egg quality and salmonella spp. growth in shell eggs packaged in

Egg Quality and Salmonella spp. Growth in Shell Eggs Packaged in Modified Atmosphere Packaging

by

Divya Aggarwal, B.S.

A Thesis

In

FOOD SCIENCE

Submitted to the Graduate Faculty of Texas Tech University in

Partial Fulfillment of the Requirements for

the Degree of

MASTER OF SCIENCE

Approved

Leslie Dawn Thompson Chairperson of the Committee

Christine Zocchi Alvarado

Mindy Brashears

Chance Brooks

Karen Killinger Mann

Fred Hartmeister Dean of the Graduate School

May, 2008

Texas Tech University, Divya Aggarwal, May 2008

ii

ACKNOWLEDGEMENT

I would first like to express my deepest gratitude to Dr. Leslie Thompson, my major

professor, for giving me this opportunity to study at Texas Tech University, and work on

this project, and for always supporting and encouraging me throughout this experience.

She has been like a family to me in US. To my family for giving me endless support

through good and bad. I would also like to express my appreciation to the other members

of my committee, Dr. Christine Alvarado, Dr. Mindy Brashears, and Dr. Chance Brooks

for their help and guidance throughout this project, Dr. Karen Killinger for her invaluable

help in planning and standardizing the microbiology procedure, Ana Marie Luna for all

her help with sample testing, data analysis, and overall support. I could not have done this

without all of you. Lastly, but not the least, I would like to thank Cal-Maine Foods Inc.

for donating the shell eggs used in this study and the USDA-ARS Nutrient Data Lab for

funding the project.


iii

TABLE OF CONTENTS

ACKNOWLEDGMENTS ii

ABSTRACT vi

LIST OF TABLES viii

LIST OF FIGURES xiii

CHAPTER

I. INTRODUCTION 1

II. LITERATURE REVIEW 6

US. Egg Industry 6

Egg Marketing 7

Egg Structure 8

Measurement of Shell Egg Quality 10

Modified Atmosphere Packaging 10

Commonly Used MAP Gases 12

Carbon dioxide in MAP 14

Effect of MAP on Microorganisms 16

Concerns about MAP 17

Egg Spoilage 18

Functional Properties of Egg 20

Hard-cooked Eggs 22

Yolk Surface Color 22

Salmonella 23


iv

III. METHODOLOGY 28

Egg Quality 28

Foam Capacity and Stability 29

Color Measurement 30

pH of Yolk, Albumen, and Whole Egg 30

Yolk Index 31

Haugh Unit 31

TBARS 31

Data Analysis 31

Microbiological Study 32

Sample Preparation 33

IV. RESULTS AND DISCUSSIONS 35

Color of Raw Albumen 35

Color of Raw Yolk 41

Color of Hard-cooked Albumen 52

Color of Hard-cooked Yolk Surface 68

Color of Inner Part of Hard-cooked Yolk 82

Peeling Property of Hard-cooked Shell Eggs 82

Foam Capacity 86

Foam Stability 87

Albumen pH 95

Whole Egg pH 101


v

Yolk pH 101

Haugh Unit 111

Yolk Index 112

Microbial Quality of Eggs 124

Thiobarbituric Acid Reactive Substances (TBARS) 132

Conclusion 136

REFERENCES 138

APPENDIX


vi

ABSTRACT

The effect of three types of modified atmosphere packaging (MAP) on the quality

attributes and Salmonella spp. growth in oiled fresh USDA Grade AA shell eggs during

storage was investigated. Shell eggs were subjected to one of four packaging treatments:

(1) control - air; (2) 20% CO2/0.4% CO/79.6% N2; (3) 20% CO2/80% O2; and (4) 20%

CO2/80% N2. Eggs were stored for up to 30 d, in a retail case at a temperature of 6 ± 1 C

(refrigerated) or on shelves at 21 ± 1 C (abusive). Eggs were packed 8 to a tray with a

tray considered as the experimental unit. Two trays per treatment per temperature per day

were prepared in each trial with a total of three trials being conducted. Packages were

opened and sampled on days (d) 1, 7, 14, 21, and 30 for determination of pH (yolk,

albumen, whole egg), color (L*, a*, b*), TBARS, foam capacity and stability, Haugh

units, and yolk index (YI). Data were analyzed by ANOVA in a 2 (temperature) x 4

(packaging treatment) x 5 (time-points) factorial design using programs in SAS. Where

appropriate, means were separated by LSMeans. Whole egg pH was lower at both

temperatures for the three MAP treatments (P < 0.05). Albumen pH for MAP treatments

was significantly lower regardless of temperature as compared to the controls (P < 0.05).

MAP treatment was effective at 6 C in maintaining lower yolk pH compared to control. A

higher Haugh unit and yolk index throughout the storage at both temperatures was

maintained by MAP treatment compared to the control treatment (P < 0.05). TBARS, and

foam stability was similar for the MAP treatments and the control. Modified atmosphere

packaging maintained higher foam capacity at both temperatures compared to control.

Modified atmosphere packaging was effective in reducing egg deterioration and loss of

functional quality during storage at refrigerated and abusive storage temperatures.


vii

For the effect of three types of MAP treatments on Salmonella spp. growth in shell

eggs, sanitized shell eggs, inoculated with 40 ul cocktail of Salmonella Enteritidis phage

13 nalidixic acid resistant, S. Heidelburg/ 3347, S. Typhimurium ATCC 14028 strains

and concentration of 1 x 104 cells were subjected to four packaging treatments. Eggs

were packed 6 to a tray and sampled on days 1, 7, and 14. Data were analyzed by

ANOVA in a 2 (temperature) x 4 (packaging treatment) x 3 (storage time) factorial

design using programs in SAS. Where appropriate, means were separated by Duncan’s

multiple range test and main effects were studied. MAP treatments were similar in their

effect on Salmonella spp. growth at both the storage temperatures in trial 1 and 2.

However, in trial 3, at refrigerated temperature, air packed eggs had the lower Salmonella

count than the three MAPs. At abusive temperature, high-ox treatment had higher

Salmonella count (P < 0.05).


viii

LIST OF TABLES

4.01 Raw albumen lightness, hue angle and chroma value ± SEM of eggs packed in modified atmosphere packaging (20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2), or air stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 16)

36

4.02 Raw albumen lightness, hue angle and chroma ± SEM of eggs packed in

20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air stored at refrigerated (6 C) and abusive (21 C) temperature (n = 20)

37

4.03 Raw yolk lightness ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4)

42

4.04 Raw yolk lightness ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature. (n = 4)

43

4.05 Raw yolk hue angle (o) and chroma ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature. (n = 16)

44

4.06 Raw yolk hue angle (o) and chroma ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 20)

45

4.07 Raw yolk chroma ± SEM of shell eggs packed in


46

4.08 Hard cooked albumen lightness ± SEM of shell eggs packed in


54


ix

4.09 Hard cooked albumen lightness ± SEM of shell eggs packed in 20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature. (n = 4)

55

4.10 Hard cooked albumen hue angle (o) ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature. (n = 4)

56

4.11 Hard cooked albumen hue angle (o) ± SEM of shell eggs packed in


57

4.12 Hard cooked albumen chroma ± SEM of shell eggs packed in


58


20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature.

59


20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 30).

60

4.15 Hard cooked yolk surface lightness ± SEM of shell eggs packed in


71

4.16 Hard cooked yolk surface lightness ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4)

72

4.17 Hard cooked yolk surface hue angle (o) ± SEM of shell eggs packed in


73


x

4.18 Hard cooked yolk surface hue angle (o) ± SEM of shell eggs packed in 20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4)

74

4.19 Hard cooked yolk surface chroma ± SEM of shell eggs packed in


75

4.20 Hard cooked yolk surface chroma ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4)

76

4.21 Lightness, hue angle (o) and chroma value of inner part of hard cooked ±

SEM egg yolk packed in 20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 16)

84

4.22 Lightness, hue angle (o) and chroma value of inner part of hard cooked ±

SEM eggs yolk packed in 20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 20)

85

4.23 Foam capacity ± SEM of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, , or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 30)

89

4.24 Foam capacity ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d

at refrigerated (6 C) and abusive (21 C) temperature (n = 24) 90

4.25 Foam stability ± SEM of shell eggs packed in 20%CO2/0.4%CO/79.6%N2

(CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6)

92

4.26 Foam stability and albumen pH ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored at refrigerated (6 C) temperature (n = 30).

93

4.27 Foam stability and albumen pH ± SEM of shell eggs packed and stored for 1,

7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 24). 94


xi

4.28 Albumen pH of ± SEM shell eggs packed in 20%CO2/0.4%CO/79.6%N2 (CO2/CO/N2), 20%CO2/80%N2 (CO2/N2), 20%CO2/80%O2 (High-ox), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6)

98

4.29 Whole egg pH value ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 30)

103

4.30 Whole egg pH ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d

at refrigerated (6 C) and abusive (21 C) temperature (n = 24) 105

4.31 Egg yolk pH ± SEM of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 30)

107

4.32 Egg yolk pH ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d at

refrigerated (6 C) and abusive (21 C) temperature (n = 24) 109

4.33 Haugh unit ± SEM of shell eggs packed in 20%CO2/0.4%CO/79.6%N2


114

4.34 Yolk index and Haugh unit ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored at refrigerated (6 C) temperature (n = 30)

116

4.35 Yolk index and Haugh unit ± SEM of shell eggs packed and stored for 1, 7,

14, 21, 30 d at refrigerated (6 C) temperature (n = 24) 117

4.36 Yolk index ± SEM of shell eggs packed in 20%CO2/0.4%CO/79.6%N2


122

4.37 Egg yolk TBARS value ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air (n = 40)

126

4.38 Egg yolk TBARS value ± SEM of shell eggs packed and stored for 1, 7, 14,

21, 30 d (n = 32) 127

4.39 Log cfu of Salmonella cocktail/g ± SEM of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air for trials 1, 2, and 3 and stored for 1, 7, 14 d at refrigerated (6 C) and abusive (21 C) temperature (n = 6)

134


xii

4.40 Log cfu of Salmonella cocktail/g ± SEM of shell eggs packed in 20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, and air for trials 1, 2, and 3 and stored for 1, 7, 14 d at refrigerated (6 C) and abusive (21 C) temperature (n = 8)

135


xiii

LIST OF FIGURES

A.01 Raw albumen lightness of eggs packed in 20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature. (n = 16)

107

A.02 Raw albumen hue angle (o) of eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature. (n = 16)

108

A.03 Raw albumen chroma of eggs packed in 20%CO2/0.4%CO/79.6%N2,


109

A.04 Raw yolk lightness of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4)

110

A.05 Raw yolk lightness of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4)

111

A.06 Raw yolk hue angle (o) of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,


112

A.07 Raw yolk chroma of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 16)

113

A.08 Raw yolk chroma of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,


114

A.09 Hard cooked albumen lightness of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4)

115

A.10 Hard cooked albumen lightness of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4)

116

A.11 Hard cooked albumen hue angle (o) of shell eggs packed in


117


xiv

A.12 Hard cooked albumen hue angle (o) of shell eggs packed in


118

A.13 Hard cooked albumen chroma of shell eggs packed in


119

A.14 Hard cooked albumen chroma shell eggs packed in


120

A.15 Hard cooked albumen chroma of shell eggs packed in


121

A.16 Hard cooked yolk surface lightness of shell eggs packed in


122

A.17 Hard cooked yolk surface lightness of shell eggs packed in


123

A.18 Hard cooked yolk surface hue angle (o) of shell eggs packed in


124

A.19 Hard cooked yolk surface hue angle (o) of shell eggs packed in


125

A.20 Hard cooked yolk surface chroma of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for days 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4)

126

A.21 Hard cooked yolk surface chroma of shell eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) treatments (n = 4)

127

A.22 Hue angle (o) of inner part of hard cooked egg yolk packed in


128


xv

A.23 Foam capacity of shell eggs packed in 20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored at refrigerated (6 C) and abusive (21 C) temperature. (n = 30)

129

A.24 Foam stability of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,


130

A.25 Albumen pH of shell eggs packed and stored for 1, 7, 14, 21, 30 d at

refrigerated (6 C) temperature (n = 24) 131

A.26 Albumen pH of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, or air and stored at refrigerated (6 C) temperature (n = 30)

132

A.27 Albumen pH of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,


133

A.28 Whole egg pH of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 30)

134

A.29 Whole egg pH of shell eggs packed and stored for 1, 7, 14, 21, 30 d at


A.30 Egg yolk pH of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,


136

A.31 Egg yolk pH of shell eggs packed and stored for 1, 7, 14, 21, 30 d at


A.32 Haugh unit of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,

20%CO2/80%N2, 20%CO2/80%O2, and air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6)

138

A.33 Haugh unit of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,


139

A.34 Haugh unit of shell eggs packed and stored for 1, 7, 14, 21, 30 d at



xvi

A.35 Yolk index of shell eggs packed in 20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air and stored at refrigerated (6 C) temperature (n = 30)

141

A.36 Yolk index of shell eggs packed and stored for 1, 7, 14, 21, 30 d at


A.37 Yolk index of shell eggs packed in 20%CO2/0.4%CO/79.6%N2,


143

A.38 Egg yolk TBARS value of shell eggs packed and stored for 1, 7, 14, 21, 30

d (n = 32) 144

A.39 Log cfu of Salmonella cocktail/g of eggs packed in

20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air for trial 1and stored for 1, 7, 14 d at abusive (21 C) temperature (n = 8)

145


20%CO2/0.4%CO/79.6%N2, 20%CO2/80%N2, 20%CO2/80%O2, or air for trial 2 and stored for 1, 7, 14 d at refrigerated (6 C) and abusive (21 C) temperature (n = 8)

146



147



148

A.43 CO2 level (%) in packages with CO2+ CO (20%CO2/0.4%CO/79.6%N2),

CO2 (20%CO2/80%N2), and high-ox (20%CO2/80%O2) treatments for trial 2 when stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 2)

149


CO2 (20%CO2/80%N2), and high-ox (20%CO2/80%O2) treatments for trial 2 when stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 2)

150


CO2 (20%CO2/80%N2), and high-ox (20%CO2/80%O2) treatments for trial 3 when stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 2)

151


xvii

A.46 CO2 level (%) in packages with CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), and high-ox (20%CO2/80%O2) treatments for trial 3 when stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 2)

152


1

CHAPTER I

INTRODUCTION

The per capita egg consumption in U.S. in 2006 was 255.7 (USDA-WAOB, 2007). In

2006, total egg production reached the record high of 90.9 billion eggs which was up by 1

percent from the previous year. Of the nearly 91 billion eggs produced, 78 billion were

eggs produced for the U.S. table egg industry. Egg industry contributes 4 billion dollars

annually to the United States (Keener et al., 2000).

Physical and chemical changes begin to occur, the moment an egg is laid, affecting

it’s quality negatively. When the eggs are exposed to elevated temperatures for an

extended period of time, microbial growth increases and egg quality decreases (Curtis et

al., 1996). Therefore, newly laid eggs must be refrigerated soon after laying and

processing.

Currently, shell egg processing practice involves conveying or manually collecting

and transferring eggs from the laying house to the egg packaging plant. After cleaning

and sorting, the eggs are packed in cartons or flats, which are packed in cases, and

palletized. The pallet is then placed under refrigeration to equilibrate to 7 C (Keener et

al., 2000). According to Anderson et al. (1992), it takes about 7 to 10 d for the eggs in the

center carton to reach 7 C when starting at an initial temperature of 25 to 30 C. The

cooling capacity of the refrigerated trucks for transporting eggs and the retail outlets is

limited. Thus during shipping, the cooling time for the eggs is further extended (Keener

et al., 2000). Most shell egg processors place a 4-week expiration date on the label since

egg quality deteriorates with prolonged storage. Currently most eggs processed in the


2

United States have an average sell by date of 30 d post processing and are sold by 19 d

post processing (Bell et al., 2001; Patterson et al., 2001).

Loss of CO2 from shell eggs occurs during storage and is a major cause for loss of

functional quality in eggs. The deterioration of eggs cannot be prevented completely.

Cold storage if done properly, is effective in preserving the egg quality, however, the

eggs deteriorate more slowly in storage if carbon dioxide is added to the air (Sharp and

Stewart, 1931). Keener et al. (2000) demonstrated that the CO2 cooling and CO2 storage

could extend the shelf life of eggs to greater than 14 weeks. The Haugh unit values for

CO2 cooled and CO2 stored eggs were higher than that of controls stored without CO2.

Curtis et al. (1995) and Jones et al. (2002) showed significant reduction in microbial load

and growth rate during extended refrigerated storage of eggs treated with CO2 as

compared to untreated controls. The study also reported increased shelf life and increased

quality of eggs from grade A to AA. Also, the effect of the presence of carbon dioxide in

air is more striking at higher temperatures where refrigeration is not operative in helping

to preserve the egg (Sharp and Stewart, 1931).

During the past two decades, eggs internally contaminated with Salmonella

Enteritidis have emerged among the leading causes of food-borne disease (CDC, 1996).

Public health surveillance data estimated 637,000 cases of human illness per year due to

the consumption of contaminated raw or undercooked eggs. Undercooked and raw shell

eggs have been identified as the most common sources of Salmonella Enteritidis

infection upon investigations of outbreaks. (CDC-MMWR, 2003).

Transovarian Salmonella Enteritidis contamination of shell eggs from Salmonella

Enteritidis-infected laying hens have been identified as a source of the problem. Freshly


3

laid eggs are rarely reported to harbor more than a few hundred S. Enteritidis cells (Chen

et al., 2002; Gast and Holt, 2000a; Humphrey et al., 1991). Prompt refrigeration of eggs

to the internal temperature of 7.2 C can inhibit the growth of Salmonella and reduce the

chances of consumers being exposed to an infective pathogen dose. Hence, the U.S.

Department of Agriculture (USDA, 1998) has established 7.2 C as the maximum

mandatory temperature for transportation and storage of shell eggs, to prevent the rapid

multiplication of small initial levels of S. Enteritidis. However, in commercial units,

cooling is slow and several days are required to reach this specific internal egg

temperature to restrict bacterial growth (Curtis et al., 1995; Thompson et al., 2000).

Modified atmosphere packaging technology enables manipulation of gases in the

environment surrounding the food within the packaging material. Specific gas mixtures

of O2, CO2, CO and N2 are commonly used. The MAP technique has several advantages

such as increased shelf life, increased safety of food, minimal handling, decreased

microbial contamination, reduction in the bacterial count of specific microorganisms. All

these benefits result in overall cost savings and consumer safety and satisfaction.

Carbon dioxide is considered to be a major antimicrobial factor in MAP. It inhibits

the growth of microorganisms by increasing the lag phase and generation time during the

logarithmic phase of growth of the organism. Sharp and Stewart (1931), in their

experiments showed that the eggs deteriorate less rapidly in storage if a little carbon

dioxide is added to the air. Sharp and Stewart (1931), suggested the use of carbon dioxide

in egg storage should receive serious consideration as a means of improving quality of

eggs during storage.


4

Modified Atmosphere Packaging may provide one of the conditions restricting the

growth of microorganisms. A study by Silliker and Wolfe (1980), indicated that carbon

dioxide is effective in reducing the growth of Salmonella species compared to food stored

in air after ten days of storage. The Salmonella count for carbon dioxide treatment was

thousand fold lower than the air stored one.

So far, studies have not been conducted on the impact of MAP technique on the

quality of and microbial growth in shell eggs. Also, no study has examined the effect of

MAP gases like CO, CO2 on the Salmonella species in shell eggs. With the difficulties in

reducing shell egg temperature for storage and a number of studies demonstrating the

benefits of modified atmosphere packaging, MAP could potentially be effective in

maintaining the functional quality of shell eggs, USDA quality grade, lower the rate of

lipid oxidation, reduce Salmonella risk, and extend the shelf life of the shell eggs during

distribution and storage, thus resulting in increased consumer safety and satisfaction. An

increase in the shelf life of eggs also would benefit the producers interested in export

opportunities (Keener et al., 2000). Organic eggs, whose production is expanding in U.S.

to meet the market demand, are included in speciality eggs (Blank, 1997). The cost of

production and the retail price of organic eggs is higher than traditional eggs. Despite the

higher cost, the organic egg market is growing (Patterson et al., 2001). This indictaes that

the consumers are willing to pay a higher price if they perceive added benefits such as

high quality standards or improved safety.

According to Stadelman and Cotterill (1986), the packaging and design of the egg

carton affects the final decision of the customer to purchase the product. Thus, a new

innovative egg package if attractive, could have certain benefits over conventional


5

packaging, like improves egg quality, could become popular among customers and could

increase the sale. However, more information is needed to provide a better understanding

of the effect of the MAP packaging of shell eggs on consumer perception and their

purchasing habits.

The first objective of the following research was to evaluate the effect of three types

of gas mixtures used in MAP systems on the quality and functional properties of the oiled

fresh USDA Quality Grade AA shell eggs during storage at refrigerated (6 C) and

abusive (21 C) temperatures. The quality and functional properties of the shell eggs

would be measured in terms of Haugh units, foaming capacity and stability, pH, color,

yolk index, and lipid oxidative stability. The second objective was to evaluate the effect

of the three gas mixtures in MAP with CO as a packaging gas, on the survival and growth

of Salmonella spp. in shell eggs stored at refrigerated and under abusive conditions.


6

CHAPTER II

LITERATURE REVIEW

U.S. Egg Industry

The per capita egg consumption in U.S. in 2006 was 255.7 (USDA-WAOB, 2007). In

2006, egg production reached record high of 90.9 billion eggs which was up by 1 percent

from the previous year, with 78 billion of these eggs used in the U.S. table egg industry.

Egg production has increased from about 60 billions eggs in 1984 (USDA-NASS, 2007).

Egg industry contributes 4 billion dollars annually to the United States economy (Keener

et al., 2000). About 70 percent of the table eggs are sold as shell eggs while the remainder

are processed into liquid, frozen or dried pasteurized egg products. Majority of the egg

products are used in institutions foodservice or are further processed into foods such as

cake mixes, pasta, ice cream, mayonnaise, and bakery goods (President’s Council on

Food Safety, 1999).

Geographically, California is the center for the western U.S. commercial egg

production. In the eastern U.S., Ohio, Indiana, and Pennsylvania are the major egg

producing states. Other states with significant egg production are Iowa, Minnesota, and

Georgia (President’s Council on Food Safety, 1999).

Currently, shell egg processing practice involves conveying or manually collecting

and transporting eggs from the laying house into the egg packaging plant. Eggs are spray

washed with a chlorinated hot water solution at 40 C and then visually inspected and

sorted. Then, the eggs are conveyed onto a computer-controlled scale, weighed, and

packed according to size into cartons. Size categories of eggs includes pee-wee, small,

medium, large, extra large, and jumbo. Cartons are packed into a case of thirty dozen, and


7

the cases are placed on a pallet of about 20 cases. The pallets are placed in a large

refrigerator to equilibrate to 7 C (Keener et al., 2000). According to Anderson et al.

(1992), it takes about 7 to 10 d for the eggs in the center carton to reach 7 C when starting

at an initial temperature of 25 to 30 C. Generally, eggs destined for retail markets are

shipped immediately after palleting. The cooling capacity of the refrigerated trucks for

transporting eggs and the retail outlets is limited. Thus during shipping, the cooling time

for the eggs further increase (Keener et al., 2000). When the eggs are exposed to

elevated temperatures for an extended period of time, especially the center ones,

microbial growth increases and egg quality decreases (Curtis et al., 1996). Most shell egg

processors put a 4 week expiration date on the label since egg quality deteriorates with

prolonged storage. Currently most eggs processed in the United States have an average

sell by date of 30 d and are sold by 19 d post processing (Bell et al., 2001; Patterson et

al., 2001). An increase in the shelf life of eggs also would benefit the produces interested

in export opportunities (Keener et al., 2000).

Egg Marketing

According to Patterson et al. (2001), egg exports have increased 23% from 1995 to

1996. Also, speciality eggs represent 3 to 5% of retail cartoned eggs in US. Speciality

eggs are uniquely different from conventional eggs as they may be fertile, have less

cholesterol, less fat, more beneficial vitamins, produced by floor or range managed hens

or any combination above. Speciality eggs may fulfill one or more specific needs, such as

quality attribute, emotional need, health benefit, or others (Looper, 1996). Also, they

offer consumer a choice (Blank, 1997). Organic eggs, whose production is expanding in

U.S. to meet the market demand, are included in speciality eggs (Blank, 1997). The cost


8

of producing organic eggs is higher than cost of producing traditional eggs. The retail

price for organic eggs is 2.9 times higher than for traditional eggs, and producers earn 2.4

times as much for organic eggs. Despite the higher cost, the organic egg market is

growing (Patterson et al., 2001).

Egg accounts for 0.7 to 1.0% of total sales in supermarkets in the U.S. Eggs are major

contributor to supermarket sales. However, this level of sales could be improved. The egg

industry has helped to promote egg sales in supermarkets within store studies of customer

shopping habits, novel packaging and special promotions. More attention is given to egg

packaging and it’s effect on consumer purchases than the egg movement in the

supermarkets. The author emphasized that an appealing egg carton in terms of color and

design determines the final decision of the customer to purchase the product (Stadelman

and Cotterill, 1986).

From the above discussion it can be concluded that a new packaging of the eggs,

which can be attractive, has certain benefits over the conventional packaging, like

improves egg quality, can become popular among customers and can increase the sale.

The consumers would be ready to pay a higher cost of the product, like they do for the

speciality eggs.

Egg Structure

The average weight of a shell egg is about 60 g (Cotterill and Geiger, 1977). The

outer most covering of the egg is the shell and it constitutes 9.5% of the total egg weight

(Chung and Stadelman, 1965; Cotterill and Geiger, 1977). It consists of calcium

carbonate (94%), magnesium carbonate (1%), calcium phosphate (1%) and organic

matter, primarily protein (4%) (Stadelman and Cotterill, 1977). Albumen accounts for


9

about 63% of the egg weight and yolk about 27.5% (Chung and Stadelman, 1965;

Cotterill and Geiger, 1977). Egg shell is made for most part with the matrix consisting of

interwoven protein fibers and spherical masses and interstitial calcite crystals (Romanoff

and Romanoff, 1949). This structure results in small pores in the shell from where gas

and moisture exchange occurs. These pores are partially sealed by keratin protein, and

allow for the loss of carbon dioxide and moisture from egg. These pores may also permit

bacterial penetration through to the shell membranes. The surface of calcified shell is

covered by a foamy layer of protein called cuticle. It covers the pores and impedes

microbial invasion of the egg contents (Board, 1968). However, during egg washing, this

layer is removed (Stadelman and Cotterill, 1986). The albumen of egg occurs in four

layers. These are the chalaziferous or inner thick white, which is adjacent to the vitelline

membrane and continuous with the chalaza. The next outer layer is the inner thin white

surrounded by the outer thick white. The outermost layer is the thin white layer. The

percentage of total white found in the four layers varies depending on the strain of the

laying hen, age of the hen and age of the egg. The next layer is the inner and outer layer

of shell membranes. Shell membranes are thin membranes and the chief defense layer

against bacterial invasion. Together, these membranes are only 0.01 to 0.02 mm thick.

The yolk of the egg consist of latebra, germinal disc and concentric layers of light and

dark yolk surrounded by vitelline membrane. The air cell in shell eggs develops as a

result of separation of the two shell membranes, usually at the large end of the egg, as the

egg content shrinks during cooling. The air cell continues to increase in size as moisture

and carbon dioxide are lost during storage of the egg (Stadelman and Cotterill, 1986).


10

Measurements of Shell Egg Quality

Shell thickness is one of the most direct measurement of egg shell quality. A shell

thickness of at least 0.33 mm is needed if the egg has to be moved through normal market

channels without breaking with a 50% chance. Haugh unit is the most widely used

measure of albumen condition (Stadelman and Cotterill, 1986). The method consists of

measuring height of the thick albumen. The Haugh unit is an expression relating egg

weight and height of thick albumen (Haugh, 1937). The higher the Haugh unit, the better

the quality of egg white. A shell egg with a Haugh unit greater than 72 is considered an

AA grade (USDA, 1990). The quality of the egg yolk is affected by several

characteristics like color, strength of the yolk membrane, spherical condition. The

spherical nature of egg yolk is expressed as yolk index (Stadelman and Cotterill, 1977).

Modified Atmosphere Packaging

The use of controlled atmosphere was recorded as early as 1927 for extending the

shelf life of apples. The apples were stored in atmospheres with reduced oxygen and

increased carbon dioxide concentrations. After a decade, modified atmosphere storage

with increased carbon dioxide concentration was used to transport beef carcasses long

distances and was shown to increase the shelf life by up to 100% (Davies, 1995). Later,

in 1970s, MAP was introduced as a packaging strategy for meat, bacon, fish, sliced

cooked meats and cooked shellfish and was successful indicating increased consumer

demand for food with longer shelf life and less preservatives (Phillips, 1996). Currently,

MAP techniques are being used in fresh or chilled foods, including raw and cooked meats

and poultry, fish, fresh pasta, fruits and vegetables, coffee, tea and even bakery products

(Phillips, 1996). Some of the advantages of MAP are increased shelf life, high quality


11

product, ease of slice separation, clear view of the product, little or no need for use of

chemical preservatives, centralized packaging. However, some drawbacks are, added

cost, increased pack volume and hence increased retail display space and transportation

cost, temperature control requirement, product safety establishment, specialized

equipment and different gas formulation requirement (Davies, 1995).

The packaging material for MAP is mostly made from one or more of the four

polymers namely polyvinylchloride (PVC), polyethylene terephthalate (PET),

polyethylene (PE) and polypropylene (PP), depending on the required features. Some of

the commonly considered factors when choosing a packaging film are barrier properties,

machine capability, sealing reliability, antifog properties. Barrier properties refer to

permeability of the film to various gases and the water vapor transmission rate. Machine

capability is the capacity for trouble-free operation. Sealing reliability is the ability to seal

to itself and to the container (Smith, 1993).

Type of material used in packaging procedure, initial gas mixture influence the

atmosphere within the product. Some packaging materials allow diffusion of gases in and

out of the package during storage. If the film is fully permeable then the atmosphere

within the packaging gradually becomes the same as the air outside. If the film is semi–

permeable, an equilibrium modified atmosphere results. Some films provide complete

barrier to the movement of gases in either direction (Phillips, 1996).

A major advancement in the modified atmosphere packaging techniques have been

the introduction of ‘intelligent’ or ‘active’ packaging systems (Phillips, 1996). The use of

oxygen scavengers together with oxygen indicators is a common example of active

packaging procedure. In active packaging, chemicals are incorporated into the packaging


12

but are not in contact with the food. These chemicals, then, interact with the packaging

atmosphere and ensure that the optimal conditions are maintained, thus enhancing the

shelf life of the product (Phillips, 1996).

Commonly Used MAP Gases

The three most commonly used gases in MAP are oxygen, nitrogen and carbon

dioxide. Recently, carbon monoxide has become another popular MAP gas, especially for

the meat industry. Some other suggested gases include nitrous and nitric oxide, sulfur

dioxide, ethane and chlorine. However, most of these gases have not been accepted

commercially due to safety, legal, consumer response and cost reasons (Church, 1993).

Oxygen in MAP has several effects on food. It is responsible for the bright red color

in meat due to the formation of oxymyoglobin. It also affects the bacterial flora on the

food product. It inhibits the growth of anaerobes but stimulates the growth of aerobic

bacteria. Low level of oxygen, less than 0.5%, results in brown to grey color formation

due to metmyoglobin in meat products (Church, 1993). Thus fresh meat is frequently

packed in 80% oxygen atmosphere in order to maintain oxygenation and the light cherry

red color (Phillips, 1996). Conversely high concentrations of oxygen may result in

rancidity particularly in high fat products due to oxidative mechanism. Such food

products like fish or unsmoked bacon are therefore packed in atmospheres without

oxygen (Phillips, 1996).

Nitrogen in MAP is an inert gas which is used as a packaging filler to prevent

package collapse. In the food products like fresh meat that are packed in high carbon

dioxide concentrations, nitrogen is used as filler gas because of it’s low solubility in


13

water and lipid. Carbon dioxide, however, is soluble in meat tissue and results in

collapsing of the package during storage (Gill and Penney, 1988).

Modified atmosphere packaging technology enables manipulation of gases in the

environment surrounding the food within the packaging material. Specific gas mixtures

of O2, CO2, CO and N2 can be used. The MAP technique has several advantages such as

increased shelf life, increased safety of food, reduced handling at retail, decreased

microbial contamination, reduction in the bacterial count of specific microorganisms. All

these benefits result in overall cost savings and consumer safety and satisfaction.

As early as 1978, Gee and Brown (1978a,b) demonstrated that microbiological and

color shelf life of ground beef patties were significantly higher when exposed to 1% CO

as compared to samples exposed to air. Later Luno et al. (1998) showed that a low

percentage of CO in modified atmosphere packaging, greatly reduced the psychrotrophic

population. Carbon monoxide has shown to reduce total aerobic population numbers,

Brochothrix thermosphacta, extend the shelf life and slow oxidative reactions. However,

it did not affect lactic acid bacteria (Luno et al., 2000). In another study by Nissen et al.

(2000), CO inhibited Yersinia enterocolitica, L. monocytogenes , controlled the growth of

E. coli O157:H7. However, the effect on Salmonella spp. was limited and laid emphasis

on the temperature control during storage. Viana et al. (2005) reported that pork loins in

1% CO MAP had highest consumer acceptance score compared to vacuum, 100% CO2,

100% O2, and 100% CO packed ones. Luno et al. (2000) reported that CO in MAP

increased the shelf life of beef steeks and slowed oxidative reactions compared to

reference containing 70% O2 + 20% CO2 + 10% N2 .


14

Carbon dioxide in MAP

Carbon dioxide is considered to be a major anti-microbial factor in MAP. It inhibits

the growth of microorganisms by increasing the lag phase and generation time during the

logarithmic phase of growth of the organism. It’s effect is influenced by the initial and

final concentration of the gas, the temperature of storage and the original population of

the organisms. According to Reddy et al. (1992), microbial growth is reduced at high

concentration of carbon dioxide in a variety of products and the effect increases as the

storage temperature decreases. The antimicrobial effect of carbon dioxide has been

summarized by Farber (1991) into some major theories like alteration of cell membrane

functioning including effects on nutrient intake and absorption, direct inhibition of

enzyme systems, penetration of membranes resulting in changes to intracellular pH,

direct changes to physico-chemical properties of proteins. According to Church (1993),

effectiveness of carbon dioxide depends on the microflora present in the food and the

product characteristics. It is most effective in foods where the normal spoilage organisms

consist of aerobic, gram negative psychrotrophic bacteria.

Sharp and Stewart (1931), in their experiments showed that the eggs deteriorate less

rapidly in storage if a little carbon dioxide is added to the air. According to the study,

carbon dioxide concentration between 0.5 and 0.6% in the air at cold storage

temperatures, retarded the development of off flavors and odors in eggs that occured due

to chemical changes in the egg contents. The eggs stored in carbon dioxide were better in

condition of thick white, yolk index, viscosity of yolk, taste, and in color of white, as

compared to eggs stored under normal conditions. Carbon dioxide very markedly

retarded the development of the yellow, orange, or pink condition of the white. In the


15

control eggs stored in the air, the thick white was flat and runny, while in carbon dioxide-

stored eggs, the thick white stood up around the yolk in more of a jelly-like condition. It

was also shown that atmospheres containing 1, 5, 15, 30, and 100 per cent carbon dioxide

maintained better odor and flavor than the normal atmosphere storage in shell eggs

(Sharp and Stewart, 1931).

At 25% carbon dioxide in the storage atmosphere, the keeping quality of shell eggs

was better than the air stored one at both 30 C and 45 C. At cold storage temperatures,

less carbon dioxide is required than the higher temperatures to achieve a similar effect.

Also, the effect of the presence of carbon dioxide in air is more striking at higher

temperatures where refrigeration is not operative in helping to preserve the egg (Sharp

and Stewart, 1931).

The deterioration of eggs cannot be prevented completely. Cold storage if done

properly, is effective in preserving egg quality, however, the eggs deteriorate less rapidly

in storage if carbon dioxide is added to the air (Sharp and Stewart, 1931). According to

Sharp and Stewart (1931), the ultimate consumer bases his opinion of the egg on it’s taste

and odor and on the appearance of opened egg, and therefore improvement in the quality

of storage eggs using carbon dioxide should receive serious consideration.

Swanson (1953) conducted a study to test the value and practicability of using carbon

dioxide as an aid in preserving the quality of cartoned eggs as they pass through normal

market channels. In the study, albumen quality scored using Haugh unit was done for the

eggs in the cartons overwrapped with a plastic bag containing solid carbon dioxide

pellets, as compared to non-carbon dioxide treated ones. The Haugh unit value for the

carbon dioxide treatment was the highest. The author emphasized the advantage of


16

adding carbon dioxide as the interval between laying and packaging increased. In the

commercial practice the time lapse between laying and cartoning is sufficiently great to

allow the escape of considerable amount of carbon dioxide from the eggs.

As early as in 1950’s, suggestions for the use of overwrap that could be heat sealed to

the carton and that could be relatively impermeable to transfer of moisture and gas was

made. It was suggested to add carbon dioxide in the solid form as a small pellet in the

pack. Often eggs are being transported at considerable distances from producer to

consumer centers, therefore making this packaging method interesting (Swanson, 1953).

The author suggested that the additional costs involved in the packaging would be offset

by improved quality preservation and that the study suggested that the plan may be

feasible.

Loss of CO2 from shell eggs occurs during storage. Storage of eggs in MAP

environment with CO2 as one of the gas, may overcome this problem. In 2000 Keener et

al. (2000) demonstrated that the CO2 cooling and CO2 storage could extend the shelf life

of eggs to greater than 14 weeks. Also, the Haugh unit values increased for these eggs.

Curtis et al. (1995) and Jones et al. (2002) showed significant reduction in microbial load

and growth rate during extended storage of eggs treated with CO2 . The studies also

reported increased shelf life and increased quality of eggs from grade A to AA.

Effect of MAP on Microorganisms

Modified Atmosphere Packaging may provide conditions restricting the growth of

microorganisms. Storage at low temperature can be another condition. According to

Leistner (1995), the combination of chill temperature and MAP generally results in a

more effective and safer storage conditions, and longer shelf life. According to (Phillips,


17

1996), Salmonella spp. and Campylobacter spp. are the two major causes of bacterial

food–borne illness in both UK and US, both of which are potential contaminants of MAP

foods such as poultry and dairy products. A study by Silliker and Wolfe (1980), indicated

that carbon dioxide was effective in reducing the growth of Salmonella spp. compared to

food stored in air after ten days of storage. The Salmonella count for carbon dioxide

treatment was thousand fold lower than the air stored one. Another study by Tassou et al.

(1996) showed that modified atmospheres containing carbon dioxide or 100% nitrogen,

have bacteriostatic and bactericidal effects on Salmonella Enteritidis, in the absence of

temperature abuse. Nychas and Tassou (1996), reported that S. Enteritidis multiplies

rapidly at 10 C in fresh fish and poultry when vacuum packaged as well as MAP

containing 100% N2, 20% CO2:80% O2, and 100% CO2. However, at 3 C, the organisms

survived but did not grow significantly. Sawaya et al. (1995) concluded that growth of

Enterobacteriaceae, including Salmonella spp. was reduced with increasing

concentration of carbon dioxide, provided storage temperature was taken into

consideration.

Salmonella are facultative anaerobes and can grow and survive in the absence of

oxygen. Research shows that atmosphere of storage can influence the growth rate of

Salmonella. Under certain circumstances, growth of Salmonella has been enhanced.

Nissen et al. (2000) found in ground beef, various Salmonella strains at 10 C increased to

higher numbers in CO2/low CO atmosphere as compared to O2 containing packaging.

Concerns about MAP

In the past, several concerns about MAP have arisen. According to Renault et al.

(1994), different storage conditions can affect the atmosphere surrounding the food


18

product. A pack which allows 10% oxygen at 10 C might theoretically induce

anaerobiosis at 20 C. As a result of the inhibition of the growth of naturally occurring

microorganisms in the food, differential multiplication of pathogenic microorganisms can

occur in the food (Hotchkiss and Banco, 1992).

The use of CO in the primary package for fresh meat has raised concern in US for

masking spoilage that could occur in fresh meat products (Eilert, 2005). Later, FDA

(2004) issued a finding that low levels of CO did not mask spoilage that could occur in a

fresh meat package. Food and Drug Administration (2004) reported that while the color

did not degrade in the package containing CO, offensive odors could still form in the

presence of CO. Study by Sorheim et al. (1999) found that low levels of CO were not

inhibitory to the growth of spoilage organisms.

Egg Spoilage

The major causes for the change in the quality of eggs during storage period are

dependent upon several factors such as microorganisms, quality of fresh eggs,

temperature of storage, passage of undesirable odors and flavors into the egg, loss of

water from the egg, and the pH of the egg contents (Sharp, 1929).

According to Sharp (1929), some of the detrimental changes that occur in eggs during

storage are the change of the thick jelly-like white surrounding the yolk of the egg to a

fluid condition; the passage of water from the white to the yolk, producing more fluid

conditions of the yolk contents; and the weakening of the yolk membrane, causing yolk

of the egg to flatten when the egg contents are removed from the shell.

Deterioration of egg quality occurs faster in higher temperatures of 30 C to 40 C than

the ambient temperature of 20 to 23 C (Chakraborty et al., 2005).


19

The pH of the white of a freshly laid egg is 7.6. This pH increases to 9.7, during

storage, due to loss of carbon dioxide (Sharp, 1929). Thus, there is an increase in the

alkalinity of approximately one hundred times. The pH of the yolk of the fresh egg is

about 6.0. During storage, yolk pH also increases to about 6.8. However, the change in

pH value of the yolk is slower than that of the white. The increase in the pH of the yolk is

due to loss of carbon dioxide from the yolk to the white which in turn is lost to the

surrounding air (Sharp, 1929). If the eggs are stored in the ordinary air, the pH of the egg

contents rise and the deterioration of egg due to decomposition of the egg proteins is

accelerated. Sharp (1929), suggested that the alkalinity in eggs due to rise in pH can be

easily neutralized and controlled by placing the eggs in atmosphere containing small

amounts of carbon dioxide. Carbon dioxide is taken up by the eggs to establish an

equilibrium between the concentration of carbon dioxide in the egg and in the air. Some

of this carbon dioxide in the egg, forms carbonic acid in the presence of water, and

neutralizes alkalinity in egg. Thus, by the use of proper concentration of carbon dioxide,

the egg can be stored to and maintained at a quality similar to a fresh egg for a longer

period of time.

According to Sharp (1929), a major deterioration factor that results from alkaline pH

of the white is the thinning of the thick and a flattening of the yolk as indicated by the

weakening and change in the permeability of yolk membrane. The keeping quality of the

yolk improves as the carbon dioxide content of the egg storage atmosphere is increased.

Another deterioration factor resulting from alkaline pH, watery white, is produced in two

ways. Firstly, the jelly-like white becomes more fluid until it cannot be recognized as

thick white. However, at relatively low pH, thick white maintains it’s jelly-like property


20

for a longer period of time. This decrease may be caused by the precipitation of a protein

the thick white. The largest amount of thick white is obtained at intermediate pH values

and thus at moderate concentrations of carbon dioxide (Sharp, 1929).

When the concentration of the carbon dioxide is relatively high, the pH of the egg is

lowered to a point near the isoelectric point of one of the proteins of the proteins of the

thick white. This is an indication of the localization of the globulin protein in the thick

white. This is marked by turbidity in the thick white. This turbidity can be made to

disappear if some of the carbon dioxide is allowed to escape from the egg. Sharp (1929),

also reported that the concentration of carbon dioxide required for the storage of eggs in

the atmosphere so that the pH of white correspond to that of the fresh eggs, lessens as the

temperature decreases. At room temperature, 10 to 12 % carbon dioxide is needed in the

atmosphere to hold the white at the pH of 7.6. Near freezing temperature, only 3 %

carbon dioxide in atmosphere is required to maintain the egg white pH. Sharp (1929)

suggested the introduction of carbon dioxide in the air atmosphere of cold storage rooms,

in appropriate concentrations, so as to retard the destructive changes in egg. He has

emphasized the importance of the control of the pH of egg contents as a factor in egg

preservation.

Functional Properties of Egg

A foam is a colloidal dispersion of gas phase dispersed in a liquid phase (Baniel et al.,

1997). Upon beating liquid albumen, air bubbles are trapped in it and a foam is formed.

During the beating of the albumen, air bubbles decrease in size and increase in number

causing the translucent albumen to take on an opaque and moist appearance. Meringues,


21

angel cake, soufflés and foamy omelets are all application of egg foaming properties

(Stadelman and Cotterill, 1986).

Foam is formed due to the unfolding of the protein molecules so that the polypeptide

chain are arranged with their long axes parallel to the surface (Griswold, 1962). As a

result of the change in the molecular configuration, loss of solubility of some albumen

occurs and collects at the liquid-air interface. This adsorption film is essential to the

stability of the foam. The major foaming components of egg white are ovomucin and

globulin. Ovomucin is responsible for the formation of insoluble film which stabilizes the

foam (MacDonnell et al., 1955). Globulins contribute to high viscosity and decreased

tendency for liquid to drain away from air bubbles. Globulins also lower surface tension

which promotes foaming (Stadelman and Cotterill, 1986).

Temperature and pH are two important factors that affect the foaming properties

(Liang and Kristinsson, 2005). Elevation of temperature results in decreased surface

tension. Thus at room temperature as compared to refrigerated temperatures, a foam is

easier to form (requires less work) and foam volume is increased. Foam stability,

however, is affected little by a change in temperature from 20 C to 34 C. At lower pHs,

the whipping time for the white increases, but the foam stability also greatly improves.

The heat stability of specific proteins is influenced by the pH and increases when pH is as

low as 6.5. At an albumen pH of 7, ovalbumin, ovomucoid and lysozyme are protected

against heat damage. Increased stability of foams allows time for heat to penetrate cakes,

and cause coagulation without collapse of air cells and thus prevent shrinkage of the cake

during baking (Stadelman and Cotterill, 1986).


22

Hard–cooked Eggs

The sale of hard cooked and peeled eggs to the retail market, food manufacturers, and

foodservice is increasing (Stadelman and Cotterill, 1986). According to Irmiter et al.

(1970), a high quality hard cooked egg peels easily without the shell adhering to the

coagulated albumen, does not have dark rings on the yolk surface and the shell does not

break during cooking.

Yolk Surface Color

The chicken egg yolk has mainly alcohol soluble xanthophylls, lutein and zeaxanthin

as the naturally occurring pigments. It also has some amount β-carotene and

cryptoxanthin. The visual impression of egg yolk color determines acceptability. Egg

yolk pigments contribute a pleasing yellow color to foods like baked products, noodles,

ice creams, custards, sauces, and omelets. Preference has been shown for the gold or

lemon colored yolks (Stadelman and Cotterill, 1977). One of the major problem when

eggs are hard cooked is that of a greenish-black discoloration on the surface of the yolk

of hard-cooked eggs. Xanthophylls are relatively stable to the normal food preparation

conditions. This greenish-black discoloration on the surface of the egg yolk is due to the

formation of FeS (Baker et al., 1967). Albumen is the source of H2S. According to Fruton

and Simmonds (1958) H2S can be released from L–cystine by the enzyme cystine

desulfhydrase. It is possible that this enzyme, and perhaps others, are responsible for the

presence of H2S in the albumen of an egg (Baker et al., 1967). It was postulated that

during heating the outward pressure of gases tend to prevent the reaction between H2S

from the albumen and Fe from the yolk. However, during cooling, the egg contents

contract and the H2S moves towards the yolk contacting it’s surface. Also, the study


23

shows that Fe is released from the yolk upon heating. Studies show that increased

alkalinity in the egg yolk favors FeS formation. The pH of the yolk has a very definite

effect upon the blackening at the interface of the yolk and the albumen. The length of

storage time before cooking also has marked effect on the yolk color. The longer the eggs

are stored, the darker the yolks become after cooking. This is because during the storage,

the pH of the yolk increases. Unlike the pH of the yolk of fresh egg which is 6.0, the

average pH of the yolks of eggs stored for 3 weeks at 2 C was 6.9 (Baker et al., 1967).

Chen and Chen (1984), found that the cooked eggs with pH around 7.5 and 7.0 had

highest H2S content. Also, on addition of acidic chelating agents like citric acid,

Na2EDTA, and polyphosphates at 0.1% or malic acid, monosodium phosphate, sorbic

acid, succinic acid, and tartaric acid at 0.5% to the raw mixtures of egg reduced the H2S

content of the cooked samples.

Another study by Gossett and Baker (1981) on the prevention of the green gray

discoloration in cooked liquid whole eggs, found that on addition of acidic chelating

agents, the discoloration was prevented.

Salmonella

Salmonella is a rod-shaped, motile bacterium with a few nonmotile exceptions,

nonsporeforming facultative anaerobe, and Gram negative. The bacteria has a

widespread occurrence in animals, especially in poultry and swine. Environmental

sources of the organism include water, soil, insects, factory surfaces, kitchen surfaces,

animal feces, raw meats, raw poultry including eggs, and raw sea foods (FDA-CFSAN,

2006). There are over 2,324 Salmonella serovars, but all have been categorized into two

species; Salmonella enterica and Salmonella bongori (Jay, 2005). Most of them are


24

classified as Salmonella enterica and are sub–categorized into five subtypes. For

serotyping Salmonella, the O antigen is used and sorts the serovars into groups A, B, C1,

C2, D, E1. This is based on the similarities and commonality of specific O antigens.

Flagellar or H antigens are used after O antigen is found. This order of classification

results in highly identified bacteria which are then called serovars (Jay, 2005).

Salmonellosis is characterized by diarrhea, abdominal cramps, nausea, vomiting,

fever, and headache. The onset of symptoms begin within 6 to 72 hours after consuming

food and last for 4 to 7 days. The symptoms are usually resolved without antibiotic

treatment for healthy individuals. However, the bacteria can enter the blood stream

leading to a severe and potentially fatal illness. The invasive, life–threatening form of the

disease is more likely to occur in highly susceptible populations, including children,

elderly, and immunocompromised people (President’s Council on Food Safety, 1999).

According to CDC (1996) report, Salmonella serotype Enteritidis accounts for an

increasing proportion of all Salmonella serotypes reported to CDC's National Salmonella

Surveillance System. Report by CDC-MMWR (1996), stated that from the years 1985 to

1995, 582 Salmonella Enteritidis outbreaks occurred and accounted for 24,058 cases of

illness, 2,290 hospitalizations, and 70 deaths. There was an 8-fold increase in infections

with Salmonella Enteritidis during the years from 1976 to 1994.

Nationwide, the number of Salmonella serotype Enteritidis isolates peaked in 1995. The

rates of Salmonella Enteritidis infection reported to CDC declined by 1999. However, it

did not decline further after 2001, and outbreaks continue to occur (CDC-MMWR, 2003).

The estimated number of human Salmonella Enteritidis cases in US alone is 200,000 to 1


25

million annually, resulting in economic losses ranging from $200 million to $1 billion

annually (Morales and McDowell, 1999).

During the past two decades, eggs internally contaminated with Salmonella

Enteritidis have emerged as a leading cause of food-borne disease (CDC-MMWR, 2003).

The Food Safety Inspection Service conducted a farm-to-table assessment of risk

associated with eggs and estimated that 2.3 million of US shell eggs contain Salmonella

Enteritidis when laid (USDA-FSIS, 1998). Public health surveillance data estimated

637,000 cases of human illness per year due to the consumption of contaminated raw or

undercooked eggs. Undercooked and raw shell eggs have been identified as the most

common sources of S. Enteritidis infection upon investigations of outbreaks (CDC-

MMWR, 2003).

Earlier, contamination of shell eggs with Salmonella was believed to occur when the

organism passed from the shell into it’s inner contents (President’s Council on Food

Safety, 1999). However, transovarian S. Enteritidis contamination of shell eggs from the

S. Enteritidis-infected laying hens have further added to the problem. It has been

estimated that the rate of transovarian egg contamination has been one S. Enteritidis

positive egg in every 20,000 eggs produced in U.S. (USDA-FSIS, 1998).

Studies show that both naturally and experimentally infected chickens produce eggs

containing S. Enteritidis in their liquid interior contents (Gast and Holt, 2000a;

Humphrey et al., 1989). Freshly laid eggs are rarely reported to harbor more than a few

hundred S. Enteritidis cells (Chen et al., 2002; Gast and Holt, 2000a; Humphrey et al.,

1991). Prompt refrigeration of the eggs to the internal temperature of 7.2 C or lower can

inhibit the growth of Salmonella and reduce the chances of consumers being exposed to


26

an infective pathogen dose. However, several factors can affect the efficacy of the

refrigeration in controlling the growth of Salmonella (Gast et al., 2005). Location of

Salmonella contaminants within eggs is one of the factors. S. Enteritidis has the ability to

colonize the reproductive tract of the hen and become entrapped in the egg albumen or

yolk of the developing eggs (Gast and Beard, 1990; Gast and Holt, 2000b; Humphrey et

al., 1989; Humphrey et al., 1991). Gast and Holt (2001) reported that the deposition of S.

Enteritidis is more often on the yolk membrane than in the interior contents of the yolk.

S. Enteritidis can survive in the albumen but is inhibited from growing for an extended

period of time due to high pH and inhibitory factors in the albumen (Baron et al., 1997;

Gast and Holt, 2000b). On the contrary, yolk is a rich microbial medium with little

capability of inhibiting S. Enteritidis. Rapid growth of S. Enteritidis, especially at storage

temperature above 20 C can occur (Bradshaw et al., 1990; Braun and Fehlhaber, 1995;

Gast and Holt, 2000b; Humphrey and Whitehead, 1993).

A study by Gast and Holt (2001) showed that Salmonella spp. can penetrate through

the yolk membrane into the yolk and multiply, therefore rapid refrigeration is necessary

to prevent bacterial penetration of the yolk and multiplication. Jones et al. (2004) stored

the eggs inoculated with S. Enteritidis at room temperature for 5 weeks. The S. Enteritidis

contamination levels determined for interior egg contents increased with storage time.

The study also demonstrated that S. Enteritidis growth can take place in the yolk during

the cooling period.

Based largely on these observations, Federal and State agencies have worked with

industry and consumers to implement farm-to-table interventions to reduce the risk of

illness from S. Enteritidis in eggs. In 1996, FSIS and the United States Department of


27

Health and Human Services (HHS) Food and Drug Administration (FDA) initiated a risk

assessment for S. Enteritidis in eggs and egg products. The results indicated multiple

interventions along the farm-to-table chain were necessary to significantly reduce the risk

of illnesses from S. Enteritidis. (President’s Council on Food Safety, 1999). Later, the

council proposed 7.2 C as a mandatory temperature for transportation and storage of

eggs, to prevent the rapid multiplication of small initial levels of S. Enteritidis. Following

this, the U.S. Department of Agriculture (1998) made it mandatory to transport and store

eggs at 7.2 C or lower. However, in the commercial units, cooling is slow and require

several days to reach this specific internal egg temperature to restrict bacterial growth

(Curtis et al., 1995; Thompson et al., 2000).

So far, studies have not been done on the impact of MAP technique on the quality

and microbial growth of shell eggs. Also, few studies have been done to examine the

effect of MAP gas like CO, CO2 on the survival and growth of Salmonella spp. in shell

eggs. With the use of low temperature for shell egg storage having several limitations and

a number of studies demonstrating the benefits of modified atmosphere packaging, MAP

could be effective in maintaining the functional quality of shell eggs, U.S.D.A. quality

grade, lower the rate of lipid oxidation, reduce Salmonella risk, and extend the shelf life

of the shell eggs during distribution and storage, thus resulting in increased consumer

satisfaction and safety. An increase in the shelf life of eggs also would benefit the

producers interested in export opportunities (Keener et al., 2000).


28

CHAPTER III

METHODOLOGY

Egg Quality

Oiled fresh USDA Quality Grade AA shell eggs were obtained from a local

commercial distributor. Upon arrival at Texas Tech University Nutrition and Food

Chemistry Laboratory, the eggs were candled to ensure that the eggs used in the study

were USDA Quality Grade AA. Three trials were conducted in total. Initially the eggs for

the trial 1 were 14 days post-lay and for trial 2 and 3, five days post-lay. Approximately,

650 eggs per trial were subjected to one of the four packaging treatments: (1) control -

air; (2) 20% CO2/0.4% CO/79.6% N2 (CO2 + CO); (3) 20% CO2/80% O2 (High-ox); and

(4) 20% CO2/80% N2 (CO2).

The eggs were packaged eight to a tray (experimental unit is a tray) in Cryovac solid

barrier polypropylene trays (Cryovac, Inc.; OTR = 0.1 cc oxygen/tray/24 h at 22.7 C and

0% relative humidity; Moisture Vapor Transmission rate = 2.0 g water vapor/64,516

cm2/24 h at 37.8 C and 100% relative humidity), flushed with gas and sealed with a high-

barrier film (LID 1050, Cryovac, Inc; OTR = < 20 cc oxygen/m2/24 h at 4.4 C and 100%

relative humidity) using a G. Mondini (model CV/VG-S, Semi-Automatic 320 × 500)

tray seal machine. The lidstock was designed with an oxygen barrier layer to maintain the

desired gas mixtures and two internal abuse layers for additional protection. The lidstock

hermetically was sealed to the preformed tray during packaging and created the desired

modified atmosphere and remained fog-free during refrigerated storage. The

polypropylene tray is multilayered consisting of coextruded barrier film laminated to the

inside surface. The transparent layer provides an oxygen and moisture barrier, and allows


29

for hermetic sealing with the lidstock. Gas mixtures were achieved using a gas mixer

(model 970260, Checkmate 9900, PBI Dansensor, Glen Rock, NJ) or certified, pre-mixed

cylinders of compressed gases (ultra-high purity) that were purchased locally. The

modified atmospheres for the egg quality study were validated by testing packages,

which were not part of the trial, at the beginning and end of each packaging treatment run

using a head-space analyzer (model 333 Pac Check®; Mocon; Minneapolis, MN ). For

the egg microbiology study, modified atmospheres were validated only in the beginning.

Packaging for the trial proceeded if packages were within ± 0.5% of the targeted oxygen,

nitrogen and carbon dioxide levels. In the CO + CO2 treatment 0.4% carbon monoxide in

the packages was verified and packages contained less than 0.5% residual oxygen.

Immediately before packaging, oxygen absorbers (Ageless Mistubishi Gas Chemicals

Inc.) were added to CO2 + CO and CO2 treatments to minimize the residual oxygen in the

packages. Eggs were stored for up to 30 d in a retail case at a temperature of 6 ± 1 C

(refrigerated) or on shelves at 21 ± 1 C (abusive). Two trays per treatment per

temperature per d were prepared in each trial with a total of three trials being conducted.

Packages were opened and sampled on d 1, 7, 14, 21, and 30.

The eggs were tested for following egg quality attributes: pH of the yolk, albumen,

and whole raw eggs; color (L*, a*, b*, chroma and hue angle) of the yolk and albumen of

raw and hard cooked eggs; thiobarbituric acid reactive substances (TBARS); foam

capacity and stability; Haugh units, and yolk index (YI).

Foam Capacity and Stability

Foam capacity and stability were expressed as a percentage and were determined

according to the method of Kitabatake and Doi (1982). The egg white was separated from


30

the yolk. Fifty-milliliters of the white were whipped in a 500-mL beaker using a

homogenizer (Biomix, Metlen, Switzerland, model # EN60335) at 10,000 rpm for 1 min.

The whipped sample was poured into a 100-mL graduated cylinder. Foam stability was

determined by measuring the amount of liquid drained from the foam, at the bottom of

the graduate cylinder, after standing for 2 h at room temperature.

Foam capacity (FC) was calculated as:

FC (%) = [(initial foam volume – initial egg white volume pre-whipped)/ initial

egg white volume pre-whipped] * 100%

Foam stability (FS) was calculated as (Matringe et al., 1999):

FS (%) = (volume of drained liquid/initial foam volume) * 100%

Color Measurement

Color of the yolk and the albumen of both raw and hard cooked egg was determined

using a colorimeter (Minolta, Ramsey, NJ, model # CR-A43), and was determined at

three different locations of the yolk and the albumen. The three values were averaged to

determine an mean value for the sample. For the hard cooked yolk, color readings were

taken for both the inner portion of the yolk when cut into halves, and the outer surface of

the yolk. Values of lightness (L*), redness (a*), and yellowness (b*) were recorded. Hue

angle was calculated using the formula tan-1 (b/a) and chroma was calculated using the

formula √ (a2 + b2).

pH of Yolk, Albumen, and Whole Egg

The pH of the yolk, albumen and whole egg was measured using a pH meter

(Accumet Basic AB–15) and low maintenance pH triode. The albumen and the yolk were

separated. Approximately, 5 g of albumen and 5 g of yolk were placed in a 400-mL


31

beaker followed by addition of 45 mL of distilled water to each of the beakers. The

mixture was mixed thoroughly with a glass rod. The pH values were recorded. The pH of

the whole egg was determined after mixing the yolk and albumen of whole egg and the

same procedure was repeated as described for albumen and the yolk.

Yolk Index

Eggs were broken out on to white Styrofoam plates. The height and width of the yolk

was measured using digital calipers (Marathon Digital Calipers). The yolk index was

calculated using the method by Stadelman and Cotterill (1986), by dividing the height of

the yolk by the width of the yolk.

Haugh Unit

Haugh units (Haugh, 1937) were determined by weighing and then breaking out the

eggs on a white Styrofoam plates. A micrometer (Ames 25M – 5 micrometer) was used

to measure albumen height. Haugh unit was calculated from albumen height using the

interior quality calculator for eggs (Brant et al., 1951).

TBARS

Thiobarbituric acid reactive substances (TBARS), for the estimation of lipid

oxidation in egg yolk, was determined using the spectrophotometric (Genesys 20

thermospectronic, Loveland, CO, model # 4001/4) method by Spanier and Traylor

(1991). A standard curve of absorbance at 532 nm versus concentration was constructed

using standard of 0, 2.5, 5, 7.5, 10 (mg malonaldehyde/mL).

Data Analysis

Initially data were analyzed by ANOVA in a 2 (temperature) x 4 (packaging

treatment) x 5 (time-points) x 3 (trial) factorial design using programs in SAS (2003). No


32

trial by treatment interactions were found, thus data from all three trials were combined.

Significant interactions between storage temperatures and treatments (packaging and time

of storage) were noted for many of the variables thus data from the two storage

temperatures were analyzed separately. If no significant packaging treatment by storage

time interactions were found then main effects means were separated by Duncan’s

multiple range test. If significant packaging treatment by storage time interactions were

noted, packaging treatment means within a storage time were separated by LSMeans. A P

value of ≤ 0.05 was considered significant.

Microbiological Study

A cocktail of Salmonella Enteritidis phage 13 nalidixic acid resistant, S. Heidelburg/

3347, S. Typhimurium ATCC 14028 strains associated previously with foodborne

outbreaks was used for conducting the microbial study. The cocktail (1 ml) was passed in

9 ml of tryptic soy broth and incubated for 24 h at 37 C. The concentration of the passed

culture was found to be 1 x 108 cfu/ml. The culture was diluted using Buffered peptone

water to adjust the concentration to 1 x 104 cells. The final suspension was used to

inoculate the eggs. The inoculation of shell eggs and standardization of the inoculum size

was done according to the methodology used by (Hammack et al., 1993). Buffered

peptone water was used as the neutral non-enriching media for preparing serial dilutions

and inoculating the eggs. Before inoculation, shell eggs were sanitized according to the

procedure described by FDA (U.S. Food and Drug Administration, 2005). The eggs were

washed with stiff brush. The eggs were then soaked in a 200-ppm of Cl- solution

containing 0.1% sodium dodecyl sulfate (SDS) for 30 min. The 200 ppm of Cl- solution

was prepared by adding 8 ml of commercial bleach (5.25% Na hypochlorite) to 992 ml of


33

distilled water (FDA-CFSAN, 2005). Eggs were removed from the sanitizing solution

and allowed to air dry. A 100-ul glass syringe was sterilized with 70% ethanol, air dried

and used to inoculate the eggs. The eggs were candled to determine the position of the

yolk and marked with the pencil on outside of the shell. A small puncture hole was made

using sterilized thumb tab, on the large end of the shell egg. The inoculum size of

Salmonella cocktail suspension of 40 ul (that had been standardized in the preliminary

studies) was injected into the egg yolk. The hole in the shell was then sealed with epoxy

glue.

Sample Preparation

After the inoculation, eggs were stored at room temperature for one day (day 0). Eggs

were stored at room temperature rather than refrigerated temperature prior to packaging

so that the eggs would not sweat when packaged. Two uninoculated and two inoculated

eggs were sampled on day 0. On day 1, the inoculated eggs were subjected to one of the

four packaging treatments: (1) control - air; (2) CO2 + CO; (3) High-ox; and (4) CO2.

Eggs were packed in modified atmosphere as described previously. Immediately before

packaging, Ageless oxygen absorbers were added to CO2 + CO and CO2 treatments to

minimize the residual oxygen in the packages. Eggs were stored for up to 14 days in a

retail case at a temperature of 6 ± 1 C (refrigerated) or on shelves at 21 ± 1 C (abusive).

Before storage eggs were packaged in a set of 6 eggs per tray (1 inoculated and 5

uninoculated) and each tray was considered as an experimental unit. Two trays per

treatment per temperature per day were prepared in each trial with a total of three trials

being conducted. Packages were opened and sampled on days 1, 7, and 14.


34

After storage for the specified time period, eggs were aseptically broken into a sterile

stomacher bag, diluted to 1:10 ratio using buffered peptone water and stomached for 1

min. Serial dilutions were made using buffered peptone water and plated in duplicate

onto XLT4 plates. The XLT4 contained proteose peptone salt to enhance the formation of

black Salmonella colonies on the plate. Plating was done using spread plate method and

aseptic techniques. The plates were incubated at 37 C for 24 h. The number of colonies

on the incubated plates were counted manually.

Data were analyzed by ANOVA in a 2 (temperature) x 4 (packaging treatment) x 3

(time-points) factorial design using programs in SAS (2003). Significant trial by

treatment interactions were found thus data from the three trials were examined

separately. A significant treatment by temperature interaction was found thus data from

the two temperatures were examined separately. Because there were no packaging by

storage time interactions differences in main effects within a temperature and trial were

examined by Duncan’s multiple range test. P ≤ 0.05 was considered significantly

different.


35

CHAPTER IV

RESULTS AND DISCUSSION

Color of Raw Albumen

Hue, lightness, and chroma are the three color attributes that when combined create a

three dimensional color solid. Hue form the outer rim of the solid, with lightness as the

center axis and chroma as the horizontal spokes. Lightness describes the brightness of a

color. It separates color into bright and dark colors. In the color wheel, lightness increases

towards the top and decreases towards the bottom. Hue is the term used to classify colors

as red, yellow, blue, etc. Chroma is the color saturation or vividness. Colors are dull near

the center of the three dimensional solid and become more vivid as values move away

from the center (Minolta, 1998).

In the L* C* h* color space that uses cylindrical coordinates, L* indicates lightness,

C* is chroma and h* is hue angle. The value of C* is 0 at the center and increases

according to the distance from the center. Hue angle of 0o is red, 90o is yellow, 180o is

green and 270o is blue.

Because there was no interaction between the packaging treatment and storage time

for raw albumen color attributes, data were pooled and examined for main effects. The

lightness of raw albumen was significantly higher (p ≤ 0.05) for day 21 and day 30, at

both 6 C and 21 C. Lightness was least on day 1 for both the storage temperatures. No

difference was seen in the lightness on days 7 and 14 for both the storage temperatures

(Figure A.01, Table 4.01). Thus with increase in storage time, the lightness of the raw

albumen increased at both the temperatures.


36

Table 4.01. Raw albumen lightness, hue angle and chroma value ± SEM of eggs packed in modified atmosphere packaging CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 16/storage time)

Storage time (d)

Storage temperature Color attribute 1

7 14 21

30

Refrigerated

Lightness 77.37c ± 0.68 80.81b ± 0.34 81.91b ± 0.50 84.39a ± 0.35 84.86a ± 0.21

Hue angle (o) 106.58c ± 0.26 109.71b ± 0.59 110.57b ± 0.59 110.16b ± 0.60 114.99a ± 0.60

Chroma 7.39a ± 0.54 5.21b ± 0.41 4.99b ± 0.19 2.81c ± 0.21 2.68c ± 0.16

Abusive

Lightness 77.69d ± 0.38 80.49c ± 0.40 81.78b ± 0.46 85.24a ± 0.28 85.19a ± 0.17

Hue angle (o) 106.86c ± 0.33 109.79b ± 0.49 111.51b ± 0.88 110.43b ± 0.78 114.65a ± 1.18

Chroma 7.90a ± 0.51 5.48b ± 0.30 4.96bc ± 0.44 2.37d ± 0.15 4.06c ± 0.45

a, b,c,dMeans with the same letter within a row were not significantly different (P ≥ 0.05).


37

Table 4.02. Raw albumen lightness, hue angle and chroma ± SEM of eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2) or air stored at refrigerated (6 C) and abusive (21 C) temperature (n = 20/ packaging treatment)

Packaging treatment

Storage temperature

Color attribute CO2 + CO CO2 High-ox Air

Refrigerated

Lightness 81.22a ± 0.67 81.80a ± 0.82 82.16a ± 0.75 82.29a ± 0.67

Hue angle (o) 109.70a ± 0.87 110.25a ± 0.71 110.86a ± 0.81 110.80a ± 0.70

Chroma 5.21a ± 0.53 4.46a ± 0.53 4.24a ± 0.45 4.55a ± 0.45

Abusive

Lightness 81.92a ± 0.78 81.76a ± 0.79 82.34a ± 0.70 82.30a ± 0.63

Hue angle (o) 111.16a ± 1.25 110.39a ± 0.86 110.37a ± 0.66 110.68a ± 0.76

Chroma 4.99a ± 0.68 5.13a ± 0.58 4.75a ± 0.45 4.93a ± 0.42

aMeans with the same letter within a row were not significantly different (P ≥ 0.05).


38

The mean hue angle of the raw albumen, was significantly higher (more green; p ≤

0.05) at day 30 for both refrigerated and abused temperatures, compared to that of day 1,

7, 14, or 21. On day 1, hue angle was significantly lower (p ≤ 0.05) than the rest of the

time periods at both temperature 6 and 21 C (Figure A.02, Table 4.01). The hue angle

increased with storage time at both the temperatures. The hue angle value ranged from

106 to 115. Hence, the color varied from yellow to light green.

The mean chroma value of 7.9 was highest on day 1 at temperature 21 C and

decreased with storage time. A similar trend was seen for the chroma value at 6 C (Figure

A.03, Table 4.01).

No significant difference (p > 0.05) was found in the lightness, hue angle and chroma

value of the raw albumen of the eggs packed in the four different packaging and stored at

both 6 C and 21 C (Table 4.02).

Color of Raw Yolk

The lightness value of the raw yolk was not significantly different (p > 0.05) for the

four packaging treatments, on day 1, and 7 at 6 C. On day 14, the lightness of raw yolk

for 20%CO2/80%N2, and high-ox treatment were significantly higher than the control (p

≤ 0.05) but not significantly different from the CO2+ CO treatment at 6 C (p > 0.05). On

day 21, the CO2+ CO, and CO2 treatments maintained significantly higher lightness value

of the raw yolk than the high-ox and control (p ≤ 0.05). On day 30, lightness value of raw

yolk for the CO2+ CO, high-ox and control were not significantly different (p > 0.05) but

the high-ox treatment and the control were significantly different (p ≤ 0.05) from the CO2

treatment, at 6 C (Figure A.04, Table 4.03).


39

Table 4.03. Raw yolk lightness ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/ storage time/packaging treatment)

Storage time (d) Packaging treatments

CO2 + CO CO2 High-ox Air

1 59.19a ± 0.33 58.82a ± 0.33 58.29a ± 0.45 59.05a ± 0.29

7 57.90a ± 0.59 58.56a ± 0.67 58.83a ± 1.08 59.21a ± 0.50

14 58.88ab ± 0.53 59.10a ± 0.32 59.37a ± 0.62 57.62b ± 0.47

21 58.99a ± 0.27 58.99a ± 0.69 57.46b ± 0.51 56.47b ± 0.56

30 57.26ab ± 0.39 58.37a ± 0.28 56.59b ± 0.30 56.59b ± 0.33

a, bLSMeans with the same letter within a row were not significantly different (P ≥ 0.05).


40

Table 4.04. Raw yolk lightness ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature. (n = 4/storage time/packaging treatments)



1 59.67a ± 0.45 58.81a ± 0.24 59.01a ± 0.37 59.85a ± 0.14

7 59.43b ± 0.66 59.27b ± 0.74 59.48b ± 0.45 61.89a ± 0.35

14 60.71b ± 1.18 60.68b ± 0.62 60.93b ± 0.87 63.60a ± 0.67

21 60.12b ± 0.71 60.20b ± 0.35 60.63b ± 0.66 63.05a ± 1.00

30 58.42b ± 0.77 58.94b ± 0.25 58.64b ± 0.96 63.11a ± 1.16



41

Table 4.05. Raw yolk hue angle (o) and chroma ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature. (n = 16/storage time)

Storage time (d)

Storage temperature

Color attribute 1

7 14 21

30

Refrigerated

Hue angle (o) 91.05a ± 0.25 90.38b ± 0.20 90.18bc ± 0.16 89.71cd ± 0.17 89.48d ± 0.20

Chroma 45.29b ± 0.59 45.96b ± 0.71 48.79a ± 0.60 46.55b ± 0.47 45.47b ± 0.40

Abusive

Hue angle (o) 90.88a ± 0.19 90.18b ± 0.19 90.13b ± 0.25 89.63b ± 0.23 88.70c ± 0.26

a, b,c,dMeans with the same letter within a row were not significantly different (P ≥ 0.05).


42

Table 4.06. Raw yolk hue angle (o) and chroma ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 20/packaging treatment)

Packaging treatments

Storage temperature

Color attribute CO2 + CO CO2 High-ox Air

Refrigerated

Hue angle (o) 46.77a ± 0.21 46.11a ± 0.13 46.10a ± 0.18 46.67a ± 0.30

Chroma 89.92a ± 0.68 90.20a ± 0.60 90.20a ± 0.54 90.32a ± 0.47

Abusive

Hue angle (o) 89.86a ± 0.26 90.14a ± 0.22 89.77a ± 0.23 89.87 ± 0.31

aMeans with the same letter within a row were not significantly different (P ≥ 0.05).


43

Table 4.07. Raw yolk chroma ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature. (n = 4/storage time/packaging treatment)



1 46.99a ± 0.96 44.84a ± 1.00 46.53a ± 1.13 45.14a ± 0.88

7 48.56a ± 0.23 48.39a ± 0.69 47.81a ± 1.06 49.91a ± 2.46

14 50.58b ± 2.94 49.74b ± 1.21 50.46b ± 0.60 56.42a ± 1.41

21 47.72b ± 1.13 47.77b ± 1.61 47.86b ± 1.60 56.89a ± 1.44

30 50.98a ± 0.97 50.69a ± 1.49 51.97a ± 1.49 53.40a ± 2.68



44

At 21 C, no significant difference was found in the lightness value of raw yolk on day

1, among the four packaging treatments (p > 0.05). On day 7, 14, 21, and 30, the three

MAP treatments maintained significantly lower lightness value of the raw yolk than the

control (p ≤ 0.05), at 21 C (Figure A.05, Table 4.04).

The hue angle of the yolk of raw egg, declined significantly (p ≤ 0.05) from day 1 to

day 30 at both 6 C and 21 C. Day 1 had the highest hue angle value while the day 30 had

the lowest significant (p ≤ 0.05) value at both the storage temperatures (Figure A.06,

Table 4.05). Thus, during storage the hue changed from yellow to somewhere between

reddish yellow. No significant difference (p > 0.05) in the hue angle of raw yolk was

observed among the four packaging treatments and at both 6 C and 21 C (Table 4.06).

No significant difference (p > 0.05) was found in the chroma value of the yolk of raw

egg at 21 C for day 1, 7, and 30, among the four packaging treatments. On day 14, and

21, chroma value was significantly lower (p ≤ 0.05) for the three MAP treatments than

the control at 21 C (Figure A.08, Table 4.07). At 6 C, day 14 had the significantly higher

(p ≤ 0.05) chroma value of the raw egg yolk compared to the other storage time periods.

No significant difference (p > 0.05) was found among the chroma values of day 1, 7, 21,

or 30 at 6 C (Table 4.05). Also, no significant difference (p > 0.05) was found in the

chroma value of raw yolk for the four packaging treatments at 6 C (Figure A.07, Table

4.06).

Color of Hard-cooked Albumen

The lightness of the hard cooked albumen, at both 6 C and 21 C, was significantly

higher for the three MAP treatments than the control, from day 1 to day 30 (p ≤ 0.05). No


45

significant difference (p > 0.05) was found in the lightness value of the hard cooked

albumen among the three MAP treatments (Figure A.09 and 4.10, Table 4.08 and 4.09).


46

Table 4.08. Hard cooked albumen lightness ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/packaging treatment/storage time)



1 95.91a ± 0.21 95.22a ± 0.30 95.70a ± 0.13 92.22b ± 0.33

7 95.87a ± 0.09 95.77a ± 0.23 95.72a ± 0.13 92.55b ± 0.21

14 95.61a ± 0.21 95.59a ± 0.18 95.61a ± 0.21 92.05b ± 0.20

21 95.38a ± 0.09 95.78a ± 0.21 95.73a ± 0.12 92.52b ± 0.52

30 95.53a ± 0.08 95.22a ± 0.15 95.20a ± 0.34 92.36b ± 0.35

a, b,cLSMeans with the same letter within a row were not significantly different (P ≥ 0.05).


47

Table 4.09. Hard cooked albumen lightness ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature. (n = 4/packaging treatment/storage time)



1 95.14a ± 0.05 95.39a ± 0.23 95.50a ± 0.20 91.54b ± 0.49

7 95.27a ± 0.19 95.51a ± 0.06 95.62a ± 0.09 90.85b ± 0.45

14 95.04a ± 0.48 95.22a ± 0.14 95.66a ± 0.18 90.89b ± 0.19

21 94.98a ± 0.12 95.38a ± 0.25 95.47a ± 0.08 90.64b ± 1.09

30 91.55b ± 3.24 94.46a ± 0.13 94.56a ± 0.24 90.67b ± 0.15



48

Table 4.10. Hard cooked albumen hue angle (o) ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature. (n = 4/packaging treatment/storage time)



1 114.15b ± 0.54 114.30b ± 0.44 114.30b ± 0.71 124.15a ± 1.58

7 113.16b ± 0.42 112.27b ± 1.37 113.44b ± 0.77 121.83a ± 0.58

14 113.12b ± 1.69 113.75b ± 0.53 111.83b ± 0.36 124.12a ± 0.94

21 111.75b ± 0.26 112.86b ± 0.62 111.63b ± 0.79 120.61a ± 2.79

30 114.25b ± 0.22 115.53b ± 0.79 115.87b ± 0.94 124.42a ± 0.64



49

Table 4.11. Hard cooked albumen hue angle (o) ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature. (n = 4/packaging treatment/storage time)



1 113.56b ± 0.51 114.15b ± 0.47 113.13b ± 0.88 128.52a ± 0.90

7 113.69b ± 0.25 113.44b ± 0.70 113.34b ± 0.46 129.85a ± 3.00

14 111.89b ± 0.98 112.82b ± 1.35 113.98b ± 0.46 125.96a ± 0.38

21 111.75b ± 0.71 112.56b ± 0.29 111.95b ± 0.38 129.56a ± 3.07

30 111.76b ± 2.17 113.99b ± 1.34 113.09b ± 1.66 125.30a ± 1.28



50

Table 4.12. Hard cooked albumen chroma ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature. (n = 4/packaging treatment/storage time)



1 12.28a ± 0.19 11.69a ± 0.09 11.41ab ± 0.34 10.53b ± 0.31

7 11.88a ± 0.44 11.51a ± 0.16 11.79a ± 0.42 10.20b ± 0.35

14 11.76a ± 0.34 11.91a ± 0.34 12.41a ± 0.44 9.70b ± 0.49

21 12.42a ± 0.28 11.80a ± 0.29 12.62a ± 0.41 10.30b ± 0.64

30 12.41a ± 0.24 12.08a ± 0.52 12.06a ± 0.26 10.98b ± 0.21



51

Table 4.13. Hard cooked albumen chroma ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature.

Storage time (d)

Chroma

1 11.28b ± 0.25

7 11.35b ± 0.23

14 12.14a ± 0.29

21 12.04a ± 0.26

30 12.11a ± 0.25

a, bMeans with the same letter within a column were not significantly different (P ≥ 0.05).


52

Table 4.14. Hard cooked albumen chroma ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 30/packaging treatment).

Packaging treatments Chroma

CO2 + CO 11.69a ± 0.23

CO2 12.18a ± 0.22

High-ox 12.20a ± 0.20

Air 11.06b ± 0.23



53

The three MAP treatments maintained significantly lower hue angle of the hard

cooked albumen, at both refrigerated and abusive temperature, than the control, from day

1 till day 30 (p ≤ 0.05). No significant difference was found in the hue angle values

among the three MAP treatments (Figure A.12 and 4.11, Table 4.10 and 4.11). The hue

angle of the air packed egg albumen was in the range of 120 to 124.5 and was more

greenish yellow than that of egg albumen packed in the three MAP. On day 1, the (CO2+

CO, and CO2 treatments maintained significantly higher chroma of the hard cooked

albumen, than the control, at 6 C (p ≤ 0.05). No significant differences were found among

the three MAP treatments. No significant difference was found between the high-ox

treatment and the control (p > 0.05). From day 7 to day 30, the three MAP treatments

maintained significantly higher chroma value (p ≤ 0.05) than the control at 6 C (Figure

A.13, Table 4.12). The hard cooked albumen chroma value was significantly lower (p ≤

0.05) for the control as compare to the three MAP treatments at 21 C (Figure A.15, Table

4.14). Thus, the color of the hard cooked albumen of shell eggs packed in control was

duller or less vivid than that of MAP treated eggs. At 21 C, day1 and 7 had significantly

lower chroma value of the hard cooked albumen compared to day 14, 21, 30 (p ≤ 0.05).

No significant difference (p > 0.05) was observed in the chroma values of day 14, 21, and

30 at 21 C (Figure A.14, Table 4.13).

A study by Sharp and Stewart (1931), showed that the eggs stored in carbon dioxide

had a whiter albumen than eggs stored under normal conditions. Carbon dioxide very

markedly retarded the development of the yellow, orange, or pink condition of the white.


54

Color of Hard-cooked Yolk Surface

The MAP treatments maintained significantly higher lightness value of the surface of

hard cooked yolks (p ≤ 0.05), compared to the control, from day 1 to day 30 at 21 C

(Figure A.17, Table 4.16). The lightness of hard cooked yolk surface was significantly

lower (p ≤ 0.05) for the control as compared to rest of the MAP treatments, at 6 C, from

day 1 to day 30. On day 1, within the three MAP treatments, high-ox treatment had

significantly higher lightness value than CO2+ CO treatment (p ≤ 0.05). The CO2

treatment was not significantly different from the CO2+ CO and high-ox treatments for

day 1, at temperature 6 C (p > 0.05). On the remaining storage time points of day7, 14,

21, and 30, no significant difference in the lightness of the hard cooked yolk surface was

observed among the three MAP treatments, at 6 C (Figure A.16, Table 4.15).

The hue angle of the hard cooked yolk surface at temperature 6 C for day 1 was not

significantly different for CO2+ CO, and CO2, and air packed eggs (p > 0.05). The control

was, however, significantly different from the high-ox packaging (p ≤ 0.05). For the rest

of the storage time from day 7 till day 30, MAP treatments maintained significantly

higher hue angle (p ≤ 0.05) of the hard cooked yolk surface (Figure A.18, Table 4.17).

The hue angle of the hard cooked yolk surface at 21 C for day 1 was not significantly

different for control, CO2+ CO, and high-ox treatments but was different for control and

CO2 treated eggs. No significant difference was observed in hue angle of hard cooked

yolk surface among the four packaging treatments for day 14 and 30 (p > 0.05). On day 7,

at 21 C, three MAP treatments maintained lower hue angle than the control (p ≤ 0.05).

On day 21, no significant difference was observed in the hue angle value among the

control, CO2, and high-ox treatments. The hue angle of CO2+ CO treated eggs was not


55

different (p > 0.05) from the other two MAPs but was significantly higher (p ≤ 0.05) than

the control at 21 C (Figure A.19, Table 4.18).

All the three MAP treatments maintained significantly higher chroma value (p ≤ 0.05)

of the outer surface of the hard cooked yolk, as compared to the control, for all the

storage time points from day 1 to day 30, at 21 C (Figure A.21, Table 4.20). A similar

trend was seen for the storage temperature 6 C (Figure A.18, Table 4.17).

Thus, the three MAP treatments maintained a more vivid and lighter yellow color of

the surface of hard cooked egg yolk than the control eggs, throughout the storage time

from day 1 to day 30 and at both refrigerated and abused temperatures.

One of the major problem when eggs are hard cooked is that of a greenish-black

discoloration on the surface of the yolk of hard-cooked eggs. This greenish-black

discoloration on the surface of the egg yolk is due to the formation of FeS (Baker et al.,

1967). Albumen is the source of H2S. According to Fruton and Simmonds (1958) H2S

can be released from L – cystine by the enzyme cystine desulfhydrase. It was postulated

that during heating the outward pressure of gases tend to prevent the reaction between

H2S from the albumen and Fe from the yolk. However, during cooling, the egg content

contracts and the H2S moves towards the yolk and comes in contact with it’s surface.

Also, the study shows that Fe is released from the yolk upon heating. Studies show that

increased alkalinity in the egg yolk favors FeS formation. The pH of the yolk has a very

definite effect upon the blackening at the interface of the yolk and the albumen. The

length of storage time before cooking also has marked effect on the yolk color.


56

Table 4.15. Hard cooked yolk surface lightness ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature. (n = 4/packaging treatment/storage time)



1 87.29b ± 3.27 90.16ab ± 0.57 91.48a ± 0.27 81.99c ± 1.04

7 90.58a ± 0.18 90.72a ± 0.02 91.40a ± 0.24 81.87b ± 2.32

14 91.44a ± 1.61 90.74a ± 0.42 90.50a ± 0.22 83.35b ± 0.12

21 90.88a ± 0.39 90.99a ± 0.08 91.12a ± 0.46 83.73b ± 0.74

30 90.67a ± 0.31 89.99a ± 0.72 89.73a ± 0.76 80.65b ± 0.58



57

Table 4.16. Hard cooked yolk surface lightness ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/packaging treatment/storage time)



1 90.14a ± 0.50 91.19a ± 0.35 91.03a ± 0.12 81.64b ± 1.84

7 90.38a ± 0.45 90.54a ± 0.55 91.74a ± 0.13 75.49b ± 0.89

14 90.53a ± 0.74 90.54a ± 0.32 91.20a ± 0.46 75.52b ± 0.11

21 90.38a ± 0.13 90.67a ± 0.25 91.16a ± 0.20 74.92b ± 0.38

30 88.71a ± 0.27 88.35a ± 0.17 90.08a ± 0.51 79.26b ± 1.87



58

Table 4.17. Hard cooked yolk surface hue angle (o) ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature. (n = 4/packaging treatment/storage time)



1 93.01ab ± 0.25 92.75ab ± 0.07 93.27a ± 0.10 92.17b ± 0.75

7 92.52a ± 0.30 92.43a ± 0.08 92.59a ± 0.15 91.57b ± 0.44

14 92.73a ± 0.40 92.82a ± 0.08 92.85a ± 0.15 91.09b ± 0.10

21 92.75a ± 0.24 92.40ab ± 0.31 92.83a ± 0.41 91.71b ± 0.40

30 93.27a ± 0.20 92.86a ± 0.50 92.86a ± 0.08 91.31b ± 0.18



59

Table 4.18. Hard cooked yolk surface hue angle (o) ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/packaging treatment/storage time)



1 92.66ab ± 0.21 93.20a ± 0.27 92.87ab ± 0.16 91.93b ± 0.32

7 92.62b ± 0.15 92.64b ± 0.26 93.06b ± 0.09 95.02a ± 0.75

14 93.20a ± 0.74 93.37a ± 0.27 92.94a ± 0.40 93.20a ± 0.32

21 92.97a ± 0.57 92.48ab ± 0.30 92.63ab ± 0.30 91.82b ± 0.26

30 92.42a ± 0.12 92.39a ± 0.57 92.40a ± 0.57 92.69a ± 0.29



60

Table 4.19. Hard cooked yolk surface chroma ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/packaging treatment/storage time)



1 46.57a ± 1.16 45.98a ± 1.32 47.01a ± 0.88 37.93b ± 1.73

7 47.88a ± 0.68 49.48a ± 0.67 49.12a ± 0.70 36.91b ± 2.87

14 48.71a ± 1.69 47.51a ± 1.08 48.36a ± 0.62 41.79b ± 0.60

21 46.43a ± 2.11 49.25a ± 0.86 46.86a ± 2.12 42.58b ± 0.56

30 47.47a ± 0.75 46.93a ± 0.81 47.45a ± 1.84 40.82b ± 1.04



61

Table 4.20. Hard cooked yolk surface chroma ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/packaging treatment/storage time)



1 45.66a ± 1.41 45.00a ± 1.03 48.50a ± 0.60 37.41b ± 1.92

7 48.24a ± 1.52 47.13a ± 0.85 47.02a ± 0.39 26.18b ± 0.87

14 47.94a ± 1.16 46.85a ± 0.38 45.97a ± 1.62 29.40b ± 1.39

21 47.49a ± 1.08 47.58a ± 1.03 47.96a ± 1.39 34.81b ± 2.18

30 49.15a ± 0.35 50.28a ± 0.72 47.18a ± 2.64 37.93b ± 1.94



62

The longer the eggs are stored, the darker the yolks become after cooking. This is

because during the storage, the pH of the yolk increases due to the loss of carbon dioxide.

Unlike the pH of the yolk of fresh egg which is 6.0, the average pH of the yolks of eggs

stored for 3 weeks at 2 C was 6.9 (Baker et al., 1967).

Thus the three MAP treatments containing 20% CO2 maintained the lower pH of the

yolk and the albumen and thus prevented the green ring formation due to formation of

FeS.

Color of Inner Part of Hard-cooked Yolk

No significant difference was observed in the lightness, hue angle and chroma value

of the inner part of the hard cooked yolk among the eggs subjected to four packaging

treatments at 21 C (p > 0.05). Similarly, no significant difference (p > 0.05) was seen in

the lightness and chroma value among the eggs subjected to four packaging treatments at

6 C (Table 4.22). The hue angle of 92.7, on day 30 was significantly higher (p ≤ 0.05)

than that of day 7 and 14 but not significantly different (p > 0.05) from day 1 and day 21

(Figure A.22, Table 4.21).

Peeling Property of Hard-cooked Shell Eggs

During the peeling of the hard cooked shell eggs for taking down the color readings,

it was observed that the peeling was harder and took more time for the three MAP treated

eggs than the air packed eggs at both 6 C and 21 C. However, no scale was designed to

score the peelability. A possible explanation for the above observation could be that the

MAP treatment lowered the pH of the shell eggs making it difficult to peel. A study by

Reinke and Spencer (1964), found that an albumen pH of 8.7 to 8.9 or higher resulted in

easy-to-peel hard cooked eggs. The shell membranes of easy-to-peel eggs were more


63

Table 4.21. Lightness, hue angle (o) and chroma value of inner part of hard cooked ± SEM egg yolk packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2(20%CO2/80%N2), High-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 16/storage time)

Storage time (d)

Storage temperature

Color attribute 1

7 14 21

30

Refrigerated

Lightness 86.41a ± 0.71 87.31a ± 0.34 87.20a ± 0.28 86.66a ± 0.36 86.45a ± 0.41

Hue angle (o) 92.46ab ± 0.12 91.93b ± 0.25 92.02b ± 0.17 92.15ab ± 0.20 92.70a ± 0.19

Chroma 48.01a ± 0.60 48.93a ± 0.74 50.39a ± 0.56 49.54a ± 0.52 49.01a ± 0.88

Abusive

Lightness 86.21a ± 0.30 87.71a ± 0.26 87.30a ± 0.36 86.61a ± 0.48 86.49a ± 0.48

Hue angle (o) 92.71a ± 0.22 92.20a ± 0.25 92.35a ± 0.19 92.39a ± 0.30 92.74a ± 0.18

Chroma 48.37a ± 0.57 49.50a ± 0.51 49.62a ± 0.68 48.51a ± 0.74 48.52a ± 0.40

a, bMeans with the same letter within a row were not significantly different (P ≥ 0.05).


64

Table 4.22. Lightness, hue angle (o) and chroma value of inner part of hard cooked ± SEM eggs yolk packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 20/packaging treatment)


Storage temperature Color attribute CO2 + CO CO2 High-ox Air

Refrigerated

Lightness 76.17a ± 0.61 76.17a ± 0.28 75.33a ± 0.25 76.17a ± 0.33

Hue angle (o) 75.75a ± 0.15 75.75a ± 0.19 76.08a ± 0.18 75.75a ± 0.21

Chroma 74.58a ± 0.64 74.58a ± 0.77 72.42a ± 0.49 74.58a ± 0.55

Abusive

Lightness 86.80a ± 0.30 86.49a ± 0.41 86.93a ± 0.37 87.23a ± 0.36

Hue angle (o) 92.59a ± 0.18 92.51a ± 0.26 92.41a ± 0.19 92.39a ± 0.20

Chroma 48.63a ± 0.44 49.29a ± 0.63 48.77a ± 0.55 48.92a ± 0.52



65

compact than that of hard-to-peel eggs. According to Swanson (1959), freshly laid eggs

were difficult to peel because of the lower pH. Hard et al. (1963) found that the eggs

treated with carbon dioxide and oil took more time to peel than the untreated ones. A

possible reason for this could be that oiling and CO2, reduced the pH of the eggs. Britton

and Hale (1975) concluded that albumen pH was the major factor to yolk discoloration

and ease of peeling.

Foam Capacity

The foam capacity of the control eggs was significantly higher (P ≤ 0.05) than that of

CO2+ CO and high-ox treated eggs at 21 C (Figure A.23, Table 4.23).

Temperature and pH are two important factors that affect the foaming properties (Liang

and Kristinsson, 2005). Elevation of temperature results in decreased surface tension.

Thus, greater volume of foam and easier the foaming at room temperature as compared to

the refrigerated one. At lower pH, the whipping time for the white increases. The MAP

treated eggs with 20% carbon dioxide lowered the pH of the shell eggs and thus reduced

the foaming capacity. No significant difference (P > 0.05) in the foam capacity was

observed among the storage time from day 1 to day 30 for refrigerated and abused

temperatures (Table 4.24). With increase in storage time, the pH of egg increases and

thus the foaming capacity should increase. However, this was not the case in the above

observations probably because MAP lowered the pH of the eggs. At 6 C, no significant

difference was observed in the foaming capacity of the eggs of the three MAP treatments

(P > 0.05). The control had significantly higher foaming capacity value than the CO2+

CO, and CO2 treatments but was not different from high-ox treatment (P ≤ 0.05), at 6 C

(Figure A.23, Table 4.23). The high-ox treated eggs had similar foaming capacity as that


66

of the control eggs even though high-ox contained 20% CO2, same as the other two MAP

treatments. This difference could be possibly explained by leakage of gas from the

package or some packaging defect.

Foam Stability

No significant difference (P > 0.05) was observed in the foam stability of the four

packaging treatments from day 1 to 30 at 21 C (Figure A.24, Table 4.25). At 6 C, no

significant difference (P > 0.05)was observed in the foam stability of shell eggs packed in

the four types of packaging and stored from day 1 to day 30 (Table 4.26 and 4. 27).

The heat stability of specific proteins is influenced by the pH and increases when pH

is as low as 6.5. At an albumen pH of 7, ovalbumin, ovomucoid and lysozyme are

protected against heat damage. At lower pH, the foam stability greatly improves. Foam

stability, however, is affected little by a change in temperature from 20 C to 34 C (Liang

and Kristinsson, 2005). However, non uniformity of this trend in the above observations

could not be explained.


67

Table 4.23. Foam capacity ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 30/packaging treatment)

Packaging treatments Storage temperature

Refrigerated Abusive

CO2 + CO 72.78b ± 3.06 76.03b ± 2.34

CO2 75.11b ± 2.30 77.83ab ± 2.18

High-ox 76.42ab ± 2.42 74.42b ± 2.28

Air 82.39a ± 1.38 82.75a ± 2.00



68

Table 4.24. Foam capacity ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 24/storage time)

Storage Time Storage temperature

(d) Refrigerated Abusive

1 75.03a ± 2.01 74.76a ± 2.54

7 75.97a ± 2.01 77.46a ± 2.05

14 72.15a ± 3.75 80.10a ± 2.70

21 80.52a ± 2.78 80.14a ± 2.56

30 79.69a ± 2.46 76.32a ± 2.71



69

Table 4.25. Foam stability ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6/storage time/packaging treatment)



1 86.94b ± 2.40 90.10b ± 1.28 88.33b ± 1.08 92.50b ± 0.81

7 90.97a ± 0.79 90.28a ± 1.07 91.67a ± 0.78 92.91a ± 0.85

14 90.69a ± 1.24 89.86a ± 0.33 89.58a ± 0.80 90.97a ± 0.62

21 93.75a ± 0.77 93.61a ± 0.28 92.08a ± 0.85 92.50a ± 1.39

30 89.17a ± 0.57 91.11a ± 1.05 90.14a ± 0.87 91.39a ± 1.11



70

Table 4.26. Foam stability and albumen pH ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) temperature (n = 30/packaging treatment).

Packaging treatments Quality attribute

Foam stability Albumen pH

CO2 + CO 91.39a ± 1.53 7.87c ± 0.05

CO2 90.33a ± 0.43 7.95bc ± 0.04

High-ox 89.80a ± 0.70 7.99b ± 0.05

Air 91.67a ± 1.01 9.19a ± 0.03

a, b,cMeans with the same letter within a column were not significantly different (P ≥

0.05).


71

Table 4.27. Foam stability and albumen pH ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 24/storage time).

Storage time Quality attribute

(d) Foam stability Albumen pH

1 89.13a ± 0.52 8.37a ± 0.13

7 91.70a ± 0.45 8.17b ± 0.13

14 91.08a ± 0.53 8.12b ± 0.12

21 90.38a ± 1.40 8.20b ± 0.12

30 91.70a ± 1.88 8.40a ± 0.11



72

Albumen pH

According to Heath (1977), Sharp and Powell (1931), the pH of albumen of a freshly

laid egg is between 7.6 and 8.5. The albumen pH increases to about 9.7 during storage.

The mean pH of albumen for day 0 was 8.9. The value was closer to the given study. At 6

C, the three MAP treatments maintained pH of the albumen in the range of 7.87 to 7.99,

while the air packed eggs had albumen pH of about 9.19 during storage. The pH of the

albumen, at 6 C was significantly higher for the control eggs as compared to the three

MAP packed ones (P ≤ 0.05). No significant difference in albumen pH between the CO2+

CO, and CO2, and between CO2 and high-ox treatment was observed (P > 0.05).

However, CO2+ CO treatment was significantly different from high-ox treatment (P ≤

0.05) in the albumen pH at 6 C (Figure A.26, Table 4.26). Thus MAP maintained the

albumen pH close to that of the fresh egg at refrigerated temperature while the pH of

control eggs increased resulting in poor quality. No significant difference (P > 0.05) in

the albumen pH between day 1 and 30 was observed, however, were significantly

different (P ≤ 0.05) from Day 7, 14, and 21 at 6 C (Figure A.25, Table 4.27).

At 21 C, MAP maintained pH of albumen in the range of 7.72 to 8.52, ie; close to that

of fresh eggs, while the albumen pH of control eggs was about 9.38. The rise in the

albumen pH of the control eggs during storage is due to loss of carbon dioxide from the

eggs through the pores in the shell. The drop in the albumen pH of the MAP treated eggs

was due to increased concentration of carbon dioxide in the environment (20%) resulting

in the higher concentration of bicarbonates in the shell eggs. At 0.03% CO2 (air) in the

environment, the pH of albumen was found to be 9.61, at 10% CO2 pH was 7.5, and at

97% CO2 albumen pH was 6.55 (Brooks and Pace, 1938). In the study by Sharp and


73

Powell (1931), the pH of albumen was 9.18 after 3 days of storage at 3 C and 9.4 after 21

days of storage irrespective of the storage temperature between 3 C and 35 C. The three

MAP treatments maintained significantly lower albumen pH (P ≤ 0.05) at 21 C for every

storage time point from day 1 to day 30, as compared to the control (Figure A.27, Table

4.28).


74

Table 4.28. Albumen pH of ± SEM shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6/packaging treatment/storage time)



1 8.52b ± 0.10 8.46b ± 0.07 8.33b ± 0.16 9.42a ± 0.04

7 8.11b ± 0.18 8.11b ± 0.08 8.13b ± 0.12 9.44a ± 0.05

14 8.04b ± 0.07 7.96b ± 0.05 7.49c ± 0.33 9.59a ± 0.03

21 7.76b ± 0.11 7.94b ± 0.07 7.72b ± 0.08 9.38a ± 0.02

30 8.30b ± 0.11 8.22b ± 0.05 8.12b ± 0.06 9.44a ± 0.02



75

Whole Egg pH

The pH of the whole egg of the air packed eggs was significantly higher than all the

three MAP treatments, at both 6 and 21 C (P ≤ 0.05). No significant difference (P > 0.05)

was found in the pH of the whole eggs among the three MAP treatments at both

refrigerated and abused temperatures (Figure A.28, Table 4.29). At 6 C, no significant

difference was observed among days 1, 7, 30 of the storage time (P ≤ 0.05). pH was

significantly lower for day 21 (Figure A.29, Table 4.30). The pH of the whole egg for day

1, 7, 14 was not significantly different (P > 0.05). The pH of 7.7 was highest for day 30 at

6 C. The mean pH of the whole egg for day 0 was 8.4. However, at 6 C and 21 C, MAP

maintained a whole egg pH of approximately 7.4. The control eggs had a whole egg pH

of 8.1. The lowering of the pH of MAP treated eggs could be explained by increase in

amount of dissolved CO2 in the egg which is dependent on the partial pressure of CO2 in

the external environment of the eggs (Brooks and Pace, 1938). The MAP treated eggs had

20 % CO2 in there external environment.

Yolk pH

At 6 C, pH of the yolk of the control was 6.32, significantly higher than the other three

MAP treatments (P ≤ 0.05). The three MAP treatments maintained pH in the range of

6.19 to 6.22. Thus, MAP maintained yolk pH values closer to that of fresh eggs. No

significant difference (P > 0.05) in the yolk pH of the four packaging treatments was

observed at 21 C (Figure A.30, Table 4.31). The pH of the yolk does not increase much

with time and may vary from 6.0 in a freshly laid egg to between 6.4 and 6.9, during

storage (Sharp and Powell, 1931). A possible reason for significant difference in the yolk


76

Table 4.29. Whole egg pH value ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 30/packaging treatment)



CO2 + CO 7.4b ± 0.04 7.4b ± 0.04

CO2 7.4b ± 0.05 7.4b ± 0.06

High-ox 7.4b ± 0.04 7.4b ± 0.05

Air 8.2a ± 0.06 8.1a ± 0.05



77

Table 4.30. Whole egg pH ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 24/storage time)

Storage time Storage temperature


1 7.7ab ± 0.1 7.6bc ± 0.1

7 7.7ab ± 0.1 7.7ab ± 0.1

14 7.5bc ± 0.1 7.6ab ± 0.1

21 7.5c ± 0.1 7.4c ± 0.1

30 7.7a ± 0.1 7.7a ± 0.1

a, b, cMeans with the same letter within a column were not significantly different (P ≥

0.05).


78

Table 4.31. Egg yolk pH ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 30/packaging treatment)



CO2 + CO 6.19b ± 0.05 6.27a ± 0.08

CO2 6.19b ± 0.03 6.25a ± 0.06

High-ox 6.22b ± 0.05 6.28a ± 0.09

Air 6.32a ± 0.04 6.39a ± 0.09



79

Table 4.32. Egg yolk pH ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 24/storage time)

Storage time Storage temperature


1 6.39a ± 0.05 6.49a ± 0.13

7 6.11b ± 0.03 6.16b ± 0.04

14 6.35a ± 0.05 6.36ab ± 0.08

21 6.12b ± 0.04 6.12b ± 0.06

30 6.19b ± 0.04 6.36ab ± 0.11



80

pH of the MAP treatments and the control at 6 C but not at 21 C could be that at

lower temperature lesser amount of CO2 is required to lower the pH. However, the

percentage of CO2 was same ie; 20% at both the storage temperatures. Thus, more CO2

was absorbed in the MAP eggs at 6 C than at 21 C. The yolk pH for day 1 and 14 was

significantly different and was significantly higher than the pH on day 7, 21, and 30 (P ≤

0.05). At 21 C, no significance difference (P > 0.05) was observed in the yolk pH at day

1 and 30 (Figure A.31, Table 4.32). The study by Sharp and Powell (1931), showed that

at storage temperature of 2 C, yolk reached a pH value of 6.4 in about 50 days and at 37

C, in 18 days.


81

Haugh Unit

The three MAP treatments maintained significantly higher Haugh unit value at 6 C

compared to the control (P ≤ 0.05). No significant difference (P > 0.05) was observed

among the three MAP treatments (Figure A.33, Table 4.34). The Haugh unit value was

significantly higher for day 1 at 6 C compare to day 7, 14, 21, and 30 (P ≤ 0.05). No

significant difference (P > 0.05) in Haugh unit was observed from day7 till day 30

(Figure A.34, Table 4.35). On day 1, at abused temperature, no significant difference was

observed in the Haugh unit values of the four packaging types (P ≤ 0.05). From day 7 till

day 30, the three MAP treatments maintained significantly higher Haugh unit (P ≤ 0.05)

than the control, at 21 C (Figure A.32, Table 4.33).

At 21 C, the three MAP treatments maintained Haugh unit values in the range of

71.42 to 79.75. The measurements above 73 is the cut off for a USDA grade AA egg. On

day 0, the average Haugh unit value was 81. Thus, MAP maintained Haugh unit value

closer to that of fresh eggs during the storage time. However, the vale declined for the

control eggs from day 7 onwards and went down to 52.92.

Thus MAP maintained a higher Haugh unit value, closer to Grade AA at both

refrigerated and abused temperature. While the value declined for air packed eggs at both

the storage temperatures. The decline was more at 21 C than at 6 C for the air packed

eggs. A possible explanation for the above observations could be that storage of eggs in

MAP containing CO2, slowed down the thinning of the egg white and thus maintained

higher Haugh unit values. Since the three MAP had same percentage of CO2, no

significant difference was observed between the three treatments. Alkaline pH of eggs

produces watery white. Firstly, the jelly like white becomes more fluid until it cannot be


82

recognized as thick white. However, at relatively low pH, thick white maintains it’s jelly

like property. The largest amount of thick white is obtained at intermediate pH values and

thus at moderate concentrations of carbon dioxide (Sharp, 1929).

According to Sharp and Stewart (1931), the eggs stored in carbon dioxide were better

in condition of thick white, yolk index, viscosity of yolk as compared to eggs stored

under normal conditions. Also, in the control eggs, the thick white was flat and flabby,

while in carbon dioxide stored eggs, the thick white stood up around the yolk in more of a

jelly-like condition.

Yolk Index

On day 1 at 21 C, no significant difference was observed among the three MAP

treatments and between the CO2+ CO, CO2, and control, for the yolk index (P > 0.05).

The yolk index of the high-ox treatment was significantly higher than the air packed eggs

at 21 C for day 1 (P ≤ 0.05). For day 7 to 30, at 21 C, the three MAP treatments

maintained significantly higher yolk index than the control (P ≤ 0.05). The mean yolk

index for day 0 was 0.43 and the three MAP treatments maintained the yolk index of the

shell eggs closer to that of day 0, throughout the storage time. While the yolk index value

for the control declined from day 1 onwards and was 0.25 on day 30 at 21 C (Table 4.36,

Figure A.37).


83

Table 4.33. Haugh unit ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6/storage time/packaging treatment)



1 79.75a ± 0.84 76.08a ± 1.80 77.58a ± 2.66 80.42a ± 1.66

7 76.17a ± 1.19 75.33a ± 2.40 75.58a ± 2.34 69.00b ± 2.35

14 75.75a ± 1.86 76.08a ± 2.60 77.58a ± 0.85 63.58b ± 3.35

21 74.58a ± 1.65 72.42a ± 2.02 71.42a ± 1.79 53.83b ± 1.80

30 73.83a ± 1.81 73.67a ± 1.19 75.42a ± 2.29 52.92b ± 1.31



84

Table 4.34. Yolk index and Haugh unit ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) temperature (n = 30/packaging treatment)

Packaging treatments Yolk index Haugh unit

CO2 + CO 0.42ab ± 0.01 76.87a ± 0.95

CO2 0.43a ± 0.01 75.35a ± 0.93

High-ox 0.44a ± 0.01 77.02a ± 0.90

Air 0.41b ± 0.01 72.38b ± 0.98



85

Table 4.35. Yolk index and Haugh unit ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 24/storage time)

Storage time Yolk index Haugh unit

(d)

1 0.45a ± 0.01 78.73a ± 1.18

7 0.42b ± 0.01 74.26b ± 1.32

14 0.42b ± 0.01 75.83b ± 0.02

21 0.42b ± 0.01 73.46b ± 0.85

30 0.41b ± 0.01 74.81b ± 0.85



86

Table 4.36. Yolk index ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6/storage time/packaging treatment)



1 0.41ab ± 0.02 0.41ab ± 0.01 0.42a ± 0.03 0.38b ± 0.01

7 0.43a ± 0.01 0.42a ± 0.01 0.43a ± 0.01 0.37b ± 0.01

14 0.41a ± 0.00 0.41a ± 0.01 0.44a ± 0.01 0.30b ± 0.01

21 0.41a ± 0.01 0.44a ± 0.01 0.43a ± 0.01 0.30b ± 0.01

30 0.40a ± 0.01 0.39a ± 0.01 0.42a ± 0.01 0.25b ± 0.01



87

A possible explanation for the above observation could be that the CO2 in the three

MAP treatments, lowered the pH of the shell eggs and slowed the protein degradation.

Also, MAP slowed down the egg quality deterioration and thus, the yolk was held firm in

the egg and had higher yolk index value. While, yolk index for the control went down

due to protein degradation, weakening of the chalazae and yolk sack.

At 6 C, the yolk index of CO2, and high-ox treatments was significantly higher than

the control (P ≤ 0.05). No significant difference for yolk index (P > 0.05) was observed

between the CO2 + CO and the control at 6 C (Figure A.37, Table 4.35). Yolk index for

day 1 was significantly higher at 6 C, compared to rest of the storage time periods from

day 7 to day 30 (P ≤ 0.05). No significant difference was observed in the yolk index (P >

0.05) among days 7, 14, 21, or 30 (Figure A.36, Table 4.35). At 6 C, the yolk index of

shell eggs was closer to that of the eggs on day 0 for all the packaging treatments and at

all the storage time points. Thus, the storage temperature and MAP both helped in

maintaining the higher yolk index value of shell eggs. According to Sharp (1929), a

major deterioration factor that results from alkaline pH of the white is the standing up

quality of the yolk as indicated by the weakening and change in the permeability of yolk

membrane. The keeping quality of the yolk of the egg improves as the carbon dioxide

content of the egg storage atmosphere is increased.

Thiobarbituric acid reactive substances (TBARS)

No significant difference (P > 0.05) between the two trials and two storage

temperatures in the TBARS value of the shell eggs was observed. Hence, the data from

the two trials was combined. Also, there was no significant interaction between the

packaging treatment and the storage time for the TBARS value. No significant effect (P >


88

0.05) of the four packaging treatments was observed on the TBARS value of the shell

eggs (Table 4.37). However, TBARS value increased significantly (P ≤ 0.05) from day 1

till day 14 and then declined from day 21 to day 30 (Figure A.38, Table 4.38). A possible

explanation for this could be that the fatty acid hydroperoxide (primary lipid oxidation

product), are produced from the PUFA in the yolk, during storage. However, as the

storage time increases, these primary oxidation products are further converted into

secondary oxidation products like malonaldehyde (MDA). The TBARS method estimate

MDA formed and hence the value increases during storage. However, after some time the

MDA further breaks down and are no longer detected by the TBA colorimetric method

(Spanier and Taylor, 1991).


89

Table 4.37. Egg yolk TBARS value ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air (n = 40/packaging treatments)

Packaging treatments TBARS

CO2 + CO 8.65a ± 0.84

CO2 8.01a ± 0.54

High-ox 7.79a ± 0.61

Air 7.34a ± 0.53

aMeans with the same letter within a column were not significantly different (P ≥ 0.05).


90

Table 4.38. Egg yolk TBARS value ± SEM of shell eggs packed and stored for 1, 7, 14, 21, 30 d (n = 32/storage time)

Storage time (d) TBARS

1 6.62b ± 0.94

7 9.50a ± 0.65

14 9.57a ± 0.77

21 6.43b ± 0.46

30 7.68ab ± 0.48



91

Microbial Quality of Eggs

The three trials in Salmonella studies had significant interaction (P > 0.05) and

therefore each trial was analyzed separately. In trial 1, no significant difference (P > 0.05)

was observed in the Salmonella count among the eggs of the four packaging treatments at

both 6 C and 21 C (Table 4.39). The storage time from day1 to day 14 had no significant

effect (P > 0.05) on the Salmonella count of the eggs at 6 C (Table 4.40). However, at 21

C, Salmonella count in eggs increased significantly (P ≤ 0.05) from day 1 to day 7 from

log 6.1 to log 8.31 cfu/ g of egg sample (Figure A.39).

In trial 2, Salmonella count in shell eggs increased significantly from log 5.45 to log

6.67 from day 7 to day 14 at 6 C (P ≤ 0.05). No significant difference in the Salmonella

count was found between day 1 and 7 (P > 0.05). At 21 C, a significant increase (P ≤

0.05) in the Salmonella count from day 1 to day 7 from log 5.74 to log 8.15 was observed

(Figure A.40). The increase was greater at abusive temperature than at refrigerated

temperature as the bacterial growth is suppressed at refrigerated temperature. However,

no significant difference in the Salmonella count in shell eggs was observed among the

four packaging treatments at both refrigerated and abusive temperature (P ≤ 0.05).

In trial 3, the Salmonella count was significantly lower for air packed eggs than the

eggs packed in three different MAP treatments at 6 C (P ≤ 0.05). At 21 C, the high-ox

treated eggs had significantly lower Salmonella count (P ≤ 0.05) than CO2+ CO, CO2,

and air packed eggs (Figure A.42, Table 4.39). A significantly higher Salmonella count

(P ≤ 0.05) was observed for day 7 and 14 at both 6 C and 21 C compared to day 1 (Figure

A.41, Table 4.40). At 21 C, day 14 had the highest Salmonella count of log 8.8 cfu/ g of

egg sample.


92

Table 4.39. Log cfu of Salmonella cocktail/g ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air for trials 1, 2, and 3 and stored for 1, 7, 14 d at refrigerated (6 C) and abusive (21 C) temperature (n = 6/packaging treatment/trial)


Storage temperature Trials CO2 + CO

CO2 High-ox Air

Refrigerated

1 6.69a ± 0.55 6.49a ± 0.46 6.81a ± 0.48 6.60a ± 0.45

2 6.30a ± 0.25 5.32a ± 0.32 6.25a ± 0.25 5.75a ± 0.52

3 7.75a ± 0.16 7.56a ± 0.12 7.49a ± 0.11 7.14b ± 0.11

Abusive

1 7.62a ± 0.65 7.57a ± 0.63 7.87a ± 0.71 7.56a ± 0.64

2 7.58a ± 0.60 7.37a ± 0.54 7.42a ± 0.58 6.74a ± 0.74

3 8.01b ± 0.28 7.86b ± 0.27 8.40a ± 0.37 8.08b ± 0.31



93

Table 4.40. Log cfu of Salmonella cocktail/g ± SEM of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air for trials 1, 2, and 3 and stored for 1, 7, 14 d at refrigerated (6 C) and abusive (21 C) temperature (n = 8/storage time/trial)

Storage time (d)

Storage temperature Trials 1

7 14

Refrigerated

1 6.10a ± 0.62 7.17a ± 0.08 6.74a ± 0.15

2 5.74b ± 0.20 5.45b ± 0.38 6.67a ± 0.16

3 7.28b ± 0.06 7.57a ± 0.15 7.61a ± 0.15

Abusive

1 6.10b ± 0.62 8.31a ± 0.12 8.57a ± 0.13

2 5.74b ± 0.20 8.15a ± 0.11 8.52a ± 0.11

3 7.28c ± 0.06 8.54b ± 0.06 8.80a ± 0.13

a, b,cMeans with the same letter within a row were not significantly different (P ≥ 0.05).


94

The yolks of inoculated eggs incubated 1 to 14 days at 21 C contained high levels of

S. Enteritidis. Eggs incubated at refrigerated temperature showed lesser growth than that

of stored at abusive temperature. Jones et al. (2004) stored the eggs inoculated with S.

Enteritidis at room temperature for 5 weeks. The S. Enteritidis contamination levels

determined for interior egg contents increased with storage time. The study also

demonstrated that S. Enteritidis growth can take place in the yolk during the cooling

period also.

The Salmonella count in shell eggs for day 0 was log 3 cfu/g egg sample for trial 1

and log 2.85 cfu/g egg sample for trial 3. No bacterial growth was seen in trial 2. A

possible explanation for this was that approximately 120 cells in 40 uL of inoculum was

injected into the yolk. A very low number of cells were inoculated to represent the actual

amount present in the eggs in nature. Freshly laid eggs are rarely reported to harbor more

than a few hundred S. Enteritidis cells (Chen et al., 2002; Gast and Holt, 2000a;

Humphrey et al., 1991). In trial 1 and 2, 1:10 ratio dilution of the inoculated egg was

plated and since the Salmonella count was low, most of the eggs did not show any

growth. However, for trial 3, 1:2 and 1:4 dilutions were plated and hence growth of

Salmonella was easily detected.

No bacterial growth was found in the uninoculated eggs. Thus most of the eggs are

sterile from inside when freshly laid and the rate of transovarian egg contamination has

been one S. Enteritidis positive egg in every 20,000 eggs produced in U.S. (FSIS June 12

1998).


95

These results demonstrate the possibility that eggs naturally contaminated with low

numbers of Salmonella organisms could sustain growth of this organism to high levels if

held at abusive temperature within 24 hrs.


96

Conclusion

MAP was effective in reducing the shell egg deterioration and maintained the

functional quality of eggs. The three MAP treatments maintained a higher Haugh unit and

yolk index at both refrigerated and abusive temperature from day 7 to 30, as compared to

the control. The effect of MAP was more prominent at abusive temperature than at

refrigerated one. A possible reason for this observation could be that MAP lowered the

pH of the albumen and thus slow down protein degradation. Thus, the thinning of egg

white and weakening of chalazae and yolk sack is slowed down and hence higher Haugh

unit and yolk index was maintained.

MAP maintained lower albumen and whole egg pH than the control at both abusive

and refrigerated temperature. The yolk pH at 6 C was lower for MAP than the control.

However, at abusive temperature similar yolk pH was observed for all the four packaging

treatments. At refrigerated temperature, lesser carbon dioxide is required to produce the

similar pH lowering effect as that of abusive temperature. Hence, the yolk pH of MAP at

6 C was lower than the control since the percentage.

The foaming capacity of air packed eggs was higher than the three MAP’s at both

refrigerated and abusive temperature. This could be explained by the fact that the three

MAP treatments lowered the pH of albumen and hence the foaming capacity decreased.

Foam stability of the MAP treatments was similar to the air packed eggs at both

refrigerated and abusive temperature.

The three MAP treatments maintained a more vivid and lighter yellow color and

prevented the green ring formation on the surface of the hard cooked egg yolk than the

control eggs, throughout the storage time from day 1 to day 30 and at both the abusive


97

and refrigerated temperature. The three MAP contained 20% carbon dioxide which

lowered the albumen pH and thus prevented the green ring of FeS formation at the

interface of the yolk and the albumen. MAP maintained a lighter, more vivid and lower

hue angle of the hard cooked albumen than the control. The air packed eggs were similar

to the MAP packed ones in the color of the inner part of the hard cooked yolk. No effect

of MAP on lightness and chroma of the raw albumen for both refrigerated and abusive

temperatures. However, as the storage time increased, lightness increased, chroma

decreased and hue angle increased for both the temperatures.

The three MAP treatments maintained the functional quality of shell eggs better than

the control. However, all the three MAPs were more or less similar in their effect and

hence it can be concluded that the same percentage of carbon dioxide in all the three

MAP was mainly responsible for maintaining the egg quality.

The TBARS value were similar for all the four packaging treatments. However,

TBARS value increased from day 1 to day 14 and then decreased from day 21 to 30.

The Salmonella count increased from day 1 to day 14 at both refrigerated and abusive

temperature. However, at abusive temperature, the bacterial growth was faster and

greater. All the fore packaging treatments were similar in their effect on bacterial growth

at both the storage temperatures in trial 1 and 2. However, in trial 3, at refrigerated

temperature, air packed eggs has the lower Salmonella count than the three MAPs. At

abusive temperature, high-ox treatment had maximum Salmonella count.


98

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106

APPENDIX


107

Storage time (d)

1 7 14 21 30

Lig

htn

ess

0.00

20.00

40.00

60.00

80.00

100.00

6 C

21 C

cd

b c b b a a a a

Figure A.01. Raw albumen lightness of eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature. (n = 16/storage temperature/storage time) a, b,c,dMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05)


108

Storage time (d)

1 7 14 21 30

Hue

ang

le (

o)

0.00

20.00

40.00

60.00

80.00

100.00

120.00

140.006 C

21 C

c cb b b b b b

a a

Figure A.02. Raw albumen hue angle (o) of eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature. (n = 16/storage temperature/storage time) a, b,cMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05)


109

storage time (d)

1 7 14 21 30

Chro

ma

0.00

2.00

4.00

6.00

8.00

10.006 C

21 Ca a

b b

b bc

c dc

c

Figure A.03. Raw albumen chroma of eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature. (n = 16/storage temperature/storage time) a, b,c,dMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05)


110

Storage time (d)

1 7 14 21 30

LS

mea

n o

f li

ghtn

ess

0.00

10.00

20.00

30.00

40.00

50.00

60.00

70.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

a a a a a a a a ab

a

ab

aa

b b ab

a

b b

Figure A.04. Raw yolk lightness of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


111

Storage time (d)

1 7 14 21 30

LS

Mea

n o

f li

ghtn

ess

0.00

10.00

20.00

30.00

40.00

50.00

60.00

70.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

a a a a

b b b

a

b b b

a

b b b

a

b b b

a

Figure A.05. Raw yolk lightness of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


112

Storage time (d)

1 7 14 21 30

Hu

e an

gle

(o)

0.00

20.00

40.00

60.00

80.00

100.00

6 C

21 C

a a b b bc b cd b d c

Figure A.06. Raw yolk hue angle (o) of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature. (n = 16/storage temperature/storage time) a, b,c,dMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05)


113

Storage time (d)

1 7 14 21 30

Ch

rom

a

0.00

10.00

20.00

30.00

40.00

50.00

60.00

b

b

a

bb

Figure A.07. Raw yolk chroma of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 16/storage time) a, bMeans with the same letter were not significantly different (P ≥ 0.05)


114

Storage time (d)

1 7 14 21 30

LS

Mea

n c

hro

ma

0.00

10.00

20.00

30.00

40.00

50.00

60.00

70.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

aa a a

a a a

a bb b

a

b b b

a

a a a

a

Figure A.08. Raw yolk chroma of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/storage time/packaging treatment) a, bMeans with the same letter within a storage time were not significantly different (P ≥

0.05)


115

Storage time (d)

1 7 14 21 30

LS

Mea

n L

*

0.00

20.00

40.00

60.00

80.00

100.00

120.00

CO2 + CO + N2

CO2 + N2

High - ox

Air

a a a

b

a a a

ba a a

ba a a

ba a a

b

Figure A.09. Hard cooked albumen lightness of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/storage time/packaging treatment) a, b,cLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


116

Storage time (d)

1 7 14 21 30

LS

Mea

n L

*

0.00

20.00

40.00

60.00

80.00

100.00

120.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

a a ab

a a a

b

a a ab

a a ab b a a

b

Figure A.10. Hard cooked albumen lightness of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


117

Storage time (d)

1 7 14 21 30

LS

Mea

n h

ue

ang

le (

o)

0.00

20.00

40.00

60.00

80.00

100.00

120.00

140.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

b b b

a

b b b

a

b b b

aa

b b b b b b

a

Figure A.11. Hard cooked albumen hue angle (o) of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


118

Storage time (d)

1 7 14 21 30

LS

Mea

n h

ue

ang

le (

o)

0.00

20.00

40.00

60.00

80.00

100.00

120.00

140.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

b b b

a

b b b

a

b b b

a

b b b

a

b b b

a

Figure A.12. Hard cooked albumen hue angle (o) of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


119

Stoarge time (d)

1 7 14 21 30

LS

Mea

n c

hro

ma

0.00

2.00

4.00

6.00

8.00

10.00

12.00

14.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

aa

ab

b

a

aa

b

a aa

b

aa

a

b

a a a

b

Figure A.13. Hard cooked albumen chroma of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


120

Storage time (d)

1 7 14 21 30

Ch

rom

a

0.00

2.00

4.00

6.00

8.00

10.00

12.00

14.00

b b

aa

a

Figure A.14. Hard cooked albumen chroma shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 16/storage time) a, bMeans with the same letter were not significantly different (P ≥ 0.05)


121

Type of packaging

Ch

rom

a

0.00

2.00

4.00

6.00

8.00

10.00

12.00

14.00

CO2 + CO + N2 CO2 + N2 High - ox Air

aa a

b

Figure A.15. Hard cooked albumen chroma of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 30/packaging type) a, bMeans with the same letter were not significantly different (P ≥ 0.05)


122

Storage time (d)

1 7 14 21 30

LS

Mea

n l

igh

tnes

s

0.00

20.00

40.00

60.00

80.00

100.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

b aba

c

a a a

b

a a a

b

a a a

b

a a a

b

Figure A.16. Hard cooked yolk surface lightness of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/storage time/packaging treatment) a, b,cLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


123

Storage time (d)

1 7 14 21 30

LS

Mea

n l

igh

tnes

s

0.00

20.00

40.00

60.00

80.00

100.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

a a a

b

a a a

b

a a a

b

a a a

b

a a a

b

Figure A.17. Hard cooked yolk surface lightness of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


124

Storage time (d)

1 7 14 21 30

LS

Mea

n h

ue

angle

(o)

0.00

20.00

40.00

60.00

80.00

100.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

ab

ab

a

ba a a

ba a a b a

ab

a ba a a

b

Figure A.18. Hard cooked yolk surface hue angle (o) of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


125

Storage time (d)

1 7 14 21 30

LS

Mea

n h

ue

ang

le (

o)

0.00

20.00

40.00

60.00

80.00

100.00

120.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

aba

abb b b b

aa a a a a ab ab b a a a a

Figure A.19. Hard cooked yolk surface hue angle (o) of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


126

Storage time (d)

1 7 14 21 30

LS

Mea

n c

hro

ma

0.00

10.00

20.00

30.00

40.00

50.00

60.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

a a a

b

a a a

b

a a a

b

a a a

b

a a a

b

Figure A.20. Hard cooked yolk surface chroma of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for days 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


127

Storage time (d)

1 7 14 21 30

LS

Mea

n c

hro

ma

0.00

10.00

20.00

30.00

40.00

50.00

60.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

a a a

b

a a a

b

a a a

b

a a a

b

a a a

b

Figure A.21. Hard cooked yolk surface chroma of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) treatments (n = 4/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


128

Storage time (d)

1 7 14 21 30

Hu

e an

gle

(o)

0.00

20.00

40.00

60.00

80.00

100.00ab

bb

ab a

Figure A.22. Hue angle (o) of inner part of hard cooked egg yolk packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 16/storage time) a, bMeans with the same letter were not significantly different (P ≥ 0.05)


129

Type of packaging

Fo

am c

apac

ity

0.00

20.00

40.00

60.00

80.00

100.00

6 C

21 C


bb ab

a

b abb

a

Figure A. 23. Foam capacity of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) and abusive (21 C) temperature. (n = 30/storage temperature/packaging treatment) a, bMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05)


130

Stoarge time (d)

1 7 14 21 30

LS

Mea

n f

oam

sta

bil

ity

0.00

20.00

40.00

60.00

80.00

100.00

120.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

b

a

b ba a

a a aa a a

aa a a a a a

a

Figure A.24. Foam stability of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


131

Storage time (d)

1 7 14 21 30

Alb

um

en p

H

0.00

2.00

4.00

6.00

8.00

10.00a

b b b a

Figure A.25. Albumen pH of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 24/storage time) a, bMeans with the same letter were not significantly different (P ≥ 0.05).


132

Type of packaging

Alb

um

en p

H

0.00

2.00

4.00

6.00

8.00

10.00


cbc

b

a

Figure A.26. Albumen pH of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) temperature (n = 30/packaging type) a, b,cMeans with the same letter were not significantly different (P ≥ 0.05).


133

Figure A.27. Albumen pH of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6/storage time/packaging treatment) a, b,cLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)

Storage time (d)

1 7 14 21 30

LS

Mea

n a

lbum

en p

H

0.00

2.00

4.00

6.00

8.00

10.00

12.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

b

a

bbb b

b

a

bb

c

a

bb b

a

b bb

a

Storage time (d)

1 7 14 21 30

LS

Mea

n a

lbum

en p

H

0.00

2.00

4.00

6.00

8.00

10.00

12.00

CO2 + CO + N2

CO2 + N2

High-ox

Air

b

a

bbb b

b

a

bb

c

a

bb b

a

b bb

a


134

Type of packaging

Wh

ole

eg

g p

H

0.00

2.00

4.00

6.00

8.00

10.00

6 C

21 C


bb b

a

b b b

a

Figure A.28 Whole egg pH of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) and abusive (21 C) temperature (n = 30/storeage temperature/packaging treatment) a, bMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05)


135

Storage time (d)

1 7 14 21 30

Whole

egg p

H

0.00

2.00

4.00

6.00

8.00

10.00

6 C

21 C

ab ab bc c abc

ab abc

a

Figure A.29. Whole egg pH of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 24/storage temperature/storage timee) a, b, cMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05)


136

Type of packaging

pH

of

yolk

0.00

1.00

2.00

3.00

4.00

5.00

6.00

7.00


b

bb

a

Figure A.30. Egg yolk pH of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) temperature (n = 30/packaging treatment) a, bMeans with the same letter were not significantly different (P ≥ 0.05)


137

Storage time (d)

1 7 14 21 30

pH

of

yo

lk

0.00

1.00

2.00

3.00

4.00

5.00

6.00

7.00

6 C

21 Ca

ba

b b

a

b

ab

bab

Figure A.31. Egg yolk pH of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) and abusive (21 C) temperature (n = 24/storage temperature/storage time) a, bMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05)


138

Storage time (d)

1 7 14 21 30

LS

Mea

n H

augh

un

it

0.00

20.00

40.00

60.00

80.00

100.00

CO2 + CO + N2

CO2 + N2

High-ox

Aira a a a

a a a

b

a a a

b

a a a

b

a a a

b

Figure A.32. Haugh unit of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), and air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


139

Type of packaging

Hau

gh

un

it

0.00

20.00

40.00

60.00

80.00

100.00

CO2 + CO + N2 CO2 + N2 HIgh - Ox Air

aa a

b

Figure A.33. Haugh unit of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) temperature (n = 30/packaging treatment) a, bMeans with the same letter were not significantly different (P ≥ 0.05)


140

Storage time (d)

1 7 14 21 30

Hau

gh

un

it

0.00

20.00

40.00

60.00

80.00

100.00

a

bb

b b

Figure A.34. Haugh unit of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 24/storage time) a, bMeans with the same letter were not significantly different (P ≥ 0.05)


141

Type of packaging

Yolk

ind

ex

0.00

0.10

0.20

0.30

0.40

0.50


aba

a

b

Figure A.35 Yolk index of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored at refrigerated (6 C) temperature (n = 30/packaging treatment) a, bMeans with the same letter were not significantly different (P ≥ 0.05).


142

Storage time (d)

1 7 14 21 30

Yolk

in

dex

0.00

0.10

0.20

0.30

0.40

0.50

a

b b bb

Figure A.36. Yolk index of shell eggs packed and stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 24/storage time) a, bMeans with the same letter were not significantly different (P ≥ 0.05).


143

Stoarage time (d)

1 7 14 21 30

LS

Mean

yolk

in

de

x

0.00

0.10

0.20

0.30

0.40

0.50

CO2 + CO + N2

CO2 + N2

High-ox

Air

abab

a

b

a a a

b

b b

b

a a a aa a

a a a

Figure A.37. Yolk index of shell eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air and stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 6/storage time/packaging treatment) a, bLSMeans with the same letter within a storage time were not significantly different (P

≥ 0.05)


144

Storage time (d)

1 7 14 21 30

TB

AR

S

0.00

2.00

4.00

6.00

8.00

10.00

12.00

b

a a

b

ab

Figure A.38. Egg yolk TBARS value of shell eggs packed and stored for 1, 7, 14, 21, 30 d (n = 32/storage time) a, bMeans with the same letter are not significantly different (P ≥ 0.05).


145

Storage time (d)

0 1 7 14

Lo

g c

fu/g

eg

g

0.00

2.00

4.00

6.00

8.00

10.00

a a

b

c

Figure A.39. Log cfu of Salmonella cocktail/g of eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air for trial 1and stored for 1, 7, 14 d at abusive (21 C) temperature (n = 8/storage time) a, bMeans with the same letter were not significantly different (P ≥ 0.05).


146

Storage time (d)

0 1 7 14

Log

cfu

/g e

gg

0.00

2.00

4.00

6.00

8.00

10.00

6 C

21 C

a

a

a

bbb

Figure A.40. Log cfu of Salmonella cocktail/g of eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air for trial 2 and stored for 1, 7, 14 d at refrigerated (6 C) and abusive (21 C) temperature (n = 8/storage tetmperature/storage time) a, bMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05).


147

Storage time (d)

0 1 7 14

Log c

fu/g

egg

0.00

2.00

4.00

6.00

8.00

10.00

6 C

21 C

a

a

b

acb

d

Figure A.41. Log cfu of Salmonella cocktail/g of eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air for trial 3 and stored for 1, 7, 14 d at refrigerated (6 C) and abusive (21 C) temperature (n = 8/storage temperature/storage time) a, bMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05).


148


Log c

fu/g

egg

0.00

2.00

4.00

6.00

8.00

10.00

6 C

21 C


a

a ab

b

b

a

b

Figure A.42. Log cfu of Salmonella cocktail/g of eggs packed in CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), high-ox (20%CO2/80%O2), or air for trial 3 and stored for 1, 7, 14 d at refrigerated (6 C) and abusive (21 C) temperature (n = 6/storage temperature/packaging treatment) a, bMeans with the same letter within a storage temperature were not significantly different (P ≥ 0.05).


149

Stoarge time (d)

1 7 14 21 30

CO

2 level in

package (

%)

0.00

2.00

4.00

6.00

8.00

10.00

12.00

14.00

16.00

18.00

CO+ CO2

CO2

High-ox

b

aa

c

b

a

c

a

ab

a

bb

a

b

Figure A.43. CO2 level (%) in packages with CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), and high-ox (20%CO2/80%O2) treatments for trial 2 when stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 2/storage temperature/packaging treatment/storage time) a, b,cMeans with the same letter within a storage time were not significantly different (P ≥

0.05).


150

Stoarge time (d)

1 7 14 21 30

CO

2 level in

package (

%)

0.00

2.00

4.00

6.00

8.00

10.00

12.00

14.00

16.00

18.00 CO+ CO2

CO2

High-ox b

aa

c

b

a

b

b

a

b

b

a

b

b

a

Figure A.44. CO2 level (%) in packages with CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), and high-ox (20%CO2/80%O2) treatments for trial 2 when stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 2/storage temperature/packaging treatment/storage time) a, b,cMeans with the same letter within a storage time were not significantly different (P ≥

0.05).


151

Storage time (d)

1 7 14 21 30

CO

2 level in

package (

%)

0.00

2.00

4.00

6.00

8.00

10.00

12.00

14.00

16.00

18.00CO+ CO2

CO2

High-ox

b

aa

b

a

a

c

b

a

c

b

a

bb

a

Figure A.45. CO2 level (%) in packages with CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), and high-ox (20%CO2/80%O2) treatments for trial 3 when stored for 1, 7, 14, 21, 30 d at refrigerated (6 C) temperature (n = 2/storage temperature/packaging treatment/storage time) a, b,cMeans with the same letter within a storage time were not significantly different (P ≥

0.05).


152

Storage time (d)

1 7 14 21 30

CO

2 le

vel in

package (

%)

0.00

2.00

4.00

6.00

8.00

10.00

12.00

14.00

16.00

18.00

CO+ CO2

CO2

High-ox

bb

aa

b

aa

b

a

b

ca

bb

a

Figure A.46. CO2 level (%) in packages with CO2+ CO (20%CO2/0.4%CO/79.6%N2), CO2 (20%CO2/80%N2), and high-ox (20%CO2/80%O2) treatments for trial 3 when stored for 1, 7, 14, 21, 30 d at abusive (21 C) temperature (n = 2/storage temperature/packaging treatment/storage time) a, b,cMeans with the same letter within a storage time were not significantly different (P ≥

0.05).

PERMISSION TO COPY

In presenting this thesis in partial fulfillment of the requirements for a master’s

degree at Texas Tech University or Texas Tech University Health Sciences Center, I

agree that the Library and my major department shall make it freely available for research

purposes. Permission to copy this thesis for scholarly purposes may be granted by the

Director of the Library or my major professor. It is understood that any copying or

publication of this thesis for financial gain shall not be allowed without my further

written permission and that any user may be liable for copyright infringement.

Agree (Permission is granted.)

Divya Aggarwal 04/10/08

________________________________________________ ________________ Student Signature Date Disagree (Permission is not granted.) _______________________________________________ _________________ Student Signature Date

egg quality and salmonella spp. growth in shell eggs packaged in

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