (Leman and
Kinsella, 1989; Swaisgood, 1985; Dalgleish, 1982). This protein has been determined to
be 199 fatty acid residues long, with a calculated molecular weight of about 23,000
daltons, depending on the variant (Swaisgood, 1985; Dalgleish, 1982). The C-terrninal is
comprised of some or—heliccs, B-sheets, B-tums, and unordered structure, while the N-
terrninal is mainly random coil in structure (Swaisgood, 1985). The secondary structure is
limited due to the presence of proline, making up about 8.5% of the residues (Kinsella,
1984)

There are two variants of this protein, with the difference being in the number of

phosphoseryl groups, either 8 or 9 (Swaisgood, 1985; Dalgleish, 1982; Fox and Mulvihill,

23
1982). The variant with 9 phosphoseryl groups is referred to as coo-casein. Even though

threonine, which is present, is capable of phospohorylation, this occurrence is uncommon
(Dalgleish, 1982). These phosphoseryl groups allow the protein to bind Ca2+. oat-casein
can bind up to 8-10 moles Cay/mole of protein under normal circumstances and up to 20
mole Cay/mole protein at high Ca2+ concentrations (Swaisgood, 1985; Dalgleish, 1982;
Fox and Mulvihill, 1982). In both variants all but one of the phosphoseryl groups are
found between residues 41 and 80. Three hydrophobic regions are present in out-casein,
from residues 1-40, 90-110, and 130-199 (Fox and Mulvihill, 1982). The overall charge
being -21mV at pH 6.8, with a hydrophobicity of 1172 cal/residue (Kinsella, 1984;
Dalgleish, 1982).

The org-caseins comprise 10-12% of the casein proteins (Leman and Kinsella,
1989; Swaisgood, 1985; Dalgleish, 1982). It consists of a 207 amino acid residue chain,
with a calculated molecular weight of 23,000-25,000 daltons, depending on the variant
(Swaisgood, 1985; Kinsella, 1984; Dalgleish, 1982). Due again to the presence of proline,
about the same amount as oat-casein, any secondary structure is limited (Kinsella, 1984;
Dalgleish, 1982).

There are 4 variants of org-casein, again differing in the number of phosphoseryl
groups, ranging ﬁ'om 10-13, termed as out-casein, org-casein, org-casein, and org-casein
respectively (Kinsella, 1984; Dalgleish, 1982; Fox and Mulvihill, 1982). These
phosphoseryl groups are fairly evenly distributed throughout the protein. However, the C-
terminal, residues 160-207, is hydrophobic. There are two cysteine residues in the
protein, which could participate in disulﬁdebonds. It is the most hydrophillic of the casein

proteins, with a hydrophobicity of 1111 cal/residue, and thus the most ionic strength

24
dependent. The overall charge ranges from -16 to -22mV at pH 6.8, depending on the

variant (Kinsella, 1984; Dalgleish, 1982).

B-casein makes up 31.3—36% of the casein proteins (Leman and Kinsella, 1989;
Swaisgood, 1985; Dalgleish, 1982). This protein is 209 residues in length, with a
molecular weight of 23,900-23,980 daltons (Swaisgood, 1985; Kinsella, 1984; Dalgleish,
1982). B-casein is comprised of 10% or-helix, 13% B-sheet, and 77% unordered structure
(Andrews er al., 1979). This random structure is again the result of the presence of 16%
proline residues (Kinsella, 1984). There are ﬁve phosphoseryl groups which are all
located in the N-terrninal. It can bind 4-5 mole Cap/mole of protein (Swaisgood, 1985).
B-casein is the most hydrophobic of all the casein proteins, with an overall hydrophobicity
of 1334 cal/residue, but the hydrophobic C-terrninal, residues 30-209, has a
hydrophobicity of 1408 cal/residue, and carries a net charge of -12mV at pH 6.8 (Kinsella,
1984; Dalgleish, 1982). It is the most temperature dependent of the caseins (Swaisgood,
1985)

There is a class of casein derived ﬁom the proteolysis of B-casein. These are
referred to as y-casein, and represent 35% of the total casein fraction (Swaisgood, 1985;
Fox and Mulvihill, 1982). The variants of this protein yt-casein, yz-casein, and yg-casein
have chain lengths of 181, 104, and 102 residues respectively, with molecular weights of
20,520, 11,822, and 11,557 Daltons (Swaisgood, 1985).

B-casein can be hydrolysed at 1 of 3 lysyl residues, 28, 104, and 106, forming 1 of
three pairs of polypeptide chains, of which one half of each pair is lost into the serum, with
the other half making up the y-caseins (Fox and Mulvihill, 1982). Of these yl-casein is the

only one containing a phosphoseryl group.

25

The ﬁnal casein protein is x-casein, which is found at levels of 10-13% (Leman and
Kinsella, 1989; Swaisgood, 1985; Dalgleish, 1982). It contains 169 fatty acid residues,
with a molecular weight of 19,000 daltons (Swaisgood, 1985; Kinsella, 1984; Dalgleish,
1982). Loucheaux-Lefebvre er a1. (1978) determined that x-casein consisted of 26% or-
helix, 31% B-sheet, and 24% B—tums. They determined that residues 105-106 probably
formed either an or-helix or B-sheet, between two stable B—tums and another B-tum at
residues 113-116, making that linkage accessible to proteolysis. There is only one
phosphoseryl group and two cysteine residues (Swaisgood, 1985; Dalgleish, 1982). x-
casein can contain from 0-3 oligosaccharide chains. The carbohydrate moiety exists as
either a tri- or tetrasaccharide, composed of N-acetyl-neuraminic acid, galactose, and N-
acetylgalatosamine. The main point of attachment is at threonine 133, with attachment
occurring at threonine 131 and threonine 135 (Fox and Mulvihill, 1982). Due to the
presence of only 1 phosphoseryl group x-casein can only bind 1-2 mole Cazilmole protein
at pH 6.8 (Swaisgood, 1985). The overall hydrophobicity is 1224 cal/residue, with an
overall charge of -4 mV (Swaisgood, 1985; Dalgleish, 1982). Due to the structure of x-
casein the bond between the phenylalanine (105) and methionine (106) residues is
susceptible to proteolysis by rennet (or chymosin). This gives rise to the hydrophobic N-
terrninal portion, known as para-x-casein, with a hydrophobicity of 1310 cal/residue and a
charge of +5 mV, and the more hydrophilic macropeptide containing the oligosaccharides,
with a hydrophobicity of 1082 cal/residue, carrying a charge of -8 mV (Dalgleish, 1982).

The macropeptide at this occurrence is then lost into the serum.

26

The casein micelle

The casein proteins along with colloidal calcium phosphate (CCP) interact with
each other to form spherical complexes known as micelles. The size of the micelles range
ﬁ'om 10-300 nm, with a molecular weight of 108 to 109 daltons (Swaisgood, 1985;
Kinsella, 1984; Dalgleish, 1982). The composition of the micelle is 92-94% casein protein
and 6-8% CCP (Swaisgood, 1985; Fox and Mulvihill, 1982). The structure of the micelle
is based on either of two generally accepted theories, both involving the concept of
subrnicelles, 1020 nm in size, aggregating to form the micelle (Swaisgood, 1985; Kinsella,
1984; Fox and Mulvihill, 1982).

The ﬁrst casein micelle model was presented by Slattery and Evard (1973) and
updated (1979). In this model the submicelle is formed containing a partially hydrophobic,
or.— and B-caseins, portion and a partially hydrophilic portion, rc-casein. It is then probable
that a tetrahedral arrangement is formed, which will keep growing until there is enough 1(-
casein on the surface to prevent any further hydrophobic interactions (Figure 3). The
micelle is stabilized by the collective presence of the hydrophobic interactions of 01.1- and
B-caseins, and through the formation of calcium phosphate salt bridges within the interior
(Slattery, 1976).

The second model was presented by Schmidt (1980). His model also uses the
submicelle theory stemming from the interaction of 01.1- and x-casein, the self-associations
of 01.1- and B—casein by hydrophobic bonding, and also through the self-association of 0.2-
casein by electrostatic interactions. The subrnicelles are spherical particles with a
hydrophobic core and a surface layer with the phosphate groups of our, our, and B-

casein and the polar macropeptide portion of x-casein. Micelle formation begins with the

27

 

115-, B-, y-caseins

- x-casein

 

 

Figure 3. The Casein Micelle (Slattery and Evard Model)

28
aggregation of tertiary colloidal calcium phosphate and the subnricelles. Growth will

continue until, as in Slattery’s model, the surface is mainly x-casein. Submicelles deﬁcient
in x-casein will be located in the core.

The CCP consists of ions of calcium, phosphate, and some magnesium and citrate,
comprising an average of 2.8-2.9, 4.3-5.2, 0.1, and 04-05% of the total micelle
respectively (Brunner, 1977; Schmidt, 1980). It is found in an apatite-like complex of
tertiary calcium phosphate, with some calcium citrate (Schmidt, 1980). Magnesium
prevents the calcium phosphate from transforming into a more stable hydroxyapatite form,
while the casein prevents ﬂocculation (Schmidt, 1980; Swaisgood, 1985). Calcium acts to
neutralize some of the repulsive electrostatic (-) charges and facilitates hydrophobic
interactions. Calcium also plays a compacting role in the micelle, through the salt bridges.

The casein micelle is rather porous and thus is highly hydrated, containing from 2-
3.7 grams H20/gram dry protein (Swaisgood, 1985; Kinsella, 1984; and Fox and
Mulvihill, 1982). The presence of some x-casein in the interior, as much as 30% of the x-
casein in the nricelle, probably allows the hydrophobic interior to stay stable (Slattery,

1976)

Casein manufacturing

The manufacturing of casein for use as a food ingredient is accomplished by one of
two ways, lactic acid or rennet precipitation.

In lactic acid casein, pastuerized skim milk is inoculated with the lactic starter
culture and allowed to incubate for 14-16 hours at 22-26°C. The fermentation of lactose

causes the pH to reduce to about 4.6, causing the casein to coagulate and form a curd.

29

With rennet casein, calf rennet or chymosin is added to pasteurized skim milk at 29°C.
This enzyme cleaves the x—casein, causing it to lose the macropeptide into the serum and
destabilizing the micelle, bringing on the clotting of the casein. This process takes about
30 minutes under a pH of 6.6 (Southward and Walker, 1980).

The remaining processing required is the same for both types of precipitated
caseins. First, the curd is cooked at 50-55°C to ﬁrm it up to withstand the rest of the
processing (Muller, 1982). The curd and whey then go through several separation and
washing stages. The casein is then dewatered, dried, milled, sieved, blended, and
packaged (Muller, 1982; Southward and Walker, 1980).

Properties of these caseins include insolubility at pH 7.0 and 4.6-4.7 for rennet
casein and lactic acid casein respectively. Acid caseins are able to be solubilized at pH
4.6-4.7 with the use of alkalis or alkaline salts, these are referred to as caseinates
(Kirkpatrick and Walker, 1985). The common alkalis and alkaline salts used are calcium,
sodium, and potassium. Both types of precipitated casein are heat stable and have good

nutritive qualities.

MATERIALS AND METHODS

Film Components and Formation

Casein samples were obtained from New Zealand Milk Products (N. America)
Inc., (Santa Rosa, CA). Lactic acid casein being Alacid 710, 30 mesh and rennet casein
being Alaren 771, 30 mesh (Table 1). Protein and ash content of the casein samples were
veriﬁed by standard methods (AOAC, 1990). Sorbitol was purchased from Sigma
Chemical Co., (St. Louis, MO), and glycerol was purchased from Mallinckrodt Specialty
Chemicals Co., (Paris, KY).

Figure 4 shows a schematic diagram of the ﬁhn forming process. The various
casein based edible ﬁlms were prepared by ﬁrst mixing lactic acid or rennet casein (3,5, or
7% w/w) with sorbitol or glycerol (5% w/w), distilled water, and 1M NaOH (to adjust pH
to 10.0) for ﬁnal mixtures of 150 g (Table 2 and Table 3). These were then heated and
stirred, using the “Magna-4” magnetic stirrer and hot plate, model 4820-4 from Cole-
Parmer (Chicago, IL), to a ﬁnal temperature of 65.5 :1: 25°C (150°F) for 30 minutes, and
then held at that temperature for 15 minutes. The ﬁnal pH of the ﬁlm forming solution
was measured using the Coming pH meter 240 (Corning, NY). These were the conditions
upon which the best solubility of the caseins were obtained.

Next, samples were ﬁltered twice, through 1 layer of cheesecloth. Then they were
stored at 20 :t 2.0°C ( 68°F) for 4 hours to allow any foaming created during the mixing
process to settle. A vacuum was applied to solutions for 30 minutes, using a hydrometric

vacuum system, to remove any residual air in the solution. The ﬁlm forming solution was

30

Table 1. Compositions of Casein Protein Powders Used1

31

 

 

 

 

 

Percent Lactic Acid Casein Rennet Casein
(%) Alacid 710 Alaren 771

Protein (N x 6.38) % 87.3 80.6
Ash % 1.8 7.8
Moisture % 9.6 11.0
Fat % 1.2 0.5
Lactose % 0.1 0.1

H (5% at 20°C) 4.6 7.1

 

' Values based on company speciﬁcations (New Zealand Milk Products (N. America) Inc.)

 

32

Mix ﬁlm forming components (150g total)
protein (3, 5, 7% w/w)
plasticizer (5% w/w)
distilled water
1M NaOl—I (adjust pH to 10.0)

1

Heat / Stir
45 min. total
(to 656°C and hold for 15 min.)

1

Filter
(cheesecloth 2X)

Equilibriate
(4 hr at 20°C)

l

Vacuum
(30 min)

1

Cast solution
(teﬂon pan)

1

Dry
(8-18 hr at 550C)

1

Peel

Test

Figure 4. Schematic Diagram of the Film Forming Process

33

Table 2. Compositions of Casein-Based Edible Films

 

Protein, Plasticizer

o/oprotein powder/

%protein powder]

 

 

 

 

Typel ‘Voplasticizer %plasticizer

wet weight dry basis

L.A. Casein, S 3.0/5.0 37.5/62.5
L.A. Casein, S 5.0/5.0 50/50

L.A. Casein, S 7.0/5.0 58.3/41.7

L.A. Casein, S 3.0/5.0 37.5/62.5
L.A. Casein, S 5.0/5.0 50/50

L.A. Casein, S 7.0/5.0 58.3/41.7

R. Casein, G 3.0/5.0 37.5/62.5
R. Casein, G 5.0/5.0 50/50

R. Casein, G 7.0/5.0 58.3/41.7

R. Casein, G 3.0/5.0 37.5/62.5
R. Casein, G 5.0/5.0 50/50

R. Casein, G 7.0/5.0 58.3/41.7

 

' L.A. Casein=Lactic acid casein, R Casein=Rennet casein; S=Sorbitol, G=Glyoerol.

 

34
Table 3. Formulations of Film Forming Solutions for Casein-Based Edible Films

 

 

 

 

 

 

 

Casein Type Protein Plasticizer Distilled H20 N aOH
(% w/w) Powder (5% w/w)
1g) 181 (Q (g)
Lactio Acid
low (3%) 4.5 7.5 133.2 4.8
medium (5%) 7.5 7.5 127.0 8.0
high (7%) 10.5 7.5 120.7 11.3
Rennot

low (3%) 4.5 7.5 136.8 1.2
medium (5%) 7.4 7.5 132.9 2.1
. high (7%) 10.5 7.5 129.1 2.9

 

 

35

then cast on a 7.5 in diameter teﬂon coated pan. A teﬂon coated pan was chosen because
upon drying on glass, ﬁlms were unable to be peeled. The amount of ﬁlm forming
Solution cast varied depending on the protein content, 52.5 i 2.5 ml of the medium and
high protein content ﬁlms and 90 i 5 ml of the low protein content ﬁlms. These casting
volumes were used to obtain dried ﬁlms with an average thickness of 0.203 mm (8 mils).
Films were dried in a gravity convection incubator, Blue M Electric Co., (Blue Island, IL),
at 55 : 2°C (130°C) until they were able to be peeled from the casting surface. The
greater amount of solution cast the longer the drying time was, because of the increased
solvent amount. Drying times were about 8 hours for the high and medium protein
content solutions and about 18 hours for the low protein content solutions. Once peeled

from the casting surface, ﬁlms were kept at 20 i 2°C (68°F) until testing was performed.

Thickness

Thickness measurements was measured using a micrometer, TMI model 549M
micrometer from Testing Machines Inc,. (Amittyville, NY). For barrier testing, thickness
was the average of 5 measurements, for mechanical testing it was the average of 3.
Barrier testing was done on ﬁlm samples of 0.203 t 0.038 mm (8.0 mils), while thickness

for mechanical testing was 0.203 i 0.089 mm (8.0 mils).

Water Vapor Permeability
WVTR was tested according to ASTM standard F 1249-90, “Water vapor
transmission rate through plastic ﬁlm and sheeting using a modulated infrared sensor.”

They were tested using the Perrnatran-W (MoCon Inc., Minneapolis, MN). The samples

36
were tested at 37.8 i: 05°C (100°F) and 90 i 3% RH. A saturated salt solution of

potassium nitrate (NH4H2P04) was used to obtain the desired RH.

Samples which were placed in a 50 cm2 difﬁrsion cell with the absorbent pad in the
bottom half of the cell soaked with the salt solution, with dry air sweeping out the top half
of the cell going to the sensor (Figure 1). The testing surface area of the casein ﬁlm
samples were reduced from 50 cm2 to 5 cm2 with the use of a foil backing. Otherwise,
they would adsorb all the water from the salt solution. Samples were conditioned for 10
hours at the testing conditions before testing was conducted. A calibration sample, with a
known WVTR , was run with all test samples. The calibration sample was 1 mil thick
Mylar, with a WVTR of 21 g H20/ m2 . day. Tests were run until steady state was
reached at which point 12 readings were taken over a 30 minute period. These readings
(in mV) were then averaged then adjusted according to the calibration sample to get
WVTR. WVP was then calculated by the equation:

WVP = WVTR x l / Ap

l = thickness of ﬁlm
Ap = partial pressure of water at test conditions

WVP values were the average of samples done in triplicate.

Oxygen Permeability

OTR was measured according to ASTM standard D-3985-81, “Oxygen gas
transmission rate through plastic ﬁlm and sheeting using a coulometric sensor.” Tests
were run at 23 i 2°C, 0 % RH., and 21% oxygen, using the Oxtran 200 (MoCon Inc.,

Minneapolis, MN). All samples were conditioned for 10 hours at the same conditions

37

prior to testing. Temperature was maintained using a water bath system, Endocal water
bath (Neslab Instruments Inc., Newington, NH).

The testing area of the sample was 50 cm2, with compressed air at 21% oxygen
sweeping the bottom half of the cell and nitrogen sweeping the top half, going to the
sensor. OP was calculated by the equation:

OP = OTR x l / Ap

l = thickness of ﬁlm
Ap = partial pressure of oxygen

OP values were the average of samples done in triplicate.

Mechanical Properties
TS and B were tested according to ASTM standard D-882-83, “Tensile properties
of thin plastic sheeting.” Tests were run using the Instron Universal Tester, model 2401
from Instron (Canton, MA), with a lkN static load cell and crosshead speed of 20 in./min.
Conditions of testing were 23 i 2°C (73.4°F) and 50 :1: 5% RH. All samples were
conditioned for 48 hours at the same conditions prior to testing (Banerjee and Chen,
1995). Testing sample size was 2 in. x 1 in. TS was determined by the equation:
TS = load / area
load = peak force
area = sample width x sample thickness
E was determined by the equation:

E = All 1 (expressed as a percentage)

38

A1 = distance sample stretched
l = original length of sample
TS and E values are the average of triplicate samples. Each sample was tested in

duplicate.

Storage Study
Two representative treatments of the casein-based edible ﬁlms developed in this research
were ﬁrrther tested in a storage study using American processed cheese slices to
investigate their effectiveness as a packaging wrap. Casein-based edible ﬁlms used for this
study were the 50% lactic acid casein/50% sorbitol and 58.3% lactic acid casein/41.7%
glycerol ﬁlms (percentages on a dry basis). These ﬁlms were chose due to their similar
barrier and mechanical properties they possessed, which were best the overall among the
treatments evaluated in this research. Thickness of the ﬁlms varied from 5.19 to 8.01 mils
(0.131 to 0.203 mm). Slices unwrapped and in the original LDPE wrapper were used as
controls. The processed American cheese slices used for this storage study, were
purchased at a local retail outlet (East Lansing, MI). Cheese slices (3.25 in. x 3.5 in.)
were placed between two layers of casein based ﬁlm, sealed together by rubber cement,
then dipped in parafﬁn wax to minimize water loss through the seal (Figure 5). The cheese
samples were stored at 2.2 i 10°C (36°F) and 88 i 5% RH. Triplicate samples of each
treatment were tested every 5 days over a 30 day storage period. Both the cheese and the
wrap were tested for color change and moisture content during storage.

Color tests were performed using the HunterLab colorimeter ﬁom (Hunter

Associates Laboratory, Inc., Reston, VA), using a black and a white standard tile for

39

l—— 3.5” ——l

Seal area
(wax and rubber cement)

 

 

3.25” Cheese

 

 

 

 

 

 

Figure 5. Packaging of Processed Cheese Slices Using Casein-Based
Edible Films.

40
calibration, and the black tile as the background when testing samples. Values of L (black

to white), a (green to red), and b (blue to yellow ) were determined. Moisture content of
the cheese was performed according to the “Standard Methods for the Examination of
Dairy Products,” (Marshall 1992). Cheese was shredded and approximately 3.0 i 0.5 g
was placed in an aluminum weighing dish and dried at 80 i 3°C (176°F) using a gravity
convection oven from Precision Instruments (Chicago, IL), until a constant weight was
reached (approximately 16 hours). After the samples were dry they were placed in a
dessicator for 30 minutes to cool and reweighed. Moisture content (%Moisture) was
calculated by the following equation:

%Moisture = [(wt. initial - wt. ﬁnal)/wt. initial] x 100
Moisture content of the ﬁlms were determined by drying 3.0 i 0.5 g of the edible ﬁlm for
16 hours at 100 i 3°C (212°F), moisture of the samples were calculated similar to the

cheese.

Statistical Analysis

Statistical analysis of the effect of protein type, plasticizer type, protein to
plasticizer concentration on ﬁlm properties, and the eﬁ‘ect of wrap type on moisture
content and color of processed cheese and their wraps during storage were made using
Sigma Stat 1.0 from the Jandel Corp, (San Rafael, CA) performing multiple comparisons

with the Student-Newman-Keuls method.

RESULTS AND DISCUSSION

Film Development

Veriﬁcation analysis of the protein powders showed that the speciﬁed values in Table 1
were within reason, protein content upon analysis being 90.18% and 83.4% for the lactic
casein powder and rennet casein powder respectively, and ash being 1.28% and 8.02%
respectively.

Upon drying all ﬁlms were smooth, ﬂexible, and transparent. However, ﬁlms
containing low protein content, 37 .5% protein and 62.5% plasticizer (all percentages in
results and discussion are on a dry basis unless otherwise speciﬁed), displayed a tackiness
that the high and medium protein content ﬁlms did not. Films made with a plasticizer
content higher than 62.5% were too tacky and fell apart upon peeling ﬁ'om the casting
surface, thus were not used in this study. The ﬁlm solution made with rennet casein and
glycerol, 58.3% protein and 41.7% plasticizer, gelled shortly after the heating process,
thus it had to be cast immediately following heating and ﬁltering. Films rrrade with any

higher protein content were too brittle and could not be peeled ﬁ'om the casting surface.

Barrier Properties
Wator vapor permeabilig (WVP)

A WVP of 34.0 g-mm/day-mz-kPa was observed with lactic acid casein and
sorbitol, 58.3% protein and 41.7% plasticizer, ﬁlms. This was the lowest WVP observed
in this study. Films made with lactic acid casein and glycerol, 50% protein and 50%
plasticizer, displayed a WVP of 59.3 gtmm/day-mz-kPa, which was the highest observed

41

42
(Table 4, 5; Figure 6). In general, ﬁlms made with sorbitol displayed lower WVP’s than

ﬁlms made with glycerol, at same protein plasticizer ratios, with a wider difference among
the lactic acid casein ﬁlms. A signiﬁcant difference (p<0.05) was witnessed among the
high protein, lactic acid casein ﬁlms, 34.0 g-mm/day-mz-kPa and 54.7 g-mm/day-mz-kPa for
ﬁlms containing sorbitol and glycerol respectively (Table 4). Protein type did not play a
signiﬁcant role in the WVP of the ﬁlms (Table 5). No trends were found pertaining to
protein concentration.

It had been shown in other studies using sorbitol and glycerol as the plasticizer that
ﬁlms made with sorbitol displayed lower WVP’s then those made with glycerol (McHugh
and Krochta 1994). This is due to the ability of glycerol to adsorb water more than
sorbitol, probably stemming from the more crystalline structure of sorbitol, making it more
stable (Sicard and Leroy, 1983). It was thought that as protein content increased in the
ﬁlms a signiﬁcant decrease in WVP would occur, due to more protein-protein interactions
in the ﬁlms matrix. Even though this trend did occur, however not statistically
signiﬁcantly, the high plasticizer content in the ﬁlms was probably high enough to
counteract the signiﬁcance of these interactions in preventing the passage of water vapor.
WVP’s didn’t vary between the two protein types, even though protein content in the
powder was substantially higher with the lactic acid casein (87.3% to 80.6% for rennet
casein). This was probably counteracted by higher portion of fat, being hydrophobic, and

ash, containing calcium that can promote crosslinking of the proteins.

43

Table 4. Effect of Plasticizer Type on Water Vapor Permeability (WVP) of Casein-Based

Edible Films (378°C, 90% RH.)

 

 

 

Treatmentl Lactic Acid Caseinz'3 Rennet Casein”
(Protein Powder%/
Plasticizer%)
S(37.5/62.5) 44.9 i 98'” 49.7 :i: 8.3'”
S(50/50) 45.0 i 9.0“ 49.6 :l: 6.6'”
S(58.3/41.7) 34.0 i 5.2" 39.6 :1: 3.6"
G(37.5/62.5) 54.9 i 1.6' 57.9 s: 4.9‘
0(50/50) 59.3 s 6.5’ 58.2 :t 2.5'
G(58.3/4l.7) 54.7 :t 6.2' 45.2 i 68'”

 

 

 

‘ Letter denotes plasticizer type: L=Lactic acid casein, R=Rennet casein; Protein powder%/p1asticizer% is

reported as a dry basis.

2 Different letters columnwise denote signiﬁcant diﬁ'erence (p<0.05).

3Mean:l:s.d.arereportedasg-mm/day-mz-kPa.

 

44

Table 5. Effect of Casein Type on Water Vapor Permeability (WVP) of Casein-Based
Edible Films (378°C, 90% RH.) '

 

 

 

 

 

 

Treatmentl Sorbitol” Glycerolz'J
(Protein Powder%/
Plasticizer%)
L(37.5/62.5) 44.9 i 9.8' 54.9 :t 1.6'
L(50/50) 45.0 i 9.0‘I 59.3 i 6.5'
L(58.3/41.7) 34.0 i 5.2'I 54.7 :1: 6.2'
R(37.5/62.5) 49.7 i 8.3' 57.9 i 4.9‘I
R(50/50) 49.6 i 6.6‘I 58.2 :1: 2.5'
R(58.3/4l.7) 39.6 :t 3.6' 45.2 i 6.8'
rLetter denotes casein type: L=Lactic acid casein, R=Rennet casein; Protein powdero/o/plasticizerf/o is
reported as a dry basis.

2 Diﬂ‘erent letters columnwise denote signiﬁcant difference (p<0.05).
3Meanisd. arereportedasg-mm/dayomztkPa.

 

45

WVP (g mm/m2 day kPa)

           

'9 A A Lactic Acid Casein Type
3 E In.
B. 8 8 C A
8 a E g g 3
m % .3 a g
Treatments 0 g

Figure 6. Water Vapor Permeability (WVP) of Casein-Based Edible Films
(378°C, 90% RH.)
Treatment: S=Sorbitol. G=Glycerol;
(Protein Pmdewpnstiem)

46

mgen permeability (OP)

Films made with lactic acid casein and sorbitol, 37 .5% protein and 62.5%
plasticizer, gave the lowest OP of 0.653 cc-um/day-mz-kPa. Films made with rennet casein
and glycerol, 37.5% protein and 62.5% plasticizer, gave an OP of 7.057
cc-um/day-mz-kPa, which was the highest OP observed (Table 6, 7; Figure 7). Plasticizer
type had an effect on OP only when rennet casein was the protein used (at low and
medium concentrations) and when glycerol was the plasticizer. These values were
signiﬁcantly higher (p<0.05) than the rest (Table 6). Films made with lactic acid casein
and glycerol, 50% protein and 50% plasticizer, displayed a signiﬁcantly higher (p<0.05)
OP than the rest of the lactic acid casein ﬁlms. Protein type had a signiﬁcant effect
(p<0.05) on OP when glycerol was used as the plasticizer. The rennet casein (low and
medium concentrations) and glycerol ﬁlms displayed higher OP’s than those made with
lactic acid casein and glycerol, and rennet casein (high protein content) and glycerol
(Table 7). Protein to plasticizer ratio did not play a signiﬁcant role in OP of the ﬁlms.

Since these tests were evaluated at 0% RH. we didn’t see a possible eﬁ‘ect of
water to act as a further plasticizer. At elevated RH.’s we could probably expect to see
glycerol effect OP more than sorbitol because of it’s greater aﬁinity towards water. Due
to these dry conditions OP differences based on protein concentration and type were not
seen because proteins are generally not reactive with oxygen. OP for the ﬁlms made with
rennet and glycerol might have been elevated due to some trapped water, because these

ﬁlms did gel quicker than the others, especially the higher protein concentration ﬁlms.

 

47
Table 6. Effect of Plasticizer Type on Oxygen Permeability (OP) of Casein-Based Edible

 

 

 

 

 

 

Films (23°C, 0% RH.)
Treatmentl Lactic Acid Caseinz‘3 Rennet Casein”
(Protein Powder%/
Plasticizer%)
S(37.5/62.5) 0.653 i 0.122‘I 0.713 i 0.155‘I
S(50/50) 0.733 t 0.133‘I 1.017 t 0.267‘
S(58.3/41.7) 0.813 :1: 0202‘ 0.963 :1: 0.153‘I
G(37.5/62.5) 0.880 :1: 1.101‘ 7.057 :1: 1831"
G(50/50) 2.177 :1: 0.544" 5.553 :1; 2842"
G(58.3/4l.7) 0.727 :t 0272‘ 1.837 t 0.791'
I Letter denotes plasticizer type: S=Sorbitol, G=Glycerol; Protein powder%/plasticizer°/o is reported as a

dry basis.
2 Different letters columnwise denote signiﬁcant diﬂ'erence (p<0.05).
3Meanis.d. arereportedascc-um/day-mz-kl’a.

48

Table 7. Effect of Protein Type on Oxygen Permeability (OP) of Casein-Based Edible
Films (23°C, 0% RH.)

 

 

 

 

 

 

Treatmentl Sorbitolz'3 Glycerol”
(Protein Powder%/

Plasticizer%)
L(37.5/62.5) 0.653 i 0.122' 0.880 i 1.101'
L(50/50) 0.733 i 0.133' 2.177 i 0.544'l
L(58.3/41.7) 0.813 i 0202‘I 0.727 2|: 0272'I
R(37.5/62.5) 0.713 3: 0.155'I 7.057 i 1.831"
R(50/50) 1.017 1- 0267‘ 5.553 i 2.842b
R(58.3/4l.7) 0.963 t 0.153' 1.837 i 0.791'

‘ Letter denotes casein type: L=Lactic acid casein, R=Rennet casein; Protein powder%/plasticizer% is
reported as a dry basis.

2 Diﬂ‘erent letters columnwise denote signiﬁcant difference (p<0.05).
3Mean:1:s.d.arereportedascc-11.111/day-m2-kPa.

 

49

 

 

 

 

 

   
   

 

OP (cc pmlmz day kPa)

Rennet

 

.................
...................
........

 
  

 
 

:7 A
S '3 e

g at g s g 6‘

w :3 g c, g

to V o h

Treatments 0 8’

Figure 7. Oxygen Permeability (OP) of Casein-Based Edible Films

(23°C, 0% RH.)
Treatments: s-Sorbitol, G=Glycerol;
(Protein Putnam/Plasticizer%)

50

Mechanical Properties
W

Upon conditioning, at 23°C and 50% RH, ﬁlms made with rennet casein and
sorbitol had a white ﬁlm layer form on their surface. It is not known what the white ﬁlm
that formed on the surface was, possibly a by-product of the hygroscopic properties of
sorbitol. However, this ﬁlm layer, did not seem to affect the TS of these ﬁlms. Films
made with rennet casein and sorbitol, 58.3% protein and 41.7% plasticizer, displayed a TS
of 15.117 MPa, which was the highest observed. Films made with lactic acid casein and
sorbitol, 37.5% protein and 62.5% plasticizer, displayed a TS of 0.415 MPa, which was
the lowest observed (Table 8, 9; Figure 8). Films made with sorbitol as the plasticizer in
all cases had signiﬁcantly higher (p<0.05) TS’s than those made with glycerol, at equal
protein to plasticizer concentrations (Table 8). Rennet casein ﬁlms tended to produce
stronger ﬁlms than lactic acid casein with either type of plasticizer, being more signiﬁcant
(p<0.05) at higher protein concentrations (Table 9). As Protein to plasticizer ratios
increased TS increased signiﬁcantly (p<0.05).

As expected ﬁlms containing sorbitol displayed a higher TS than ﬁlms formulated
with glycerol, at the same protein to plasticizer contents. This is probably due to their
greater crystallinity, and higher viscosity which it possesses, and the ability of glycerol to
hold more water at equivalent RH.’s, decreasing protein-protein interactions (Sicard and
Leroy, 1983). More calcium crosslinking, possibly being stronger than the direct protein-
protein bonding, in the rennet casein ﬁlms, because of the larger ash content, could be the

main factor in the higher TS of rennet ﬁlms compared to lactic acid ﬁlms. The increase in

51

Table 8. Effect of Plasticizer Type on Tensile Strength (TS) of Casein-Based Edible Films
(23°C, 50% RH.)

 

 

 

 

 

 

TreatmentI Lactic Acid Casein"3 Rennet Casein”
(Protein Powder%/

Plasticizer%)
S(37.5/62.5) 2.427 :t 00751" 3.827 :1: 0.307‘
S(50/50) 7.483 i 0.7457” 9.527 :1: 1.405b
S(58.3/41.7) 11.647 t 0.3800" 15.117 i 2.270c
G(37.5/62.5) 0.415 1 0.0603‘l 0.830 :1: 0.290‘l
G(50/50) 1.243 :1: 0.0273° 2.423 :1: 0.166“I
G(58.3/41.7) 2.507 i 0.0666' 4.497 a 0.698'

ILetter denotes plasticizer type: S=Sorbitol, G=Glyccrol; Protein powdeﬁ/dplasticizeﬂ/o is reported as a
dry basis.

2 Different letters columnwise denote signiﬁcant difference (p<0.05).
3 Mean 1: s.d. are reported as MP3.

 

52

Table 9. Effect of Protein Type on Tensile Strength (TS) of Casein-Based Edible Films
(23°C, 50% RH.)

 

 

 

 

 

 

Treatment’ Sorbitol” Glycerol”
(Protein Powder%/

Plasticizer%)
L(37.5/62.5) 2.427 a 0.075“ 0.415 i 0060‘
150/50) 7.483 i 0.746” 1.243 i 0.047“
L(58.3/41.7) 11.647 : 0.380c 2.507 :1: 0.067c
R(37.5/62.5) 3.827 a 0307' 0.830 i 0.290“
R(50/50) 9.527 i 1.405“ 2.423 :1: 0.166c
R(58.3/41.7) 15.117 3: 2.270“ 4.497 :1; 0.698“

Letter denotes casein type: L=Lactic acid casein, R=Rennet casein; Protein powdeﬁ/dplasticizef/o is
reported as a dry basis.

2 Different letters columnwise denote signiﬁcant difference (p<0.05).
3 Mean 1 s.d. are reported as MPa.

 

53

Lactic Acid Casein Type

            

 

6.86.50

Haw” A8 anvmv

E $5.86

. , 38‘3an

89¢me

Treatments

4. .................................... . . ANFVB.$Vm

Anus: wk

Figure 8. Tensile Strength (TS) of Casein-Based Edible Films

(23°C, 50% RH.)

Treatments: S=Sorbitol. G=Glycerol;

(Protein Pwder‘lb/Plasticizer‘lt)

54

TS as protein content increased is most likely due to an increase in protein-protein

interactions.

Elongation (E%)

Again ﬁlms made from rennet casein and sorbitol had a white ﬁlm form on their
surface upon conditioning, this appeared to make the ﬁlm brittle reducing the E%. Films
made with lactic acid casein and sorbitol, 50% protein and 50% plasticizer, displayed an
E of 253.6%, which was the highest observed. Those made with rennet casein and
sorbitol, 58.3% protein and 41.7% plasticizer, displayed an E of 17.9%, being the lowest
observed (Table 10, 11; Figure 9). Films made with glycerol displayed signiﬁcantly higher
(p<0.05) E%’s than those made with sorbitol at the same protein to plasticizer ratio,
except for the lactic acid casein (low protein content) and glycerol ﬁlms which had a lower
E% than lactic acid casein (low protein content) and sorbitol ﬁlms (Table 10). Protein
type did not contribute signiﬁcantly to the E%, except when sorbitol was used. This being
due to the white ﬁlm that formed on the surface of the rennet casein and sorbitol ﬁlms
(Table 11). No signiﬁcant trends in E% were present based on protein to plasticizer ratio.

Films containing glycerol had the higher E%, as expected, again because of the
ability of glycerol to absorb more water than sorbitol, acting to further plasticize the ﬁlms,
making them less brittle. Due to the higher crystallinity present in sorbitol would tend to
make them more rigid. However, as protein content increased a decrease in E% was not
observed, which was not expected. It was thought that E% would decrease because of
the increased protein-protein interactions increase, making them ﬁlm more resistant to

stretching.

 

55

Table 10. Effect of Plasticizer Type on Elongation (E%) of Casein-Based Edible Films
(23°C, 50% RH.)

 

 

 

 

 

 

Treatmentl Lactic Acid Casein2’3 Rennet Casein2'3
(Protein Powder%/

Plasticizer%)
S(37.5/62.S) 170.7 a 2.0“ 4.9 :1: 9.8‘
S(50/50) 156.0 :1: 6.1' 7.6 :1: 22.5“
S(58.3/41.7) 50.6 :1: 7.5“ 17.9 :1: 4.6'
G(37.5/62.5) 121.4 i 10.2“ 123.2 :1: 22.4“
G(50/50) 253.6 :1; 16.3“ 185.4 i 22.8“
G(58.3/41.7) 194.1 s. 20.6“ 223.5 :1: 22.7“

Letter denotes plasticizer type: S=Sorbitol, G=Glycerol; Protein powder%lplasticizeP/o is reported as a
dry basis.

2 Diﬂ‘erent letters columnwise denote signiﬁcant diﬁerence (p<0.05).
3 Mean :t s.d. are reported as %.

 

56

Table 11. Effect of Protein Type on Elongation (E%) of Casein-Based Edible Films
(23°C, 50% RH.)

 

 

 

 

 

 

Treatmentl Sorbitol2'3 Glycerol2'3
(Protein Powder%/

Plasticizer%)
L(37.5/62.5) 170.7 i 2.0' 121.4 3: 10.2“
L(50/50) 156.0 : 6.1‘I 253.6 s 16.3“
L(58.3/41.7) 150.6 i 7.5' 194.1 i 20.6““
R(37.5/62.5) 34.9 i 9.8b 123.2 a 22.3“
R(50/50) 77.6 a 22.5“ 185.4 i 22.8““
R(58.3/41.7) 17.9 :1: 4.6" 223.5 1“ 22.7““

TLetter denotes casein type: L=Lactic acid casein, R=Rennet casein; Protein powdeﬁ/dplasticizeﬁ/o is
reported as a dry basis.

2 Diﬂ‘erent letters columnwise denote signiﬁcant difference (p<0.05).
3 Mean i s.d. are reported as %.

  

 

 
   

E A -- Rennet Casein Type
S S 3 r: A
13' ‘15 3. ; fr?
Dr «'3 3 g
«71’ 18 6 h
Treatments 5 8

Figure 9. Elongation (E%) of Casein-Based Edible Films
(23°C, 50% RH.)
Treatment: S=Sorbitol, G=Glycerol;
(Protein Powder%lPlesticizer‘lt)

58

Comparisons to Other Films

Properties of casein-based edible ﬁlms developed in this study compared favorably
to synthetic polymers in some aspects, while in others they were inferior (Table 12). The
casein-based ﬁlms from this study included in Table 12 were chosen because they
possessed good properties in the various categories. These casein-based ﬁlms also
compared favorably to other protein-based ﬁlms that have been developed (Table 13).
Casein-based ﬁlms were very poor water vapor barriers as compared to synthetic ﬁlms.
Low density polyethylene (LDPE) is considered a good water barrier, while nylon 6 is
considered rather poor one. The WVP of casein-based edible ﬁlms is approximately 7
times greater than that of the nylon 6. This is most likely due to the hydrophilic
characteristics that proteins possess. The water barrier properties of these ﬁlms compared
rather favorably to other edible protein ﬁlms that have been developed. However, direct
comparisons cannot be made due to the different experimental conditions, ﬁlm
composition, and thickness. The only protein-based ﬁlms reported that possessed lower
WVP’s than our ﬁlms were the caseinate ﬁlms developed by Banerjee and Chen (1995)
and Park and Chinnan (1990). However, ﬁlms from those studies contained higher protein
concentrations and tested under less severe conditions. Typically, as temperature or
relative humidity rise so will the WVP.

The casein-based ﬁlms developed in this study possessed good oxygen barrier
properties compared to both synthetic and other protein-based edible ﬁlms. Ethylene vinyl
alcohol (EVOH) is considered a very good oxygen barrier and nylon 6 a good barrier.
The OP of our casein—based ﬁlms were comparable to that of EVOH. The OP’s of our

ﬁlms were also lower than any of the values reported for other protein-based ﬁlms. The

 

59

Table 12. Comparison of Selected Casein-Based Edible Films and Synthetic Polymers

 

 

Protein- Thickness WVP2 0P3 TS" E4 Reference
Plasticizerl (mm) (g-mm/ (cc-um/ (MPa) (%)
m2-d-kPg m2-d-kPa)
LA-S Film 0.203 45.03 0.71 7.48 156.0 Present
(50/50)
LA-G Film 0.203 54.69 0.77 2.51 194.1 Present
(58.3/41.7)
R-S Film 0.203 49.68 0.71 3.83 34.9 Present
(37.5/62.5)
R-G Film 0.203 58.15 3.95 2.42 185.4 Present
(so/50)
Synthetic
Polymers
LDPE 0.0254 - 1870“ 8.6-17 500 Salame
(1986)
HDPE 0.0254 0.02 427“ 17-35 300 Smith
(1986)
EVOH 0.0254 - 0.066 39.2-68.7 235-325 Foster
(56% von) (1986)
Nylon 6 0.0254 7.1 10.1 69-828 400-500 Tubrity &
Sibilia
(1986)

 

 

 

 

 

 

 

' Numbers in parenthesis denotes protein powder%lplasticizer %; L.A.=Lactic Acid Casein, R=Rennet
Casein, S=Sorbitol, G=Glycerol, LDPE=Low density polyethylene, HDPE=High density

polyethylene, EVOH=Ethylene vinyl alcohol (VOH-Vinyl alcohol).

2 Evaluated at 378°C and 90% RH.

3 Evaluated at 23°C and 0% RH.

‘ Evaluated at 23°C and 50% RH.

 

Table 13. Comparisons of Various Protein-Based Edible Fihns

6O

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Film TypeT Thickness WVP OP T83 Ez Reference
(Protein- (mm) (g-mm/ (cc-um/ (MPa) (%)
Plasticizer) m2-d-kPa) mz-d-kPa)
c'z-c 7.69-11.49 13.0-44.9 - - Park and
(SIM/16.9) (21°C, (30°C, Chinnan (1990)
85% 11.11.) 0% 11.11.)
srr-c 0.064 - 4.75 3.13-5.23 66.5-90.3 Brandenburg et
(sac/37.0) (25°C, 01. (1993)
0% 11.11.)
WG-6 3.82 Gennadios et
(HA/28.6) 108.4 (23°C, al.(l993b)
0.140 (378°C, 0% 1111.) 1.8 25 Aydt er al.
100% RH.) (1991)
NFDM-G 0.069 81.0 - 5.1 12.2 Maynes and
lactose (30°C, Krochta(l994)
extracted 61% RH.)
(mo/25.0)
NFDM-G 0.071 70.3 - 9.1 5.2 Maynes and
ultra-ﬁltered (30°C, Krochta (1994)
(75.01250) 65% 11.11.)
WPl-G 0.121 119.8 . - - McHugh and
«2.5/37.5) (25°C, Krochta (1994)
65% 11.11.)
Tvrr-s 0.129 61.92 - - - McHugh and
(625/375) (25°C, Krochta(l994)
_ 79% 1111.)
WPl-G 0.110 - 61.92 13.9 30.8 McHugh and
(70.0/30.0) (23°C, Krochta (1994)
50% RH.)
wrr-s 0.110 - 8.3 14.7 8.7 McHugh and
(sac/50.0) (25°C, Krochta (1994)
79% 11.11.)
801; 0.109 12.90 - 2.98 29.89 Banerjee and
(66.7333) (23°C, Chen (1995)
72% RH.)
cc-c 0.105 7.91 - 4.25 1.45 Banerjee and
(66.71333) (23°C, Chen (1995)
72% 11.11.)
(an-casein - - - 4.1 38.0 Motoki er al.
-G (1987)
(988/20)
tau-casein - - - 10.6 77.0 Motoki er al.
(1987)
transglnt.
(sen/2.0)

 

' Numbers in parenthesis denotes protein %lplasticizer %; CZ=Com Zein, SPI=Soy protein isolate,
WG=Wheat gluten, NFDM=Non fat dry milk, WPI=Whey protein isolate, SC=Sodium caseinate,
CC=Calcium caseinate, G=Glycerol, S=Sorbitol, n'ansglut.=transglutaminase.

2 Evaluated at 23°C and 50% 11.11.

 

61
TS and E% of these casein-based ﬁlms were considerably lower than the synthetic

polymers. TS’s were approximately 2 to 10 times weaker than the synthetic polymers,
while the E% was approximately 1.5 to 2 times lower. These casein-based ﬁlms were very
comparable to the other protein-based ﬁlms in TS, while they possessed much higher

E%’s than any of the reported protein-based edible ﬁlms.

Storage Study
Moisture Content

The casein-based edible ﬁlms proved to be ineffective in preventing moisture loss
in processed cheese slices. Moisture content of the cheese began at 39.32%, and those
slices wrapped in LDPE did not lose any signiﬁcant amount of moisture over the duration
of storage. However, cheese slices wrapped in the casein-based ﬁlms and the unwrapped
controls lost a signiﬁcant amount (p<0.05) of moisture, dropping to about 30% (Table
14). In fact, these ﬁlms didn’t even delay the loss of moisture. Both casein-based wraps
performed similarly to unwrapped controls, in retaining moisture in the cheese slices,
throughout storage (Table 15). This can be attributed to the poor moisture barrier
properties these ﬁlms possess. Also, this was probably a worst case scenario for
evaluating these ﬁlms due to the high moisture content of the cheese, and because of the
high RH. of the storage environment, which was 88%.
There was a dramatic increase in the moisture content of the casein-based ﬁlms, when
used as a cheese wrap (Table 16). The lactic acid casein and sorbitol ﬁlm had a moisture

content of 9.02%, while the lactic acid casein and glycerol ﬁlm had an initial moisture

62

Table 14. Effect of Storage Time on Moisture Content of Processed Cheese Slices

Packaged in Various Wraps (22°C, 88% RH.)

 

Percent Moisture

 

 

 

 

Wrap 'ije
Storage LDPEW L.A.-S Film L.A.-G Film No Wrap “3““
(Days) (so/50) ““5 (58.3/41.7) 1.2.3
0 39.32 a 0.64“ 39.32 a 0.64“ 39.32 a 0.64“ 39.32 i 0.64“
5 39.62 a 4.77“ 31.10 : 2.23b 34.04 i 0.54b 30.75 s 4.77“
10 41.41 i 1.39“ 28.19 i 0.44““ 32.32 s 0.48““ 29.39 a 088"
15 41.86 1. 0.39“ 28.01 a 2.39““ 27.73 i 1.20“ 29.69 1: 0.22“
20 40.29 1 1.00“ 26.48 s. 0.75“ 31.55 i 0.69““ 28.24 i 1.71“
25 39.59 :t 0.23“ 25.70 1 0.64“ 29.30 1: 0.91“ 28.86 1 0.77“
30 42.75 i 0.15“ 28.92 i 2.18““ 29.85 :r 1.35“ 30.77 a 0.51“

 

 

 

 

 

‘ LDPE=Low density polyethylene, L.A.=Lactic acid casein, S=Sorbitol, G=Glycero1; numbers in
parenthesis (protein powdeﬂ/dplasticizeﬁ/o).
2 Different letters columnwise denotes a signiﬁcant diﬁerence (p<0.05).

3 Mean :t s.d.

 

 

Table 15. Effect of Wrap Type on Moisture Content of Processed Cheese Slices

63

During Storage (22°C , 88% RH.)

 

Percent Moisture

 

 

 

 

Storage Time (Days)
Wrap 0"3 5"“ 10’“ 15 2'“ 20 2“ 25 2’ 30 ’3
1

Type
LDPE 39.32 1 0.64“ 39.62 1 4.77“ 41.41 1 1.39“ 41.86 1 0.39“ 40.29 1 1.00“ 39.59 1 0.23“ 42.75 1 0.15“
L. A.-S 39.32 1 0.64“ 31.10 1 2.23“ 28.19 1 0.44“ 28.01 1 2.39“ 26.48 1 0.75“ 25.70 1 0.64“ 28.92 1 2.18“
Film
(SO/50)
L. A.-G 39.32 1 0.64“ 34.04 1 0.54“ 32.32 1 0.48“ 27.73 1 1.20“ 31.37 1 0.69‘ 29.30 1 0.91“ 28.85 1 1.35“
Film
(58.3l4r.7)
N0 Wrap 39.32 1 0.64“ 30.75 1 1.03“ 29.39 1 0.88“ 29.69 1 0.22“ 28.24 1 1.71“ 28.86 1 0.77“ 30.77 1 0.51“

 

 

 

 

 

 

 

 

3 LDPE=Low density polyethylene, L.A.=Lactic acid casein, S=Sorbitol, G=Glycerol; numbers in

parenthesis (protein powder°/n/plasticizer°/o).
2 Different letters columnwise denotes a signiﬁcant difference (p<0.05).
3 Mean 1 s.d.

 

 

64

Table 16. Effect of Storage Time on Moisture Content of Edible Films Used as a Wrap
for Processed Cheese Slices (22°C, 88% RH.)

 

Percent Moisture

 

 

 

 

Wrap Type

Storage L.A.-S Film L.A.-G Film

(Days) (50/50) W gas/41.7) W
0 9.02 11.21“ 17.80 1 3.86“
5 37.60 1 2.58“ 45.47 1 1.59“
10 37.85 1 3.76“ 42.73 1 1.43““
15 36.69 1 2.73“ 37.46 1 0.72“
20 37.96 11.08“ 44.12 11.71““
25 37.29 1 0.12“ 43.10 1 0.24““
30 35.72 14.16“ 38.14 14.27“

 

 

 

 

I L.A.=Lactic acid casein, S=Sorbitol, G=Glycerol; numbers in parenthesis
(protein powder%/ plasticize1%).

2 Different letters columnwise denotes a signiﬁcant difference (p<0.05).

3 Means i s.d.

65
content of 17.80%. In the ﬁrst 5 days of storage these quickly rose to 37.60% and

45.47% for the casein and sorbitol, and casein and glycerol ﬁlms respectively, then stayed
relatively constant throughout the rest of storage. Which is due to the hydrophilic nature
of both the casein and the plasticizer (Swaisgood, 1985; Sicard, 1982). This was the same
trend that was observed with the moisture loss of the cheese slices. If we look at the
weight change of the cheese slices and wrap (package system) over the period of storage
we see that the water lost by the cheese in retained in the ﬁlm (Table 17). The package
system of cheese slices with the casein and sorbitol ﬁlm maintained a constant weight
throughout storage, while the package system of the cheese slices wrapped with the casein
and glycerol ﬁlm actually gained weight during storage. This is most likely attributed to

the ability of glycerol to absorb moisture from the air (Merck, 1989).

 

Casein-based ﬁlms were ineffective at retaining color in processed cheese slices
also (Table 18). Cheese slices wrapped in LDPE retained their original, creamy orange,
appearance throughout the duration of storage. A signiﬁcant change (p<0.05) was
observed in the Hunter L-Value for cheese slices wrapped in the casein-based ﬁlms and
the unwrapped controls occurring shortly after being packaged (Table 18a). This value
shows a darkening of the cheese, decreasing in value ﬁom 71.07 to about 56.0. There
was no signiﬁcant difference in L-Value among the cheese slices wrapped in the casein-
based ﬁlms and the unwrapped slices. The unwrapped and casein-based ﬁlm wrapped
slices also witnessed a signiﬁcant change (p<0.05) in redness during storage (Table 18b).

This is shown by the a-value increasing from 8.60 to around 11.0, indicating a slight

66

Table 17. Weight Gain of Processed Cheese Slices and Wrap During Storage
(22°C, 88% RH.)

 

 

 

Storage (Days) LDPELZ-3 L.A.-S Film L.A.-G Film
(so/so) u: (58.3/4l.7) “3"
0 0.000 1 0.000“ 0.000 1 0.00“ 0.000 1 0.00“
5 0.016 1 0.002“ 0.731 1 0.17“ 3.342 1 0.76“
10 0.017 1 0.003“ -0054 1 0.37“ 2.791 1 0.77“
15 0.005 1 0.002“ 0.383 11.31“ 3.587 1 1.59“
20 -0.026 1 0.035“ 0.995 1 0.03“ 4.148 1 0.45“
25 0.020 1 0.017“ 0.206 1 0.39“ 2.856 1 1.02“
30 -0.006 1 0.004“ 0.600 1 0.94“ 1.916 1 1.06“

 

 

 

 

' LDPE=Low density polyethylene, L.A.=Lactic acid casein, S=Sorbitol, G=Glycerol; numbers in
parenthesis (protein powdeﬂ/dplasticizeﬁ/o).

2 Diﬁ'erent letters columnwise denotes a signiﬁcant difference (p<0.05).

3 Means :t s.d. reported in grams.

 

67

Table 18. Effect of Wrap Type on Color Changes in Processed Cheese Slices
During Storage (22°C, 88% RH.)

11) L-Value (0 black to 100 white)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Storage Time (Days)
Wrap 0"“ 5"“ 10“ 157““ 20"“ 25"“ 30"
Type'
LDPE 71.07 1 70.80 1 71.33 1 71.37 1 69.97 1 69.80 1 69.70 1
0.60“ 0.95“ 1.36“ 0.50“ 0.15“ 0.35“ 0.60“
L.A.-S 71 .07 1 59.73 1 57.87 1 58.07 1 56.20 1 54.93 1 55.53 1
Film 0.60“ 1.50“ 0.47“ 3.89“ 1.32“ 1.10“ 3.36“
(50/50)
L.A.-G 71.07 1 67.17 1 63.70 1 57.33 1 63.33 1 60.90 1 58.13 1
Film 0.60“ 1.19“ 1.22“ 2.82“ 2.1 1“ 1.56“ 2.42“
(583/417)
No Wrap 71.07 1 61.30 1 60.53 1 60.33 1 56.10 1 55.67 1 55.23 1
0.60“ 0.60“ 1.21““ 0.32“ 2.86“ 0.80“ 1.45“
b) a-Valne (- green to + red)
Storage Time (Days)
Wrap 0"“ 5"“ 10"“ ' 15"“ 20“” 25““ 30““
Type1
LDPE 8.60 1 9.10 1 8.93 1 8.33 1 9.00 1 8.70 1 8.10 1
0.35“ 0.36“ 0.38“ 0.61“ 0.70“ 0.10“ 0.72“
L.A.-S 8.80 1 10.67 1 11.57 1 10.57 1 10.03 1 11.47 1 10.93 1
Film 0.35“ 1.04“ 0.15“ 0.55“ 0.21“ 0.75“ 0.61“
(50/50)
L.A.-G 8.60 1 10.43 1 10.33 1 11.43 1 9.60 1 11.13 1 10.63 1
Film 0.35“ 0.31“ 0.47“ 0.65“ 0.27“ 0.47“ 0.21“
(SSS/41.7)
No Wrap 8.60 1 11.30 1 11.30 1 10.73 1 11.67 1 12.10 1 11.43 1
0.35“ 0.44“ 0.76“ 0.31“ 0.81“ 0.46“ 0.61“
c) b-Value (- blue to + yellow)
Storagé Time (Days)
Wrap 0"“ 5"“ 10" 15"“ 20" 25““ 30"“
Type'
LDPE 33.93 1 34.60 1 34.87 1 33.90 1 34.30 1 34.37 1 34.77 1
0.58“ 0.17“ 0.55“ 0.27“ 0.10“ 0.06“ 0.49“
L.A.-S 33.93 1 33.50 1 32.47 1 32.00 1 31.30 1 30.87 1 31.10 1
Film 0.58“ 0.87“ 0.65“ 1.83“ 0.61“ 0.74“ 1.74“
(50150)
L.A.-G 33.93 1 35.30 1 33.73 1 31.77 1 33.40 1 33.27 1 32.20 1
Film 0.58“ 0.53“ 1.42“ 1.30“ 1.32““ 0.71““ 1.14““
(583/417)
No Wrap 33.93 1 33.73 1 34.10 1 33.77 1 31.77 1 32.13 1 31.77 1
0.58“ 1.59“ 1.33“ 0.59“ 1.16“ 0.71““ 0.67“

 

 

 

 

 

 

 

 

I LDPE=Low density polyethylene, L.A.=Lactic acid casein, S=Sorbitol, G=Glycerol; numbers in

parenthesis (protein powdeﬁ/dplasticizer‘h).
2 Different letters columnwise denotes a signiﬁcant difference (p<0.05).
3 Means :1: s.d.

 

 

 

 

68

reddening of the cheese. No signiﬁcant difference was observed between the cheese slices
wrapped in the casein-based ﬁlm or the unwrapped slices, again occurring shortly after
being packaged. A signiﬁcant change (p<0.05) in Hunter b-value occurred towards the
end of storage in cheese slices wrapped in the casein-based ﬁlms and the unwrapped
controls (Table 18c). Values decreased from 33.93 to around 32.0 at the 20 day period
and remaining relatively constant, indicating a slight loss of yellowness in the cheese. This
color change is attributed to the loss of moisture in the cheese slices, because this color
change (L-value and a-value) occurred at the same rate as did moisture loss. As with
moisture loss following the initial change values tended to remain constant for the rest of
storage.

The color of the casein-based ﬁlm itself aﬁer being used to wrap processed cheese
slices changed signiﬁcantly (p<0.05) during storage (Table 19). Casein-based ﬁlms were
signiﬁcantly more transparent (p<0.05) than the LDPE ﬁlm. This is shown by the Hunter
L-value at day 0 of storage, where LDPE has a value of 23.63 and casein-based ﬁlms have
values of 13.40 and 11.83 for the ﬁlms containing sorbitol and glycerol, respectively
(Table 19a), values closer to 0 being more transparent since the black tile was used as the
background tile. However, by the end of storage there was no diﬁerence in transparency
among the ﬁlms. Hunter a-value were similar for all the ﬁlms and remained constant
throughout storage (Table 19b). Films at the beginning of storage were similar (being
slightly bluish), with the casein-based ﬁlms signiﬁcantly changing (p<0.05) to a slight
yellowish color during storage, while the LDPE ﬁlms remained unchanged (Table 19c).

This is observed in the similar changes of the Hunter b-values of the casein-based ﬁlms

 

Table 19. Comparison of Color Changes of Wraps After Storage on Processed

a) L-Value (0 transparent to 100 white)

Cheese Slices (22°C, 88% RH.)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Storage Time (Days)
Wrap 0"" 5“ 10"" 15"" 20"" 25"J 30“
Type1
LDPE 23.63 :t - - - - - 24.47 :1:
0.35'I 0.75'
L.A.-S 13.40 i 20.60 :t 21.93 i 21.93 :1: 20.33 :1: 20.93 i 20.83 1
Film 1.64“ 1.05“ 2.12“ 1.35“ 1.52“ 0.67“ 2.97“
(SO/50)
L.A.-G 11.83i 21.17: 22.9031: 19.17: 20.703: 21.67: 21.6721:
Film 1.07“ 1.37“ 1.77“ 3.19“ 1.18“ 2.71“ 3.36“
(58.3/41.7)
b) a-Value (- green to + red)
Storage Time (Days)
Wrap 0“ 5“ 10‘: 15“ 20“ 25“ 30“
Type1
LDPE 0.13 :1: - - - - - 0.67 :1:
1.25' 1.36'
L.A.-S -0.80 i -0.30 i -0.60 :t -0.567i 0.67 d: 0.33 i -0.37 :1:
Film 0.36'I 0.20'I 0.85' 0.32' 0.29'I 0.21' 0.50II
(50150)
L.A.-G -0.50 :t -0.367 i -0.53 i -0.87 i -0.33 i -0.73 :l: -0.23 1
Film 030' 0.45' 045| 057' 0.25' 0.16' 0.58'
(58.31417)
c) b-Value (- blue to + yellow)
Storage Time (Days)
Wrap 0“ 5"J 10” 15“ 20“ 25" 30"
Type1
LDPE -1.47 1 - - - - - -1.73 1
0.85' 0.90'
L.A.-S -1.87 i 0.50 i 0.30 i 0.60 i -0.60 :1: 0.10 i 0.63 :1:
Film 0.85“ 0.20“ 0.446“ 0.44“ 0.61“ 0.36“ 0.51“
(50/50)
L.A.-G -l.50 i 0.50 :1: 0.73 :t 0.57 i 0.30 i 0.73 i 0.67 1
Film 0.78'I 0.56' 0.60'l 0.35' 0.20'I 0.21'| 0.21"
(583I4l.7)

 

 

 

 

 

 

 

 

LDPE=Low density polyethylene, L.A.=Lactic acid casein, S=Sorbitol, G=Glycerol; numbers in

parenthesis (protein powder°/o/plasticizer%).

2 Diﬁ‘erent letters columnwise denotes a signiﬁcant difference (p<0.05).
3 Means i s.d.

 

 

 

 

70
from about -1.50 to about 0.65. These color changes in the ﬁlm might be attributed to

some residual cheese sticking to the ﬁlm.

CONCLUSIONS

. Casein-based ﬁlms developed in this study possess poor water barrier properties.

. They do possess good oxygen barrier properties, similar to synthetic polymers with
good oxygen barrier properties.

Sorbitol used as a plasticizer will provide ﬁlms with better overall properties than if
glycerol were used as the plasticizer.

. Casein-based ﬁlms tend to have inferior properties compared to synthetic ﬁlms, except
for oxygen barrier properties.

. Overall properties of casein-based ﬁlms from this study compare favorably to
properties of other protein-based ﬁlms.

. These casein-based ﬁlms did not act as good wrap for processed cheese slices.

71

RECOMMENDATIONS

Work needs to be done on these ﬁlms to improve their WVP properties. This
could be accomplished with the use of crosslinking agents, or the incorporation of lipids
into the ﬁlms or as part of a bilayer ﬁlm. Studies need to be done to determine the
properties of these ﬁlms at different environmental conditions, temperature and RH.
Biodegradation studies should be conducted to determine biodegradability of these ﬁlms
and to establish testing methods for this. Further studies need to be done incorporating
these ﬁlms into other food systems, especially oxygen sensitive food items where these
ﬁlms could provide their greatest utility. Processes to seal these ﬁlms must also be

developed if they are to be used as an alternative packaging material.

72

 

LIST OF REFERENCES

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AOAC 1990. Meat and Meat Products. In “Oﬁcial Methods of Analysis,” 15th ed.
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Avena-Bustillos, R., and Krochta, J .M. 1993a. Water vapor permeability of caseinate-
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Avena-Bustillos, R., Cisneros-Zevallos, L.A., and Krochta, J .M. 1993b. Optimization of
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74

Cuq, B., Gontard, N., and Guilbert, S. 1995. Edible ﬁlms and coatings as active layers.
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Dalgleish, DO. 1982. Milk proteins - chemistry and physics. In “Food Proteins,” ed. P.F.
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