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Optimization of a Pretreatment to Reduce Oil Absorption in Fully Fried, Battered and

Breaded Chicken Using Whey Protein Isolate as a Postbreading Dip

A thesis presented to

the faculty of

the College of Health and Human Services of Ohio University

In partial fulfillment

of the requirements for the degree

Master of Science

Eunice Mah

June 2008

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This thesis titled

Optimization of a Pretreatment to Reduce Oil Absorption in Fully Fried, Battered, and

Breaded Chickens Using Whey Protein Isolate as a Postbreading Dip

by

EUNICE MAH

has been approved for

the School of Human and Consumer Sciences

and the College of Health and Human Services by

Robert G. Brannan

Assistant Professor of Human and Consumer Sciences

Gary S. Neiman

Dean, College of Health and Human Services

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ABSTRACT

MAH, EUNICE, M.S., June 2008, Food and Nutrition

Optimization of a Pretreatment to Reduce Oil Absorption in Fully-Fried, Battered, and

Breaded Chicken Using Whey Protein Isolate as a Postbreading Dip (193 pp.)

Director of Thesis: Robert G. Brannan

As consumers become more aware of the deleterious effects of a high-fat diet,

there are increased efforts to lower fat content in foods. The effectiveness of whey

protein isolate (WPI) solution as a postbreading dip to reduce oil absorption in deep-fried,

battered, and breaded chicken patties and its effect on sensory properties was

investigated. Chicken patties were battered, breaded with either crackermeal or Japanese breadcrumbs, and dipped in WPI solutions prepared at four different protein concentrations (0%, 2.5%, 5%, and 10% w/w WPI) that were adjusted to pH 2, 3, and 8 before being deep-fried. Undipped chicken patties served as the control. Overall, the most effective treatment was observed for WPI solutions made at high concentrations (5% and

10% WPI) at low pH levels (pH 2 and 3). The highest lipid reduction was 31.2% for

patties breaded with crackermeal (CMP) at pH 2 with 5% WPI and 37.5% for patties

breaded with Japanese breadcrumbs (JBP) at pH 2 with 10% WPI. The only perceivable

sensory changes in treated patties were related to color, hardness, and crunchiness.

Increasing WPI concentration caused darkening of JBP but made CMP lighter while

patties treated at pH 8 were significantly darker across all WPI concentrations. The

presence of WPI increased perceived hardness and crunchiness for CMP but only

increased perceived hardness for JBP. Variations in pH levels did not affect texture for

both breading systems. JBP that showed the highest lipid reduction (10% WPI at pH 2)

4 were observed to be darker, less yellow, but did not produce any perceivable changes in hardness or crunchiness, while CMP with the lowest lipid content (5% WPI at pH 2) were darker, more yellow, harder, and crunchier. These results suggest that WPI exhibits oil- barrier properties that do not significantly affect the flavor of the product, irregardless of breading type, thus making it a promising alternative in lowering fat content of fried foods.

Approved: ______

Robert G. Brannan

Assistant Professor of Human and Consumer Sciences

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ACKNOWLEDGMENTS

I would like to acknowledge the following amazing individuals who have supported me directly and indirectly throughout my M. S. studies and in the production of this thesis.

Thank you so much for making me the person that I am.

Dr. Robert G. Brannan Joshua Bear Dr. Darlene Berryman Joshua O’Donnel Dr. Elizabeth Crockett Julian Price Dr. Ann Paulins Katie Horn Amanda Palmer Keely Trisel (Ashley) Beamish Ken McLean Ashley Zurmehly Lisa Dael Beth Wiseman Dr. Margaret Manoogian Bobbi Conliffe Rachel Bisset Chris Sandford Sarah Diamond Crystal Hazen Simona Allen Daria Janssen Svetha Swaminathan David Holben Dr. Sky Cone Dr. Deb Murray Vishakha Magon Ms. Diana Manchester Doug Grammer All my other friends, classmates, and Dr. Fang Meng professors at Ohio University Gary Saum Gerard Akindes (and gang) The funding agency: Dr. Grace Brannan National Dairy Council Discovery Pilot Program Grant Harris

Dr. Greg Janson

Jane Boney And last but not least, Jessica Grey My mum, dad, brother, and sister who are Jody Grenert always there for me.

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TABLE OF CONTENTS

Page

Abstract ...... 3

Acknowledgments...... 5

List of Tables ...... 11

List of Figures ...... 13

Chapter 1: Introduction ...... 14

Statement of the Problem ...... 16

Research Questions ...... 17

Significance of Study ...... 17

Delimitations and Limitations...... 18

Definition of Terms...... 20

List of Abbreviations and Acronyms ...... 21

Chapter 2: Review of the Literature ...... 22

Introduction ...... 22

Consumption of Fried Foods and Fat Intake...... 22

Battered and Breaded Foods ...... 24

Theories of Oil Uptake in Fried Items ...... 26

Stages of Deep-Fat Frying ...... 26

Mechanisms of Oil Absorption ...... 27

Water Replacement ...... 27

Cooling-Phase Effect ...... 27

Surfactant Theory of Oil Absorption ...... 31

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Factors Affecting Oil Absorption ...... 32

External Factors Affecting Oil Absorption ...... 33

Composition of the Frying Oil ...... 33

Frying Time and Temperature ...... 33

Postfry Handling ...... 34

Intrinsic Factors Affecting Oil Absorption ...... 34

Methods for Reduction of Oil Absorption ...... 35

Nonprotein-Based Coatings or Films...... 37

Protein-Based Coatings or Films ...... 42

Protein Gelation ...... 45

Factors affecting protein gelation ...... 45

Whey Protein ...... 48

Gelation of Whey Protein ...... 49

Effect of pH on whey protein gelation and oil uptake...... 51

Effect of concentration on whey protein gelation and oil

uptake...... 53

Sensory Evaluation of Foods ...... 55

Conclusion ...... 57

Chapter 3: Methodology ...... 60

Overview of Approach ...... 60

Data Collection and Analysis...... 64

Materials ...... 64

Preparation of Deep Fried Samples ...... 64

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Objective Analysis ...... 65

Oil degradation...... 65

Lipid analysis...... 65

Moisture analysis...... 66

Texture and color analysis...... 66

Sensory Analysis ...... 68

Panelist selection and training...... 68

Evaluation of samples...... 70

Statistical Analysis ...... 70

Chapter 4: Results ...... 72

Oil Degradation ...... 72

Coating Pickup ...... 73

Lipid and Moisture Content ...... 75

Surface Appearance ...... 85

Texture ...... 89

Mouth Feel Sensation and Flavor ...... 94

Chapter 5: Discussion and conclusion ...... 98

Oil Degradation ...... 98

Effect of WPI Treatment on Lipid Content ...... 98

Effect of WPI Treatment on Moisture Content ...... 101

Effect of WPI Treatment on Organoleptic Properties of Fried Chicken Patties . 102

Surface Appearance: Color ...... 102

Texture: Hardness and Crunchiness...... 105

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Mouth Feel Sensation and Flavor ...... 107

Conclusion ...... 108

Future Studies ...... 109

References ...... 111

Appendix A: IRB Form ...... 132

Appendix B: Timeline for Sensory Selection, Training, and Sampling ...... 146

Appendix C: Sensory Panel Questionnaire ...... 149

Appendix D: Consent Form ...... 151

Appendix E: Training Session 1 ...... 153

Appendix F: Training Session 2 ...... 156

Appendix G: Training Session 3 ...... 158

Appendix H: Standard Intensity Hardness Scale ...... 160

Appendix I: Standard Intensity Crispness Scale ...... 161

Appendix J: Standard Intensity Juiciness Scale ...... 162

Appendix K: Training Session 4 ...... 163

Appendix L: Training Session 5 ...... 165

Appendix M: Training Session 6 ...... 167

Appendix N: Training Session 7 ...... 169

Appendix O: Training Session 8 ...... 172

Appendix P: Training Session 9 ...... 174

Appendix Q: Training Session 10 ...... 176

Appendix R: Training Session 11 ...... 180

Appendix S: Training Session 12 ...... 182

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Appendix T: Sensory Attributes for Cooked Ground Chicken ...... 184

Appendix U: Ballot for Cooked Ground Chicken ...... 185

Appendix V: Sensory Attributes for Fried, Battered, and Breaded Chicken Patties ...... 186

Appendix W: Ballot for Crackermeal-Coated Patties ...... 188

Appendix X: Ballot for Japanese Breadcrumb-Coated Patties ...... 190

Appendix Y: Sampling Code and Order for Crackermeal-Coated Patties ...... 192

Appendix Z: Sampling Code and Order for Japanese Breadcrumb-Coated Patties ...... 193

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LIST OF TABLES

Page

Table 1: Examples of Nonprotein Ingredients Used in Formulations for Reduction of

Oil Absorption in Fried Foods ...... 41

Table 2: Examples of Protein Ingredients Used in Formulations in Reduction of Oil

Absorption in Fried Foods ...... 44

Table 3: Effects of pH Level Variations on Whey Protein Gelation ...... 52

Table 4: Timeline for Thesis ...... 63

Table 5: Mean Values for Weights for Raw Patty, Coating Pickup, Pre and

Postfrying, and Weight Difference for Deep-Fried, Battered, and Breaded Chicken

Patties ...... 74

Table 6: Main Effect Analysis for Lipid Content (%) for Deep-Fried, Battered, and

Breaded Chicken Patties ...... 76

Table 7: Main Effect Analysis for Moisture Content (%) for Deep-Fried, Battered, and Breaded Chicken Patties ...... 81

Table 8: Mean Values for Sensory Color, Evenness of Color, and Greasiness of

Surface Rating and Instrumental Color Values for Deep-Fried, Battered, and Breaded

Chicken Patties...... 86

Table 9: Mean Values for Sensory Hardness and Crunchiness, Crust Thickness, and

Instrumental Hardness, Crust Fracture, and Crust Work for Deep-Fried, Battered, and

Breaded Chicken Patties ...... 91

Table 10: Mean Values for Rating of Mouth Feel Attributes for Deep-Fried, Battered, and Breaded Chicken Patties ...... 95

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Table 11: Mean Values for Rating of Basic Tastes for Deep-Fried, Battered, and

Breaded Chicken Patties ...... 96

Table 12: Mean Values for Rating of Flavor Attributes for Deep-Fried, Battered, and

Breaded Chicken Patties ...... 97

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LIST OF FIGURES

Page

Figure 1: Diagram of liquid in a pore...... 29

Figure 2: The CIELAB color space representing the three color coordinates L*, a*, and b*...... 57

Figure 3: Flowchart of methodology for optimization of whey protein isolate (WPI) solution in reducing oil absorption in batter and breaded fried chicken when used as a postbreading dip...... 62

Figure 4: Average total polar material (%) of oil samples during the frying process with relation to the number of patties fried...... 72

Figure 5: Final lipid content (%) for crackermeal-coated patties (CMP) treated at various pH levels and whey protein isolate (WPI) concentrations...... 78

Figure 6: Final lipid content (%) for Japanese breadcrumb-coated patties (JBP) treated at various pH levels and whey protein isolate (WPI) concentrations...... 79

Figure 7: Final moisture content (%) for crackermeal-coated patties (CMP) treated at various pH levels and whey protein isolate (WPI) concentrations...... 83

Figure 8: Final moisture content (%) for Japanese breadcrumb-coated patties treated at various pH levels and whey protein isolate (WPI) concentrations...... 84

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CHAPTER 1: INTRODUCTION

This study investigated the effectiveness of a treatment using whey protein isolate

(WPI) as a postbreading dip to reduce the fat content of deep-fried, battered, and breaded chicken patties. Research towards the reduction of fat content in foods are important because it could simultaneously fulfill the steady demand for fried foods and contribute to the growing efforts of Americans to consume less fat.

The Institute of Medicine’s (IOM’s) Acceptable Macronutrient Distribution

Range (AMDR) for fat intake for adults is 20 to 35% of total calories per day. Although the mean percent of daily calories contributed by fats for all ages is within this range

(32.7%; Breifel & Johnson, 2004), they are on the high end. Since total energy intake has increased (Gebhardt, et al., 2006), the absolute level of fat intake has risen from 73.4 g in

1989-1991 to 76.4 g in 1994-1996 (Breifel & Johnson, 2004; Chanmugam et al., 2003).

High intake of fat is associated with increased risk for many chronic diseases and health complications such as cardiovascular disease (Oh, Hu, Manson, Stampfer, &

Willett, 2005), high blood cholesterol levels (Albert & Mittal, 2002), obesity (Howarth,

Huang, Roberts, & McCrory, 2005), diabetes (Thanopoulou, et al., 2003), and some types of cancer such as breast cancer (Smith-Warner & Stampfer, 2007) and prostate cancer

(Fradet, Meyer, Bairati, Shadmani, & Moore, 1999). The high level of fat intake may be closely related to the increase of convenient fried food products in the market. Fried foods contain significant amounts of fats, reaching in some cases one third of the total food product by weight (Mellema, 2003). As consumers become increasingly aware of the need to lower their daily fat intake, there is a push towards low- or reduced-fat foods and beverages (Calorie Control Council, 2006). In light of the demands for low-fat foods,

15 research into reducing fat/oil absorption during deep-fat frying may be a strategy that can help achieve calorie reduction in the American diet.

Methods currently employed to reduce fat absorption in fried foods include modifications of frying techniques such as shaking and draining of fried foods (Bouchon,

Aguilera, & Pyle, 2003; Mellema, 2003), careful monitoring of frying temperature and oil degradation (Math, Velu, Nagender, & Rao, 2004), and altering the surface of the food by reducing the surface area (Goni, Bravo, Larrauri, & Calixto, 1997; Moreira, Sun, & Chen,

1997) or covering the surface with lipid barriers. Most barriers that are used in the commercial production of fried foods are made from proteins or nonprotein hydrocolloids such as corn zein, soy protein, albumin, cellulose, and gums (Gennadios, Weller, Hanna,

& Froning, 1996; Mellema, 2003). The mechanisms by which these ingredients are responsible for oil inhibition are varied and include an increase in water holding in the product, which reduces the likelihood of steam escape during frying, as observed when using curdlan (T. Funami, M. Funami, Tawada, & Nakao, 1999), the alteration of surface hydrophobicity of the product being deep fried (Annapure, Singhal, & Kulkarni, 1999), and the creation of a hurdle to moisture release and subsequent oil absorption, usually via the formation of thermally induced gels such as those formed by whey proteins.

Whey protein could potentially be used as a lipid barrier due to its ability to form thermally induced gels. It is composed of various proteins, the main protein of which is

β-lactoglobulin (β-lac), a globular protein with an isoelectric point of 5.1. This protein makes up 50-55% of the total protein contained in whey. Other important globular proteins of whey are α-lactalbumin (α-lac) and bovine serum albumin (BSA). All of these proteins, particularly β-lac, can form thermally-induced gels that alter the porosity of the

16 product, thus lowering moisture loss due to evaporation and subsequent oil absorption into the fried food when used as a coating (Dogan, Sahin, & Sumnu, 2005; Van Vliet,

Lakemond, & Visschers, 2004). The usage of whey protein as a food coating has long been investigated and applied (Gennadios, Hanna, & Kurth, 1997; Mellema, 2003).

However, the extent of commercial application of whey protein coatings as oil barriers has been limited to separating oil-rich products, such as nuts, from other components of heterogeneous foods such as cereal (Haines, 2004). Hence, this research seeks to expand the usage of whey protein as a lipid barrier by examining the effectiveness of whey protein in reducing oil absorption in deep-fried, battered, and breaded chicken patties.

Statement of the Problem

The overall objective of this research is the optimization of a pretreatment to reduce oil absorption in fully fried, battered, and breaded products by utilizing a solution of WPI as a postbreading dip. Whey protein coatings may form gels when heated, such as that during deep frying. Because temperature, concentration of protein, pH, and ionic strength affect gelation of whey protein due to influences on the rate of denaturation and aggregation (Belitz & Grosch, 1999), it may be beneficial to study the effects of these factors on the ability of WPI coatings to reduce fat uptake in fried foods. This current research focused on the effects of whey protein concentration and pH on fat and moisture content and organoleptic properties (appearance, taste, texture, and flavor) in fried foods and determined an optimal pH and protein concentration for the development of a WPI coating.

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Research Questions

1. What is the optimal pH and concentration of whey protein isolate (WPI) solution

necessary in a postbreading dip to reduce fat uptake in fully fried, battered and

breaded chicken patties?

2. How does the coating affect the chemical (fat and moisture content) and organoleptic

properties (appearance, taste, texture, and flavor) of the product?

Significance of Study

The usage of whey protein as food coatings has long been investigated and applied (Gennadios et al., 1996; Haines, 2004; Mellema, 2003). However, the extent of commercial application of whey protein coatings as oil barriers is limited to separating oil-rich products, such as nuts, from other components of heterogeneous foods such as cereal (Haines, 2004). Furthermore, application of other types of coatings to reduce oil uptake are not commercially feasible due to physical, chemical or economical constraints.

A search of available patents on oil-barriers in foods revealed that the technology involves multiple steps that are time consuming (long postdipping drying time), and use non-natural chemical additives, or employ treatments that potentially compromise the quality of the finished product (U.S. Patent No. 4,917,908, 1990; U.S. Patent No.

5,126,152, 1992). The usage of WPI as a postbreading solution requires optimization of various factors such as pH level and protein concentration. In addition, the final product should be nutritious and appealing to the senses. The results of this research will contribute to existing studies on the oil-barrier capabilities of whey protein films and may become the stepping stone to the commercial application of these coatings on fried foods.

Because whey has many disposal issues, increased utilization of whey will not only

18 benefit the dairy industry financially, but also will help decrease the amount of whey that is being disposed. Fried food manufacturers may also benefit by being able to promote their foods as being low- or reduced-fat products. Finally, the commercial application of reduced-fat-frozen-prefried foods might contribute to the reduction of fat intake in the

American diet.

Delimitations and Limitations

There are many factors apart from the independent variables of this study (pH levels and WPI concentration) that may affect oil absorption in the fried chicken patties, such as oil quality and composition (Fillion & Henry, 1998), frying temperature and time

(Sahin, Sumnu, & Altunakar, 2005), product composition (Sahin et al., 2005), moisture content (Sjöqvist & Gatenholm, 2005), shape (Mellema, 2003), porosity (Gamble, Rice,

& Selman, 1987), prefrying treatment (Krokida, Oreopoulou, Maroulis, & Marinos-

Kouris, 2001), surface treatments (Krokida, Oreopoulou, & Maroulis, 2000), initial interfacial tension (Pinthus & Saguy, 1994), and crust size (Maskat & Kerr, 2002).

Efforts were made to minimize the influence of these factors on the end results and these included standardization of batter composition and viscosity, and keeping as many frying parameters constant as possible such as oil degradation, patty weight, batter, breading, and whey pickup, and frying temperature. Nothing was deliberately done to alter the product composition (pure chicken breast meat), moisture content, shape, and porosity of the samples. During statistical analysis, all factors that were suspected to influence the measured variables were treated as covariants and analyzed using analysis of covariance

(ANCOVA).

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All instrumental analyses were individually performed on randomly selected patties from each treatment, thus resulting in the large standard deviations due to preanalytical errors that affect individual patties in addition to errors during analysis.

During statistical analysis of these measurements, all outliers were removed from the results to account for these errors. Sensory analysis results may be affected by psychological errors that influenced individual responses. Some of these errors may include those due to expectation, preference, environment, and fatigue (Stone & Sidel,

1985). The panel was screened and trained for a total of 17, 50-minute sessions to reduce occurrence of these errors during sampling. In addition, sampling was limited to six samples per session to minimize the potential for fatigue (Stone & Sidel, 1985). All samples were randomly selected and served to each panelist immediately upon being reheated to an internal temperature of 74 °C to minimize variations due to rethermalization.

Despite the many factors that may influence the oil absorption and organoleptic properties of the fried chicken patties, this study only focused on the effect of pH and protein concentration on final lipid content, moisture content, and sensory attributes. This study used ground chicken breast as the sample medium and results may not be applicable to other food products such as vegetables due to differences in moisture content and food matrix structure.

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Definition of Terms

α-lactalbumin: Second most abundant (20-25%) protein fraction in whey, globular protein with isoelectric point between 4.2 and 4.5.

β-lactoglobulin: Major protein fraction (50-55%) in whey, globular protein with isoelectric point at around 5.1.

Deep-frying: Frying by submerging food into hot frying medium.

Edible film: Continuous barrier that is formed on to the surface of the foodstuff, uses ingredients that are safe to eat or Generally Recognized as Safe (GRAS) by FDA.

Globular proteins: A class of protein that is usually spherical in shape, may have multiple domains, contain hydrophobic core, and have multiple functions (in constrast to fibrillar proteins that have a structural function).

Isoelectric point: pH at which protein molecules have no net charge and are least soluble.

Low-fat: Contains less than 3 g of fat and contributes less than 30% of calories from fat per serving.

Organoleptic: Includes sensory properties of a product, which may involving taste, color, aroma, and feel.

Par-frying: Blanching or half-frying to an internal temperature of 71 °C, then cooled and stored.

Reduced-fat: Contains 35% less fat compared to the original version.

Total polar material: Total amount of compounds that are the result of the breakdown of triglycerides (e.g., diglycerides, monoglycerides, fatty acids, and other non-triglyceride compounds), measure of oil degradation.

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Whey protein isolate: Whey which was processed to remove most of the lactose and fat and contains at least 90% protein.

Thermogelation: Gelation induced by heating. Pertaining to protein solutions: As heat is increased, denaturation occurs and lowers viscosity of the protein solution. This is followed by aggregation, which increases viscosity, and the resulting product is a gel.

Umami: The fifth basic taste. Applies to the sensation of savoriness, specifically to the detection of the natural amino acid, glutamic acid, or glutamates.

List of Abbreviations and Acronyms

β-lac: β-lactoglobulin

ANCOVA: Analysis of Covariance

ANOVA: Analysis of Variance

CMP: Crackermeal-coated patties

JBP: Japanese breadcrumb-coated patties

± s. d.: Standard deviation

TPM: Total polar material

WPI: Whey protein isolate

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CHAPTER 2: REVIEW OF THE LITERATURE

Introduction

Consumption of Fried Foods and Fat Intake

The Institute of Medicine’s (IOM’s) Acceptable Macronutrient Distribution Range

(AMDR) for fat intake for adults is 20 to 35% of total calories per day. According to data obtained from the Third National Health and Nutrition Examination Survey (NHANES

III) and from NHANES 1999–2000, the mean percent of daily calories contributed by fats for all ages is 32.7% with mean fat intake for ages 2 to 19 being 33.5% (Breifel &

Johnson, 2004; Troiano, Briefel, Carroll, & Bialostosky, 2000). Although these values show that the average fat intake is within the recommended range, they are on the high end. In addition, studies using the results from the Continuing Survey of Food Intake by

Individuals (CSFII) for 1994–1996 and 1998) show that fewer than 5% of children and adults have intakes less than 20% of calories from fats while about 25% of them have intakes greater than 35% of calories from fats (IOM, 2002). Despite reports that the total fat intake has decreased from 36% of calories in 1971-1974 to 33% of calories in 1999-

2000, the absolute level of fat intake has increased from 73.4 g in 1989-1991 to 76.4 g in

1994-1996 (Breifel & Johnson, 2004; Chanmugam, et al., 2003). It was suggested that the decrease of percent of calories from fat is caused by the concurrent increase in total energy intake (Gebhardt, et al., 2006).

Poor diet and/or unhealthy lifestyles contribute to various chronic health conditions that negatively affect the quality of life. Three of the top ten leading causes of death are diet-related, including coronary heart disease (contributes 21.1% of all deaths), cancer (23.4% of deaths), stroke (6.7% of deaths), and diabetes mellitus (2.5% of deaths;

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Singh, Kochanek, & MacDorman, 1996). All these complications are highly correlated to fat intake (Gebhardt, et al., 2006). High saturated fats in the diet may lead to an increase in low-density lipoprotein (LDL) cholesterol concentrations which increases the risk for coronary heart disease. In addition, diets higher than 35% calories in fat have been observed to increase the risk of developing breast and colorectal cancer and diabetes

(Bingham & Riboli, 2004; Boyd, et al., 2003; Gebhardt, et al., 2006). Furthermore, the prevalence of excess body weight, particularly obesity, has long been attributed to high fat intake.

Recent research has shown that Americans, especially children, are becoming heavier. Correlation studies conclude that overweight prevalence is related to consumption of food prepared away from the home (Guthrie, Lin, & Frazão, 2002).

There is also a positive correlation between the number of restaurants per capita and high body mass index and obesity levels (Chou, Grossman, & Saffer, 2004). Foods consumed away from home have a higher fat content (Lin, Frazão, & Guthrie, 1999; Nielsen, Seiga-

Riz, & Popkin, 2002) and are most likely to be fried (Taveras, et al., 2005), thus contributing to the increasing absolute fat intake for the average American. Fried foods contain significant amounts of fats, reaching in some cases up to a third of the total food product by weight (Mellema, 2003).

Consumers increasingly are aware of the need to lower their daily fat intake.

According to a national survey conducted by the Calorie Control Council (2006), 188 million adult Americans (88% of the adult U.S. population) were reported to consume low- or reduced-fat foods and beverages and two-thirds of adults believe there is a need for food ingredients that can replace the fat in food products. The food industry has

24 responded to consumer demand by offering an ever-increasing variety of low-fat eating choices. In 2001, 7.4% of new food products launched worldwide claimed to contain reduced levels of fat, and this rose to 10.4% in 2005 indicating an increase in low- or reduced-fat food demand (Datamonitor Plc., 2006). Examples of low- or reduced-fat foods include ice creams made with skim milk instead of whole milk, leaner cuts of meats used in frozen entrees, and potato chips and other salty snacks that are baked instead of fried. Despite the trend towards lower fat foods, health problems related to high fat intake are still on the rise. This is probably due to the increasing trend to patronize fast food restaurants where deep-fat frying is one of the most popular methods of cooking. In addition to frequent visits to fast food outlets and restaurants, the consumption of snack foods which include deep fried foods, are high (Kuchler, Tegene,

& Harris, 2004). Kuchler et al. (2004) reported that up to 95.5% of household surveyed purchase and consume chips (potato, corn, and tortilla). This totals to 16.34 lbs of deep- fried chips per year. Due to the growing effort of Americans to consume less fats and the increasing demand for fried foods, research into reducing fat/oil absorption during deep- fat frying might be a strategy to achieve calorie reduction in the American diet.

Battered and Breaded Foods

The popularity of battered and breaded food products has risen worldwide. For example, according to a study done by Leatherhead Foods, a UK based research firm, demand for coated foods in Europe is worth €3.46 billion and increased an estimated

18% between 2004 and 2008 (Patton, 2005). Preparation of battered and breaded fried foods involves a series of steps. After the substrate (e.g., chicken) is washed, the product may be predusted. Predusts are usually made up of flour to absorb moisture on the

25 surface of the product, thus allowing better adhesion of the batter (Yang & Chen, 1979).

In addition, gums, starches and proteins used alone or in combination can also absorb moisture from the surface of the product (Kuntz, 1997). Next, the food product is coated with liquid batter and then placed into contact with a breading material such as breadcrumbs or flour. The food product may be packaged raw, par-fried or fully fried

(U.S. Patent No. 5,126,152).

Breading differ in size, composition, and texture. Typical breading types that are currently used in the food industry are sheeted breadcrumb products such as crackermeal, white breaders, and colored breaders, American style breadcrumbs, and Japanese breadcrumbs or Panko. Sheeted breadcrumb products are produced by rolling out dough that are then baked before being ground to produce firm and dense particles. The composition of the dough varies according to the type of breading. For example, crackermeal only consists of flour and water, while white breaders may also include browning agents, leavening agents, salt, oil, and flavorings. American style breadcrumbs are made from dried bread. The crumbs are spiracle, dense, and crunchy. Japanese breadcrumbs are produced from loaves of dough that are baked in a unique process known as dielectric baking where electric current is passed through the loaves to generate heat. The resulting crumbs are porous, pale in color, and light. Japanese breadcrumbs are generally larger compared to American style breadcrumbs and sheeted breadcrumb products (U.S. Patent No. 4,943,438).

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Theories of Oil Uptake in Fried Items

Stages of Deep-Fat Frying

Deep fat-frying can be described in four stages (Farkas, Singh, & Rumsey, 1996).

The first stage is known as the initial heating stage. During this stage, the temperature of the surface of the food rises to the boiling temperature of the surface water. This stage is short, lasting for about 10 s, and is marked by negligible lost of mass (water) and heat is transferred via natural convection. The second stage is known as surface heating. The heat transfer mechanism changes from natural convection to forced convection as surface water reaches boiling temperature and is turned to vapor. The change to forced convection increases the heat transfer coefficient and thus, heat transfer into the food is faster. This stage marks the beginning of crust formation. The next stage is the falling rate stage and is the longest of all the stages. Most of the moisture is lost in this stage as the core region approaches boiling point of water. Towards the end of this stage, the rate of vapor mass transfer steadily decreases due to the low amount of free water and continued thickening of the crust that acts as a barrier for rapid vapor release. The final stage is the bubble end point. This stage is characterized by the apparent cessation of moisture loss from the food during frying. This may be caused by several factors ranging from the complete removal of all liquid water in the sample, such as with potato chips, to a reduction in heat transfer to the crust/core interface. The dryness and porosity of the crust lowers its thermal conductivity, which decreases heat transfer into the food.

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Mechanisms of Oil Absorption

According to Dana and Saguy (2006), there are three proposed mechanisms explaining oil uptake in fried foods, namely water replacement, cooling-phase effect, and surfactant theory of frying.

Water Replacement

According to this mechanism, oil absorption is the result of the replacement of evaporated water with cooking oil during the frying process. When food is immersed in oil that is heated to above the boiling temperature of water, moisture near the surface of the food is almost instantly converted to steam. This leaves pores on the surface of the food. When the void is large enough, positive vapor pressure is very low and this allows oil to enter into the food. However, this is restricted to large voids near the surface of the food (i.e., the crust region). This mechanism is able to explain the direct relationship observed between water loss and oil uptake observed during the frying process (Pinthus

& Saguy, 1994; Rice & Gamble, 1989). However, many studies have shown that oil absorption occurs mainly during the cooling phase (Bouchon et al., 2003; Moreira et al.,

1997; Pinthus & Saguy, 1994) and thus, this mechanism taken alone is inadequate in explaining the totality of oil absorption in fried foods.

Cooling-Phase Effect

After the food is fried and removed from the hot oil, the food cools and this results in water vapor condensation which decreases the internal pressure of the pores that develop during the frying process. The sudden drop in pore pressure creates a vacuum effect that sucks the oil that adhered to the surface into the pores. This mechanism relies heavily on the relationship between oil uptake and the surface of the

28 food. Oil intake is restricted to the immediate crust and product surface and is largely dependent on crust microstructure (Pinthus & Saguy, 1994) and oil viscosity (Dana &

Saguy, 2006). Since oil uptake via the cooling-phase effect is highly dependent on the surface of the food, this mechanism could be further explained using surface chemistry, particularly wetting, capillary penetration and displacement (Pinthus & Saguy, 1994).

Wetting deals with the affinity of a fluid for a solid: Fluid with low affinity will form beads on the surface of the solid, and fluid with high affinity will form a film on the surface (Miller & Neogi, 1985). Wettability is dependent on the contact angle, interfacial tension, and the radius of the pores. The relationship between these three properties with capillary pressure is represented in Equation 1 and is shown in Figure 1 (Adamson &

Gast, 1997).

2γ cosθ P = P − P = (1) c nw w r

The wetting fluid, which in this case is the frying medium, will continue to enter a pore until there is an opposing pressure, the pressure from the non-wetting fluid (Pnw), that matches that of the pressure exerted by the wetting fluid (Pw). An example of an

opposing pressure would be the pressure from the trapped vapor in the pore. As described

by the capillary pressure (Pc) equation, this is dependent on the interfacial tension

between the gas or air and liquid (γ), contact angle between the surface and the liquid (θ),

and the inner radius of the pore (r). If the contact angle is greater than 90°, the liquid does not wet the solid and tends to move about on the surface and not enter the capillary pores.

The liquid completely wets a solid only when the angle is zero (Adamson & Gast, 1997).

29

Figure 1. Diagram of liquid in a pore. In this diagram, the meniscus is part of a circle

with radius R. The capillary pressure is the net pressure contributed by the opposing

pressure of the non-wetting phase (Pnw) and the wetting phase (Pw). In deep-fat frying, the

non-wetting phase is the water vapor while the wetting phase is the frying medium.

______Note. From Physical Chemistry of Surfaces (p. 12), by A. W. Adamson and A. P. Gast, 1997, New York: Wiley Interscience. Copyright 1997 by John Wiley & Sons, Inc. Adapted with permission.

In addition to contact angle values, the oil would more likely enter a pore if the

radius of the pore is large and/or if the interfacial tension is low. Upon removal of the

food from the frying oil, the interfacial tension between oil and air within the pores

increases as temperature decreases and causes surface oil to flow rapidly into pores

(Moreira et al., 1997). Bouchon et al. (2003) studied the relationship between the amount of oil that penetrated the surface postfrying and oil left on the surface and found that the

amount of surface oil that enters the food increased with frying time, mostly probably due

30 to the increased formation of pores within the crust. While oil absorption was restricted in regions closer to the surface, the distribution of oil absorption was not even, reflecting the probability of the influence of local microstructural differences (Saguy, Gremaud,

Gloria, & Turesky, 1997). Based on these results, it was suggested that the microstructure

(mean pore size, connectedness, permeability) of the crust region is the single most important product-related determinant of oil uptake into the food.

Although the cooling-phase effect mechanism only deals with oil absorption postfrying, many studies have shown that oil absorption is the highest during the cooling phase, although it does not account for all the oil absorption in fried foods. In fact, 80% of oil absorbed in fried tortilla, frozen, par-fried potatoes during refrying and in french fries (Aguilera & Gloria-Hernandez, 2000; Moreira et al., 1997; Saguy et al., 1997;

Ufheil & Escher, 1996). This may be due to the slower pressure build-up when frying at a lower temperature, which facilitates capillary diffusion of oil. During cooling, fries cooled at a lower temperature (25 °C) absorbed oil faster than those cooled at a higher temperature (80 °C) because higher cooling temperature induces a smaller capillary pressure difference (Yamsaengsung & Moreira, 2002).

Since the characteristics of the surface of the fried product is important in determining the amount of oil that enters the product in accordance to the cooling-phase mechanism, any modifications of the surface microstructure will affect oil absorption.

These modifications will have a significant impact on oil absorption because many studies have shown that the highest oil uptake is during the cooling phase (Aguilera &

Gloria-Hernandez, 2000; Moreira et al., 1997; Saguy et al., 1997; Ufheil & Escher,

1996).

31

Surfactant Theory of Oil Absorption

The surfactant theory attributes increased oil uptake to the generation of surfactants especially in aging oil (Blumenthal & Stier, 1991). Oil absorption during frying or postfrying is largely dependent on interfacial tension between oil and water and contact angle (Dana & Saguy, 2006). As the frying process progresses, the oil degrades and this changes the composition of oil from 96 to 99% triglycerides to a mixture of compounds (e.g., diglycerides, monoglycerides, free fatty acids, and glycerol; Paul &

Mittal, 1997). These breakdown compounds act as surface-active agents that serve to lower the interfacial tension between the oil and the water (Dana & Saguy, 2006).

In support of this mechanism, it has been observed that these breakdown compounds (especially monoglycerides) cause a significant decrease in interfacial tension in oil/water systems between frying oil and water due to their degree of unsaturation and molecular structure (Feuge, 1947; Gaonkar, 1989; Gaonkar & Borwankar, 1991). Gil and

Handel (1995) did a study on the effect of monoglycerides of differing degrees of saturation on interfacial tension in oil and water systems and found that interfacial tension decreased as the degree of saturation increased. They concluded that monostearin, which has no double bond, can align at the interface more compactly due to an absence of a kink in the structure as compared to monoolein and monolinolein, which have one double bond and two double bonds, respectively. While some investigators have shown that oil absorption increased with degradation (Dana & Saguy, 2006), others have observed either no significant increase in oil absorption (Dobarganes, Márquez-Ruiz, &

Velasco, 2000) or a decrease in oil uptake (Moreira et al., 1997). The inconsistent results obtained by various studies suggest that the surfactant theory may only apply for certain

32 oil and product conditions such as oil type and composition of fried food (Dana & Saguy,

2006). Nevertheless, this mechanism cannot be ignored, because it was observed to have an effect on oil absorption in some studies.

In conclusion, none of the mechanisms, taken alone, adequately explain the complexity of oil absorption in fried foods. While many studies showed that most of the oil absorption occurs during the cooling phase, they also show that some of the oil is taken up during frying (water replacement mechanism), and that there is a correlation between degradation of oil and oil uptake (surfactant theory of oil absorption). In addition, all of these mechanisms show the strong dependence of oil absorption on the microstructure and surface characteristics of the product.

Factors Affecting Oil Absorption

There are many factors affecting oil absorption in fried foods. These can be roughly classified into two categories: (a) those that are intrinsic properties of the food being fried such as product composition, moisture content, shape, porosity, initial interfacial tension, and crust size, and (b) those that are part of the frying process such as oil quality and composition, frying temperature and time, and frying methods (Albert &

Mittal, 2002; Fillion & Henry, 1998; Gamble et al.,1987; Krokida et al., 2000; Krokida et al., 2001; Maskat & Kerr, 2002; Mellema, 2003; Pinthus & Saguy, 1994; Sahin et al.,

2005). Identifying these factors would help in the development of processes that would lead to a lower final fat content.

33

External Factors Affecting Oil Absorption

Composition of the Frying Oil

Frying oil may differ in degree of saturation and fatty acid composition.

Nutritionally, it is recommended that frying fats should be low in saturated fats and trans- fat and high in unsaturated fats. However, saturated fats are more stable and are preferred in deep-fat frying to maintain food quality (Stier, 2004). Choosing a suitable frying medium is important in controlling fat absorption in fried foods although it is a minor factor in oil absorption compared to the intrinsic properties of the product that is being fried (Dobarganes et al., 2000). Based on the surfactant theory of oil absorption, a suitable frying oil that will reduce the final fat uptake will be one that reduces lipid oxidation since this will lessen the degree of oil degradation, such as oils that are rich in oleic acid, a monounsaturated fatty acid (Abdulkarim, Long, Lai, Muhammad, &

Ghazali, 2005; Ngadi, Li, & Oluka, 2007; Romero, Cuesta, & Sanchez-Muniz, 2000).

However, Krokida et al. (2000) found no difference in fat uptake when french fries were fried in oil with different concentrations of hydrogenated oil. The inconsistency of the effect of the degree of hydrogenation of frying oil suggests that the properties of the frying medium may not have a significant impact on oil absorption.

Frying Time and Temperature

Monitoring frying time and temperature has been practiced in the frying industry to ensure the consistent quality of fried foods. This may be attributed to the increased formation of surfactants due to oil degradation and its relation to oil uptake. Fat content was shown to increase with frying time in both coated and uncoated foods (Makinson,

Greenfield, Wong, & Wills, 1987) until it reaches an equilibrium (Krokida et al., 2000).

34

Foods fried at high temperatures have been observed to have less oil uptake and this may be due to the quick pressure buildup when frying at high temperatures, which prevented capillary diffusion of oil (Yamsaengsung & Moreira, 2002).

Postfry Handling

As mentioned before, oil is actively absorbed into the product once it is removed from the frying medium. Oil absorption postfrying is related to the amount of oil remaining on the food surface and can also be affected by cooling temperatures. More viscous oil (due to degradation) tends to cling better to food surfaces and increase the chances of being absorbed into foods (Mackay, 1999) while cooling fried foods at low temperatures promotes fat uptake (Yamsaengsung & Moreira, 2002). Thus, practices such as shaking or draining fried foods which physically remove the oil from the food surface might help reduce oil absorption.

Intrinsic Factors Affecting Oil Absorption

In general, foods that have lower initial moisture content will result in a higher final fat content (Gamble et al., 1987). Many studies have shown the dependence of fat uptake on initial moisture content and loss (Gamble et al.; Mellema, 2003). Another related factor is the initial fat content of the food being fried. This is because there is a need for a concentration gradient for fat absorption to take place from the surroundings into the product (Ateba & Mittal, 1994). According to a study done by Mai, Shimp,

Weihrauch, and Kinsella (1978), the higher the fat content of the fish (e.g., trout), the less lipid change is induced by frying. Makinson et al. (1987) showed that higher amounts of fat were absorbed if the substrate is of a plant origin that naturally has a higher moisture content and lower fat content.

35

Components of foods that have been shown to have a significant impact on fat uptake are protein level, type of starch, and the presence of hydrocolloids. Overall, higher protein content results in lower final fat content due to the ability of protein to retain moisture (Salvador, Sanz, & Fiszman, 2005). The effect of protein is dependent on the nature of the major protein fraction. More lipophobic proteins, such as ovalbumin, may decrease oil absorption while more lipophilic proteins, such as lipoproteins and phosphoproteins found in egg yolk, may emulsify oil and water into the fried product

(Mohamed, N. A. Hamid, & M. A. Hamid, 1998). Even though addition of emulsifiers in batters increased oil absorption, excess amounts were actually shown to decrease oil uptake (Mohamed et al., 1998). Batters made with soy, egg albumin, and whey protein were observed to reduce oil absorption either due to improved water vapor barrier properties and/or through imparting lipophobic properties to the batter (Dogan et al.,

2005). Fat uptake in starchy foods depends on the amylose/amylopectin ratio of the flours used and particle size since they influence porosity and moisture content. Flour with smaller particle size tends to form a thick crust very early in the frying process, thereby impeding the transfer of heat to inner portions of the fried foods and less moisture is evaporated (Singh, Hung, Phillips, Chinnan, & McWatters, 2004). Furthermore, smaller particles form smaller pores with higher capillary pressure, leading to a reduction in oil uptake (Moreira et al., 1997).

Methods for Reduction of Oil Absorption

Because initial moisture content is a factor in the absorption of oil during frying, prefry drying is a method that has been shown to reduce final fat content in fried foods

(Debnath, Bhat, & Rastogi, 2003; Pedreschi & Moyano, 2005). In a study on fried potato

36 chips, Lamberg, Hallstorm, and Olsson (1990) found that drying the potato chips prior to deep-fat frying reduced the fat uptake up to 49%. However, variations in prefry drying methods also play a role in determining final oil uptake. Prefry drying via microwave or hot air drying was observed to lower oil absorption but freeze drying increased oil uptake

(Gamble et al., 1987). Freeze drying causes an even distribution of moisture and when freeze-dried food is fried, moisture loss occurs evenly over the surface and results in even oil covering (Gamble et al., 1987). Microwave drying and air drying gives a heterogeneous moisture distribution, and oil absorption is only limited to areas with low moisture content. Methods that increase starch granule integrity have been shown to reduce oil uptake in starchy fried foods. Blanching potato with calcium chloride was observed to lower oil absorption because pectic enzymes in the potato react with calcium ions to create more rigid structure and increases the firmness of the cell wall. This decreases leaching of starch granule content and reduces oil absorption into the granules

(Rimac-Brnčic´, Rade, & Šimundic´, 2004).

Research in the development of lower fat products has focused on physical modification of the food product (Mellema, 2003) such as decreasing surface area and coatings. In general, products with a larger surface area have a higher final fat content compared to products with smaller surface area. Greenfield, Makinson, and Wills (1992) found that decreased fries size significantly increased fat content of fries in a linear fashion due to a higher surface area. Potato chips that were cut thin and had rough surfaces had a higher fat uptake than potato chips that were thicker and had smoother surfaces (Goni et al., 1997; Keller, Escher, & Solms, 1990). Other modifications that have been done on fried food surface include application of coatings or batter using

37 various compositions (Gennadios et al., 1997). According to Makinson et al. (1987), batter-coated products absorbed less oil than uncoated fish sticks, while breading alone did not significantly change the total fat uptake of the fish. Presence of batter apparently resulted in the formation of a hard crust, which was impervious to the movement of water and fat. Hence, water loss and fat absorption were reduced during frying (Fillion &

Henry, 1998). Properties of coatings that are beneficial to reducing fat intake include low moisture content, low moisture permeability, thermogelling or crosslinking. Coatings should have low moisture content because oil uptake is dependent on the moisture content of the surface of the fried food (Lamberg et al., 1990; Moreira et al., 1997). Low moisture permeability in coatings helps to reduce water loss and thus, reduces oil uptake.

However, too much of this property may result in soggy foods. (Mellema, 2003).

Thermogelling or crosslinking results in high gel strength and leads to lower water diffusivity. Thermogelling or crosslinking also promotes the formation of wide punctures with low capillary pressures (Mellema, 2003).

Nonprotein-Based Coatings or Films

Another method of modifying the surface of fried products is the application of edible films and coatings. Both can be applied to the product via spraying, immersion, or direct application (Rayner, Ciolfi, Maves, Stedman, & Mittal, 2000). Edible films and coatings have been used commercially as coverings on fresh produce, candy, meats, nuts, cereal, and so forth (Albert & Mittal, 2002; Gennadios et al., 1997). According to a review done by Gennadios et al. (1997) on the application of edible films and coatings on meats, the use of coatings can help prevent moisture loss during storage of fresh meats, reduce lipid oxidation, minimize load of spoilage and pathogenic microorganism, restrict

38 odor pick-up, and reduce oil uptake in battered and breaded fried products. The most common nonprotein edible films and coatings are made up of lipids such as waxes and glycerides, and polysaccharides (Gennadios et al., 1997; Morillon, Debeaufort, Blond,

Capelle, & Voilley, 2002) or a combination of both (Coughlan, Shaw, J. F. Kerry, & J. P.

Kerry, 2004; Phan, Debeaufort, Luu, & Voiley, 2005).

While lipid-based films and coatings are not typically used as lipid barriers, polysaccharides are probably the most studied edible films for reduction of oil absorption. Garcia, Ferrero, Bértola, Martino, and Zaritzky (2002) did a study using methylcellulose (MC) and hydroxypropylmethylcellulose (HPMC) as an edible coating to reduce oil uptake in deep fried potato strips. They found that a mixture of MC with sorbitol was effective in reducing oil absorption by 40.6% compared to uncoated potato strips. Williams and Mittal (1999) compared the oil barrier properties of gellan gum, MC and HPMC and found that MC was most effective in reducing oil absorption (91% reduction). They also tested the effect of film thickness of gellan gum on moisture and fat content. They observed that moisture loss decreased as film thickness increased. Fat absorption was reduced overall compared to uncoated samples. However, the effect of film thickness was not apparent.

Alginates, extracted from seaweed, are salts of alginic acid and are linear co- polymers of D-mannuronic and L-guluronic acid monomers (Gennadios et al., 1997).

Unfortunately, alginates exhibit high water vapor permeability because they are hydrophilic polysaccharides and for practical purposes, are often mixed with calcium, which acts as a gelling agent. Krochta and de Mulder-Johnston (1997) studied the oil- barrier property of alginates mixed with calcium chloride (CaCl2) and found that it was

39 impervious to oils. Another extract of seaweed that could be used as an oil-barrier is carrageenan, a galactose polymer. Annapure et al. (1999) found that a 2% carrageenan coating for chick pea dough showed a 17% reduction in oil uptake. They ranked the ability for fat reduction of several non-protein hydrocolloids: gum arabic > carrageenan > gum karaya > guar gum > carboxymethylcellulose > hydroxypropylmethyl cellulose where hydroxypropylmethyl cellulose such as xanthan and locust bean gum hardly showed any reduction in fat absorption at all.

Gellan gum is a polysaccharide manufactured by microbial fermentation of the

Sphingomonas elodea microorganism. When used as a mixture with soy protein isolate, the film was found to reduce fat absorption of fried doughnut mix by 55% (Rayner et al.,

2000). Rayner et al. (2000) also did a texture analysis and found no significant difference between the coated and uncoated fried food. Bajaj and Singhal (2007) observed a reduction of fat uptake from 37% to 28% when batter is mixed with gellan gum at 0.25%

(w/w). William and Mittal (1999) prepared a coating made up of 2% gellan gum and

0.5% CaCl2. They observed a reduction of 60% when pastry flour mix product is coated

with this solution and deep-fried.

Nonprotein coatings have been used in industry longer than protein-based

coatings. An example of a commercial edible oil-barrier that is in the market is

Kelcogel® which is produced by CP Kelco (Decision News Media, 2004). This product

is made from gellan gum which is an FDA-approved stabilizer and thickener. When

mixed with mono- or divalent salts, they form films that have good barrier capabilities.

Other nonprotein based oil barriers that are in the market now are methylcellulose and

40 hydroxypropyl cellulose, manufactured by Dow Chemical Co. and Watson Foods, and they have been used to decrease oil absorption in french fries and onion rings.

Many studies have been conducted to compare the properties of different compositions of films and coatings (Bozdemir & Tutas, 2003; Coughlan et al., 2004;

Morillon et al., 2002). However, the requirements of edible films and coatings are the same regardless of material; they must provide a good barrier against oxygen, have good mechanical properties to allow various manipulations of the food product, and have low water-vapor permeability (Gennadios et al., 1997). In fried foods, edible films or coatings must not only fulfill all these requirements, but also are expected to have the ability to reduce fat uptake. A list of research on the effectiveness of hydrocolloids an oil-barrier in fried foods can be found in Table 1.

Table 1 (continued on page 42)

Examples of Nonprotein Ingredients Used in Formulations for Reduction of Oil

Absorption in Fried Foods

Fat Ingredient Usage Product reduction Reference (%) * Cellulose 2% in batter Fried falafel 8.5 Pinthus, Weinberg, balls and Saguy (1993)

Methylcellulose 1% in batter Fried falafel 46.4 Pinthus et al. (1993) (MC) balls

2% in batter Fried, breaded, 6.5 Holownia, Chinnan, and marinated Erickson, and chicken strips Mallikarjunan (2000)

1% MC + 0.75% Fried wheat 29.9 Suárez, Campañone, sorbitol coating dough balls García, and Zaritzky, solution (2008)

2% MC + 0.2% Fried mashed 83.6 Mallikarjunan, polyethylene potato balls Chinnan, glycol film Balasubramaniam, and Phillips (1997)

Carboxy- 0.25-2% in Deep-fat fried 9.8-3.0 Annapure et al. methylcellulose batter foods (1999)

0.25-1% in Fried papad 1.7-19.5 Patil, Singhal, and batter Kulkarni (2001)

2% in tortilla Tortilla chips 5-40 Esturk, Kayacier, and chips Singh (2000)

Hydroxypropyl- 0.25-2% in Deep-fat fried 12.6-7.8 Annapure et al. methylcellulose batter foods (1999) (HPMC) 2% in batter Fried breaded 4.8 Holownia et al. marinated (2000) chicken strips

0.25-1% in Fried papad 17.2-2.4 Patil et al. (2001) batter

2.2% HPMC + Fried chicken Up to Balasubramaniam, 0.5% balls 33.7 Chinnan, polyethylene Mallikarjunan, and glycol film Phillips (1997)

2% HPMC + Fried mashed 31.4 Malikarjunan et al. 0.2% potato balls (1997) polyethylene glycol film

Guar gum 0.25-2% in Deep-fat fried 9.7-8.2 Annapure et al. batter foods (1999)

0.25-1% in Fried papad 9.7-22.0 Patil et al. (2001) batter

Locust bean 0.25-2% in Deep-fat fried 3.4-4.8 Annapure et al. gum batter foods (1999)

Carrageenan 0.25-2% in Deep-fat fried 3.4-15.7 Annapure et al. batter foods (1999)

0.25-1% in Fried papad 2.8-12.0 Patil et al. (2001) batter

Xanthan gum 0.25-2% in Deep-fat fried 8.4-4.4 Annapure et al. batter foods (1999)

Note. *Fat reduction compared to control.

Protein-Based Coatings or Films

Coatings or films made from protein are good barriers to oxygen and lipid, but

they exhibit relatively high water vapor permeability values (Gennadios et al., 1997;

Ustunol & Mert, 2004). The limited resistance of protein films and coatings to water

vapor transmission is attributed to the hydrophilic nature of proteins and to substantial amounts of hydrophilic plasticizers, such as glycerin and sorbitol, incorporated into films to impart adequate flexibility (Gennadios et al., 1997; McHugh, 2000). However, moisture-barrier properties of protein films and coatings can be improved by incorporating hydrophobic materials such as lipids to produce protein-lipid emulsion films. When forming these films or coatings to be used as lipid barriers, it should be noted that excess protein may reduce crispness and increase oil uptake by emulsifying more oil and water into the fried product (Mohamed et al., 1998).

Corn zein and soy protein are examples of coating ingredients from plant sources that have been used as oil barriers. Malikarjunan et al. (1997) saw a 59% reduction in potato balls coated with corn zein solution (15% corn zein in ethanol). Another example of plant protein used as an edible film in inhibiting oil absorption is gluten (Gennadios,

Weller, & Testin, 1993; Park & Chinnan, 1995). Whey protein and other milk proteins, such as casein, are examples of coating ingredients that have been shown to have oil- barrier capabilities. Banerjee and Chen (1995) showed that whey protein concentrate gels had good water vapor barrier (better than that made from WPI and casein) and mechanical properties (stretchable), while addition of monoglycerides decreased water vapor permeability by 200% (Anker, Brensten, Hermansson, & Stading, 2002). In addition, the monoglyceride acted as a plasticizer, which created a desirable increase in the fracturability of the film. Dough balls that were dipped in soy protein isolate (SPI) and whey protein isolate (WPI) mixed solution and dried to create a layer of protein film surrounding the product were observed to have the highest fat uptake reduction (99.8%) compared to dough balls treated with other combinations of proteins including gelatin,

gellan gum, κ-carrageenan-konjac-blend, locust bean gum, methyl cellulose, microcrystalline cellulose, pectin, sodium caseinate, and vital wheat gluten. Purified whey protein adjusted to pH 7.0 using sodium hydroxide was observed to reduce fat uptake by 5% when applied on blanched potato chips before being deep-fried (Aminlari,

Ramezani, & Khalili, 2005). Other sources of animal protein-based coatings are gelatin

(Arvanitoyannis, Psomiadou, Nakayama, Aiba, & Yamamoto, 1997) and egg albumin

(Gennadios et al., 1996; Handa, Gennadios, Hanna, Weller, & Kuroda, 1999). Some examples of protein films or coatings that were used as lipid barriers in fried foods as found in literature are listed in Table 2.

Table 2

Examples of Protein Ingredients Used in Formulations in Reduction of Oil Absorption in

Fried Foods

Fat reduction Coating Formulation Food model Reference (%)* Soy 10% soy protein + Fried doughnut 55.12 Rayner et al. protein 0.05% gellan gum disks (2000) film

Corn 14% corn zein + Fried mashed 59 Mallikarjunan et Zein 2.8% glycerin film potato balls al. (1997)

Casein 3% sodium caseinate Potato chips 13.6 Aminlari et al. protein film (2005)

Whey 3% whey protein Potato chips 4.8 Aminlari et al. protein film (2005)

Albumin 3% albumin film Potato chips 12.1 Aminlari et al. (2005) Note. *Fat reduction compared to control.

Protein Gelation

In order for a protein to undergo gelation, it must first go through denaturation.

Denaturation is used to describe a reversible or irreversible change of molecular structure of a protein without cleavage of covalent bonds except for the disulfide bridges (Belitz &

Grosch, 1999). This can be initiated by changes in temperature and pH, increases in the interface area, or the addition of organic solvents, salts, urea, guanidine hydrochloride or detergents (Belitz & Grosch, 1999). During denaturation, side chains of the amino acid are exposed and undergo intermolecular interactions. They form small spherical aggregates which combine into linear strands that interact to create a gel network

(Fitzsimons, Mulvihill, & Morris, 2007). If the unfolding of the peptide chain is stabilized by interaction with other chains such as interactions between exposed reactive groups (e.g., thiol groups), the denaturation process would be irreversible (Belitz &

Grosch, 1999). The ratio of the rate of denaturation with the rate of aggregation determines the gel characteristics (Belitz & Grosch, 1999). If the rate of aggregation is slower in comparison with the rate of denaturation, the resulting gel will be a finer network of protein chains, less opaque, and less capable of holding on to water due to smaller voids between molecules (Gossett, Rizvi, & Baker, 1984).

Factors affecting protein gelation. Gelation of protein is affected by factors that influence the rate of denaturation and aggregation such as temperature, concentration of protein, pH and ionic strength (Belitz & Grosch, 1999). Temperature and heating rate can affect both the rate of denaturation and the rate of protein-protein interaction (Belitz &

Grosch, 1999). The temperature above which a gel will not form is known as the critical gelation temperature, which is directly proportional to heating rate (Li, Ould Eleya, &

Gunasekaran, 2006; Tosh & Marangoni, 2004). While there is a critical (maximum) gelation temperature, there is no indication for the existence of a minimal temperature of gelation (Le Bon, Nicolai, & Durand, 1999). However, below critical gelation temperatures, the time required for the formation of the gel will decrease (Tosh &

Marangoni, 2004). In the case of forming gels for coatings, the aggregation rate should be slower than the unfolding rate to avoid forming coarse and unstructured gels (Belitz &

Grosch, 1999).

Besides heat, protein concentration also determines the characteristics of the gel and the likelihood of it being formed. Generally, the elastic properties of the gel network vary depending on protein concentration (Puyol, Perez, & Horne, 2001). When the level of protein is too low, a protein network is difficult to establish because protein-protein interactions tend to occur within molecules rather than between molecules (Belitz &

Grosch, 1999). As the protein content increases, the likelihood of intermolecular crosslinks increases and gelation is more likely to occur. The critical protein concentration for gelation is dependent on pH and ionic strength. For example, the critical concentration for β-lac is a minimum at the isoelectric point (5.1) and is independent of ionic strength. At other pH values, the critical protein concentration varies inversely with ionic strength (Puyol et al., 2001).

As mentioned before, pH can have a marked effect on the structure of proteins. At pH levels far away from the isoelectric point, the protein is highly charged (Belitz &

Grosch, 1999). Hence, the association or aggregation of the protein molecules is difficult to achieve due to electrostatic repulsion even upon heating. However, aggregation and gelation can occur at high enough protein concentrations and/or high ionic strength

(Schokker, Singh, Pinder, & Creamer, 2000). If the pH is adjusted towards the isoelectric point, the charge on the protein molecules will be reduced, promoting aggregation

(Fitzsimons et al., 2007). However, these aggregates are colloidally stable at ambient temperature; thus, formation of gels at the isoelectric point can only be achieved when heated (Ju & Kilara, 1998). Under conditions of strong electrostatic repulsion (away from

isoelectric point), whey protein gels are transparent, and have a fine-stranded structure. In

conditions of weak electrostatic repulsion, whey protein gels become more opaque and

coarser with bigger pores (Puyol et al., 2001). Differences in gel structure not only are influenced by electrostatic interactions, but also may be due to the dependence of rate of denaturation and aggregation on pH level. Thus, pH of the protein solution should be adjusted to achieve proper balance between the rate of denaturation and aggregation that

is needed for gel formation (Belitz & Grosch, 1999).

Salts in general can affect the structure of protein molecules as well as the nature

of protein-water interactions by changing the ionic strength of the protein solution

(Chantrapornchai & McClements, 2002). These effects influence both the solubility of

protein and their rate of thermal denaturation. As with temperature and concentration,

there is generally an optimal level of salt that favors gel formation (Belitz & Grosch,

1999). In their study on the effect of salt concentration in heat induced whey protein gels,

Chantrapornchai and McClements (2002) found that at neutral pH (pH 7), the gels went

from fairly fine grained and homogeneous to coarse grained and porous with an increase

in salt concentration. The gels formed at high salt concentrations exhibited increased

water loss. The lack of gelation at low salt concentration at neutral pH may be due to the

high electrostatic repulsion between highly negatively charged molecules (Bryant &

McClements, 1998). The addition of salt increases the ionic strength of the solution which promotes interaction of the charged protein molecules through charge shielding

(Belitz & Grosch, 1999). However, salt only has a significant effect on properties of protein gels when it is added prior to heating (Boye, Alli, & Ismail, 1996; Foegeding,

Bowland, & Hardin, 1995; Langton & Hermansson, 1992).

Whey Protein

Whey is the watery portion of milk remaining after milk coagulation and removal of the curd. Whey can be obtained by acid, heat, and rennet coagulation of milk. There are two kinds of whey: Sweet whey and acid whey. Sweet whey is manufactured during making of rennet type hard cheese like cheddar or Swiss cheese and has a pH level of more than 5.6. On the other hand, acid whey or sour whey is obtained during making of acid type of cheese such as cottage cheese and has a pH level of less than 5.1. Both types

of whey contain about 0.7 to 0.8% protein on a liquid basis (Dairy Management, Inc.,

2005). On average, about 90% of the milk used for cheesemaking ends up as whey (S.

Bhattacharjee, C. Bhattacharjee, & Datta, 2006,). It is estimated that the world production

of whey is about 104 billion kg per year with the USA producing about 30 billion kg per

year (Saddoud, Hassairi, & Sayadi, 2007; Cheryan, 1998). However, about 30 to 47% of the total amount of whey available worldwide is not being used (Hutchinson, Balagtas,

Krochta, & Sumner, 2003; Saddoud et al., 2007). Using whey is difficult because it has low solid content (Bhattacharjee et al., 2006; Cheryan, 1998) while its high biological oxygen demand (around 32,000 to 60,000 ppm) creates a severe disposal problem

(Cheryan, 1998). Because whey has to be treated before being released into the environment, this adds an extra burden on waste treatment plants and requires the cheese

industry to build additional treatment facilities (Tuchenhagen South Africa, Ltd., n.d.).

Thus, there is a push to decrease the amount of whey that has to be disposed of by increasing its utilization.

Whey protein consists of a number of individual protein components. The two most abundant proteins are β-lac (50-55%) and α-lac (20-25%). β-Lac has a molar mass

of 18.3 kDa and diameter of about 2 nm. The isoelectric point is 5.1 and the denaturation

temperature is 78 °C (Sagis, Ganzevles, Ramaekers, Bolder, & van der Linden, 2002). β-

Lac is largely responsible for solubility, gelation, foaming, emulsification, and flavor

binding of whey protein. Because β-lac is the most abundant protein in whey, it has been

suggested to be one of the main determinants of the properties of whey protein gels (Van

Vliet et al., 2004).

Gelation of Whey Protein

As mentioned before, whey protein coatings act as oil barriers by forming a gel

coating around the food. The characteristic of the gel that is formed determines the ability

of the whey protein film to reduce the oil uptake of fried foods. Gelation of whey protein

gels can be affected by numerous factors such as temperature, concentration of protein,

pH, and ionic strength (Belitz & Grosch, 1999). Gelation of whey proteins is the result of

an aggregation process that occurs through adhesion of exposed hydrophobic regions to

form aggregates that are then stabilized, at least above a certain temperature threshold, by

intermolecular disulfide exchange (Renard & Lefévre, 1992). β-Lac forms gels when the

protein is dissolved in an aqueous solution and heated above the denaturation

temperature. The formation of intermediate aggregates by β-lac involves two broad types

of bonding: Covalent and noncovalent bonding (Galani & Apenten, 1999). β-Lac

contains two disulfide bridges and a free thiol or sulfhydryl group (-SH group). At room temperature, β-lac exists mostly as a dimer although it may dissociate into monomers at higher temperatures (Hoffmann & van Mill, 1997). The polymerization reaction leading to gelation of β-lac is initiated by the dissociation of the dimers through heating.

According to Iametti, de Gregori, Vecchio, and Bonomi (1996), the protein unfolds, resulting in a molten-globule-like structure, with increased exposure of the previously buried inner hydrophobic groups and the thiol group. Hydrophobic interactions between the exposed groups can cause aggregation of the protein molecules while still in the molten-globule state. The thiol group in the modified monomer is capable of building oligomers by disulfide bond switch with one of the two disulfide bridges in β-lac, leading to the formation of disulfide-linked aggregates (Bauer, Carotta, Rischel, & Øgendal,

2000; Iametti et al., 1996). The exposed -SH group initiates sulfhydryl/disulfide (SH/S-S) interchange reactions, leading to irreversible aggregation/polymerization (Galani &

Apenten, 1999). It is generally accepted that these thiol/disulfide exchange reactions, leading to the formation of intermolecular disulfide bonds, play a role in the heat-induced aggregation and gelation of β-lac

(Hoffmann & van Mill, 1997).

Apart from the disulfide cross-linked aggregates, noncovalently driven association occurs within the aggregates (Galani & Apenten, 1999). These noncovalent interactions include hydrophobic, ionic, and hydrogen bonding and other weak interactions that also contribute to the formation of aggregates and a gel network

(Hoffmann & van Mill, 1997). Furthermore, these noncovalent bonds are believed to be

responsible for aggregate formation (Bauer et al., 2000; Manderson, Hardmann, &

Creamer, 1998). The relative contribution of noncovalent interactions to the overall β-lac aggregation mechanism varies with initial protein concentration, temperature, and pH

(Galani & Apenten, 1999; Verheul & Roefs, 1998).

Effect of pH on whey protein gelation and oil uptake. The reactivity of the free thiol, exposed after denaturation, has attracted a number of experiments over the years

(Raso, et al., 2005). The reactivity is pH dependent, being unreactive at low pH but becoming much more reactive at pH levels that are above the isoelectric point (Sawyer &

Kontopidis, 2000; Verheul & Roefs, 1998). At low pH, the thiol groups are buried in the

β-lac dimers, presumably in the region of contact between the monomer subunits

(Hoffmann & van Mill, 1997). Due to the limited exposure of the reactive thiol group,

aggregation and gelation is more dependent on protein concentration, ionic strength, and

heating temperatures at low pH levels compared to those above the isoelectric point

(Schokker et al., 2000). On the other hand, at higher pH, the protein unfolds, exposing the thiol group and thus increasing its reactivity. A large number of dimers, trimers, and tetramers appear due to the increase in the number of reactive intermediates with an

exposed, reactive thiol group. This leads to the increased probability of termination of

reactive sulfhydryl groups resulting in the formation of smaller disulfide-linked β-lac aggregates without a reactive thiol group (Roefs & de Kruif, 1994; Verheul & Roefs,

1998). The large aggregates observed at pH slightly higher than the isoelectric point may be formed by secondary, noncovalent interactions of primary, disulfide-linked aggregates

(Hoffmann & van Mill, 1997). In their study on the microstructure of gels formed by β- lac, Boye, Ma, Ismail, Harwalkar, and Kalab (1997) observed that the compact protein globules formed at acidic pH of 3 and 5 were either loosely aggregated or fused into

chains or clusters of several globules. Aggregation of protein into these structures affected the initially uniform distribution of proteins and created large voids that were filled with the liquid phase of the gel. Gels produced under neutral (pH 7.0) and alkaline

(pH 8.6) conditions had microstructural features completely different from those of the gels made under acidic conditions; the proteins were more evenly distributed in these gels than in the gels made at lower pH. Since their clusters were connected through narrow bridges, void spaces between the clusters were considerably smaller in these gels than in the acidic gels. A summary of the effects of pH on whey protein gel characteristics based on studies found in the literature is shown in Table 3.

Table 3 (continued on page 53)

Effects of pH Level Variations on Whey Protein Gelation

pH level Effect on whey protein gelation References pH 2 • No aggregates were formed when the protein Kavanagh, Clark, dispersion was left unheated and Ross- • Upon heating, rod-like structures formed Murphy (2000) • The rods were thought to occur when disulfide exchange was inhibited by repulsive electrostatic interactions

pH 2.5 • Mixture of short and long linear aggregates was Kavanagh et al. formed. (2000)

pH 3 • Loosely aggregated or fused into chains or clusters of Boye et al. several globules (1997) • Large spaces void of proteins which were filled with the liquid phase of the gel • Aggregates formed at pH 3 are smaller than that of pH 5 • β-lac was shown to form octamers

pH 4 • Coarse aggregates were uniformly distributed in ‘fine- Kavanagh et al.

stranded’ network (2000) • β-lac was shown to form octamers pH 4.5 • Coarse, uneven aggregates are formed Kavanagh et al. • β-lac was shown to form octamers (2000) pH 5 • Loosely aggregated or fused into chains or clusters of Boye et al. several globules (1997) • Large spaces void of proteins filled with the liquid phase of the gel • Aggregates formed at pH 5 are larger than that of pH 3 pH 7 • Proteins were more evenly distributed than in the gels Boye et al. made at lower pH. (1997) • Void spaces were considerably smaller in these gels than in the acidic gels. • Have better water holding capacity than the acidic gels. • Aggregates formed were in the nanometer region (aggregates formed at pH 3-5 are in micrometer) pH 8-8.6 • Proteins were more evenly distributed in these gels Boye et al. than in the gels made at lower pH. (1997) • Void spaces were considerably smaller in these gels than in the acidic gels. Hoffmann and • Have better water holding capacity than the acidic van Mill (1997) gels. • The aggregates formed were in the nanometer region (aggregates formed at pH 3-5 are in micrometer) • Aggregates formed at pH 8 are purely made up of thiol/disulfide bonds • Thiol reactivity at pH 8 was reported to be as high as 90% and reacts even at room temperature. Some thiol groups have disappeared via oxidation and thiol/disulfide exchange reactions and are involved in intermolecular disulfide bonds

Note. β-lac = β-lactoglobulin.

Effect of concentration on whey protein gelation and oil uptake. Because it is believed that inhibition of oil absorption in fried food is dependent on gel formation of

whey protein isolates, the understanding of the dependence of gel structure on the initial protein concentration of the coating is crucial in developing this coating. All proteins gel above a minimum concentration known as the critical gelation concentration (Kavanagh et al., 2000; Mleko, 1999; Renard & Lefévre, 1992; Vardhanabhuti & Foegeding, 1999).

Preparation of whey protein isolate coatings should be done at or above the critical gelation concentration to ensure the formation of gels. According to a study done by

Renard and Lefévre (1992), the critical gelation concentration remains at around 1%

(w/w) irrespective of ionic strength at the isoelectric point of β-lac (pH 5.1). The value increases the more the pH is displaced from the isoelectric point of the protein. At extreme pH values of pH 2 and 9, Renard and Lefévre (1992) found that the critical

concentration was as high as 8% (w/w). Similar results were found by Kavanagh et al.

who observed that the critical concentration was as low as 1% between pH 4.5 and 5.6

and the critical concentration increased to 5% at lower pH values of 2.3 and 3 and

increased further to 10% when the pH was 7. In a study done by Mleko (1999), it was shown that the increase in protein concentration that was heated up to produce a heat-set gel led to a rise in the gel permeability coefficient. This increase suggests that a higher protein concentration increased the size of the aggregates which formed a gel matrix with a larger pore size. Ju and Kilara (1998) observed that increasing whey protein concentration (1–9%) increased the size and amount of the aggregates and resulted in whey protein polymers that are larger and/or have more asymmetrical shapes

(Vardhanabhuti & Foegeding, 1999). In short, higher protein concentration not only increased the size and number of the aggregates formed but also increased the pore size

of the gel (Verheul & Roefs, 1998), thereby affecting the gel characteristics of the whey protein coating and determining its effectiveness as a lipid barrier.

Sensory Evaluation of Foods

The main role of sensory analysis is to obtain information on the sensory characteristics of the product. In the development of new food products, this process is important because it affects the decisions involved in materialization of the final product and its success. Up to 80-90% of new food products fail within a year of production, resulting in an estimated loss of up to 20 billion dollars. Much of it is due to a flawed process of product development which includes lack of sensory analysis (Moskowitz,

Beckley, & Resurreccion, 2006). Sensory testing of foods can be grouped into two general categories: The first category measures sensory responses to the product and these include discrimination, acceptance, and preference testing; the second category focuses on characterizing the product by using descriptive analysis testing which may be accompanied by physicochemical measurements or objective measurements of identified attributes (Moskowitz et al., 2006). Consumer testing, which looks at the consumer acceptance of the product, may or may not employ untrained panelists. For example, acceptance tests would employ consumers of the particular product while discrimination testing includes both users and nonusers of the product (Stone & Sidel, 1985). Either way, these tests require a large number of participants, which may range from 25 to 100 or more (Moskowitz et al., 2006; Stone & Sidel, 1985). On the other hand, descriptive sensory analysis involves rigorous training of a smaller panel of about 10 to 12 selected consumers (Stone & Sidel, 1985). The panel is trained to generate a lexicon or list of

attributes that are important characteristics of the product, and is required to evaluate these attributes with an analytical and objective approach (Moskowitz et al., 2006).

Objective or physicochemical analyses rely on analytical measurements of

components that are believed to make up the sensory characteristics. For example, one

method of characterizing color is by using the CIELAB system where three components

of color are used as specified by the Commission Internationale d'Eclairage (CIE) or

International Commission of Illumination. This model is based on the color perception of

92% of the population that does not have vision deficiencies (Hutchings, 1999;

Hutchings, Luo, & Ji, 2002). The whiteness or blackness is represented by L*, redness is

represented by +a*, greenness is represented by –a*, yellowness by +b*, and blueness by

–b* (Jones, 1943) as shown in Figure 2. As for crunchiness, many methods have been

suggested to measure and characterize this property. These include a combination of

mechanical and acoustic measurements (Vickers, 1987), acoustical and force-deformation

measurements, and employment of ultrasonic techniques (Antonova, Mallikarjunan, &

Duncan, 2003).

Figure 2. The CIELAB color space representing the three color coordinates L*, a*, and b*. ______Note. From Konica Minolta Sensing USA. (2005, June 27). Precise Color Communications. In Educational Booklet on Color Communication (p. 19). Retrieved May 12, 2008, from http://se.konicaminolta.us/support/product_applications/pdf/colorcommunications_app.p df. Reprinted with permission.

Conclusion

Oil absorption in deep-fat frying is a complex process that involves various factors related to the food, such as composition and surface characteristics, and factors related to the frying process such as frying medium, temperature, time, and postfrying handling.

The oil taken up by battered and breaded fried foods during immersion frying can occur during the frying process via water replacement or immediately after frying due to the cooling phase effect. Throughout the process, the degree of oil absorption is affected by the continuous degradation of the frying oil, as described by the surfactant theory. A variety of ingredients have been employed that retard oil absorption to varying degrees during immersion frying of battered and breaded fried foods. The majority of these ingredients are film-forming or aqueous solutions of proteins or nonprotein hydrocolloids that are either added to the batter or breading, or applied as a postbreading dip.

Whey protein is a potential gel-forming ingredient that can be used as a lipid barrier in fried foods. It is composed of various proteins, the main protein of which is β-lac, a

globular protein with an isoelectric point of 5.1. This protein makes up 50-55% of the

total protein contained in whey. Other important globular proteins of whey are α-lac and

BSA. All of these proteins, particularly β-lac, can form thermally-induced gels that alter

the porosity of the product, thus lowering both moisture loss due to evaporation and

subsequent oil absorption into the fried food when used as a coating (Dogan et al., 2005,

Van Vliet et al., 2004). The usage of whey protein as food coatings has long been

investigated and applied (Gennadios et al., 1997, Mellema 2003). However, the extent of

commercial application of whey protein coatings as oil barriers has been limited to

separating oil-rich products, such as nuts, from other components of heterogeneous foods

such as cereal (Haines, 2004).

The use of WPI to reduce oil absorption in fried foods, not as a film that requires a

setting time before frying but rather as a postbreading dip after which the product can be

immediately fried, requires optimization of various factors such as pH level and WPI

concentration. In addition, the final product should be nutritious and appealing to the senses. Since temperature, concentration of protein, pH and ionic strength have a

significant effect on the gelation of whey protein (Belitz & Grosch, 1999), it would be

beneficial to study the effects of these factors on the ability of whey protein isolate

coatings to reduce fat uptake in fried foods. From an industrial standpoint, the whey

protein isolate coating must produce at least a 25% fat reduction compared to the original

reference to meet the labeling criteria for reduced-fat. In addition, the product has to be

deemed acceptable by consumers and this involves the challenge or retaining the

appearance, texture, and flavor of the product.

CHAPTER 3: METHODOLOGY

Overview of Approach

Samples were prepared using the standard commercial breading procedure and treated with whey protein isolate (WPI) solutions of different concentrations and pH levels. A 3 x 4 x 2 full factorial design was constructed with three factors, pH level (2, 3,

8), whey protein solution concentration (0%, 2.5%, 5%, 10%), and breading

(crackermeal, Japanese breadcrumb). The complete design was replicated three times.

All samples were then tested for lipid content, moisture content, and texture attributes.

Finally,a descriptive sensory analysis was conducted on the samples. The lipid and moisture content of the samples determined the effectiveness of the WPI coating in reducing oil uptake and the effect on moisture content. Instrumental texture analyses were used in relation to the sensory analysis to determine the effect of the treatment on appearance, texture, mouth feel, and flavor. The results were then analyzed using analysis of variance and/or covariance and post hoc means separation was achieved using

Duncan’s multiple range test. Pearson correlation analysis was performed wherever necessary. A flowchart of the methodology is shown in Figure 3 and the timeline of this study is shown in Table 4.

This research was funded by the National Dairy Council Discovery Pilot Program, and the proposal that was approved by the National Dairy Council was written based on a previous study that looked at the efficacy of WPI, soy protein, and egg albumin as a postbreading dip in reducing oil absorption in battered and breaded fried chicken patties

(Brannan & Teyke, 2006). It was observed that 10% WPI solutions made at pH 6.1, 5, and 3, reduced oil absorption by 20.8%, 1.6%, and 68.4%, respectively. The current study

looked at the efficacy of WPI when used at a lower pH level (i.e., pH 2) and at a higher pH level (i.e., pH 8) compared to the pH levels in the previous study. Treatment was repeated at pH 3 since it showed the highest lipid reduction in the previous study

(Brannan & Teyke, 2006). In addition, the effect of WPI concentration variations which was not examined in the previous study was investigated here. Again, WPI at 10% concentration was repeated in the current study because it was observed to produce lipid reduction in the previous study.

62

Whey protein Mixing Whey protein solution Three pH levels (pH 2, 3, 8) Crackermeal Four WPI concentration (0%, 2.5%, 5%, 10%) or Japanese breadcrumbs

Deionized water

48.75% flour Lipid analysis Weighing 1% xanthan gum Mixing Dipping Moisture analysis 48.75% corn flour Battering Breading Deep-frying Texture analysis 1% baking powder Weighing At 191°C to Weighing Weighing internal T of Weighing 0.5% salt 74°C Color analysis Chicken Grinding Forming Holding Oil degradation Sensory analysis 20 g 2-in ≤ 24 hrs at - measuring diameter patties 18°C Frying oil

Figure 3. Flowchart of methodology for optimization of whey protein isolate (WPI) solution in reducing oil absorption in batter and breaded fried chicken when used as a postbreading dip.

Table 4

Timeline for Thesis

Feb Mar Apr May Jun July Aug Sept Oct Nov Dec Jan Feb Mar Apr May Sample preparation

Lipid and moisture analysis

Instrumental texture and color analysis

Sensory recruitment and training

Sensory sampling

Data analysis

Thesis proposal defense

Thesis defense

Data Collection and Analysis

Materials

All chemicals used in the objective analyses, which included chloroform, methanol, and sodium chloride, were obtained from Thermo/Fisher (Waltham, MA). Food ingredients such as chicken breast, batter ingredients, distilled water, and sensory references during training and sampling were purchased from local retailers. Japanese breadcrumbs, crackermeal, and frying oil were purchased from Foodservicedirect Inc. (Hampton, VA).

WPI was given by Volac International Ltd. (Orwell, UK) and sodium bisulfate (pHase®) was given by Jones-Hamilton Co. (Walbridge, OH).

Preparation of Deep Fried Samples

Fresh chicken breast was washed and cut into approximately 4 cm cubes after all visible fat was removed. The meat was then ground once using a stand mixer with a food grinder attachment with coarse grinding plate (model K45SS/250W, KitchenAid®,

Whirpool Corporation, MI). The ground chicken was formed into 2-in diameter patties using a mold and each patty weighed approximately 20 ± 2 g. Due to the large number of patties needed for this study, patties were stored frozen (-18 °C) for up to 24 hrs before being coated.

The standard batter formulation was based on the work of Sahin et al. (2005) and consisted of 48.75% (w/w) wheat flour, 48.75% corn flour, 1.0% xanthan gum, 1.0% salt,

0.5% baking powder and deionized water. The ratio of dry ingredients to water is adjusted to a viscosity of 3062.5 centipoise (cps). The batter was made fresh on the day it was used and stored refrigerated (4 °C). WPI solutions (0%, 2.5%, 5%, and 10%) were prepared by mixing WPI with distilled water. The pH of the solution (pH 2, 3, and 8) was

adjusted using a low flavor impact, food grade acidulant (sodium bisulfate) or baking soda (sodium bicarbonate). All solutions were prepared 24 hrs in advance, stored refrigerated (4 °C), and allowed to come to room temperature before use. Patties (in groups of four or five) were weighed and dipped into batter with the excess shaken off and weighed again before being coated with breading and weighed. Treated patties were dipped in WPI solution, weighed, and immediately fried in 191 °C frying oil (canola oil with added dimethylpolysiloxane) in a deep fryer (Presto® Dual ProFryTM/1800W,

National Presto Industries Inc., WI) until an internal temperature of 74 °C was achieved.

Control patties were not dipped in WPI solution and were directly placed in the frying oil.

After frying, all patties were weighed again and allowed to cool to room temperature for

30 mins before being placed in labeled freezer bags. They were then stored frozen (-18

°C) until analyzed.

Objective Analysis

Oil degradation. Total polar material (TPM) in the frying oil was monitored

throughout the process. An oil sample was taken from the deep fryer before and after

frying of each treatment (12-15 patties per fryer). The samples (1 ml) were microfiltered

(0.45 μm) and equilibrated to 60 °C before the absorbance (ABS) at 490 nm was read.

TPM was calculated using Equation 2 (Xu, 2000):

TPM = -2.7865(ABS)2 + 23.782(ABS) + 1.0309 (2)

Lipid analysis. Analysis on the lipid content of deep-fried samples was accomplished according to a modified Folch method (Folch, Lees, & Sloane Stanley,

1957). A sample was selected randomly from a treatment and finely ground using a food processor (Osterizer ®, Jarden Corporation, Boca Raton, FL) until a homogeneous

mixture of coating and chicken was obtained. Lipid from the sample (1 g) was extracted with three successive washings of chloroform: methanol (2:1 v/v) at 10, 5 and 5 ml respectively. A volume of 7.5 ml 0.5% NaCl solution was added to assist in separation of the aqueous layer. The supernatant was removed and evaporated to dryness and the purified lipid was measured to calculate the final lipid content. The final lipid content

(wet basis) was calculated using Equation 3:

lipid purified lipid weight (g) (%)content lipid (%)content = × %100 (3) sample initial sample weight (g)

Lipid extractions were duplicated for each patty and a total of three patties were analyzed from each treatment, including the control (n = 6). This procedure was repeated for each of the different breading systems (Japanese breadcrumbs and crackermeal).

Moisture analysis. Moisture of deep fried samples was determined by oven drying. Finely ground samples were weighed (5 g) into tubes and placed in an incubator at 80 °C. The weights of the samples were measured once a day until a constant weight was obtained. The moisture content was calculated using Equation 4:

sample final sample weight initial-(g) sample weight (g) (%)content moisture (%)content = × %100 (4) sample initial sample weight (g)

As with lipid analysis, moisture analysis was duplicated for each patty and a total of three patties were analyzed from each treatment, including the control (n = 6), for both breading systems.

Texture and color analysis. Penetrometry tests using a Ta-XT2i Texture

Analyzer (Texture Technologies Corp., Scarsdale NY/Stable Micro Systems, Godalming,

Surrey UK) were used to evaluate the texture of the samples. Instrumental texture measurements were made using a 70-mm knife-blade probe on a solid flat platform at a

crosshead speed of 10 mm/s. The depth of penetration of the probe was set at 10 mm to ensure that the probe fully penetrated the coating system into the substrate matrix.

Samples were positioned such that only 25 mm (1 in) of the sample measured from the edge was penetrated by the probe surface. This was to assure that only a single edge of the crust was penetrated uniformly across samples of different shapes. The Texture

Analyzer was controlled via Texture Expert Software and this package was used to record data and generate force-determination curves. The textural attributes that were determined were (a) crust fracture work, (b) crust fracture force, (c) total work, (d) hardness, and (e) resistance (Brannan, 2008). Duplicate readings were taken from one patty and a total of five patties were measured for each treatment for each breading system. The thickness of the coating was determined by using calibrated calipers. Each sample was cut in half through the vertical axis and the height of the upper crust was measured at three separate points on the cross-section. The average of the three readings was taken as the thickness of the crust. The CIE L*, a*, and b* values for color was measured using a Konica BC-10 (Konica Minolta Sensing Americas Inc., Ramsey, NJ).

Three readings were obtained from three different positions on the surface of the patties and the average of these readings was taken as the color of the samples. A total of five patties were evaluated for crust thickness and color for each treatment for each breading system. All instrumental texture analysis, crust thickness, and color measurements were performed on samples that were reheated in a 191 °C oven until an internal temperature of 74 °C was reached.

Sensory Analysis

Descriptive sensory analysis was performed to obtain information on the appearance, mouthfeel sensation, texture, and flavor of the samples. Approval from the

Ohio University Institutional Review Board was obtained for the use of human subjects prior to the start of the sensory analysis (see Appendix A). The timeline for all sensory sessions including informational sessions, training sessions, and sampling sessions is listed in Appendix B.

Panelist selection and training. Recruitment for the sensory panel was done through five informational sessions. During these sessions, potential panelists were given a basic introduction to sensory testing, specifically descriptive sensory testing. They were

then asked to fill in a questionnaire asking about known food allergies, food preferences,

medical conditions related to diet, and availability (see Appendix C). Panelists (n = 6; 3

females, 3 males) were chosen based on their answers on the questionnaire. Once they

were chosen, they were asked to fill in an informed consent form (see Appendix D) as

required by the Institutional Review Board of Ohio University.

Panelist training and subsequent testing of samples followed the SpectrumTM

Method (Meilgaard, Civille, & Carr, 1999). Panelists participated in 17, 50-min training sessions over the period of six months during which training in identifying and rating various attributes pertaining to flavor and texture were completed. During the first two training sessions, panelists were trained to identify and discriminate between the five basic tastes; sweet, sour, salty, bitter, and umami (see Appendixes E and F). Next, panelists were introduced to methods of lexicon development (see Appendix G). The next

14 training sessions focused on introducing sensory definitions and techniques for

different texture attributes, including hardness, crispness, and juiciness (see Table 4) in addition to continued practice in the generation of lexicon and identification of basic tastes. Panelists were also introduced to standard hardness, crispness, and juiciness scale

(see Appendixes H, I, and J). Instructions and scoresheets used throughout these eight training sessions can be found in Appendixes K through S.

Six of the training sessions were dedicated to sample lexicon development and fine-tuning for the samples. The first three sessions focused on introducing the concept of lexicon development to the panel which included discriminating and defining attributes, and standardizing techniques to identify them. In the next two sessions involving lexicon

development, the panel was asked to identify attributes related to cooked ground chicken

(see Appendix T) and was introduced to using a 15-cm line scale (see Appendix U).

Finally, the panel was asked to identify attributes that pertained to deep-fried, battered,

and breaded chicken patties. Identification of the preliminary attributes was done based on commercial fried chicken products (Banquet Chicken Breast Nuggets® and Banquet

Chicken Breast Patties®) and fine-tuning of these attributes was done on chicken patties that were prepared in a similar manner to the crackermeal undipped controls later utilized in this study. The final lexicon for the samples is shown in Appendix V.

Following the training sessions, two sessions were used to calibrate the panel against these sensory attributes for crackermeal-coated patties (CMP). A warm-up sample, which was a battered and breaded patty (with crackermeal) without WPI solution was prepared in advance and was used as one of the anchors of the 15-cm line scale for

CMP sensory analysis. Panelists participated in another calibration session for Japanese breadcrumb-coated patties (JBP) after sampling of CMP and before sampling of JBP.

During this calibration session, the same attributes were used, but panelists were required to re-anchor the new reference sample which was a battered and breaded patty (with

Japanese breadcrumbs) without WPI solution. After each of the calibration sessions for

CMP and JBP, a scoresheet for sampling was generated for each of the breading systems and these are shown in Appendix W for CMP and X for JBP.

Evaluation of samples. A total of three sampling sessions were held for CMP and a total of five sampling sessions were held for JBP. In each sampling session, panelists were supplied with all the anchored references for each attribute that were marked on the scoresheets (see Appendix W and X). They were also supplied with sterilized water, a spit-cup, napkins, and unsalted crackers. Each panelist performed independent evaluations, rating a total of six samples for the various sensory attributes. Panelists were seated apart from each other in a room under fluorescent light. At the start of the sampling session, each panelist was given a warm-up sample which was removed before the real sampling began. Samples were served to each panelist at random, in paper plates coded with 3-digit random numbers (see Appendix Y and Z). A 5-min break was given after the first three samples to minimize fatigue and to allow the experimenter to reheat the next three samples. All samples were reheated in an oven at 191 °C to an internal temperature of 74 °C and held at that temperature throughout the sampling session.

Statistical Analysis

Data obtained from the study were analyzed using the Statistical Package for the

Social Sciences program (SPSS, version 14.0. 2005, SPSS Inc., Chicago, IL). For each replication, means for lipid and moisture content were generated from three patties, with duplicate measurements performed on each patty (n = 6). Means for color, crust

thickness, and texture data were generated from five patties with duplicate measurements taken for each patty for texture and averaged to give one value for each patty (n = 5). For color and crust thickness, three measurements were taken for each patty for color and crust thickness and averaged to give one value per patty (n = 5). Analysis of variance was used to analyze differences between treatments and post hoc means separation was achieved using Duncan’s Multiple Range test. Any suspected covariants were analyzed for significance using analysis of covariance (ANCOVA). Pearson correlation was also used wherever necessary. Significant differences were determined at the confidence level of p < 0.05.

To identify outliers, boxplots for all the measured variables were generated within a breading system, replication, pH, and WPI concentration using Explore on SPSS (e.g., lipid content vs. crackermeal × replication 1 × pH 2 × 10% WPI). Values that are not located within the top, bottom, or interquartile range were identified as outliers. All outliers that were identified for lipid and moisture content were removed before further analysis. Outliers within the sensory and instrumental measurements involving organoleptic properties (appearance, texture, tastes, and flavors) were removed according to the discretion of the author, due to the smaller sample size for each variable (n = 6 for sensory analysis data, n = 10 for instrumental texture data, and n = 5 for instrumental color data and crust thickness measurement). For further statistical analysis, pairwise deletion of data was used to handle missing data to minimize the loss of data for each variable.

CHAPTER 4: RESULTS

Oil Degradation

Degradation of the frying medium throughout the frying process, expressed as

total polar material (TPM), is caused by hydrolytic reactions of triglycerides resulting in

the formation of diglycerides, monoglycerides, free fatty acids, and glycerol (O’Brien,

1998). The degradation of the frying medium as reflected in the changes in TPM

throughout the frying process is shown in Figure 4.

30

25

20 Crackermeal

15 Japanese breadcrumb Recommended maximum 10 Total polar material (%) 5

0 0 20 40 60 80 20 40 60 80 100 120 100 120 140 change Number of patties fried

Figure 4. Average total polar material (%) of oil samples during the frying process with relation to the number of patties fried. The oil is changed once during a frying session as indicated by change on the x-axis. The recommended maximum for total polar material

(TPM) in the food industry is 24% TPM.

Coating Pickup

The mean values for the weights of the raw patty, batter, breading, and whey pickup, and pre- and postfrying are listed in Table 5. There were no significant differences between the measured variables within a breading system. When

measurements were compared between crackermeal-coated patties (CMP) and Japanese

breadcrumb-coated patties (JBP), the breading pickup for JBP was significantly higher (p

< 0.05) compared to those for CMP. However, whey pickup and net loss were not

significantly affected by the difference in breading pickup.

Table 5

Mean Values for Weights for Raw Patty, Coating Pickup, Pre- and Postfrying, and

Weight Difference for Deep-Fried, Battered, and Breaded Chicken Patties

Patty Batter Breading Whey Total Weight Weight Weight Variable weight pickup pickup pickup pickup prefrying postfrying difference (g) (g) (g) (g) (g) (g) (g) (g)

Japanese breadcrumb-coated patties

pH level Control 20.9 8.9 5.1 0.0 14.0 34.9 29.8 -5.1 pH 2 20.4 8.0 5.5 4.4 17.9 38.4 28.0 -10.3 pH 3 20.4 8.5 5.8 4.4 18.6 39.2 28.6 -10.5 pH 8 20.3 5.4 5.9 4.4 18.6 35.9 28.8 -7.1

WPI concentration Control 20.9 8.9 5.1 0.0 14.0 34.9 29.8 -5.1 0% 20.5 7.5 5.7 3.9 16.7 37.6 27.4 -10.2 2.5% 20.3 9.0 5.7 4.2 18.8 39.1 28.4 -10.7 5% 20.4 8.7 6.0 4.3 19.0 39.4 29.6 -9.8 10% 20.4 8.0 5.7 5.0 18.7 39.0 28.6 -10.4 Crackermeal-coated patties

pH level Control 20.1 8.0 3.4 0.0 10.2 31.4 26.2 -5.2 pH 2 20.5 7.8 1.8 3.9 13.1 34.0 26.8 -7.2 pH 3 20.6 8.0 2.1 3.7 13.6 34.4 27.0 -7.4 pH 8 20.5 8.1 2.7 3.3 13.8 34.6 27.8 -6.8

WPI concentration Control 20.1 8.0 3.4 0.0 10.2 31.4 26.2 -5.2 0% 20.6 7.9 1.3 3.4 11.9 33.2 26.0 -7.2 2.5% 20.2 7.9 3.3 3.1 13.7 34.5 27.2 -7.3 5% 20.9 7.9 2.0 3.8 13.4 34.6 27.7 -6.9 10% 20.2 8.2 2.4 4.1 14.7 34.9 28.1 -6.8

Note. Control indicates patties that are not dipped in WPI solution.

Lipid and Moisture Content

The results of the univariate analysis of the main effects (pH level, WPI concentration) on lipid content of both CMP and JBP are shown in Table 6. The final lipid content for both CMP and JBP were significantly affected by the pH level of the post-breading dips in the order of control > pH 8 > pH 2 = pH 3 for CMP and control = pH 8 > pH 2 = pH 3 for JBP. For CMP, lipid reduction ranged from 11.4% for pH 8 to

24.3% for pH 3 while JBP lipid reduction ranged from 12.4% for pH 8 to 12.8% for pH

2. In both breading systems, there were no significant differences between patties treated

with WPI dips at pH 2 and pH 3.

The WPI concentration of the post breading dip also significantly affected final lipid level for both CMP and JBP (see Table 6). Overall, the 10% WPI solution resulted

in patties with significantly lower lipid levels than undipped control patties. However,

WPI concentration affected the two breading systems differently. All CMP exhibited significantly lower lipid levels than the control, with the lowest lipid content observed for patties treated at 5% and 10% WPI (21.4% and 24.3% reduction, respectively). Patties treated with 0% and 2.5% WPI solution also showed significantly lower final lipid content compared to the control. On the other hand, only JBP patties treated with 10%

WPI showed a significant lipid reduction (16.8%) compared to the control.

Table 6

Main Effect Analysis for Lipid Content (%) for Deep-Fried, Battered, and Breaded

Chicken Patties

Crackermeal-coated patties Japanese breadcrumb-coated

Variable (CMP) patties (JBP)

Lipid content ± s.d. Lipid content ± s.d.

pH level1

2 Control 13.32a 0.68 14.80a 0.73

pH 2 10.39b 0.39 12.90b 0.65

pH 3 9.78b 0.28 12.95b 0.39

pH 8 13.18a 0.31 14.41a 0.34

WPI concentration3

2 Control 13.32a 0.68 14.79a 0.73

0 % 11.46b 0.40 14.42a 0.56

2.5 % 11.56b 0.36 13.62ab 0.39

5 % 11.17b 0.37 13.21ab 0.65

10 % 10.26c 0.41 12.30b 0.61

Note. CMP = crackermeal-coated patties, JBP = Japanese breadcrumb-coated patties. ± s.d. = standard deviation. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a breading system and pH or WPI concentration variation for a particular measured variable. 1n = 72 for each group. 2n = 16. 3n = 54 for each group.

The results of the two-way univariate analysis for lipid content for CMP and JBP are shown in Figures 5 and 6, respectively. The highest lipid reduction was observed for

CMP treated with 5% and 10% WPI at pH 2 and at 10% WPI at pH 3. For JBP, the highest lipid reduction was seen in JBP treated with 10% WPI at pH 3 and pH 2. In both breading systems, patties treated at pH 8 with 2.5%, 5%, and 10% WPI were not significantly different compared to the control. While the results for CMP at pH 3 across

all WPI concentrations showed significantly lower lipid content compared to the control,

the results for JBP were less consistent since only JBP treated with 10% WPI at pH 3 was

significantly lower compared to the control.

15 0% WPI 2.5% WPI 5% WPI 10% WPI a 14 ab abcd abc 13 abcd bcde 12 cde cde cde de 11 e e e 10

Lipid content (%) content Lipid 9

8

7 control pH 2 pH 3 pH 8

Figure 5. Final lipid content (%) for crackermeal-coated patties (CMP) treated at various pH levels and whey protein isolate (WPI) concentrations. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p <

0.05 within a treatment (pH x WPI concentration). ncontrol = 16. ntreatment = 18 for each group.

17 0% WPI 2.5% WPI 5% WPI 10% WPI a

15 ab ab bc abc abc cd bcd bc c 13 cd cd d 11 d 9 Lipid content (%) content Lipid

7

5 control pH 2 pH 3 pH 8

Figure 6. Final lipid content (%) for Japanese breadcrumb-coated patties (JBP) treated at

various pH levels and whey protein isolate (WPI) concentrations. Control indicates

patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a treatment (pH x WPI concentration). ncontrol = 16. ntreatment

= 18 for each group.

When univariate analysis was performed on final moisture content of both CMP and JBP, only CMP were observed to be significantly affected by pH level and WPI concentration (p < 0.05). Results for the analysis on the main effects for CMP are shown in Table 7. The results for affect of pH level on final moisture content mimics that observed with final lipid content. CMP treated at pH 8 was not significantly different compared to the control. However, CMP treated at pH 2 and 3 had significantly lower moisture content ranging from 42.59% to 43.10% when compared to the control. For the effect of WPI concentration, CMP treated with all levels of concentration (0%, 2.5%, 5%, and 10% WPI) showed significantly different moisture content compared to the control with CMP at 5% WPI having the highest moisture reduction of 8.46%.

Table 7

Main Effect Analysis for Moisture Content (%) for Deep-Fried, Battered, and Breaded

Chicken Patties

Crackermeal-coated patties Japanese breadcrumb-coated

Variable (CMP) patties (JBP) Moisture content ± s.d. Moisture content ± s.d.

pH level1

2 Control 46.51a 0.94 51.34 0.47

pH 2 43.10b 0.37 49.82 0.57

pH 3 42.59b 0.48 50.31 0.42

pH 8 45.37a 0.31 51.24 0.50

WPI concentration3

2 Control 46.51a 0.94 51.34 0.47

0 % 44.28b 0.28 50.78 0.77

2.5 % 44.50b 0.45 50.43 0.40

5 % 42.57c 0.61 50.01 0.47

10 % 43.26bc 0.43 50.56 0.61

Note. ± s.d. = standard deviation. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a breading system and pH or WPI concentration variation for a particular measured variable. 1n = 72 for each group. 2n = 16. 3n = 54 for each group.

The results for the two-way univariate analysis for CMP and JBP for moisture content are shown in Figures 7 and 8, respectively. As mentioned earlier, while there were no significant differences observed for the main effects for moisture content for

JBP, the interaction between pH level and WPI concentration showed significant

differences. The highest moisture reduction for JBP was observed for patties treated with

2.5% WPI at pH 2. This treatment was the only treatment for JBP that was significantly

different compared to the control. For CMP, the lowest moisture content was observed at

pH 3 and 5% WPI.

53 0% WPI 2.5% WPI 5% WPI 10% WPI

51 a 49 ab bcd abc cd 47 cd cdef bcd cde e def cdef 45 f ef 43

41

Moisture content (%) content Moisture 39

37

35 control pH 2 pH 3 pH 8

Figure 7. Final moisture content (%) for crackermeal-coated patties (CMP) treated at various pH levels and whey protein isolate (WPI) concentrations. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a treatment (pH x WPI concentration). ncontrol = 16. ntreatment

= 18 for each group.

55 0% WPI 2.5% WPI 5% WPI 10% WPI ab 53 ab a a abc ab ab ab abc ab 51 abc c bc 49 c

47 Moisture content (%)

45

43 control pH 2 pH 3 pH 8

Figure 8. Final moisture content (%) for Japanese breadcrumb-coated patties treated at various pH levels and whey protein isolate (WPI) concentrations. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a treatment (pH x WPI concentration). ncontrol = 16. ntreatment

= 18 for each group.

Surface Appearance

There was an obvious difference in appearance between patties treated with WPI dips at pH 8 compared to the control and those at pH 2 and pH 3. The samples treated with WPI at pH 8, regardless of WPI concentration, looked burned and unappealing. As shown in Table 8, lightness (L*) and both chromaticity coordinates (a*, b*) were significantly different for pH 8 than for the other samples. For JBP, L*, a*, and b* were all lower for pH 8 than in the control or at pH 2 or 3. In CMP, L* and b* were lower while a* was higher for pH 8 than in the control or at pH 2 or 3.

Panelists rated the appearance of the patties for color, evenness of color, and surface greasiness on an anchored 15-cm line scale (see Table 8) for patties treated across all WPI concentrations at pH 2 and 3. As shown in Table 8, no significant differences were observed for any of these attributes between patties treated at pH 2 and 3 and with the undipped control. However, instrumental color analysis showed that JBP patties treated with WPI at pH 2 were slightly but significantly darker compared to the control patties. This effect was not observed for patties treated with WPI at pH 3. No differences were observed between the control and patties treated with WPI at pH 2 or pH 3 for either of the chromaticity coordinates. In CMP, patties treated with WPI at pH 2 and 3 were significantly lighter (L*) and exhibited a significantly lower a* value than the undipped control. Analysis of b* values shows that patties treated at pH 2 exhibited a significantly higher b* than the control, while patties treated at pH 3 exhibited a significantly lower b* than the control. These results suggest that pH has more of an effect on CMP than on JBP.

86

Table 8 (continues on pages 87-88)

Mean Values for Sensory Color, Evenness of Color, and Greasiness of Surface Rating and Instrumental Color Values for Deep-Fried,

Battered, and Breaded Chicken Patties

Sensory Instrumental

Variable Evenness Greasiness Color ± s.d. ± s.d. ± s.d. L* ± s.d. a* ± s.d. b* ± s.d. of color of surface

Crackermeal-coated patties (CMP)

pH level1

2 Control 8.7 0.7 8.6 0.7 6.2 0.1 50.9b 1.0 12.7b 0.8 32.6a 0.6

pH 2 6.5 0.3 7.8 0.2 6.3 0.2 54.1a 0.6 9.6c 0.4 57.9c 0.6

pH 3 6.8 0.3 8.7 0.3 6.1 0.1 53.7a 0.6 9.6c 0.4 29.8b 0.6

pH 8 ------37.1c 1.5 13.9a 0.4 24.2d 0.7

87

WPI concentration3

2 Control 8.5a 0.7 8.6 0.7 6.2 0.1 50.9b 1.0 12.7ab 0.8 32.6a 0.6

0 % 5.2c 0.3 8.2 0.3 6.5 0.1 56.0a 0.7 7.7c 0.4 27.7bc 0.6

2.5 % 7.1b 0.5 8.0 0.3 6.0 0.2 46.9c 1.6 12.0b 0.5 28.4b 0.7

5 % 6.6b 0.4 7.9 0.5 6.4 0.2 45.6cd 1.8 12.3ab 0.5 26.7bc 0.9

10 % 7.5b 0.4 8.9 0.2 6.1 0.2 45.2d 1.7 12.9a 0.5 26.3c 1.1

Japanese breadcrumb-coated patties (JBP)

pH level1

2 Control 10.1 0.3 9.4 0.5 6.7 0.2 43.7a 1.1 12.4a 0.7 18.9a 1.7

pH 2 9.8 0.2 8.5 0.2 6.9 0.1 41.8b 1.2 12.0a 0.5 18.4a 0.6

pH 3 9.7 0.3 8.6 0.2 6.8 0.1 42.8ab 1.0 12.7a 0.4 18.9a 0.6

pH 8 ------35.8c 1.4 9.6b 0.4 11.3b 0.9

88

WPI concentration3

2 Control 10.1b 0.3 9.4 0.5 6.7 0.2 43.7b 1.1 12.4a 0.7 18.9b 1.7

0 % 8.8c 0.3 8.7 0.2 6.7 0.1 45.2a 1.1 11.6abc 0.5 21.0a 0.7

2.5 % 10.6ab 0.4 8.2 0.3 7.1 0.1 38.7c 1.3 11.9ab 0.5 15.1c 0.9

5 % 11.4a 0.4 9.1 0.3 6.9 0.2 37.7c 1.6 11.4bc 0.5 14.6c 1.0

10 % 11.3a 0.3 8.4 0.3 6.7 0.2 38.1c 1.5 10.9c 0.6 14.1c 0.9

Note. * Position on 15-cm line scale. ± s.d. = standard deviation. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a breading system and pH or WPI concentration variation for a particular measured variable. For sensory: 1n = 24 for each group. 2n = 6. 3n = 18 for each group. For instrumental: 1n = 20 for each group. 2n = 5. 3n = 15 for each group.

The concentration of the WPI in the dips did not have an affect on evenness of color or greasiness of the surface of the patties for either breading system, but did influence color (see Table 8). In JBP patties, color of the patties was observed as 0% WPI

< control < 2.5% WPI = 5% WPI = 10% WPI. Thus, patties dipped in water at varying pH levels, i.e. 0% WPI, were more yellow than the control while patties treated with WPI were darker brown. This trend was reflected in the chromaticity coordinate b*, for which

an increasing positive value indicates a more yellow color. In CMP, the color of the

patties was observed as 0% WPI < 2.5% WPI = 5% WPI = 10% WPI < control. Thus, in

CMP, all dipped patties were less dark brown (more yellow) than the control; however,

the opposite trend was observed for the chromaticity coordinate b*, because all WPI

treated samples were significantly less yellow than the control although all of the WPI-

treated patties were lighter (L*) than the control.

Texture

The only texture sensory attributes that showed significant differences for main

effect were hardness and crunchiness that were observed when WPI concentrations were

varied for CMP (see Table 9). Hardness was defined as the force required to bite through

a sample (Meilgaard et al., 1999) while crunchiness reflects the amount of sound and

fracturability detected when the sample is chewed once with the molars (Vickers, 1987).

Hardness for CMP was rated in the order of control = 0% = 5% > 2.5% > 10% WPI.

When the results for objective values for hardness were analyzed, it was observed that

CMP in the presence of WPI were significantly harder compared to the control. Results

from the sensory and instrumental measurements of hardness suggest that while

variations in the pH levels of the dip does affect the hardness of the patties, changes in

hardness can only be perceived in the presence of WPI. As for crunchiness, CMP treated with 10% WPI was rated as having the highest intensity. Patties treated at other WPI concentrations were not significantly different compared to the control. An instrumental measurement that may correspond to perceived crunchiness is crust fracture (Brannan,

2008) which represents the peak force at the point of crust thickness. Thus, higher crust fracture values imply a crunchier product. CMP treated with 2.5%, 5%, and 10% WPI were observed to have significantly higher crust fracture values compared to the control

(see Table 9). Similar to the results observed with instrumental hardness, CMP treated with 0% WPI (i.e., water) was not significantly different compared to the control. In addition to hardness and crust fracture, crust work, which is the work done until the crust fracture occurs, was also observed to show significant differences. CMP treated at 0% and 2.5% WPI had significantly lower crust work compared to the control.

91

Table 9 (continued on pages 92-93)

Mean Values for Sensory Hardness and Crunchiness, Crust Thickness, and Instrumental Hardness, Crust Fracture, and Crust Work for Deep-Fried, Battered, and Breaded Chicken Patties

Sensory Instrumental

Crust Crust Total Crust Hardness Crunchiness Hardness ± s.d. ± s.d. thickness ± s.d. ± s.d. fracture ± s.d. work ± s.d. work ± s.d. (cm) (cm) (g) (mm) (g) (g) (g)

Crackermeal-coated patties (CMP) pH level1

2 Control 5.8 0.2 5.8 0.5 3.04b 0.32 651.53 55.00 194.63 24.71 930.51 17.36 59.25 14.91

pH 2 6.2 0.1 6.0 0.3 3.73a 0.12 839.41 28.86 259.98 12.56 397.58 12.10 44.36 2.9

pH 3 6.4 0.1 6.1 0.2 2.92b 0.11 862.80 29.53 247.29 20.01 423.93 14.54 45.32 5.28

pH 8 - - - - 3.16b 0.12 858.04 28.32 264.92 16.38 422.18 14.57 47.91 4.00

92

WPI concentration3

2 Control 5.8b 0.2 5.8b 0.5 3.04cd 0.32 651.53c 55.00 194.63b 24.71 930.51 17.36 59.25a 14.91

0% 6.1b 0.1 5.1b 0.2 2.74d 0.12 915.93a 38.34 209.37b 15.63 419.87 15.57 34.37b 4.19

2.50% 6.5a 0.1 6.1b 0.5 3.21bc 0.14 851.08ab 31.66 259.64a 22.12 406.87 15.77 46.82b 5.61

5% 6.0b 0.1 5.6b 0.2 3.49ab 0.13 813.22b 24.23 267.12a 15.94 396.63 12.34 50.37a 4.52

10% 6.6a 0.1 7.4a 0.3 3.64a 0.15 850.65ab 37.07 291.65a 19.87 436.09 19.33 50.95a 4.26

Japanese breadcrumb-coated patties (JBP) pH level1

Control2 5.6 0.3 7.2 0.5 4.67 0.47 943.08 83.82 423.98 70.85 471.42 10.59 98.61 37.00

pH 2 5.2 0.1 6.7 0.3 4.36 0.24 945.31 26.35 409.93 15.90 477.56 4.23 90.35 14.43

pH 3 5.4 0.1 6.8 0.2 4.05 0.16 1004.06 32.74 404.09 24.81 492.53 4.64 92.09 13.74

pH 8 - - - - 4.24 0.15 904.00 21.60 386.26 18.11 450.37 3.17 87.71 9.80

93

WPI concentration3

2 Control 5.6a 0.3 7.2 0.5 4.67 0.47 943.08 83.82 423.9870.85 471.42 10.59 98.61 37.00

0% 5.0b 0.1 7.1 0.3 4.22 0.32 993.39 28.69 391.0823.30 493.05 4.44 84.48 9.81

2.50% 5.4a 0.2 6.9 0.4 4.38 0.19 936.44 39.13 420.3025.45 473.10 5.96 100.158.96

5% 5.2ab 0.2 7.0 0.3 4.13 0.18 932.66 31.69 385.5824.44 460.23 4.70 87.06 7.70

10% 5.5ab 0.2 6.6 0.3 4.13 0.15 942.00 27.07 403.4218.54 467.70 3.42 88.50 6.21

Note. * Position on 15-cm line scale. ± s.d. = standard deviation. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a breading system and pH or WPI concentration variation for a particular measured variable. For sensory: 1n = 24 for each group. 2n = 6. 3n = 18 for each group. For instrumental: 1n = 20 for each group. 2n = 5. 3n = 15 for each group.

Mouth Feel Sensation and Flavor

The results for the sensory attributes that describe mouth feel sensation and flavor are shown in Table 10. None of these attributes were observed to be significantly different compared to the control and between treatments except for bitterness in JBP as WPI concentration is varied. Panelists rated those treated with 10% WPI to be significantly more bitter compared to the control.

Table 10

Mean Values for Rating of Mouth Feel Attributes for Deep-Fried, Battered, and Breaded

Chicken Patties

Variable Moisture release ± s.d. Oil mouth coating ± s.d.

Crackermeal-coated patties (CMP) pH level1 Control2 6.8 0.5 4.3 0.1 pH 2 6.8 0.3 4.1 0.1 pH 3 7.4 0.3 4.1 0.1

WPI concentration3 Control2 6.8 0.5 4.3 0.1 0% 6.9 0.3 4.1 0.1 2.5% 7.0 0.4 4.1 0.1 5% 7.9 0.4 4.2 0.1 10% 6.8 0.4 4.0 0.2 Japanese breadcrumb-coated patties (JBP) pH level1 Control2 5.0 0.3 4.1 0.2 pH 2 4.4 0.1 3.8 0.1 pH 3 4.2 0.2 3.7 0.1

WPI concentration3 Control2 5.0 0.3 4.1 0.2 0% 4.4 0.1 3.8 0.1 2.5% 4.3 0.2 3.8 0.1 5% 4.4 0.2 3.8 0.1 10% 4.2 0.2 3.6 0.1

Note. ± s.d. = standard deviation. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a breading system and pH or WPI concentration variation for a particular measured variable. 1n = 24. 2n = 6. 3n = 18.

Table 11

Mean Values for Rating of Basic Tastes for Deep-Fried, Battered, and Breaded Chicken

Patties

± ± ± ± ± Variable Salty Sweet Sour Bitter Umami s.d. s.d. s.d. s.d. s.d.

Crackermeal-coated patties (CMP) pH level1 Control2 2.4 0.1 1.2 0.1 0.0 0.0 0.0 0.0 0.9 0.2 pH 2 2.2 0.1 1.1 0.1 0.1 0.0 0.1 0.0 0.7 0.1 pH 3 2.3 0.1 1.1 0.1 0.1 0.0 0.1 0.0 0.9 0.1

WPI concentration3 Control2 2.4 0.1 1.2 0.1 0.0 0.0 0.0 0.0 0.9 0.2 0% 2.2 0.1 1.1 0.1 0.1 0.0 0.1 0.0 0.8 0.1 2.5% 2.1 0.1 0.9 0.1 0.2 0.1 0.1 0.0 0.7 0.1 5% 2.5 0.1 1.1 0.1 0.0 0.0 0.0 0.0 1.0 0.1 10% 2.1 0.1 1.2 0.3 0.1 0.0 0.1 0.0 0.8 0.1

Japanese breadcrumb-coated patties (JBP)

pH level1 Control2 1.1 0.2 0.1 0.0 0.1 0.0 0.0 0.0 0.2 0.1 pH 2 1.0 0.1 0.1 0.0 0.1 0.0 0.1 0.0 0.2 0.0 pH 3 1.3 0.2 0.1 0.0 0.1 0.0 0.1 0.0 0.2 0.0

WPI concentration3 2 Control 1.1 0.2 0.1 0.0 0.1 0.0 0.0b 0.0 0.2 0.1

0% 1.0 0.1 0.1 0.0 0.1 0.0 0.0b 0.0 0.2 0.1

2.5% 1.5 0.4 0.1 0.0 0.1 0.0 0.1b 0.0 0.2 0.1

5% 1.1 0.1 0.1 0.0 0.1 0.0 0.1ab 0.0 0.1 0.0

10% 1.4 0.3 0.1 0.0 0.1 0.0 0.2a 0.1 0.2 0.0

Note. ± s.d. = standard deviation. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a breading system and pH or WPI concentration variation for a particular measured variable. 1n = 24. 2n = 6. 3n = 18.

Table 12

Mean Values for Rating of Flavor Attributes for Deep-Fried, Battered, and Breaded

Chicken Patties

Cooking ± Chicken ± ± ± Variable Chickeny Whey oil s.d. fat s.d. s.d. s.d.

Crackermeal-coated patties (CMP) pH level1 Control2 2.8 0.2 3.6 0.2 5.6 0.2 1.8 0.3 pH 2 2.5 0.1 3.6 0.1 5.2 0.1 1.5 0.2 pH 3 2.4 0.1 3.5 0.1 5.3 0.1 1.5 0.2

WPI concentration3 Control2 2.8 0.2 3.6 0.2 5.6 0.2 1.8 0.3 0% 2.5 0.2 3.7 0.1 5.2 0.1 1.5 0.2 2.5% 2.3 0.2 3.3 0.1 5.1 0.2 1.6 0.3 5% 2.3 0.1 3.7 0.1 5.2 0.2 1.7 0.2 10% 2.7 0.2 3.4 0.1 5.5 0.2 1.2 0.3 Japanese breadcrumb-coated patties (JBP) pH level1 Control2 3.8 0.3 3.7 0.2 6.3 0.3 1.4 0.3 pH 2 3.9 0.2 4.4 0.8 5.9 0.2 1.5 0.2 pH 3 4.0 0.2 3.7 0.1 6.1 0.1 1.7 0.2

WPI concentration3 Control2 3.8 0.3 3.7 0.2 6.3 0.3 1.4 0.3 0% 3.9 0.2 4.8 0.9 6.3 0.1 1.3 0.2 2.5% 3.8 0.2 3.8 0.2 5.8 0.2 1.8 0.3 5% 4.2 0.3 3.7 0.1 5.9 0.1 1.8 0.3 10% 3.7 0.2 3.6 0.1 5.7 0.2 1.6 0.2

Note. ± s.d. = standard deviation. Control indicates patties that are not dipped in WPI solution. Different letters indicate significant differences at p < 0.05 within a breading system and pH or WPI concentration variation for a particular measured variable. 1n = 24. 2n = 6. 3n = 18.

CHAPTER 5: DISCUSSION AND CONCLUSION

Oil Degradation

The increase in total polar material (TPM) wass expected to increase the oil absorption of the fried foods due to the surfactant nature of these breakdown materials

(Blumenthal & Stier, 1991). However, despite a maximum TPM measurement of 8% (see

Figure 2), this was not observed to affect oil absorption (data not shown). The TPM that was generated throughout the frying process might have been be too low to produce any significant increase in oil absorption since it has been observed that interfacial tension only decreases once a certain critical level of TPM is generated (Dana & Saguy, 2006).

Furthermore, the maximum TPM observed (8%) was well below 24% maximal level of

TPM for frying oil as recommended by the delegates of the 3rd International Symposium on Deep-Fat Frying (Deutsche Gesellschaft für Fettwissenschaft, 2000). In short, the absence of the relation between TPM and oil absorption implies that oil absorption may be only affected by degradation of the frying medium above a critical level.

Effect of WPI Treatment on Lipid Content

The analysis of the main effects of WPI concentration on lipid content showed that the lowest final lipid content for CMP and JBP was observed for patties treated with

10% WPI (see Table 6). It is likely that protein concentration affects the lipid barrier properties by influencing the structure of the protein gel. When the level of protein is too low, a protein gel is difficult to establish because intermolecular interactions tend to occur rather than intramolecular interactions (Belitz & Grosch, 1999). As the protein content increases, the likelihood of intermolecular crosslinks increases and gelation can occur. The conversion of monomers of β-lac to fibrils and subsequent aggregation

increases with protein concentration (Bolder, Vasbinder, Sagis, & van der Linden, 2007).

However, some studies have shown that gels formed with higher WPI concentrations have larger aggregates and smaller pores (Mleko, 1999; Verheul & Roefs, 1998), while others suggest that gel structure is independent of protein concentration (Lefévre &

Subirade, 1999; Le Bon, Nicolai, & Durand, 1999). Our results demonstrated that while postbreading WPI dips did inhibit oil absorption, especially at 5% and 10% WPI, there was no difference between the amount of final lipid content between patties treated with solutions containing 2.5% WPI and those treated with 0% WPI. Because a reduction in lipid content was observed for CMP at 0% WPI compared to the control, this implied that the pH of the WPI-containing postbreading dips has a larger influence on the amount of oil absorbed into fried patties than the WPI concentration of the dips.

The analysis of the main effects of pH level on lipid content showed that the highest lipid reduction was observed at low pH, such as pH 2 and 3, for both CMP and

JBP (see Table 6). The importance of pH level on the fat reducing properties of the WPI dips could be inferred from the influence that ionic strength has on protein gelation

(Durand, Gimel, & Nicolai, 2002; Sagis et al., 2002; Schokker et al., 2000). The presence of charged molecules in a protein solution either enhances or decreases electrostatic repulsion between protein monomers. At low pH (e.g., pH 2 and 3), aggregates are formed almost exclusively via noncovalent bonding while at higher pH levels (e.g., pH

7), disulfide interactions between protein molecules are present (Kavanagh et al., 2000).

While increased protein concentration promotes random aggregation due to the close contact between monomers, this does not matter as much if the monomers repel each other due to similar charges. Hence, the observed reduction in oil absorption at high

protein concentration (e.g., 10% WPI) may be due to the influence of random aggregation that, as the protein concentration decreases, increases the role of pH due to its influence on protein charges (Schokker et al., 2000).

For all WPI concentrations, the lipid contents of patties treated at pH 8 were mostly higher compared to those at pH 2 and 3 for both breading systems. If the lipid inhibition observed for WPI dips at low pH is the result of thermally induced gels of one or more of the whey protein fractions, then gels produced at low pH levels have better lipid barrier properties compared to those which form at high pH levels. When the structures of heat-induced globular gels were analyzed, it was observed that gels formed at lower pH levels consist of large, fibrillar aggregates (Durand et al., 2002, Schokker et al., 2000). There were also large voids between the chains of globules (Boye et al., 1997).

On the other hand, at pH levels higher than the isoelectric point, the gels consist of smaller, particulate aggregates that are more evenly distributed and the void spaces are much smaller compared to those observed at pH levels below the isoelectric point (Boye et al.; Hoffman & van Mill, 1998). The difference in pore formation of acidic and alkaline gels may account for the results observed for final lipid content in this study. The water replacement mechanism of oil uptake suggests that as steam escapes during the frying process, it forms large voids with low positive vapor pressure through which oil enters the product (Dana & Saguy, 2006). Acidic gels tend to form larger pores, and thus, according to the water replacement mechanism, more oil is absorbed during the frying stage. However, after removal from the frying medium, the smaller pores formed by the alkaline gels may encourage uptake of oil during cooling since the sudden drop of positive vapor pressure sucks in oil that is present on the food surface (Dana & Saguy,

2006). Because oil uptake during the cooling phase has been shown to account for the

bulk of the oil absorption in fried foods (Moreira et al., 1997), factors that increase oil

absorption through this mechanism would ultimately increase final oil content of the fried

food. This is observed with the higher final lipid content of patties dipped in WPI

solutions at pH 8 compared to those at pH 2 and 3.

Effect of WPI Treatment on Moisture Content

The analysis on the main effects of WPI concentration and pH levels on final

moisture content indicated that WPI concentration had a different effect on CMP

compared to JBP (see Table 7). The highest reduction in moisture content was for CMP

treated with high WPI concentrations, that is, 5% and 10% WPI, and at low pH levels,

that is, pH 2 and 3, while none of the WPI concentration and pH variations affected

moisture content for JBP. CMP that exhibited the highest moisture reduction coincides

with those that have the highest lipid reduction (see Table 6).

While it has been suggested that high moisture retention would lead to lower oil

reduction (Moreira et al., 1997; Pinthus et al., 1993), the results from both CMP and JBP

suggested that oil uptake was not only affected by the direct exchange between water and

oil, but other factors as well, such as those mentioned previously (see Factors Affecting

Oil Absorption). While the formation of pores through the evaporation of water is

involved in oil uptake, there was no direct relationship between moisture loss and oil

absorption that was observed in this study. This points to the complexity of the dynamics

of moisture and oil migration during the frying process as mentioned by the water

replacement mechanism (Pinthus & Saguy, 1994; Rice & Gamble, 1989) and after

removal from the frying medium via the cooling phase effect (Moreira et al., 1997). In

addition, while moisture loss is continuous throughout the frying process, oil uptake reaches a maximum level and remains constant until removal of the fried food from the frying medium (Yamsaengsung & Moreira, 2002). Nevertheless, moisture loss is still related to oil uptake and factors that affect oil uptake would ultimately play a role in determining the movement of moisture. Gels that have large pores allow moisture to vaporize more easily compared to those with smaller pores and hence, gels at lower pH levels would be expected to show lower moisture content compared to gels at higher pH levels. This may explain why moisture content was less at low pH levels.

Effect of WPI Treatment on Organoleptic Properties of Fried Chicken Patties

In addition to lowering the fat content in fried foods, product developers must also ensure that the product is accepted by consumers. In other words, all sensory characteristics associated with the food such as flavor, color, and texture must be met

(Fuller, 2004). For example, a desirable and expected textural property in fried foods is crunchiness or crispness. Fried foods that lack this attribute are usually seen as being of poor quality and will not appeal to the majority of consumers. In addition, consumers often use color as a quick and often effective parameter in determining the quality of foods.

Surface Appearance: Color

Surface color is a quick and often effective parameter in determining the quality

of fried foods. As shown in Table 8, lightness (L*) and both chromaticity coordinates (a*,

b*) were significantly different for pH 8 than for the other samples. For JBP, L*, a*, and

b* were all lower for pH 8 than in the control or at pH 2 or 3. In CMP, L* and b* were

lower while a* was higher for pH 8 than in the control or at pH 2 or 3. Fried foods often

exhibit increased values for a* and lower values for L* and b* when they are fried under nonoptimum conditions such as long frying time, in highly degraded oil, and at very high temperatures (Krokida et al., 2001; Ngadi et al., 2007). For this reason and because patties treated with WPI at pH 8 exhibited minimal oil inhibition properties (see Table 6), patties treated with WPI at pH 8 were not further analyzed by sensory analysis.

As shown in Table 8, no perceivable significant differences were observed for color, evenness of color, and greasiness of surface between patties treated at pH 2 and 3 and with the undipped control. However, when instrumental measurement of surface color was performed, it was observed that JBP at pH 2 were significantly darker compared to the control patties. This effect was not observed for patties treated with WPI at pH 3. No differences were observed between the control and patties treated with WPI at pH 2 or pH 3 for either a* or b* values. In CMP, patties treated with WPI at pH 2 and

3 were significantly lighter (higher L* value) and less red (lower a* value) than the undipped control. A significantly higher b* values for patties treated at pH 2 indicated that treating CMP at pH 2 produced a more yellow patty while the significantly lower b* value for CMP treated at pH 3 means that this pH level made the patty less yellow compared to the control. These results suggest that pH has more of an effect on the color of CMP than on JBP.

The concentration of the WPI in the dips did not have an effect on evenness of

color or greasiness of the surface of the patties for either breading system, but did

influence color (see Table 8). In JBP patties, color of the patties was rated as 0% WPI <

control = 2.5% WPI = 5% WPI = 10% WPI. Thus, patties dipped in water at varying pH

levels, that is, 0% WPI, were more yellow than the control while patties treated with WPI

were darker brown. This trend was reflected in the chromaticity coordinate b* as only

JBP treated with 0% WPI showed a significantly higher b* value, indicating a more yellow color compared to the control. In CMP, the color of the patties was rated as 0%

WPI < 2.5% WPI = 5% WPI = 10% WPI < control. Thus in CMP, the presence of WPI resulted in patties that were less dark brown (more yellow) than the control. However, the opposite trend was observed for the chromaticity coordinate b*, where JBP across all

WPI concentration had a lower b* value (less yellow) compared to the control. In addition, L* values were lower for JBP treated with WPI. These results suggest that while treating the patties with acidic solutions, i.e. pH 2 and 3, made the patty more yellow and

bright, the presence of WPI darkened the color of the patties and downplayed the

“yellowing” and “brightening” effect of low pH levels.

When correlation analysis was done on the sensory color, L*, a*, and b* for

CMP, it was observed that the L* values were negatively correlated with color (r = -

0.480, p < 0.01) while a* values were positively correlated to color (r = 0.508, p < 0.01).

Surprisingly, b* values (indicating yellowness) was weakly correlated with color (r =

0.237, p < 0.05). The results of the correlation analysis suggest that the panel relied more

on the lightness and the redness of the patties in rating the color of CMP compared to the

yellowness even though the scale was set to range from yellow to dark brown. On the

other hand, when a similar correlation analysis was done on JBP, none of the

instrumental color values were significantly correlated with color rating of the panel. The

inconsistencies between the results from the univariate and correlation analysis of

perceived color for each breading system imply the complexity of the perception of color

because it is dependent on various other factors such as (a) variations in the food product

such as particle size (crackermeal vs. Japanese breadcrumbs), (b) differences in concentrations and distributions of light-absorbing pigments (Little, 1973), and (c) variations in the psychology of the consumers (Churchland, 2007). In foods, colorimetry is not used for the strict reproduction of the product, but more as an indication of the range of acceptability of the product as it pertains to consumer appeal (Little, 1973).

Thus, both color and instrumental measurements of the different treatments for JBP and

CMP should be taken into consideration when fine-tuning the parameters of this treatment to ensure consumer acceptability.

Texture: Hardness and Crunchiness

As shown in Table 9, only variations within the WPI concentrations produced significant differences in texture, specifically sensory and instrumental hardness, crunchiness, and crust work. However, CMP alone showed significant differences in all the aforementioned attributes while only the sensory hardness for JBP was affected by variations in WPI concentration.

Hardness was defined as the force required to bite through a sample (Meilgaard et al., 1999) while crunchiness reflects the amount of sound and fracturability detected when the sample is chewed once with the molars (Vickers, 1987). The panel rated JBP treated with 0% WPI to be significantly softer compared to control and other WPI concentrations while the sensory hardness for CMP was rated in the order of control =

0% = 5% > 2.5% = 10% WPI. While the instrumental measurements did not show any differences for hardness for JBP, it showed that CMP were significantly harder and have higher crust fracture in the presence of WPI. The sensory and instrumental measurements of crunchiness suggest that treating patties with WPI increases the crunchiness of the

patties independent of pH levels, but this effect could only be perceived at high WPI concentrations (e.g., 10% WPI). The inconsistency could be attributed to the fact that unlike the instrumental measurement of crunchiness, sensory crunchiness was judged not only based on the force it took to bite through the sample, but also the amount of sound produced. Various studies have shown that the amplitude of sound generated when biting through a sample correlated well with perceived crunchiness (Antonova et al., 2003;

Vickers, 1987).

Because the instrumental measurements of texture involves the penetration of the crust region, it is suspected that the components of the crust such as the amount of batter, breading, and/or whey pickup and crust thickness affected these textural properties.

When ANCOVA was used to analyze the effects of these factors on CMP instrumental hardness and crust fracture, it was observed that only crust thickness significantly affected both instrumental hardness and crust fracture (p < 0.05). When the effects of these factors were analyzed for CMP crust fracture, it was also observed that only crust thickness significantly affected the variable (p < 0.05). Correlation analysis on crust thickness and instrumental hardness and crust fracture showed that crust thickness was positively correlated with crust fracture (r = 0.585, p < 0.01) but was not significantly correlated with instrumental hardness. These observations may explain why JBP did not show any significant differences for hardness and crust fracture (see Table 9).

Another factor that may contribute to the difference in crust fracture or crunchiness of the patties is the moisture content (Antonova et al., 2003). Moisture content was observed to be significantly lower in CMP treated with 5% and 10% WPI.

This may contribute to the significantly higher crust fracture for CMP treated with 5%

and 10% (see Table 7). In addition, the sensory panel rated CMP treated with 10% WPI to be the crunchiest of all the treatments. Moisture reduces the crunchiness of a food

product by weakening the solid matrix (Katz & Labuza, 1981; Van Vliet et al., 2007) due

to breakage of the macromolecular interactions of the food structure by water-water interactions. This causes the macromolecules to be more mobile and slide against each other and this is perceived as a reduction in crunchiness (Katz & Labuza, 1981). Thus,

the results suggest that treatments that lead to lowering of moisture content (e.g., at 10%

WPI) increase crunchiness of the battered and breaded product.

Mouth Feel Sensation and Flavor

The results for the sensory attributes that describe mouth feel sensation and flavor

are shown in Table 10. None of these attributes were observed to be significantly different when compared to the control and between the treatments except for bitterness in JBP as WPI concentration is varied. Panelists rated those treated with 10% WPI to be significantly more bitter compared to the control. However, since no other independent variable variations for both JBP and CMP produced a significant difference in bitterness, the observed result may be due to the psychological errors of the panelists, and these errors might include those due to expectation, preference, environment, and fatigue

(Stone & Sidel, 1985). Overall, treating patties with WPI solutions at various pH levels and WPI concentrations did not significantly affect the general mouth feel sensation and flavor of the patties.

Conclusion

WPI exhibits lipid barrier properties that are dependent on both pH and protein

concentration when used as a postbreading dip in battered and breaded fried chicken

patties. The highest lipid reduction was obtained when 5% and 10% WPI solutions were

used at low pH levels (pH 2 and 3) for both breading systems. Lipid reduction ranged

from 31.24% for CMP at pH 2 with 5% WPI and 37.5% for JBP at pH 2 with 10% WPI.

Treatment of deep-fried, battered, and breaded chicken patties with WPI did not

cause any perceivable changes in the flavor of the product although color, perceived

hardness, and perceived crunchiness were significantly affected. WPI concentrations

were observed to have different effects on the two breading systems where JBP were

darker and less yellow when treated with 5% and 10% WPI while CMP were lighter and

more yellow compared to the control across all WPI concentrations. Patties treated at pH

2 and 3 did not produce any perceivable color changes but had significantly lower L* and

b* values and higher a* values compared to the other treatments. As for hardness, patties treated with the WPI dip were either perceived to be harder or similar in hardness compared to the control. Instrumental measurements confirms the sensory evaluations by showing a significant increase in hardness and crust fracture for patties treated with 2.5%,

5%, and 10% WPI. Thus, JBP that showed the highest lipid reduction (10% WPI at pH 2) were observed to be darker, less yellow, more bitter, but did not produce any perceivable changes in hardness, crunchiness, mouth feel, and flavor while CMP with the lowest lipid content (5% WPI at pH 2) were darker, more yellow, and are perceived to be crunchier, but were not perceived to be different in mouth feel, taste, and flavor.

The usage of WPI as a postbreading dip is a promising alternative in reducing fat content in fried foods since it could simultaneously fulfill the steady demand for fried foods and contribute to the growing effort of Americans to consume less fat. In addition, this treatment was observed to be versatile due to the comparable effectiveness in reducing fat uptake for both breading systems despite the difference between the composition, structure, and size of the breadings. Despite the significant effect of the

WPI postbreading dip on the color, hardness, and crunchiness of the deep-fried, battered, and breaded chicken patties, these changes may not deter consumers who place more emphasis on reducing their fat consumption. In short, while there is still room for improving this treatment to minimize the effect on color and texture, the usage of WPI as a postbreading dip is a promising alternative in reducing fat content in fried foods since it does not alter the flavor profile of a full-fat product.

Future Studies

The results of this research suggests the need for further investigation to further the understanding of the mechanism and versatility of WPI as a postbreading dip in reducing oil absorption. Specific to whey, the role of each protein fraction in contributing to the lipid barrier properties of whey protein isolate should be investigated. These fractions could be separated out and combinations of protein fractions could be used to make the postbreading dip and the effects on final lipid content could then be analyzed.

In addition, future studies could focus on the optimization of the WPI treatment in reducing oil absorption in fried foods by investigating (a) effects of variations of the substrate that is being fried, particularly vegetable vs. meat products, (b) variations in product composition (e.g., batter formulation, breading types, battered and breaded vs.

not battered and breaded), and (c) variations in processing methods such as par-fried vs. fully fried, differences in rethermalization methods such as oven baking vs. microwave heating, and various storage conditions (e.g., frozen par-fried, frozen fully-fried, changes throughout refrigerated and frozen storage). In relation to the food industry, consumer sensory tests should be conducted to investigate the overall acceptability of the treatment and the product that is being treated, in addition to continuous descriptive sensory analysis.

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APPENDIX A: IRB FORM

APPENDIX B: TIMELINE FOR SENSORY SELECTION, TRAINING, AND

SAMPLING

Session Date Description

Info session 2/1/07 • Giving out information concerning basic sensory 2/7/07 analysis. 3/6/07 • Those interested were asked to fill in a 3/7/07 questionnaire (see Appendix C) 3/8/07

Training session 1 4/26/07 • Panel were asked to sign informed consent form (see Appendix D) • Identification and rating of basic tastes (see Appendix E)

Training session 2 5/10/07 • Identification and rating of mixtures of basic tastes (see Appendix F)

Training session 3 5/15/07 • Introduction to lexicon development: cookie variations (see Appendix G)

Training session 4 5/22/07 • Introduction to standard intensity scale for hardness (see Appendix H) • Continuation of lexicon development : fried oat bran patties (see Appendix K)

Training session 5 5/24/07 • Introduction to standard intensity scale for crispness (see Appendix I) • Continuation of rating of hardness using standard references (see Appendix L)

Training session 6 5/31/07 • Continuation of rating of crispness using standard references (see Appendix M)

Training session 7 7/10/07 • Continuation of rating of hardness and crispness using standard references (see Appendix N)

Training session 8 7/12/07 • Continuation of lexicon development: fried dough balls (see Appendix O)

Training session 9 7/17/07 • Continuation of rating of crispness using standard references (see Appendix P)

Training session 10 7/19/07 • Review of basic taste identification and rating for hardness and crispness using standard references (see Appendix Q)

Training session 11 7/24/07 • Introduction to standard intensity scale for juiciness (see Appendix J and R)

Training session 12 7/26/07 • Continuation of rating of juiciness using standard references (see Appendix S)

Training session 13 8/14/07 • Lexicon development for cooked ground chicken (see Appendix T)

Training session 14 8/16/07 • Introduction to rating using a 15-cm line scale for attributes for cooked ground chicken (see Appendix U)

Training session 15 9/20/07 • Lexicon development for fried, battered, and breaded chicken products (see Appendix V)

Training session 16 9/25/07 • Continuation of lexicon development for fried, battered, and breaded chicken products

Training session 17 9/27/07 • Fine-tuning of lexicon for fried, battered, and breaded chicken patties using the 15-cm line scale

Calibration for 10/2/07 • Calibration of attribute rating with warm-up CMP CMP sample

Calibration for 10/4/07 • Calibration of attribute rating with warm-up CMP CMP sample

CMP sampling 10/9/07 • Sampling of CMP session 1 • Ballot (see Appendix W) • Order or sampling (see Appendix Y) CMP sampling 10/11/07 session 2

CMP sampling 10/12/07 session 3

Calibration for JBP 10/23/07 • Calibration of attribute rating with warm-up JBP sample

JBP sampling 10/30/07 • Sampling of JBP session 1 • Ballot (see Appendix X) • Order or sampling (see Appendix Z) JBP sampling 11/1/07 session 2

JBP sampling 11/6/07 session 3

JBP sampling 11/8/07 session 4

JBP sampling 11/9/07 session 5

APPENDIX C: SENSORY PANEL QUESTIONNAIRE

HISTORY: Name: ______Address: ______Phone (home and business): ______Email: ______

TIME: 1. Are there any weekdays (M-F) that you will not be available on a regular basis?______2. How many weeks of vacation do you plan to take in 2007? ______

HEALTH: 1. Do you have any of the following? Dentures _____ Diabetes _____ Oral or gum disease _____ Hypoglycemia _____ Food allergies _____ Hypertension _____

2. Do you take any medications that affect your senses, especially taste and smell? ______

FOOD HABITS: 1. Are you currently on a restricted diet? If yes, explain ______2. How often do you eat out in a month? ______3. How often do you eat fast food in a month? ______4. What is (are) your favorite food(s)? ______5. What is (are) your least favorite food(s)? ______6. What is (are) your least favorite food(s)? ______7. What foods can you not eat? ______8. What foods do you not like to eat? ______9. If you ability to distinguish smell and tastes:

SMELL TASTE Better than average ______Average ______Worse than average ______

10. Does anyone in your immediate family work for a food company? ______11. Does anyone in your immediate family work for an advertising company or a marketing research agency? ______

QUICK QUIZ:

1. How would you describe the difference between flavor and aroma? ______

2. What is the best one or two word description of grated Italian cheese (Parmesan or Romano)? ______

3. Describe some of the noticeable flavors in cola ______

APPENDIX D: CONSENT FORM

OHIO UNIVERSITY CONSENT FORM Optimization of the Pre-treatment to Reduce Oil Absorption in Frozen Pre-fried Breaded Products Using Whey Protein Isolate You are being asked to participate in a research study. Participation in this study is completely voluntary. Please read the information below and ask questions about anything that you do not understand before deciding if you want to participate. A researcher listed below will be available to answer your questions.

RESEARCH TEAM AND SPONSORS

Principal Investigator: Dr. Robert G. Brannan Associate Professor Department of Human and Consumer Sciences (740) 593-2879

Co-Investigator Eunice Mah Department of Human and Consumer Sciences (740) 590-1871

PURPOSE OF STUDY The purpose of this research study is to study the effectiveness of whey protein isolate coating in reducing oil absorption into fully battered and breaded deep-fried foods when applied post-breading. ELIGIBILITY Potential participants will be screened based on the information obtained from the attached questionnaire.

PROCEDURES If you agree to participate you will be involved in a series of training for several sessions spanning several weeks where you will be familiarized with the methods used in sensory analysis of foods. Training will involve tasting various food products to test for texture and flavor acuity and developing terminology to describe the food product. After completing the training session, you will be called upon to taste and describe deep fried chicken patties that treated with the whey protein isolate coating. Ballots containing terminologies that were developed will be used as guidelines. You will be working as a group throughout the duration of the study. Each training and tasting session is estimated to be no more than an hour and will be scheduled according to the availability of the group.

RISKS, STRESS, OR DISCOMFORT Risks associated with participating in this study will be minimal to none. You will be screened according to the food allergies or medical conditions that you will be asked to list in the attached questionnaire.

UNKNOWN RISKS There may be risks to being in this study that we don't know about now. You will be informed of any changes in the way the study will be done and any additional identified risks to which you may be exposed.

BENEFITS The level of fat intake for the average American is four percent higher than the maximum recommended total dietary fat. High intake of fat is associated with increased risk for many diseases and therefore, consumers are encouraged to adopt a low- or reduced-fat diet. Research into reducing fat/oil absorption during deep-fat frying using natural ingredients may offer more varieties of low- or reduced-fat foods in the market.

______Printed name of researcher Signature of researcher Date

Subject’s statement This study has been explained to me. I volunteer to take part in this research. I have had a chance to ask questions. If I have general questions about the research, I can ask one of the researchers listed above. If I have questions regarding my rights as a participant, I can call Jo Ellen Sherow, Director of Research Compliance, Ohio University, (740)593-0664. This project has been reviewed and approved for human participation by the Ohio University IRB. I will receive a copy of this consent form.

______Printed name of subject Signature of subject Date

APPENDIX E: TRAINING SESSION 1

Date: April 26, 2007 Venue: Grover Center W120

Objectives: o Basic panel screening and calibration o Developing skills of rating taste intensities without the distraction of aromatics

Test Design: o Panel will familiarize themselves with the reference set o Panel will then taste the evaluation set and record their impressions using the score sheet provided. o Reference set: All 6 samples are clearly labeled and placed on individual serving trays (10 ml per sample). o Evaluation set: All 9 samples are clearly labeled with codes and placed on individual serving trays in random order (10 ml per sample). o The scores will be averaged and analyzed.

Materials: Assume 20 participants and 10ml serving size

o 300* plain plastic serving cups, 2-oz size o 20 individual serving trays o 20 large plastic cups with lid (spit cups), 16-oz size o 20 water rinse cups, 6-oz size o 5 water serving pitchers o 1 packet napkins

*(6 reference solution + 9 evaluation solution) x 20 = 300 samples

Evaluation set 1 Label Content ( in 250 ml Code water) Salty 0.10 g salt 852 Salty 0.15 g salt 796 Salty 0.25 g salt 936 Sweet 0.5 g sugar 374 Sweet 1 g sugar 710 Sweet 2 g sugar 611 Sour 0.05 g citric acid 485 Sour 0.07 g citric acid 918 Sour 0.10 g citric acid 660 Umami 0.025 g umami 991 Umami 0.05 g umami 384 Umami 0.1 g umami 968

Preparation of solution: o Bulk solution can be prepared one day ahead and refrigerated. o For evaluation set, solutions are prepared by mixing equal quantities of the appropriate reference solutions. o On the day of the test, all samples are allowed to warm to room temperature

`BASIC TASTE SCORECARD

Date:

Name:

Characteristic studied: Basic taste

Instructions: You have 3 samples on the tray in front of you. Taste the samples from left to right. Identify the taste (sweet, sour, salty, or umami). Rank the taste intensity by recording the codes in the appropriate space (1 being the weakest and 3 being the strongest).

Set 1

Basic taste: ______

Intensity rank: 1. ______2. ______3. ______

Set 2

Basic taste: ______

Intensity rank: 1. ______2. ______3. ______

Set 3

Basic taste: ______

Intensity rank: 1. ______2. ______3. ______

Set 4

Basic taste: ______

Intensity rank: 1. ______2. ______3. ______

BASIC TASTE SCORECARD

Date:

Name:

Characteristic studied: Basic taste

Instructions: You have four samples on the tray in front of you. Record down the sample code before tasting the sample. Taste the samples from left to right and indicate the taste by checking the appropriate space. Do not repeat the evaluation of previous samples.

Sample Code Sweet Sour Salty Umami

______

APPENDIX F: TRAINING SESSION 2

Date: May 10th, 2007 Venue: Grover Center W313

Objectives: o Basic panel screening and calibration o Developing skills of rating taste intensities without the distraction of aromatics

Test Design: o Panel will familiarize themselves with the reference set o Panel will then taste the evaluation set and record their impressions using the score sheet provided. o Reference set: All 6 samples are clearly labeled and placed on individual serving trays (10 ml per sample). o Evaluation set: All 9 samples are clearly labeled with codes and placed on individual serving trays in random order (10 ml per sample). o The scores will be averaged and analyzed.

Materials: Assume 20 participants and 10ml serving size

o 300* plain plastic serving cups, 2-oz size o 20 individual serving trays o 20 large plastic cups with lid (spit cups), 16-oz size o 20 water rinse cups, 6-oz size o 5 water serving pitchers o 1 packet napkins

*(6 reference solution + 9 evaluation solution) x 20 = 300 samples

Evaluation set Content Code

Sucrose/citric acid 246 Sucrose/NaCl 425 Sucrose/umami 132 Citric acid/umami 861 Citric acid/NaCl 388 NaCl/umami 258

BASIC TASTE SCORECARD

Date:

Name:

Characteristic studied: Basic taste

Instructions: You have six samples on the tray in front of you. Record down the sample code before tasting the sample. Taste the samples from left to right and indicate the taste by checking the appropriate space. Do not repeat the evaluation of previous samples.

Sample Code Sweet Sour Salty Umami

______

APPENDIX G: TRAINING SESSION 3

Date: May 15th, 2007 Venue: Grover Center E120

Objectives: o Introduce lexicon development to panel

Test Design: o Panel will be asked to generate a lexicon for different cookie variations (refer to table below).

Code Cookie Variation Ingredients 2 ½ C flour 762 Flour, water margarine 1 C water 2 ½ C flour 806 Flour, water, sugar 1 C water ½ C + 2 T margarine 2 ½ C flour 1 C water 427 Flour, water, margarine, sugar ½ C + 2 T margarine 1 C white granulated sugar 2 ½ C flour 1 C water 1 C white granulated sugar 800 Flour, water, sugar, egg, margarine, vanilla 1 egg ½ C + 2 T margarine 1 t pure vanilla extract

o Method: o Prepare each recipe as shown in table. o Spread dough into 9 x 13 oblong non-stick baking pan. Precut into squares before baking. o Bake at 350 for 35 minutes o Store in airtight containers.

Cookie Variation Exercise Worksheet

Date:

Name:

Instructions: You have four samples on the tray in front of you. Record down the sample code before tasting the sample. Taste the samples and describe the flavor, texture, and appearance.

Code Description

APPENDIX H: STANDARD INTENSITY HARDNESS SCALE

Scale value Product Brand/Type/Manufacturer Sample size

1 Cream cheese Kraft/Philadelphia light ½ in cube

2.5 Egg white Hard cooked ½ in cube

Yellow American pasteurized 4.5 Cheese ½ in cube process

1 olive pimento, 6 Olives Goya foods/giant size, stuffed removed

Large, cooked 5 min/Hebrew 7 Frankfurter ½ in slice national

Cocktail type in vacuum 9.5 Peanuts 1 nut, whole tin/Planters

11 Carrots Uncooked, fresh, unpeeled ½ in slice

14.5 Hard candy Lifesavers 3 pieces, one color

APPENDIX I: STANDARD INTENSITY CRISPNESS SCALE

Scale value Product Brand/Type/Manufacturer Sample size

2 Granola bar Quaker Low Fat Chewy Chunk 1/3 bar

5 Club cracker Keebler Partner Club Cracker ½ cracker

6.5 Graham cracker Honey Maid 1 in square

7 Oat cereal Cheerios 1 oz

9.5 Bran flakes Kellogg’s Bran Flakes Cereal 1 oz

Cheese crackers Pepperidge Farm Cheddar Cheese 11 1 oz goldfish Crackers

14 Corn flakes Kellogg’s Corn Flakes Cereal 1 oz

APPENDIX J: STANDARD INTENSITY JUICINESS SCALE

Scale value Product Brand/Type/Manufacturer Sample size

1 Banana Raw ½ inch slice

2 Carrot Raw ½ inch slice

4 Mushroom Raw ½ inch slice

7 Snap beans Raw 5 pieces

8 Cucumber Raw ½ inch slice

10 Apple Raw ½ inch wedge

12 Honeydew melon Raw ½ inch cube

15 Watermelon Raw ½ inch cube

APPENDIX K: TRAINING SESSION 4

Date: May 22, 2007 Venue: Grover Center W320

Objectives: o Familiarize panel with intensity scale values using references o Practice lexicon development

Test Design: o Panel will be asked to taste references for hardness. o Panel will be asked to come up with a lexicon for fried oat bran patties

Standard reference for hardness

Scale value Sample brand Sample size 1 Cream cheese Kraft/Philadelphia light ½ in cube 2.5 Egg white Hard cooked ½ in cube Cheese Yellow American pasteurized ½ in cube 4.5 process/ land O’lakes Olives Goya foods/giant size, stuffed 1 olive pimento, 6 removed Frankfurter Large, cooked 5 min/Hebrew ½ in slice 7 national Peanuts Cocktail type in vacuum 1 nut, whole 9.5 tin/Planters 11 Carrots Uncooked, fresh, unpeeled ½ in slice 14.5 Hard candy lifesavers 3 pieces, one color

o Panelists will be presented with all the reference samples and asked to familiarize themselves with the scale values. o Prepare out bran patties (instructions to follow) and present it to the panel to form a lexicon for the product. Group discussion.

o Oat bran patties: Ingredients: 1 C Kroger’s Oat Bran Muffin Mix ® ¼ C water Method: Mix the two ingredients together to form a paste. Drop a tablespoon of batter into a pre-heated deep fryer (375F) and fry until golden brown. Drain excess oil with paper towel. Serve at room temperature

Lexicon Exercise Worksheet

Name:

Instructions: You have a sample on the tray in front of you. Record down the sample code before tasting the sample. Taste and describe the sample according to the instructions provided.

1. Look at the sample (without touching) and describe the appearance. Give attention to characteristics such as surface moisture, surface roughness (smoothness), and color.

2. Place sample between incisors and bite down evenly. Evaluate the force required to bite through the food. Refer to the reference scale for hardness.

3. Describe the flavor and aroma of the product. Include perception of basic tastes (sweet, sour, salty, bitter, umami)

APPENDIX L: TRAINING SESSION 5

Date: May 24, 2007 Venue: Grover Center W320

Objectives: o Familiarize panel with intensity scale values using references

Test Design: o Panel will be asked to taste references for crispness. o Panel will be evaluated for rating of hardness intensity

o Standard references for crispness scale: Scale reference brand Sample size value 2 Granola bar Quaker low fat chewy chunk 1/3 bar 5 Club cracker Keebler partner club cracker ½ cracker 6.5 Graham cracker Honey maid 1 in square 7 Oat cereal Cheerios 1 oz 9.5 Bran flakes Kellogg’s bran flakes cereal 1 oz 11 Cheese crackers Pepperidge farm cheddar cheese 1 oz goldfish crackers 14 Corn flakes Kellogg’s corn flakes cereal 1 oz o Panelists will be presented with all the reference samples and asked to familiarize themselves with the scale values.

o Evaluation for hardness intensity Code Sample brand Sample size 816 Cream cheese Kraft/Philadelphia light ½ in cube 327 Egg white Hard cooked ½ in cube 762 Cheese Yellow American pasteurized ½ in cube process/ land O’lakes 841 Olives Goya foods/giant size, stuffed 1 olive pimento, removed 994 Frankfurter Large, cooked 5 min/Hebrew ½ in slice national 953 Peanuts Cocktail type in vacuum 1 nut, whole tin/Planters 615 Carrots Uncooked, fresh, unpeeled ½ in slice 756 Hard candy lifesavers 3 pieces, one color o Panelists will be presented with all the samples and asked to rate the hardness intensity of the samples. Answers will be compared to the standard references for hardness (refer to training session 4).

Hardness Intensity Exercise Worksheet

Date:

Name:

Instructions: Record down the sample code before tasting the sample. Taste the samples and rate the crispness of the samples. Scale values 1 and 14.5 have been identified for you. Do not discuss with other panel members.

Code Scale value

816 1

756 14.5

______

______

______

______

______

______

APPENDIX M: TRAINING SESSION 6

Date: May 31, 2007 Venue: Grover Center W320

Objectives: o Familiarize panel with intensity scale values using references

Test Design: o Evaluation of panel’s perception of crispness.

Evaluation of panel’s perception on crispness:

Code reference brand Sample size 451 Granola bar Quaker low fat chewy chunk 1/3 bar 124 Club cracker Keebler partner club cracker ½ cracker 682 Graham Honey maid 1 in square cracker 914 Oat cereal Cheerios 1 oz 694 Bran flakes Kellogg’s bran flakes cereal 1 oz 385 Cheese Pepperidge farm cheddar cheese 1 oz crackers crackers goldfish 685 Corn flakes Kellogg’s corn flakes cereal 1 oz

o Panel will be presented with the samples. o Panel will be asked to rank the crispness of the samples using a 15-point scale (1 = not crisp/soggy, 15 = very crisp). o Results will be compared to the reference set (refer to training session 5)

Crispness Intensity Exercise Worksheet

Date:

Name:

Instructions: Record down the sample code before tasting the sample. Taste the samples and rate the crispness of the samples. Scale value 1 and 15 have been identified for you. Do not discuss with other panel members.

Code Scale value

451 1

685 15

______

______

______

______

______

APPENDIX N: TRAINING SESSION 7

Date: July 10th, 2007 Venue: Grover Center E120

Objectives: o Re-familiarize panel with intensity scale values using references

Test Design: o Panel will be asked to taste references for hardness and crispness.

o Standard hardness scale reference: Scale Reference brand Sample size value 1 Cream cheese Kraft/Philadelphia light ½ in cube 2.5 Egg white Hard cooked ½ in cube 4.5 Cheese Yellow American pasteurized ½ in cube process/ land O’lakes 6 Olives Goya foods/giant size, stuffed 1 olive pimento, removed 7 Frankfurter Large, cooked 5 min/Hebrew ½ in slice national 9.5 Peanuts Cocktail type in vacuum 1 nut, whole tin/Planters 11 Carrots Uncooked, fresh, unpeeled ½ in slice 14.5 Hard candy lifesavers 3 pieces, one color

o Standard crispness scale reference: Scale Reference brand Sample size value 2 Granola bar Quaker low fat chewy chunk 1/3 bar 5 Club cracker Keebler partner club cracker ½ cracker 6.5 Graham cracker Honey maid 1 in square 7 Oat cereal Cheerios 1 oz 9.5 Bran flakes Kellogg’s bran flakes cereal 1 oz 11 Cheese crackers Pepperidge farm cheddar cheese 1 oz goldfish crackers 14 Corn flakes Kellogg’s corn flakes cereal 1 oz

o Present the standards to panel and ask them to re-familiarize themselves with the scale. o Practice using the scale with the following product: o Apple (113) o Ginger snaps (260) o Saltine crackers (185) o Lima beans (336)

Scorecard 7a (hardness)

Name:

Instructions: You have a sample on the tray in front of you. Record down the sample code before tasting the sample. Follow the instructions.

Sample code: ______

Place sample between your front teeth and bite through the sample once. Evaluate the hardness. Refer to the reference scale for hardness.

Sample code: ______

Place sample between your front teeth and bite through the sample once. Evaluate the hardness. Refer to the reference scale for hardness.

Sample code: ______

Place sample between your front teeth and bite through the sample once. Evaluate the hardness. Refer to the reference scale for hardness.

Sample code: ______

Place sample between your front teeth and bite through the sample once. Evaluate the hardness. Refer to the reference scale for hardness.

Scorecard 7b (crispness)

Name:

Instructions: You have a sample on the tray in front of you. Record down the sample code before tasting the sample. Follow the instructions.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the crispness. Refer to the reference scale for crispness.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the crispness. Refer to the reference scale for crispness.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the crispness. Refer to the reference scale for crispness.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the crispness. Refer to the reference scale for crispness.

APPENDIX O: TRAINING SESSION 8

Date: July 12, 2007 Venue: Grover Center E120

Objectives: o Familiarize panel with intensity scale values using references

Test Design: o Panel will be asked to generate a lexicon for fried dough balls (preparation instructions follows)

Instructions: o Prepare the following product:

Fried dough balls

Ingredients: • 1 cup water • 2 1/2 tablespoons white sugar • 1/2 teaspoon salt • 2 tablespoons vegetable oil • 1 cup all-purpose flour • 2 quarts oil for frying

Methods: o In a small saucepan over medium heat, combine water, 2 1/2 tablespoons sugar, salt and 2 tablespoons vegetable oil. Bring to a boil and remove from heat. Stir in flour until mixture forms a ball. o Heat oil for frying in deep-fryer or deep skillet to 375 degrees F (190 degrees C). Drop balls of dough into hot oil. Fry until golden; drain on paper towels.

o Sample is coded 312 and presented to the panel. o Panelists were asked to describe the product.

Lexicon Exercise Worksheet

Name:

Instructions: You have a sample on the tray in front of you. Record down the sample code before tasting the sample. Taste and describe the sample according to the instructions provided.

Sample code: ______

1. Look at the sample (without touching) and describe the appearance. Give attention to characteristics such as surface moisture, surface roughness (smoothness), and color. Try to rate the hardness and crispness of the product.

APPENDIX P: TRAINING SESSION 9

Date: July 17, 2007 Venue: Grover Center W320

Objectives: o Familiarize panel with intensity scale values using references o Generate lexicon for cooked ground chicken

Test Design: o Evaluation of panel’s perception of crispness.

Code reference brand Sample size 451 Granola bar Quaker low fat chewy chunk 1/3 bar 124 Club cracker Keebler partner club cracker ½ cracker 682 Graham cracker Honey maid 1 in square 914 Oat cereal Cheerios 1 oz 694 Bran flakes Kellogg’s bran flakes cereal 1 oz 385 Cheese crackers Pepperidge farm cheddar cheese 1 oz goldfish crackers 685 Corn flakes Kellogg’s corn flakes cereal 1 oz

o Panel will be presented with the samples. o Panel will be asked to rank the hardness of the samples using a 15-point scale (1 = not crisp/soggy, 15 = very crisp). o Results will be compared to the reference set (refer to training session 4)

o Generate lexicon for cooked ground chicken o Panel will be presented with cooked ground chicken (ground chicken cooked in a bag in boiling water for 15 minutes. o Record and discuss attributes that are listed by panelists.

Crispness Intensity Exercise Worksheet

Date:

Name:

Instructions: Record down the sample code before tasting the sample. Taste the samples and rate the crispness of the samples. Scale value 1 and 15 have been identified for you. Do not discuss with other panel members.

Code Scale value

451 1

685 15

______

______

______

______

______

APPENDIX Q: TRAINING SESSION 10

Date: July 19, 2007 Venue: Grover Center E120

Objectives: o Practice of basic taste recognition o Practice of usage of intensity scale values using references for hardness and crispness

Test Design: o Panel will go through basic taste recognition. o Reference set: Label Content ( in 250 ml water) Salty 0.10 g salt Sweet 0.5 g sugar Sour 0.05 g citric acid Bitter Umami 0.025 g umami

o Practice set: Label Code Sweet + sour 226 Salty + bitter 933 Sweet + umami 693 Sweet + bitter 148

o Panel will be asked to taste references for hardness and crispness

o Standard hardness scale reference: Scale reference brand Sample size value 1 Cream cheese Kraft/Philadelphia light ½ in cube 2.5 Egg white Hard cooked ½ in cube 4.5 Cheese Yellow American pasteurized process/ ½ in cube land O’lakes 6 Olives Goya foods/giant size, stuffed 1 olive pimento, removed 7 Frankfurter Large, cooked 5 min/Hebrew national ½ in slice 9.5 Peanuts Cocktail type in vacuum tin/Planters 1 nut, whole 11 Carrots Uncooked, fresh, unpeeled ½ in slice 14.5 Hard candy lifesavers 3 pieces, one color

o Standard crispness scale reference: Scale reference brand Sample size value 2 Granola bar Quaker low fat chewy chunk 1/3 bar 5 Club cracker Keebler partner club cracker ½ cracker 6.5 Graham cracker Honey maid 1 in square 7 Oat cereal Cheerios 1 oz 9.5 Bran flakes Kellogg’s bran flakes cereal 1 oz 11 Cheese crackers Pepperidge farm cheddar cheese 1 oz goldfish crackers 14 Corn flakes Kellogg’s corn flakes cereal 1 oz

o Present samples to panelists in order of scale value. o Allow panelists to familiarize themselves with the scale. o Practice applying the hardness reference scale on: ƒ Yellow cake (254) ƒ Lima beans (353) o Practice applying the crispness reference scale on: ƒ Melba toast (376) ƒ popcorn (981)

Scorecard 10a (basic tastes)

Name:

Instructions: You have four samples on the tray in front of you. Record down the sample code before tasting the sample. Taste the samples from left to right and indicate the taste by checking the appropriate space. Do not repeat the evaluation of previous samples.

Sample Code Sweet Sour Salty Bitter Umami

______

Scorecard 10b (hardness)

Name:

Instructions: You have a sample on the tray in front of you. Record down the sample code before tasting the sample. Follow the instructions.

Sample code: ______

Place sample between your front teeth and bite through the sample once. Evaluate the hardness. Refer to the reference scale for hardness.

Sample code: ______

Place sample between your front teeth and bite through the sample once. Evaluate the hardness. Refer to the reference scale for hardness.

Scorecard 10c (crispness)

Name:

Instructions: You have a sample on the tray in front of you. Record down the sample code before tasting the sample. Follow the instructions.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the crispness. Refer to the reference scale for crispness.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the crispness. Refer to the reference scale for crispness.

APPENDIX R: TRAINING SESSION 11

Date: July 24, 2007 Venue: Grover Center E120

Objectives: o Practice of usage of intensity scale values using references for juiciness

Test Design: o Panel will be asked to taste references juiciness o Panel will practice on chicken products and frankfurter.

o Standard juiciness scale reference: Scale value Reference Type Sample size 1 Banana Raw ½ inch slice 2 Carrot Raw ½ inch slice 4 Mushroom Raw ½ inch slice 7 Snap beans Raw 5 pieces 8 Cucumber Raw ½ inch slice 10 Apple Raw ½ inch wedge 12 Honeydew melon Raw ½ inch cube 15 Watermelon Raw ½ inch cube

o Present samples to panelists in order of scale value. o Allow panelists to familiarize themselves with the scale. o Practice applying the juiciness reference scale on the products prepared: o Banquet Chicken Breast Nuggets® o Banquet Chicken Breast Patties® o Hebrew National frankfurter®

Scorecard 11

Name:

Instructions: You have three samples on the tray in front of you. Record down the sample code before tasting the sample. Taste the samples from left to right. Follow the instructions.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the juiciness. Refer to the reference scale for juiciness.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the juiciness. Refer to the reference scale for juiciness.

Sample code: ______

Place sample between your molars and bite through twice. Evaluate the juiciness. Refer to the reference scale for juiciness.

APPENDIX S: TRAINING SESSION 12

Date: July 26, 2007 Venue: Grover Center E120

Objectives: o Practice of usage of intensity scale values using references for juiciness

Test Design: o Panel will be asked to taste references juiciness o Panel will practice on chicken products and frankfurter.

o Standard juiciness scale reference: Code Reference Type Sample size 163 Banana Raw ½ inch slice 401 Carrot Raw ½ inch slice 947 Mushroom Raw ½ inch slice 755 Snap beans Raw 5 pieces 261 Cucumber Raw ½ inch slice 615 Apple Raw ½ inch wedge 630 Honeydew melon Raw ½ inch cube 435 Watermelon Raw ½ inch cube

o Panel will be presented with the samples. o Panel will be asked to rank the hardness of the samples using a 15-point scale (1 = not juicy, 15 = very juicy). o Results will be compared to the reference set (refer to training session 11)

Juiciness Intensity Exercise Worksheet

Date:

Name:

Instructions: Record down the sample code before tasting the sample. Taste the samples and rate the juiciness of the samples. Scale value 1 and 15 have been identified for you. Do not discuss with other panel members.

Code Scale value

163 1

435 15

______

______

______

______

______

______

APPENDIX T: SENSORY ATTRIBUTES FOR COOKED GROUND CHICKEN

Sensory attributes and its description and anchored references, and their position on the 15-cm line scale as identified for cooked ground chicken.

Position Attribute Description Reference/Brand/Preparation (cm) Color Color of the outer surface of the sample Boiled chicken breast 1 Boiled beef 15

Chicken brothy Aromatics associated with chicken broth Chicken broth 9.9

Fishy Aromatics associated with cooked fish Boiled fish 9.9

Sulfury Aromatics associated with boiled egg yolk Boiled egg yolk 5.0

Musty Aromatics associated with wet cardboard Wet cardboard 4.0

Sweet The amount of sweet taste detected from 5% sucrose solution 5.0 the sample as it is being chewed before being swallowed or expectorated.

Sour The amount of sweet taste detected from 0.8% citric acid solution 5.0 the sample as it is being chewed before being swallowed or expectorated.

Salty The amount of salty taste detected from 0.5% NaCl solution 5.0 the sample as it is being chewed before being swallowed or expectorated.

Bitter The amount of bitter taste detected from 0.8% caffeine solution 5.0 the sample as it is being chewed before being swallowed or expectorated.

Umami Flavor associated with monosodium 0.5% monosodium glutamate 7.6 glutamate. solution

Serumy/metallic Flavor associated with blood or rare meat Rare beef 3.0

Cooked chicken Flavor associated with cooked white Boiled chicken breast 8.0 chicken meat

Fatty Flavor associated with animal fat Chicken fat 8.0

Fishy Flavor associated with cooked fish Boiled fish 11.0

Rancid Flavor associated with rancid/oxidized oil Oxidized oil 6.0

APPENDIX U: BALLOT FOR COOKED GROUND CHICKEN

APPENDIX V: SENSORY ATTRIBUTES FOR FRIED, BATTERED, AND

BREADED CHICKEN PATTIES

Sensory attributes and its description and anchored references, and their position on the 15 cm line scale as identified for deep-fried, battered, and breaded chicken patties.

Position Attribute Description Reference/Brand/Preparation (cm) 1 Color Color of the top surface of the sample CMP warm-up sample 5.7 (0 – yellow, 15 – dark brown)2 JBP warm-up sample 10.4

Evenness of Evenness of the color of the top CMP warm-up sample 7.9 color surface of the sample JBP warm-up sample 8.3 (0 – even, 15 – not even/blotchy)2

Greasiness The amount of grease that is CMP warm-up sample 6.6 of surface perceived from looking at the top JBP warm-up sample 7.0 surface of the sample (0 – not greasy, 15 – greasy)2

Hardness The force that is required to bite Cheese/pasteurized American/ 1/2 in 4.5 through the sample with incisors 3 slice CMP warm-up sample 4.8 JBP warm-up sample 5.4 Olive/Goya foods®/one giant size 6.0 Frankfurter/ Hebrew national®/large, 7.0 cooked 5 min/ ½ in slice Peanuts/ Planters®/cocktail type in vacuum tin 9.5

Crunchiness The force and noise with which a Defrosted fries/Ore-Ida® Golden 1.2 product breaks or fractures (rather Fries/brought to room temperature than deforms) when chewed with the CMP warm-up sample 6.5 molar teeth 3 JBP warm-up sample 8.9 Cereal/Quaker® Oatmeal squares 12.7

Moisture The amount of moisture released Carrot/1 inch cubes 2.0 release during a predetermined number of Mushroom/button/quartered 4.0 chews 3 JBP warm-up sample 4.0 Snap beans/1/2 inch pieces 7.0 CMP warm-up sample 10.0

Oily mouth The amount of oily coating that is JBP warm-up sample 3.7 coating perceived in the mouth cavity after CMP warm-up sample 4.1 the sample has been swallowed or Cold fries/Ore-Ida® Golden Fries/deep- 6.0 expectorated 2 fried, cooled to room temperature

Salty The amount of salty taste detected JBP warm-up sample 1.3 from the sample as it is being CMP warm-up sample 2.2 chewed before being swallowed or 0.5% NaCl solution 5.0 expectorated 2

Sweet The amount of sweet taste detected JBP warm-up sample 0.2 from the sample as it is being CMP warm-up sample 1.0 chewed before being swallowed or 5% sucrose solution 5.0 expectorated 2

Sour The amount of sour taste detected CMP warm-up sample 0.1

from the sample as it is being JBP warm-up sample 0.2 chewed before being swallowed or 0.8% citric acid solution 5.0 expectorated 2

Bitter The amount of bitter taste detected CMP warm-up sample 0.0 from the sample as it is being JBP warm-up sample 0.0 chewed before being swallowed or 0.8% caffeine solution 5.0 expectorated 2

Umami The amount of umami taste detected JBP warm-up sample 0.0 from the sample as it is being CMP warm-up sample 0.6 chewed before being swallowed or 0.5% monosodium glutamate solution 7.0 expectorated 2

Chicken fat The amount of chicken fat flavor CMP warm-up sample 3.7 flavor detected from the sample as it is JBP warm-up sample 3.9 being chewed before being Render chicken fat 9.6 swallowed or expectorated 2

Chickeny The amount of chicken flavor CMP warm-up sample 5.1 detected from the sample as it is JBP warm-up sample 6.4 being chewed before being Chicken breast/cooked in bag in boiling 10.1 swallowed or expectorated 2 water for 15 min

Cooking oil The amount of cooking oil flavor CMP warm-up sample 2.3 flavor detected from the sample as it is JBP warm-up sample 4.9 being chewed before being Used frying oil/Canola oil with added 9.7 swallowed or expectorated 2 dimethylpolysiloxane

Whey flavor The amount of cooking oil flavor CMP warm-up sample 0.8 detected from the sample as it is JBP warm-up sample 1.1 being chewed before being 5% whey solution 9.7 swallowed or expectorated 2 1 Position on 15-cm line scale 2 Generated by descriptive analysis panel 3 Adapted from Meilgaard et al. (1999)

APPENDIX W: BALLOT FOR CRACKERMEAL-COATED PATTIES

APPENDIX X: BALLOT FOR JAPANESE BREADCRUMB-COATED PATTIES

APPENDIX Y: SAMPLING CODE AND ORDER FOR CRACKERMEAL-

COATED PATTIES

WPI concentration Code Replication pH level (% w/w) Session 1 106 2 3 0 156 2 3 5 286 2 2 2.5 328 1 2 10 336 1 3 0 445 2 2 10 Session 2 524 2 2 0 607 2 3 2.5 686 1 control control 723 2 control control 725 1 3 5 768 1 3 2.5 Session 3 826 2 2 5 868 1 2 2.5 876 1 3 10 927 2 3 10 929 1 2 5 972 1 2 0

APPENDIX Z: SAMPLING CODE AND ORDER FOR JAPANESE

BREADCRUMB-COATED PATTIES

WPI concentration Code Replication pH level (% w/w) Session 1 112 2 3 10 167 2 3 5 221 1 3 5 233 1 3 0 254 1 2 10 Session 2 300 1 8 5 377 2 8 2.5 389 2 2 10 415 2 control 423 2 3 0 471 1 2 5 Session 3 473 1 3 10 519 1 8 10 562 2 2 5 653 1 control 761 2 8 5 783 1 2 0 Session 4 808 2 3 2.5 823 2 8 10 828 1 3 2.5 Session 5 886 2 8 0 945 1 8 0 951 1 8 2.5 962 2 2 2.5 978 2 2 0 984 1 2 2.5