UNIVERSITI PUTRA MALAYSIA PRODUCTION OF PROCESSED … · SFC Solid Fat Content SMP Slip Melting...

UNIVERSITI PUTRA MALAYSIA

PRODUCTION OF PROCESSED CHEESE USING LIPASE-CATALYZED TRANSESTERIFIED PALM KERNEL OLEIN

AND ANHYDROUS MILK FAT MIXTURES

MARIAM MOHAMED ISMAIL

FSMB 1999 2

PRODUCTION OF PROCESSED CHEESE USING LIPASE-CATALYZED TRANSESTERIFIED PALM KERNEL OLEIN AND ANHYDROUS MILK

FAT MIXTURES


MASTER OF SCIENCE UNIVERSITI PUTRA MALAYSIA

1999


FAT MIXTURES

By


Thesis submitted in Fulfillment of the Requirements for the Degree of Master of Science in the Faculty of

Food Science and Biotechnology U niversiti Putra Malaysia

July 1999

To those who are proud of my success

my life partner,

Fareg

and my beloved daughter,

Eman

ACKNOWLEDGEMENT

All praise due to Allah, the Most Gracious and Merciful, for giving me the

strength, health and determination to complete my study.

Then, I would like to express my deepest gratitude, honor and sincere

appreciation to my supervisor Professor Dr. Hasanah Mohd Ghazali for her invaluable

guidance, encouragement, constructive suggestions throughout the study period, and for

giving me total freedom to explore my research. Her enthusiasm and patience have left a

feeling of indebtedness that can not be fully expressed. I would like to thank her for

sponsoring my study.

My heartfelt appreciation and gratitude go to the members of my supervisory

committee, Dr. Mohd. Yazid Abd. Manap, Associate Professor Dr. Salmah Yusof, Dr.

Kamariah Long and Mr. Dzulkifly Mat Hashim for their generous guidance, valuable

comments during the study and for reviewing my manuscript and making valuable

comments from time to time.

My special appreciation is also extended to Dr. Lai Oai Meng for her warm

friendship and helpful comments while she was a graduate student at the department of

Biotechnology, Faculty of Food Science and Biotechnology. I wish to express my

sincere appreciation to my colleagues, namely, Maryam, Hanan, Tri Haryati, Gabby,

Tan Chin Ping, Shuhaimi and Khalid for their friendship and support.

111

My deep appreciation is accord to the Dean, staff members, technicians and my

colleagues, Faculty of Food Science and Biotechnology for their support and

cooperation. Special thanks to Mr. Mohd Amran bin Suratman, Food Processing Lab.

for his assistance in the formulation of plastic casein. Mrs. Asmarani Abdullah at the

Deputy Dean's Office for her assistance.

I also wish to extend my sincere gratitude and thanks to the staff of the Palm Oil

Research Institute Malaysia (PORIM) for allowing me to formulate the experimental

processed cheese at their labs. The assistance and support by Dr. Chong Chiew, Mrs.

Karimah Ahmad and their co-workers at the Physics and Processing Labs are deeply

appreciated.

Most of all I am grateful to my husband, Fareg Mohamed Aldebrez and my

daughter Eman for their encouragement, endless patience, and sacrifice all the way

throughout this work. Without the benefit of their loving support and dedication, I

would not have had the wonderful opportunities that I have experienced.

Last but not least, heartfelt appreciation goes to the Government of Malaysia and

Universiti Putra Malaysia for providing the financial support for this study through the

IRP A program (No. : 03-02-04-0025) awarded to Prof Dr. Hasanah Mohd Ghazali.

IV

TABLE OF CONTENTS

Page

ACKNOWLEDGEMENTS... . . . ... ... ... ... ... ... ... ... ... ... ... ... ... ... ... ..... 111 LIST OF TABLES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . vu LIST OF FIGURES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IX LIST OF PLATES . . . ... . .. ... ... . .. ... . .. . . . . . . . .. ... ... .. . . . . ... ... . . . . . . ... .. ... Xl LIST OF ABBREVIATIONS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xu ABSTRACT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xv ABSTRAK . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . XVll

CHAPTER

I

II

ill

INTRODUCTION ....................................................... . 1

LITERATURE RE�W .............................................. . 6

Fats and Oils . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . .. 6 Structure and Composition of Fats and Oils . .. . . . . . . ... . . . . . . .. 6 Types of Fats and Oils . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 Reactions of Fats and Oils . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16

Lipases . . . ........ ... ... ... ...... ... ... ... ... ... ... ... ... ... ... ... ... ... 17 Definition of Lipases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 17 Lipase Selectivity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20 Reactions of Lipases . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . 23 Applications ofLipases . .. . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . 29

Cheese . . . . . . . . . ... . . . .. . ... ... ... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . 32 History and Background . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . 32 Definition of Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32 Types of Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33 Techniques and Procedures Used in Cheese Production . .. ... 34 Factors Affecting The Physical Properties of Cheese . . . . . . .. . . . 35 Processed Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36

Sensory Evaluation and Texture Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40 Sensory Evaluation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 40 Texture Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42

MATERIALS AND METHODS 44

Enzymatic Transesterification of Palm Kernel Olein and Anhydrous Milk Fat . .. ... . . . . . . . . . . . . . . . . . . . . . . . 44

Materials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44 Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45

v

IV

v

Formulation of Processed Cheese Using Transesterified Fat . . . . . . . . . . . 5 1 Preparation of Starter Culture . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 1 Preparation of Plastic Casein . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 1 Formulation of Processed Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59

Chemical Analyses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62 Analysis of Plastic Casein . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62 Analysis of Produced Processed Cheese Blends . . . . . . . . . . . 67

Sensory Evaluation and Texture Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67 Sensory Evaluation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67 Texture Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69

RESULTS AND DISCUSSION 71

Enzymatic Transesterification of Palm Kernel Olein and Anhydrous Milk Fat . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71

Triglyceride Compositions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 1 Degree of Hydrolysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77 Slip Melting Point . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79 Solid Fat Content . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . , . . . . . . . . , 8 1 Crystal Forms of Trans esterified PKO and AMP . . . . . . . . . . . 84 Thermal Behaviour . . . . . . . . . . .. .. ... . . . . . . . . . . .. . .. . . . . . .... . . . . . , . 89

Production of Processed Cheese Using Transesterified Fat . . . . . . . . . . . 99 Plastic Casein . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99 Processed Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 102

Sensory Evaluation and Texture Analysis of Produced Processed Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 106

Sensory Evaluation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 106 Texture Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 107 Relating Sensory and Non-Sensory Data . . . . . . . . . . . . . . . . . . . , 1 1 0

CONCLUSION AND RECOMMENDATIONS 1 12

Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 12 Recommendations for Further Study . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 1 5

BmLOGRAPHY 1 1 6

APPENDIX

APPENDIX A 137

APPENDIXB 141

APPENDIX C 144

VITA 147

VI

Table

1

2

3

4

5

6

7

8

9

1 0

1 1

12

1 3

1 4

1 5

1 6

LIST O F TABLES

Chemical Characteristics of Palm Kernel Olein

Physical Properties of Palm Kernel Olein

Fatty Acids Composition of Anhydrous Milk Fat . . . . . . . . . . . . . . . . . . . . . .

Physical and Chemical Properties of Anhydrous Milk Fat

Amount of Ingredients in the Final Product . . . . . . . . . . . . . . . . . . . . . . . . . . .

X-Ray Diffiaction Analysis of Pure PKO and AMF

X-Ray Diffiaction Analysis of the 3 : 7 PKO:AMF

X-Ray Diffiaction Analysis of the 1 : 1 PKO:AMF

X-Ray Diffiaction Analysis of the 7 :3 PKO:AMF

Last Melting Peak (LMP) and Slip Melting Point (SMP) of Pure PKO and AMF Fats . . . . . . . . . . . . . . . . , . . . . . . . . . . . . . . , . . . . . . . . . . . . .

Last Melting Peak (LMP) and Slip Melting Point (SMP) of the 3 :7 PKO:AMF Reaction Mixture . . . . . . . . . . . . . . . . . . . . . . . . . . .

Last Melting Peak (LMP) and Slip Melting Point (SMP) of the 1: 1 PKO:AMF Reaction Mixture . . . . . . . . . . . . . . . . . . . . . . . . . . .

Last Melting Peak (LMP) and Slip Melting Point (SMP) of the 7:3 PKO:AMF Reaction Mixture . . . . . . , . . . . . . . . . . . . . . , . . . . . .

Chemical Analyses of Commercial Cheddar Cheese and Prepared Plastic Casein . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Chemical Analysis of Processed Cheese Blends

Texture Analysis of Processed Cheese Blends

V11

Page

10

1 1

14

15

60

86

86

87

87

96

97

97

98

1 00

104

1 10

1 7 Sensory Data Vs Texture Analyzer Data of Processed Cheese Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1 8 Degree of Hydrolysis of the Palm Kernel Olein and Anhydrous Milk Fat and their Reaction Mixtures before and after

1 9

20

2 1

22

23

24

Transesterification

Slip Melting Point of Palm Kernel Olein and Anhydrous Milk Fat and their Reaction Mixtures before and after Transesterification

Solid Fat Content of Pure PKO and AMF

Solid Fat Content of the 3 : 7 PKO:AMF Reaction Mixture


Solid Fat Content of the 7 :3 PKO:AMF Reaction Mixture

Amount of Ingredients in the Final Product . . . . . . . . . . . . . . . . . . . . . . . . . . .

viii

1 1 1

137

1 38

1 38

1 39

1 39

140

143

Figure

1

2

3

4

5

6

7

8

9

1 0

1 1

12

1 3

1 4

1 5

1 6

LIST OF FIGURES

The Lipase Reaction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Positional Specificity of Microbial Lipase . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Flow Chart of the Preparation Process of Plastic Casein

Determination of Total Protein Using Micro Kjeldahl Method . . . . . . .

Triglyceride Profile of Palm Kernel Olein

Triglyceride Profile of Anhydrous Milk Fat

Triglyceride Profile of the 3 : 7 PKO:AMF Reaction Mixture before and after Transesterification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Triglyceride Profile of the 1 : 1 PKO:AMF Reaction Mixture before and after Transesterification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Triglyceride Profile of the 7:3 PKO:AMF Reaction Mixture before and after Transesterification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Free Fatty Acid Content ofPKO, AMF and their Reaction Mixtures before and after Transesterification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Slip Melting Point ofPKO&AMF and their Reaction Mixtures . . . . . .

Solid Fat Content of Pure PKO and AMF Fats



Solid Fat Content of the 7:3 PKO:AMF Reaction Mixture

Heating Thermogram ofPKO . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

lX

Page

1 8

2 1

5 8

65

72

72

73

74

75

78

80

82

82

83

83

90

1 7

1 8

1 9

20

2 1

22

23

24

25

26

27

28

29

Heating Thermogram of AMF

Heating Thermogram of the 3 : 7 PKO:AMF before and after Transesterification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Heating Thermogram of the 1 : 1 PKO:AMF before and after Transesterification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Heating Thermogram of the 7 :3 PKO:AMF before and after Transesterification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Titrable Acidity of Plastic Casein during Preparation Process . . . . . . . . .

Multiple Comparison Test for Processed Cheese Sample

Texture Profile of Internal Control Processed Cheese

Texture Profile of Non-Transesterified Fat Processed Cheese . . . . . . . . .

Texture Profile of Transesterified Fat Processed Cheese

Texture Profile of Commercial Processed Cheese

Sensory Data Vs Texture Analyzer Measurement Relationship . . . . . . .

Panelists to Numerical Scores Conversion Scale

Schematic Diagram of Texture Profile of the Cheese Sample

x

9 1

93

94

95

101

1 07

1 08

108

109

109

1 1 1

146

146

Plate

1

2

3

4

5

6

7

8

9

1 0

1 1

12

1 3

14

1 5

1 6

1 7

LIST OF PLATES

Prepared Starter Sub-Culture . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Ingredients of Plastic Casein . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Pasteurization of Reconstituted Skimmed Milk Powder

Addition of Sub-Culture

Addition of Enzyme . . . . . . . . . . . . . . . . . . . . . . . . . . . . . , . . . . . . . . . . . . . . . . . . . . . ' "

Cutting the Curd with Cheese Knife . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Cheddaring of the Curd . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Molding the Curd with a Muslin Cloth before Pressing

Pressing the Plastic Casein using Manual Screw Press

Major Ingredients of Processed Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . .

Stephan Direct Steam Injector Stainless Steel Cooker . . . . . . . . .

Paired Comparison Test of Processed Cheese Samples

Multiple Comparison Test of Processed Cheese

Stable Micro System (SMS) Texture Analyzer

Internal Control Prepared Processed Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Transesterified Fat Processed Cheese . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Non-Transesterified Fat Processed Cheese

Xl

Page

53

54

54

55

55

56

56

57

57

6 1

61

68

69

70

1 04

105

1 05

LIST OF ABBREVIATIONS

Symbol Meaning

AMF Anhydrous Milk Fat

AOCS American Oil Chemist Society

BS British Standard

C 12 :0 Lauric Acid

C 14:0 Myristic Acid

C 1 6:0 Palmitic Acid

C I 6 : I Palmitoleic Acid

C 1 8 :0 Stearic Acid

C I 8 : I Oleic Acid

C 1 8 :2 Linoleic Acid

C 1 8 : 3 Linolenic Acid

C22 : I Erucic Acid

Ca2+ Calcium

CBS Cocoa Butter Substitute

DG Diglycerides

DSC Differential Scanning Calometry

FA Fatty Acids

FFA Free Fatty Acids

xu

HMG

HPLC

IDF

IV

LMG

LMP

LOO

LOP

MG

MMG

Na+

N a3C6Hs07

NBD

NMR

NTF

000

pH

PKO

POP

PORIM

POS

PPP

High Melting Glycerides

High Performance Liquid Chromatography

International Dairy Federation

Iodine Value

Low Melting Glycerides

Last Melting Peak

l -linoleoyl-dioleoyl glycerol

1 -linoleoyl-2-oleoyl-palmitoyl glycerol

Monoglycerides

Medium Melting Glycerides

Sodium

Sodium Citrate

Neutralized Bleached Deodorized

Nuclear Magnetic Resonance

Non Transesterified Fat

Triolein

Hydrogen Ion

Palm Kernel Olein

1 ,3-dipalmitoyl -2-oleoyl glycerol

Palm Oil Research Institute of Malaysia

I -palmitoyl-2-oleoyl-stearoyl glycerol

Tripalmitin

Xlll

PPS 1,2-dipalmitoyl-stearoyl glycerol

r Correlation Coefficient

RBD Refined Bleached Deodorized

rpm Revelation Per Minute

SFC Solid Fat Content

SMP Slip Melting Point

SMS Stable Micro System

Sp Species

TA Titrable Acidity

TF Transesterified PKO and AMF Fat

TG Triglyceride

TLC Thin Layer Chromatography

v/v Volume Per Volume

w/w Weight Per Weight

XRD X-Ray Diffraction

XIV

Abstract of thesis presented to the Senate ofUniversiti Putra Malaysia in fulfillment of the requirements for the degree of Master of Science.


FAT MIXTURES

By


July 1999

Chairman: Professor Hasanah Mohd. Ghazali, Ph.D.

Faculty: Food Science and Biotechnology

In recent years, lipase-catalyzed transesterification has received considerable

attention from the fats and oils industry due to its potential to modify the physical and

chemical properties of edible fats and oils. In this study, enzymatic transesterification

of three different ratios of palm kernel olein (PKO) and anhydrous milk fat (AMF)

was carried out using immobilized lipase from Rhizomucor miehei (Lipozyme 1M 60)

to modify their chemical and physical properties, such that they can be used in the

production of processed cheese.

After transesterification for 8 hours, there were either increases or decreases

in the concentrations of several existing triglycerides (TG) of the reaction mixtures

and, in some cases, the formation of new peaks. The results also showed that

increasing the reaction time, increased the % free fatty acid (FF A) between 1 to 2%.

xv

The slip melting points (SMP) of the fat mixtures, which were determined after

removal of free fatty acids, declined after the transesterification process. The solid fat

content (SFC) decreased with respect to the control samples. At 40°C, the PKO:AMF

(7:3) transesterified fat mixture was completely melted, making it a potential

candidate in the formulation of processed cheese. Thermal profiles showed that the

original peaks of the transesterified mixtures have either shifted or completely

changed and some new peaks were also formed. It was also found that the

transesterified fat mixtures were in the �'

form, compared to a predominantly � form

before transesterification.

After the preparation of experimental processed cheese, several chemical and

physical analyses of this final product were carried out. The results showed that the

experimental processed cheese had a satisfactory aroma and texture compared to the

commercial sample. It also recorded a high total solid content and a low amount of

moisture, with a good workability and overall general behaviour at cool, ambient, and

elevated temperatures.

Sensory evaluation and texture analysis studies were also conducted on the

final product. The experimental processed cheese samples achieved an acceptable

response from the panelists, where there was no significant difference between the

samples in terms of colour and texture. The texture profiles indicated that the

experimental processed cheese samples exhibited close texture profiles compared to

their counterpart commercial sample.

XVI

Abstrak tesis yang dikemukakan kepada Senat Universiti Putra Malaysia sebagai memenuhi keperluan untuk ijazah Master Sains.

PENGHASILAN KEJU TERPROSES MENGGUNAKAN CAMPURAN OLEIN ISIRONG KELAPA SA WIT DAN LEMAK SUSU TANPA AIR YANG

DITRANSESTERIFKASI MELALUI PEMANGKINAN BERLIPASE

Oleh


Julai 1999

Pengerusi : Profesor Hasanah Mohd. Ghazali, Ph.D.

Faculti : Sains Makanan dan Bioteknologi

Pada masa kini, transesterifikasi yang dimangkin oleh lipase telah menerima

perhatian dari industri minyak dan lemak disebabkan potensinya untuk mengubah

ciri-ciri fizikal dan kimia minyak dan lemak. Dalam kajian ini. transesterifikasi

secara ber enzim di antara campuran minyak sawit isirong olein (PKO) dan lemak

susu anhidrus (AMF) pada tiga nisbah yang berbeza telah dijalankan menggunakan

lipase tersekat-gerak dari Rhizomucor miehei (Lipozyme 1M 60) untuk mengubah

ciri-ciri kimia dan fizikal dalam campuran untuk membolehkan campuran minyak

untuk menghasilkan keju terproses.

XVll

Selepas transesterifikasi selama 8 hours, terdapat pengurangan atau

penambahan kepekatan beberapa trigliserida yang menang wujud dalam campuran

yang bertindakbalas, dan pembentukan puncak-puncak baru dalam kes-kes tertentu.

Keputusan juga menunjukkan bahawa peningkatan masa tindakbalas telah

menyebabkan penambahan % asid lemak bebas di antara 1 hingga 2%. Titik lebur

(SMP) campuran-campuran lemak yang ditentukan setelah dikeluarkan asid-asid

lemak bebas didapati menurun selepas proses transesterifikasi. Kandungan pepejal

lemak (SFC) telah berkurangan berbanding dengan sampel-sampel kawalan. Pada

suhu 40°C, campuran lemak PKO:AMF (7:3) tertransesterifikasi telah lebur

sepenuhnya. 1ni menjadikan campuran tersebut berpotensi dalam formulasi keju

terproses. Gambarajah terma menunjukkan bahawa puncak-puncak asal dalam

campuran-campuran tertransesterifikasi telah samada beralih tempat atau berubah

sepenuhnya dan beberapa puncak baru telah terbentuk. Selain itu, keputusan juga

menunjukkan bahawa campuran-campuran lemak tertransesterifikasi menghasilkan

bentuk kristal jenis f3', dimana bentuk f3 adalah dominan sebelum transesterifikasi.

Selepas penyediaan keju terproses, beberapa analisis kimia dan fizikal produk

akhir telah dilakukan. Keputusan yang diperolehi menunjukkan behawa keju terproses

yang disediakan mempunyai aroma yang memuaskan dan tekstur yang setanding

dengan sampel komersial. Produk juga merekodkan kandungan pepejal yang tinggi

dan kuantiti lembapan yang rendah dengan kebolehan keIja yang baik dan sifat umum

keseluruhan pada suhu-suhu sejuk, ambien dan tinggi.

XVI11

Kajian penilaian deria dan analisis tekstur juga dijalankan pada produk akhir.

Sarnpel-sarnpel keju terproses yang diediakan telah mencapai respon yang boleh

diterima daripada para ahli panel dari segi warna dan tekstur sifat sarnpel-sampel ini

tidak menunjukkan perbezaan yang bererti. Sarnpel-sampel keju terproses

menunjukkan sifat tekstur yang hampir sarna dengan sarnpel komersial.

XIX

CHAPTER I

INTRODUCTION

Fats and oils are complex mixtures of simple and mixed triglycerides that

function as energy reserves. Triglycerides differ according to the types and composition

of fatty acids and their distribution along the glycerol backbone. These characteristics

play an important role in the physical, rheological, and nutritional properties of fats and

oils.

In recent years, the use of lipases as catalysts has received considerable attention

because of its potential ability to modify the physical and chemical properties of edible

fats and oils (Kyotani et al. 1 988; Yamane et al. 1989; Miller et aI . , 1 99 1 ; Choo and

Rhee, 1 993 ; Foglia et aI . , 1 993 ; Ghazali et aI . , 1 995b; Lai et aI., 1 998). The main

technical application of lipases is to modify the fatty acid compositions of triglycerides

by interesterification.

Lipases are a group of enzymes that preferentially catalyze the hydrolysis and

synthesis of triglycerides. Immobilized lipases show many advantages over traditional

inorganic catalysts (Eigtved et at . , 1 986; Hansen et at . , 1 986; Miller et aI . , 1 988). They

1

2

have large catalytic activity under mild operating conditions. They show large selectivity

to the desired product with no significant side reactions, leading to products of high

purity. They are easily recovered from the reaction mixture and can be reused. There is

no contamination of the final product, saving time and cost in the purification stage

(Coteron et aI. , 1998).

Transesterification with lipase provides a useful way to improve the properties of

triglyceride. Through enzymatic transesterification, it is possible to incorporate a desired

acyl group onto specific positions of the glycerol, whereas chemical transesterification

does not possess this regiospecificity due to the random nature of the reaction

(Sreenivasa, 1978; Norris, 1982; Gunstone and Norris, 1983). Thus, lipase-catalyzed

transesterification can provide regio- or sterospecific structured lipids for nutritional,

medical, and food applications (Akoh, 1995 ; Lee et aI. , 1997; 1998).

Transesterification reaction is most often confirmed by closely monitoring

changes in the product's physico-chemical properties (Goh et aI. , 1993; Kerry, et aI. ,

1993). Physical properties such as melting characteristics, crystallization, and thermal

behaviour are commonly used as specifications for fats and oils. The melting

characteristics including melting point and solid fat content of fat are among the most

important predictors of its functionality used in industry today, and many fat ingredients

are selected for applications based on these characteristics (Kaylegian et aI. , 1993). The

SFC profile over a given temperature range is an important characteristic in fat

functionality that can be used to predict how the fat will behave at different stages of

manufacturing (rolling and baking of doughs). Another important indicator of fat

3

functionality is its crystallization behaviour, which dictates the crystal form present and

the respective stabilities of crystals. Fats, in particular their component triglyceride

molecules, can crystallize in more than one polymorphic forms which differ from each

other in terms of melting point, heat of fusion and crystallization, and density. The a

form is a form that crystallises from the melt, has a strong single short spacing at

0.4 1 5nm and exhibits an enantiotropic transition on further cooling Hagemann ( 1 988).

According to Hagemann ( 1988), D'Souza et ai. ( 1990) and Timms ( 1 991 ), the �' form is

a form exhibiting two strong short spacing bands at 0 .42 and 0.38nm, and the � form

exhibits three strong short spacings bands at 0.46, 0.39 and 0.37nm. Thermal analyses of

fats and oils are also indicators of their physical properties. Differential scanning

calorimetry (DSC) is the most convenient method for determining the thermal behaviour

of fats and oils (Hagemann, 1 988; Che Man et aI. , 1 995; Haryati et aI. , 1996). It can

provide much information on the behaviour of fats and oils.

Cheese is a commercial product, which is widely consumed and almost the one

food of good keeping ability, high in protein with fat, calcium phosphorus, riboflavin,

and other vitamins, in a concentrated form (Sukumar, 1 980). Cheeses of various kinds

always have been important sources of nutrients wherever milk-producing animals could

be raised. It has been an art handed down from generation to generation and was known

as a staple food with bread since 6000BC (Scott, 1 986). Natural cheese is made from the

curd of the milk of cows and other animals, the curd being obtained by the coagulation

of milk casein with an enzyme (usually rennin), an acid (usually lactic acid), and with or

4

without further treatment of the curd by heat, pressure, salt, and ripening with selected

microorganisms (Sukumar, 1 980).

Progress in the development of cheese production, combined with a considerable

increase in the quality of the finished product, made its appearance at the end of the 19th

century (Kosikowski, 1 982). However, there are some constraints (from a nutritional

viewpoint) on the use of cheese depending on the interpretation of the word 'cheese'

(Meyer, 1 973). Therefore, some fresh curds still hold lactose in the free whey and this

may cause allergic reactions in susceptible persons, while riper cheese with no free

lactose is safe. Extremely ripe cheese, where the protein is well broken down to

peptones and amino acids, can be absorbed very quickly and so create difficulty in some

persons, who should, therefore, avoid overripe cheese. Although these difficulties when

consuming cheese do exist for a minority of the population, the individual has usually a

wide assortment of varieties from which to choose so that some of the difficulties can be

avoided (Potter, 1 978) . However, the cheese may reach a condition that makes it unfit

for consumption. This is due to uncontrollable protein decomposition, where all types of

cheese, including the hard cheeses, are unsuitable for long storage, in the sense of

forming an "iron ration". In addition, the harder cheeses are also dependent on

temperature, for instance, it is a well-known fact that hard and semi-hard cheese in

tropical countries become unfavorably altered in flavour and texture (Meyer, 1 973).

To overcome the shortcoming of the natural cheese, the processed cheese

becomes a solution. The invention of processed cheese in 1 9 1 1 has been a very

important step forward in the dairy field (Kosikowski, 1 982). Processed cheese is the

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