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EFFECTS OF PROCESSING METHODS ON COMPOSITION, ANTINUTRIENTS, AND PHYSICOCHEMICAL PROPERTIES OF PALM

KERNEL CAKE

by

THARINDU DANANJA PEIRIS GALLAGE

Thesis submitted in fulfilment of the requirements for the degree of

Master of Science

July 2011

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ACKNOWLEDGEMENTS

First and foremost, I would like to express my sincere gratitude to my supervisor, Prof. Abd Karim bin Alias, whose guidance and generosity in providing ideas and knowledge has supported and encouraged me throughout both the experimental work and compiling the thesis. I would like to also thank my co-supervisor Dr. Fazilah Ariffin for precious guidance and support for the research. I would like to acknowledge the Institute of Postgraduate Studies for the funding of USM fellowship.

I am grateful to staff of Doping Control Centre, USM for their assistance and guidance in using LC/MS for vitamin analysis.

I would like to thank Chew Shio Heong, members of lab 215, lab 232, lab 233, and my apartment mates (Dr. Abu Tariq, Dr. Anis Khan, Dr. Irshad Bhat, Dr. Showkat Ahamed) and Dr. Aamir Bhat for their help along the way towards the accomplishment of my research study.

Last but not least, I want to express my eternal gratitude to my family for their continuous understanding, moral support and unconditional love I could not have done without.

Tharindu Dananja Peiris Gallage

School of Industrial Technology, USM

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TABLE OF CONTENTS

Page

Acknowledgement ii

Table of Contents iii

List of Tables viii

List of Figures ix

List of Abbreviations x

Abstrak xii

Abstract xv

CHAPTER 1 - INTRODUCTION 1

1.1 Introduction 1

1.2 Rationale of the Research 2

1.3 Hypothesis of the Research 4

1.4 Objectives 4

CHAPTER 2 - LITERATURE REVIEW 5

2.1 Palm Kernel Cake (PKC) 5

2.1.1 The Production Process of PKC 5

2.1.2 PKC as an Animal Feed 6

2.1.3 The Composition of PKC 6

2.1.3.1 Proximate Composition 6

2.1.3.2 Dietary Fiber Contents 11 2.1.3.3 Protein and Amino Acid Contents 14 2.1.3.4 Mineral Contents 17 2.1.3.5 Other Constituents 20 2.2 Antinutrients 22

2.2.1 α-amylase Inhibitory Activity 23

2.2.2 Hydrogen Cyanide 26

2.2.3 Oxalate 28

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2.2.4 Phytic Acid 30

2.2.5 Tannin 34

2.2.6 Saponin 39

2.3 Various Means, Applied to Reduce Antinutrient Contents in Plant 41 Products

2.4 Dietary Fiber 43

2.4.1 Non-starch Polysaccharide 43

2.4.1.1 Cellulose 43

2.4.1.2 Hemicellulose 45

2.4.2 Lignin 48

2.5 Hydrogen Peroxide and Uses 50

CHAPTER 3 - MATERIALS AND METHODS 53

3.1 Materials 53

3.2 The Nutritional Composition of PKC 53

3.2.1 Proximate Composition 53

3.2.1.1 The Determination of Moisture Contents 53 3.2.1.2 The Determination of Crude Protein Contents 54 3.2.1.3 The Determination of Crude Fat Contents 54 3.2.1.4 The Determination of Crude Fibre Contents 54 3.2.1.5 The Determination of Ash Contents 55 3.2.2 The Determination of Free Sugar Contents 55 3.2.3 The Determination of Free Amino Acid Contents 56 3.2.4 The Determination of Fatty Acid Contents 57

3.2.4.1 Sample Extraction 57

3.2.4.2 Preparation of FAME 57

3.2.4.3 GC-FID Conditions 57

3.2.5 The Determination of Mineral Contents 58

3.2.5.1 Sample Preparation 58

3.2.5.2 Instrumentation 58

3.2.6 The Determination of Vitamin Contents 58

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3.2.6.1 Standard Preparation 58

3.2.6.2 Sample Preparation 59

a. Water Soluble Vitamin Extraction 59

b. Fat Soluble Vitamin Extraction 59

3.2.6.3 LC/MS Conditions 60

3.3 Antinutritional Composition of PKC 61

3.3.1 α-amylase Inhibitory Activity 61

3.3.2 Hydrogen Cyanide Contents 62

3.3.3 Total and Soluble Oxalate Contents 62

3.3.4 Phytic Acid Contents 62

3.3.5 Tannin Contents 63

3.3.6 Total Saponin Contents 63

3.4 Dietary Fibre (DF) Contents 64

3.4.1 Non-starch Polysaccharide (NSP) Contents 64 3.4.1.1 Sugar Analysis in Non-starch Polysaccharide 65

a. Sample Preparation 65

b. HPLC Conditions for Sugar Analysis 65

3.4.2 Klason Lignin Contents 65

3.5 Physicochemical Treatments on PKC 67

3.5.1 Hydrogen Chloride Treatment 67

3.5.2 Hydrogen Peroxide Treatment 67

3.5.3 Heat Treatment (Boiling) 67

3.5.4 Removal of Residual Fat 68

3.5.5 Autoclaving 68

3.6 Enzymatic Treatments on PKC 68

3.6.1 Cellulase 68

3.6.2 Hemicellulase 68

3.6.3 Xylanase 69

3.7 Functional Properties 69

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3.8 Colour Change of PKC 69

3.9 Experimental Design and Statistical Analysis 70

CHAPTER 4 - RESULTS AND DISCUSSIONS 72

4.1 The Nutritional Composition of PKC 72

4.1.1 Proximate Composition 72

4.1.2. Vitamin Contents of PKC 73

4.1.3 Mineral Contents of PKC 74

4.1.4 Fatty Acid Contents of PKC 76

4.2 Antinutritional Factors in PKC 77

4.2.1 Phytic Acid Contents 77

4.2.2. Tannin Contents 79

4.2.3 Total, Soluble, and Insoluble Oxalate Contents 79

4.2.4 α-amylase Inhibitory Activity 81

4.2.5 Total Saponin Contents 81

4.2.6 Hydrogen Cyanide Contents 82

4.3 Effects of Physicochemical Treatments on Antinutrient Contents of PKC 82

4.3.1 Tannin Contents 82

4.3.2 Total Saponin Contents 86

4.3.3 Phytic Acid Contents 87

4.3.4 Hydrogen Cyanide Contents 88

4.3.5 Total, Soluble, and Insoluble Oxalate Contents 90

4.3.6 α-amylase Inhibitory Activity 92

4.4 Effects of Physicochemical and Enzymatic Treatments on Dietary Fiber 93 Contents of PKC

4.4.1 Effect on Insoluble Dietary Fiber 93

4.4.2 Effect on Soluble Dietary Fiber 97

4.4.3 Effect on Soluble and Insoluble Dietary Fiber Ratio (SDF: IDF) 99 4.4.4 Enhancement of Enzymatic Activity due to Physicochemical Pre 102

-treatments

4.4.5 Effects of Physicochemical Treatments on Lignin Contents 103

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4.5 Effects on Color of PKC due to Physicochemical Treatments 105

4.6 Effects on Functional Properties of PKC upon Physicochemical 107

Treatments 4.6.1 Effects on Water Holding Capacity (WHC) 107

4.6.2 Effects on Oil Holding Capacity (OHC) 112

CHAPTER 5 - CONCLUSION AND RECOMMENDATIONS 114

5.1 Conclusions 116

5.2 Recommendations 117

REFERENCES 118 APPENDICES

Appendix A: Standard curve for free sugar analysis (Glucose standard)

Appendix B: Standard curve for free amino acid analysis (Glutamic acid standard)

Appendix C: Standard curve for phytic acid analysis (Calcium phytate

standard)

Appendix D: Standard curve for total saponin analysis

Appendix E: Standard curve for tannin analysis (Catechin standard)

Appendix F: Standard curve for oxalate analysis (Sodium oxalate standard)

Appendix G: Fat soluble vitamin availability in PKC; (a) Vitamin A (b) Vitamin E (c) Vitamin K; chromatograms of the standards and the samples are aligned vertically for the each vitamin mentioned

Appendix H: Water soluble vitamin availability in PKC; (a) Vitamin B1

(b) Vitamin B2 (C) Vitamin B3 (d) Vitamin B5 (e) Vitamin B6 (f) Vitamin B7 (g) Vitamin B12; chromatograms for the standards and the samples are aligned vertically for the each vitamin mentioned

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LIST OF TABLES

Page

Table 2.1 Inclusion levels of PKC ruminants and non-ruminants feed 7 Table 2.2 A summary on proximate composition and chemical 9

compounds of PKC

Table 2.3 A summary of fibre contents in PKC 11

Table 2.4 Sugar contents of soluble and insoluble NSP in PKC 12 Table 2.5 A summary of amino acid contents in PKC (mg/g protein) 16

Table 2.6 A summary of mineral contents in PKC 18

Table 3.1 Instrumental parameters for the ICP-OES analysis 59 Table 4.1 Proximate composition and chemical compounds of PKC 73 Table 4.2 Mineral contents of PKC on dry basis (n = 3; mean ± SD) 75 Table 4.3 Fatty acids composition of PKC compared with fatty acids 78

contents of palm kernel

Table 4.4 Antinutrient contents of PKC on dry basis (n = 4; mean ± SD) 80 Table 4.5 Phytic acid, HCN, and α-amylase inhibitory activity of 89

physicochemically treated PKC samples

Table 4.6 Contents of insoluble, soluble, and total dietary fiber of PKC 95 and effects of physicochemical and enzymatic treatments on dietary fiber contents

Table 4.7 Effects of pre-treatments on soluble dietary fiber and enzymatic 104 activity

Table 4.8 Lignin and CIELAB values for treated PKC (n = 3; mean ± SD) 105 Table 4.9 WHC of different dietary fiber sources 112

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LIST OF FIGURES

Page

Figure 2.1 The reaction of myo-inositol and phosphoric acid to form phytic 31 acid

Figure 2.2 p-Penta-O-galloyl-D-glucose structure 36

Figure 2.3 Structure of flavan: The basic structure of condensed tannin 37 Figure 2.4 The structure of a cellulose polymer (Collinson and Thielemans, 44

2010)

Figure 3.1 Scheme of isolation and characterization of dietary fiber-rich 66 material from PKC.

Figure 3.2 Experimental design for the physicochemical and enzymatic 71 treatments of PKC.

Figure 4.1 Saponin and tannin contents of PKC with respect to different 84 physical and chemical treatments.

Figure 4.2 Total, soluble, and insoluble oxalate contents of PKC with 91 respect to different physicochemical treatments (n = 4).

Figure 4.3 Change in insoluble dietary fiber contents of PKC respect to 98 different combinations of pre-treatments and enzymatic

treatments.

Figure 4.4 Variations in soluble dietary fiber (SDF) contents of 100 PKC with respect to different physical, chemical, and enzymatic treatments

Figure 4.5 Distribution of SDF: IDF ratios related to different 101 combinations of pre-treatments and enzymatic treatments

Figure 4.6 Color change in untreated PKC (a) and PKC treated with 107 4.5% H2O2 (b)

Figure 4.7 WHC of PKC with respect to different physicochemical 110 treatments

Figure 4.8 OHC of PKC with respect to different physicochemical 114 treatments.

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LIST OF ABBREVIATIONS

BDH Rochelle salt

BSA Bis (Trimethylsilyl) Acetamide CGTase Cyclodextrin glycosyltransferase

CT Condensed tannin

DF Dietary fiber

EDTA Ethylenediaminetetraacetic acid EGTA Ethyleneglycoltetraacetic acid FAME Fatty acid methyl ester

GC-FID Gas chromatography-Flame ionization detector

HCN Hydrogen cyanide

HDL High density lipoprotein

HT Hydrolysable tannin

ICP-OES Inductively coupled plasma-Optical emission spectrometry

IDF Insoluble dietary fiber

IP6 1,2,3,4,5,6-hexakisphosphate

Ka Disassociation constant

LC-APCI-MS Liquid chromatography-Atomic pressure chemical ionization- Mass spectrometry

LC-ESI-MS Liquid chromatography-Electrospray ionization-Mass spectrometry

LDL Low-density lipoprotein

Mr Relative molecular mass

NFE Nitrogen free extract

NSP Non-starch polysaccharide

OHC Oil holding capacity

PA Phytic acid

PKC Palm kernel cake

SDF Soluble dietary fiber

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SNP Soluble non-starch polysaccharide TAN Tropical ataxic neuropathy

TDF Total dietary fiber

WHC Water holding capacity

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KESAN KAEDAH PEMPROSESAN TERHADAP KOMPOSISI, ANTINUTRIEN DAN SIFAT FIZIKOKIMIA KERNEL KEK

SAWIT

ABSTRAK

Kernel kek sawit (BIS) merupakan bahan sampingan pemprosesan minyak kernel sawit. BIS digunakan sebagai bahan makanan ruminan kerana kos yang rendah dan bahan ini boleh diperolehi dengan mudah jika dibandingkan dengan bahan yang lain.

Walaubagaimanapun, penggunaan BIS terhad dalam diet haiwan bukan ruminan akibat kandungan serat yang tinggi dan kehadiran antinutrien. Faktor nutrisi dan antinutrisi BIS telah dikaji. Komposisi proksimat BIS turut dianalisa. Vitamin B2

(1.55 ± 0.03 mg/g), B5 (13.70 ± 0.02 mg/g), B6 (0.50 ± 0.003 mg/g) and E (3.98 ± 0.05 mg/g) telah dikesan dalam BIS. Asid laurik (2.91 mg/g), asid lignoserik (1.86 mg/g) dan asid sterik (1.17 mg/g) merupakan asid-asid lemak utama dalam BIS.

sulfur (28.59 g/kg), fosforus (14.44 g/kg), kalsium (9.63 g/kg) dan magnesium (2.55 g/kg) merupakan mineral utama yang ditemui dalam BIS. Asid fitik (4.16 ± 0.007 mg/g), tanin (0.30 ± 0.06 mg/g), saponin (0.27 ± 0.002 mg/g), kandungan oksalat keseluruhan (7.38 ± 0.04 mg/g) dan terlarut(0.48 ± 0.01 mg/g), aktiviti perencatan α- amilase (98%), hydrogen sianida (0.40 ± 0.14 mg/g) dan polisakarida bukan-kanji (45.79 ± 2.07%) turut diuji untuk antinutrien. Kandungan serat yang tinggi dan kehadiran antinutrien dalam BIS mengurangkan potensi BIS sebagai sumber serat makanan. Pelbagai kaedah fizikokimia dan enzimatiktelah dijalankan untuk menyingkirkan faktor-faktor ini. Keberkesanan rawatan fizikokimia berbeza berdasarkan jenis antinutrien. Hidrogen peroksida (H2O2) telah mengurangkan kandungan saponin, tanin dan aktiviti perencatan α-amilase dengan lebih baik berbanding rawatan fizikokimia yang lain. Kandungan oksalat keseluruhan dan

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tidak-larut berkurang melalui penggunaan asid hidroklorik. Pendidihan telah mengurangkan kandungan oksalat tidak-larut dengan lebih baik berbanding kaedah yang lain. HCl dan H2O2 telah menghilangkan kandungan hidrogen sianida. Tidak ada kaedah yang menunjukkan kesan yang signifikan terhadap kandungan asid fitik.

Sampel yang dirawat dengan H2O2 menunjukkan penurunan tertinggi dalam kandungan serat makanan tidak larut (SMT). Kombinasi H2O2 dan enzim selulase menghasilkan pengurangan maksimum dalam kandungan SMT berbanding kombinasi lain. Kandungan serat makanan terlarut (SML) adalah tertinggi untuk rawatan dengan H2O2 berbanding rawatan lain. Kombinasi H2O2 4.5% dan enzim hemiselulase menunjukkan kandungan SML yang tertinggi di antara semua kombinasi. Kandungan liginin dalam BIS berkurang sebanyak 60% apabila dirawat dengan H2O2 4.5%. Nisbah optimum SML:SMT (1:2) diperoleh apabila sampel BIS dirawat dengan hemiselulase selepas pra-rawatan dengan hydrogen peroksida. Semua aktiviti enzim dipertingkatkan melalui pra-rawatan. Kegiatan enzim xilanase menunjukkan peningkatan yang tertinggi apabila melalui pra-rawatan berbanding enzim-enzim yang lain. Rawatan dengan 4.5% H2O2 meningkatkan kegiatan hemiselulase dan selulase manakala pendidihan selama 60 minit paling meningkatkan aktiviti xilanase. Rawatan dengan H2O2 bermanfaat dalam menghilangkan antinutrien, memperbaiki nisbah SML:SMT dan sebagai kaedah pra- rawatan untuk enzim. SIfat berfungsi BIS turut ditingkatkan secara siginifikan melalui rawatan fizikokimia. Rawatan dengan H2O2 meningkatkan WHC dan OHC, sekaligus mengoptimumkan nisbah SML:SMT. Walaubagaimanapun WHC BIS yang rendah selapas rawtan H2O2 menjadikannya sumber yang kurang sesuai untuk diterapkan sebagai bahan menghentikan sineresis dalam makanan manakala BIS yang dirawat dengan H2O2 (4.5%) boleh digunakan dengan jayanya sebagai bahan

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penstabil makanan dengan peratusan lemak dan emulsi yang tinggi. Secara kesimpulannya, kajian ini melaporkan kandungan antinutrien BIS buat pertama kalinya dan kaedah kimia dan fizikal untuk mengurangkan antinutrien. Di samping itu, BIS telah diperbaiki sebagai makanan haiwan bukan ruminan kerana kandungan polisakarida bukan-kanji tidak larut yang rendah dan kandungan polisakarida bukan- kanji larut yang lebih tinggi. Peningkatan SML dan nisbah SML: SMT untuk BIS yang dirawat menjadi sumber serat makanan baru yang diperlukan dalam kajian haiwan dan keselamatan.

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EFFECTS OF PROCESSING METHODS ON COMPOSITION, ANTINUTRIENTS, AND PHYSICOCHEMICAL PROPERTIES

OF PALM KERNEL CAKE

ABSTRACT

Palm kernel cake (PKC) is a by-product of palm kernel oil milling process. It has been used as a feed ingredient for ruminants due to its relatively low cost and availability. However, PKC application is impeded in non-ruminant diets due to its high fiber contents and the presence of antinutrients. The nutritional and antinutritional factors of PKC were determined in this study. The proximate composition of PKC was analyzed. PKC was treated with physical (Removal of fat, boiling, and autoclaving), chemical (HCl and H2O2), and enzymatic (cellulase, hemicellulase, and xylanase) treatments. The effectiveness of the physicochemical treatment as a pre-treatment for the enzymes is also studied. The moisture content, crude protein content, crude fiber content, crude fat content, and ash content are 9.60

± 0.05%, 12.72 ± 0.53%, 24.11 ± 2.61%, 11.10 ± 0.73%, and 8.18 ± 0.58%, respectively. Vitamin B2 (1.55 ± 0.03 mg/g), B5 (13.70 ± 0.02 mg/g), B6 (0.50 ± 0.003 mg/g), and E (3.98 ± 0.05 mg/g) are detected in PKC. Lauric acid (2.91 mg/g), lignocereic acid (1.86 mg/g), and stearic acid (1.17 mg/g) are the main fatty acids found in PKC. Sulfur (28.59 g/kg), phosphorus (14.44 g/kg), calcium (9.63 g/kg), and magnesium (2.55 g/kg) are the major minerals detected in PKC. Phytic acid (4.16 ± 0.007 mg/g), tannin (0.30 ± 0.06 mg/g), saponin (0.27 ± 0.002 mg/g), total oxalate (7.38 ± 0.04 mg/g) and soluble oxalate (0.48 ± 0.01 mg/g), α-amylase inhibitory activity (98%), hydrogen cyanide (0.40 ± 0.14 mg/g), and non-starch polysaccharide (45.79 ± 2.07%) were tested for antinutrients. A high amount of

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fibers and antinutrient contents present in PKC reduced its potential as a dietary fiber source. The effectiveness of the physicochemical treatments was different according to the type of antinutrient. Saponin, tannin contents and α-amylase inhibitory activity were reduced by hydrogen peroxide (H2O2) treatments better than the other physicochemical treatments. Total and insoluble oxalate contents were reduced under the HCl treatments. Boiling treatments reduced the insoluble oxalate contents over the other treatments. HCl and H2O2 treatments eliminated the hydrogen cyanide (HCN) contents. No treatment showed a significant effect on phytic acid contents.

Among the physicochemical treatments, the highest reduction in insoluble dietary fiber contents was achieved for the sample treated with H2O2. The combine treatment of H2O2 and cellulase enzyme denoted the maximum reduction in insoluble dietary fiber (IDF) contents compared to the all treatment combinations. Soluble dietary fiber (SDF) contents were highest for the H2O2 treated PKC. The combination of 4.5% H2O2 and hemicellulase enzyme treatment exhibited the highest soluble dietary fiber contents. Lignin contents were reduced by 60% in PKC, treated with 4.5%

H2O2. The optimum SDF: IDF ratio (1: 2) exhibited for the hydrogen peroxide pre- treated and hemicellulase treated PKC samples. All the enzyme activities were enhanced by the pre-treatments. Among the enzymes employed, xylanase denoted the highest enhancement in activity due to the pre-treatments. The treatment of 4.5%

H2O2 enhanced the activities of hemicellulase and cellulase whereas boiling for 60 min enhanced xylanase activity, the most. H2O2 treatments were beneficial in removing antinutrients, improving SDF: IDF and as a pre-treatment method for enzymes. The functional properties of PKC were improved significantly after the physicochemical treatments. H2O2 treatment increased the WHC and OHC, simultaneously optimized the SDF: IDF ratio. But, low WHC of H2O2 treated PKC

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makes it a poor source to be applied as an ingredient to stop syneresis in food whereas H2O2 treated (4.5%) PKC can be a successful candidate after in-vivo and in- vitro studies to stabilize food with a high percentage of fat and emulsion. As a summary, the study reported antinutrient content for the first time for PKC and the means of chemical and physical methods to reduce it. Further, the treated PKC is improved as a non-ruminant feed due to its lower insoluble non-starch polysaccharides and higher soluble non-starch polysaccharides. With increased amount of SDF and SDF: IDF, treated PKC become a possible new candidate for a dietary fiber source which needs an animal and safety study.

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CHAPTER 1

INTRODUCTION

1.1 Introduction

Oil palm is a leading commercial, perennial crop in Malaysia, providing a large proportion for the revenue of the country. Palm oil, palm kernel oil, oleochemicals, bio-fuel and palm kernel cake are the major derivatives of oil palm industry.

Palm kernel cake (PKC) is a by-product of palm kernel oil extraction process. Palm kernel expeller and palm kernel meal are synonyms for PKC. Palm kernel oil extraction is done by either mechanical pressing or solvent extraction method, which produces mechanical pressed PKC or solvent extracted PKC. Thus, the solvent extraction method is less popular in the industry due to its high cost in the production of the oil (Sue and Teoh, 1985; Awaludin, 2001; Alimon, 2004).

Most of PKC (70%) in the world is produced by Malaysia and Indonesia (Sundu and Dingle, 2005) and the production has increased by 15% in the last two decade. In 2008 and 2009 Malaysia produced 2.3 million tonnes of PKC, where, 2.2 million was exported in 2008 and 2.3 million was exported in 2009 (MPOB, 2009). The exported PKC was used as a component in animal feed.

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2 1.2 Rationale of the Research

PKC has become a popular feed source in an animal husbandry due to its higher availability (Babatunde et al., 1975) and low price (Babatunde et al., 1975; Jaafar and Hamali, 1989; Orunmuyi et al., 2006). Other frequently used feed sources, such as soybean, groundnut, cottonseed (Ojewola and Ozuo, 2006), and maize (Rhule, 1996) are seasonal and their availability is insufficient during off-seasons, which increases the price of animal feed. In many developing countries, the rising price of livestock feed and the growing scarcity of fish meal and soy meal have forced animal nutritionists to seek alternatives (Babatunde et al., 1975). This quest directed scientist towards the use of fibrous by-products such as PKC as an animal feed.

The use of PKC in animal feed was first reported in 1915 for cattle (Anonymous, 1915). Ever since, it has been using as a ruminant and non-ruminant feed, although the feed cannot be used solely for non-ruminants own to high fiber contents, antinutrient contents and the putrid odor (butyric odor) of PKC. Studies performed using PKC as a non-ruminant feed (poultry and fish) showed unsatisfactory outcomes in all contexts. The growth rate of animals was compromised. However, for ruminants, there were no such incidences observed.

Non-ruminant health was impaired with PKC application in their diet due to high fiber contents. Humans are also considered as non-ruminants. Therefore, high contents of fiber in PKC are a major hurdle in the application of PKC for human food. High fiber diets are known in the prevention and treatment of some diseases such as constipation, diverticular disease, colonic cancer, coronary heart disease and

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diabetes (Grigelmo-Miguel et al, 1999). These indigestible portions in fiber by human enzymes are referred as dietary fiber (DF).

Dietary fiber is categorized as insoluble and soluble according to the solubility in water. The insoluble part of DF is related to water absorption and intestinal regulation. The soluble fraction is associate with the reduction of cholesterol in blood and the diminution in the intestinal absorption of glucose. In the terms of health benefits, the both kinds of fiber complement each other and a 50 - 70% insoluble and 30 - 50% soluble DF considers a well-balanced proportion (Grigelmo-Miguel et al, 1999), although, PKC contains less soluble dietary fiber. The reduction of insoluble dietary fiber and increment in soluble dietary fiber will enhance the application and properties of PKC on human food.

Antinutrients hinder mineral absorption (phytic acid, tannin, and oxalate), digestive enzyme systems (phytic acid and tannin), bind protein and reduce the absorption (phytic acid, tannin) and denote hemolytic activities (saponin). Few studies have been reported on the analysis of antinutrient contents and the ways to reduce antinutrients contents in PKC. The health condition and simultaneously the nutritive value of PKC are affected by the action of antinutrients in a diet. Therefore, data on antinutrient contents in PKC will be useful when it is tested for both human and animal applications.

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4 1.3 Hypothesis of the Research

To overcome the mentioned antinutritive compounds and unsatisfactory IDF and SDF contents, PKC was treated with physical (boiled, autoclaved, and defatted), chemical (H2O2, HCl), and enzymatic (cellulase, hemicellulase, and xylanase) treatments.

1.4 Objectives

The general objective of the research was to evaluate the potential for the applicability of PKC to human foods. Screening the nutritional and antinutritional compounds in PKC is vital in applying it to human foods. The knowledge on chemical constituents is beneficial on selecting the proper treatment for PKC to make it more suitable for non-ruminant feed applications. Furthermore, this information would contribute to the development of a low cost dietary fiber source for human food application and improve the applicability of PKC as an animal feed. To achieve the main objective and address the above factors, specific objectives were formulated,

1. To determine the nutritional composition of PKC.

2. To determine and reduce the antinutrient contents in PKC by physical and chemical treatments.

3. To determine water holding capacity, oil holding capacity, and color of PKC and improve the properties through physicochemical treatments.

4. To determine the effect of physicochemical and enzymatic treatments of PKC on the insoluble and soluble dietary fibre content.

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CHAPTER 2

LITERATURE REVIEW

2.1 Palm Kernel Cake (PKC)

2.1.1 The Production Process of PKC

The oil palm (Elaeis guineensis Jacq) is an important oil crop that can be found in four tropical regions of the world (Sundu and Dingle, 2005): Africa, Southeast Asia, Latin America and the South Pacific. Palm oil and palm kernel oil are the major products of the oil palm industry.

Palm kernel cake (PKC) is a by-product of palm kernel oil processing, and is generated after the oil has been extracted from palm kernels. PKC is also known as palm kernel meal and palm kernel expeller. However, Okeudo et al. (2005) defined PKC as the by-product of mechanical expression of palm kernel oil while palm kernel meal is the solvent-extracted by-product. The mechanical expeller method and the solvent (hexane) extraction method are the most commonly used procedures in the palm oil industry to extract oil, though solvent extraction process is elusive, owing to a high cost of production (Sue and Teoh, 1985; Awaludin, 2001; Alimon, 2004). Henceforth, PKC is defined as the by-product generated after oil has been extracted by the mechanical expeller method from palm kernels.

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6 2.1.2 PKC as an Animal Feed

PKC has been used as a feed for ruminants (cattle, sheep) and non-ruminants (pigs, poultry, fish). The incorporation level is different for ruminants and non-ruminants (Table 2.1). The use of PKC in animal feed was first reported in 1915 for the cattle feed (Anonymous, 1915). Since then, many studies have been conducted to test the incorporation of PKC for ruminants and non-ruminants. An understanding of the composition and chemical nature of PKC is vital for enhancing the inclusion level of PKC in the diet of ruminants and non-ruminants. Hence, a considerable amount of researches have been conducted to examine the properties of PKC. The analyzed nutritional and antinutritional properties described by various authors are reviewed in the next section.

2.1.3 The Composition of PKC

2.1.3.1 Proximate Composition

Studies on the proximate composition of PKC are summarized in table 2.2. It is evident that the variability of the data from different sources is high. The chemical constituents and quality of PKC vary widely according to the degree of oil extracted from palm kernels (Sundu and Dingle, 2005), the nature of the raw materials (Akpanabiatu et al., 2001), storage conditions and the amount of shell materials removed.

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PKC has been reported to contain a mediocre amount of protein (8 - 20%) compared to other common feed sources (Balogun, 1982; Düsterhöft and Voragen, 1991; Chin, 2001)

Table 2.1: Inclusion levels of PKC in ruminants and non-ruminants feed

Animal Inclusion level Authors

Ruminants

Cattle 60% Umunna et al., 1980

Feedlot cattle 100% Zahari and Alimon, 2004 Dairy cattle 30% - 50% Zahari and Alimon, 2004

Sheep 30% Zahari and Alimon, 2004

Goat 50% Zahari and Alimon, 2004

Non-ruminants

Pig 20 - 25% Saad et al., 1997; Ng and Chong, 2002;

Zahari and Alimon, 2004 20% - 30% Siew, 1989

30% Adesehinwa, 2007

34.5% Okai et al., 2006

Poultry 5% Ojewola and Ozuo, 2006

20% Saad et al., 1997; Ng and Chong, 2002;

Zahari and Alimon, 2004

Layers 25% Zahari and Alimon, 2004

Rabbit 30% Orunmuyi et al., 2006

Duck 30% Zahari and Alimon, 2004

Red hybrid tilapia 20% Ng et al., 2002; Zahari and Alimon, 2004 O. mossambicus 30% Lim et al., 2001

Cat fish 30% Zahari and Alimon, 2004

Fish 30% Saad et al., 1997; Ng and Chong, 2002

It is not as rich in protein as fish meal. Further, hexane-extracted PKC has a higher protein concentration than PKC derived via the mechanical method (Akpanabiatu et al., 2001). However, Chin (2001) mentioned that there was no significant difference in the crude protein contents of solvent-extracted PKC and the mechanically expeller-pressed PKC. Nevertheless, the fat and ash percentages are lower in solvent- extracted PKC than in the mechanically expeller-pressed PKC, as the solvent

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extraction method removes more oil from palm kernel, thus comparatively concentrating the other available nutrients (Ezieshi and Olomu, 2007).

Although the quality of PKC is expressed in terms of crude protein, the actual nutritive value of the protein should determined by comparing the amino acid contents in PKC with the essential amino acid profile required by a consumer. If the tested amino acid profile is proximate to the essential amino acid contents, then the feed is considered to be a good protein source. Further, crude protein usually includes non-protein nitrogen which is contributed by nucleic acid components without any nutritive value (Iluyemi et al., 2006). Therefore, for a better understanding of protein quality, the individual amino acid contents must express instead of crude protein contents.

PKC contains a negligible amount (1 g/kg) of starch (Düsterhöft and Voragen, 1991).

Crude fiber contents in PKC have been reported to be in the range of 12% to 18%

which is relatively high compared to other oil cakes (Awaludin, 2001). Crude fiber refers to the cellulose and lignin contents of a measured commodity (Joslyn, 1970).

The amount of crude fiber represents only a part of indigestible matter in PKC.

Ezieshi and Olomu (2007) claimed that crude fiber contents are higher in mechanically expeller-pressed PKC (17.96%) than in solvent-extracted PKC.

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Table 2.2: A summary on proximate composition and chemical compounds of PKC

Components Value (%) Authors

Moisture 9.00 - 58.92 Onuora & King, 1985; Sue & Teoh, 1985; Rhule, 1996; Akpanabiatu et al., 2001; Lim et al., 2001; Ng, 2004; Kolade et al., 2005; Adesehinwa, 2007; Ezieshi & Olomu, 2007

Dry matter 87.50 - 94.50 Babatunde et al., 1975; Kuan et al., 1982; Jaafar & Hamali, 1989; Siew, 1989; Mustaffa et al., 1991; Agunbiade et al., 1999; O’Mara et al., 1999; Perez et al., 1999; Awaludin, 2001; Chin, 2001; Alimon, 2004; Orunmuyi et al., 2006; Gill & Hill, 2008

Carbohydrate 41.20 - 62.00 Onuora & King, 1985; Sue & Teoh, 1985; Gill & Hill, 2008

Crude fiber 6.02 - 24.90 Babatunde et al., 1975; Umunna et al., 1980; Balogun, 1982; Kuan et al., 1982; Onuora & King, 1985; Sue & Teoh, 1985; Jaafar & Hamali, 1989; Siew, 1989; Mustaffa et al., 1991; Rhule, 1996; Agunbiade et al., 1999; O’Mara et al., 1999; Perez et al., 1999; Akpanabiatu et al., 2001;

Awaludin, 2001; Chin, 2001; Lim et al., 2001; Alimon, 2004; Ng, 2004; Marini et al., 2005; Orunmuyi et al., 2006; Adesehinwa, 2007; Ezieshi

& Olomu, 2007; Gill & Hill, 2008

Crude fat 6.39 - 13.42 Balogun, 1982; Sue & Teoh, 1985; Lim et al., 2001; Ng, 2004; Ezieshi & Olomu, 2007; Gill & Hill, 2008

Ash 2.90 - 12.00 Umunna et al., 1980; Kuan et al., 1982; Balogun, 1982; Onuora & King, 1985; Sue & Teoh, 1985; Jaafar & Hamali, 1989; Siew, 1989;

Mustaffa et al., 1991; Rhule, 1996; Agunbiade et al., 1999; O’Mara et al., 1999; Perez et al., 1999; Akpanabiatu et al., 2001; Awaludin, 2001;

Chin, 2001; Lim et al., 2001; Alimon, 2004; Ng, 2004; Orunmuyi et al., 2006; Adesehinwa, 2007; Ezieshi & Olomu, 2007

Crude protein 7.70 - 20.30 Lyman et al., 1956, 1958; Babatunde et al., 1975; Umunna et al., 1980; Balogun, 1982; Kuan et al., 1982; Onuora & King, 1985; Sue & Teoh, 1985; Jaafar & Hamali, 1989; Siew, 1989; Mustaffa et al., 1991; Düsterhöft et al., 1992; Rhule, 1996; Agunbiade et al., 1999; O’Mara et al., 1999; Perez et al., 1999; Akpanabiatu et al., 2001; Awaludin, 2001; Chin, 2001; Lim et al., 2001; Omoregie, 2001; Alimon, 2004; Atil, 2004;

Ng, 2004; Marini et al., 2005; Illuyemi et al., 2006; Orunmuyi et al., 2006; Adesehinwa, 2007; Ezieshi & Olomu, 2007; Gill & Hill, 2008;

Sekoni et al., 2008 Nitrogen Free

Extract

46.70 - 63.50 Balogun, 1982; Kuan et al., 1982; Jaafar & Hamali, 1989; Siew, 1989; Awaludin, 2001; Chin, 2001; Lim et al., 2001; Alimon, 2004; Ng, 2004;

Orunmuyi et al., 2006; Ezieshi & Olomu, 2007

Reducing Sugars 0.29 Ng et al., 2002

NSP 46.60 - 50.00 Düsterhöft & Voragen, 1991; Knudsen, 1997

Ether extract 0.80 - 19.50 Babatunde et al., 1975; Umunna et al., 1980; Kuan et al., 1982; Onuora & King, 1985; Jaafar & Hamali, 1989; Siew, 1989; Mustaffa et al., 1991; Rhule, 1996; Agunbiade et al., 1999; O’Mara et al., 1999; Perez et al., 1999; Awaludin, 2001; Chin, 2001; Alimon, 2004; Orunmuyi et al., 2006; Adesehinwa, 2007

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Nitrogen-free extract (NFE) represents the soluble carbohydrate in a diet (Sekoni et al., 2008). According to table 2.2, the total carbohydrate contents, the NFE portion and the non-starch polysaccharide (NSP) amount fluctuates around 50%; hence, the proximate analysis of PKC is not giving the actual valuation of the distribution of carbohydrate, NSP, and NFE.

Although PKC is considered as a good source of protein for ruminants and non- ruminants, the availability of protein and digestibility of the feed plays a vital role.

As discussed in the preceding section, substitution of PKC in non-ruminant feed is generally limited to 30%, beyond which there will be a growth reduction (Ng et al., 2002; Zahari and Alimon, 2004). Ruminants can digest PKC via their gut microflora, but non-ruminants are not capable of such a process. Siew (1989) mentioned that the difficulties in digestion for non-ruminant may be due to the fiber contents and grittiness of PKC. Hence, to improve the amount of PKC inclusion and nutrients availability, the indigestible matter must be converted to a digestible form. The indigestible portion mainly consists of NSP, or fiber. NSPs in PKC consist mostly of hemicellulose (mannan and xylan) and cellulose. Lignin contents also play a vital role in digestion. Therefore a detailed analysis, rather than a mere measurement of the crude fiber amount is necessary for an understanding of the fiber contents of PKC which will be a critical factor for the improvement of PKC as a feed and food ingredient.

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11 2.1.3.2 Dietary Fiber Contents

The term “dietary fiber” encompasses many polymers such as cellulose, hemicellulose, pectin, and lignin that are not digestible by human digestive system (Sundu and Dingle, 2005). Hemicellulose portion is constituted mainly of mannan and xylan. The dietary fiber content of PKC is summarized in table 2.3.

Table 2.3: A summary of fiber contents in PKC Hemicellulose (%)

Cellulose (%) Lignin (%) Authors Mannan Xylan

57.80 3.70 11.60 - Ong et al., 2004

37.03 27.86 - Iluyemi et al., 2006

- - 7.30 13.60 Knudsen, 1997

34.80 2.40 7.20 - Daud and Jarvis, 1992

- - - 18.09 O'Mara et al., 1999

39.00 1.50 6.00 12.00 Düsterhöft et al., 1992

Some authors have analyzed the acid and neutral detergent fiber contents which were 31.00 - 54.33% and 66.40 - 80.11%, respectively (Agunbiade et al., 1999; O’Mara et al., 1999; Chin, 2001; Alimon, 2004; Marini et al., 2005); these values are important for animal nutrition. Joslyn (1970) performed a detailed sugar contents analysis for the insoluble and soluble NSP (Table 2.4). According to the study, the most of NSP in PKC are insoluble and represent almost 50% of the total weight of PKC [Alimon (2004), Palm kernel cell walls are made up of cellulose, hemicellulose, lignin and pectin substances].

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Table 2.4: Sugar contents of soluble and insoluble NSP in PKC Sugar Insoluble NSP (%) Soluble NSP (%)

Arabinose 0.9 0.3

Galactose 1.2 0.3

Glucose 0.4 0.3

Mannose 29.3 1.6

Uronic acid 1.2 0.7

Xylose 3.1 -

Source: (Knudsen, 1997)

As mentioned earlier, the inclusion levels of PKC in animal feed were impaired mainly by the fiber contents or fibrous nature of the feed. PKC is not a good feed source for weaners due to its fibrous nature and low digestibility (Babatunde et al., 1975). The limits of incorporation of PKC into fish diets were also low due to its low protein contents and the presence of a high level of NSP in PKC cell wall materials (Ng et al., 2002). Agunbiade et al. (1999) mentioned, the limitation of PKC is a consequence of its fibrous nature and the decrease in digestibility may be due to a high fiber level of palm kernel products. Moreover, a high dietary fiber, grittiness (Sekoni et al., 2008) and low digestibility of protein (Balogun, 1982) have precluded the inclusion of PKC in broiler diets. Sekoni et al. (2008) claimed that PKC is inferior in quality compared to concentrated conophor seed meal, soybean meal, and groundnut cake.

It is patently clear from the above-mentioned studies that NSP contents can be considered as the limiting factor for PKC inclusion in non-ruminant diets. NSPs are

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known to impair the digestion and utilization of nutrients present in PKC by increasing the viscosity of the intestinal contents, thus reducing the rate of hydrolysis and the absorption of nutrients (Ng et al., 2002). On the contrary, galactomannan or mannan in PKC has a low water absorption capacity and thus may not greatly contribute to the viscosity (Sundu and Dingle, 2005). Further, Perez et al. (1999) indicated that high dietary fiber contents can provoke slogging of intestinal epithelial cells, causing an increase in mucosal secretion into the intestine which leads to a loss of endogenous amino acids. The indigestible property of dietary fiber originates from the β-glucosidase linkage that makes up 90% of the linkages in dietary fiber (Sundu and Dingle, 2005). Ruminants are able to cleave this linkage, but monogastric animals or non-ruminants are incapable of that. Therefore, NSP must be broken down to small fragments to ease the digestion for non-ruminants.

PKC galactomannan is a hard, crystalline, and has a high mannose: galactose ratio (Sundu and Dingle, 2005). It is possible to increase the linkage between galactomannan and cellulose with decreased galactose contents (Whitney et al., 1998). Therefore, PKC galactomannan also has a considerable amount of linkage with cellulose. This property of galactomannan may be the reason that the commercially available enzyme can only hydrolyze a limited amount of mannan in PKC (Zahari and Alimon, 2004).

Solid-state fermentation of raw PKC with Aspergillus flavus increases the crude protein contents of the final product to 22.3% from the initial protein contents of 16.5% (Lim et al., 2001). This suggests that a nexus exists between protein and

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lignocellulosic fiber in PKC, and when the links are degraded, protein content is increased.

As stated earlier, NSP contents in PKC is an antinutritional factor that hinders the digestibility and palatability of the feed. To reduce the antinutritional effect of the NSP, degradation of these components would be desirable. Extensive studies on the degradation of NSP into oligomers have been carried out with enzymes such as mannase, cellulase and xylanase. The digestibility of NSP can be further improved using several degrading enzymes simultaneously (Sundu and Dingle, 2005).

Therefore, a thorough understanding of the chemical composition of the NSP contents in PKC is necessary to allow selection of an appropriate combination of enzymes that can cleave PKC mannan or galactomannan efficiently. The stereochemistry and molecular structure of the polymers must be explored to allow for its enzymatic degradation, which would make PKC a highly desirable component of animal feed as well as human food.

2.1.3.3 Protein and Amino Acid Contents

PKC contains 19% protein which classifies it as a medium-protein feed (Balogun, 1982). Studies on the amino acid contents of PKC have been carried out by some authors with contradictory results. Amino acid availabilities in PKC have been reported as 85% (Nwokolo et al., 1976), 74.4% (Rhule, 1996), 67.85% (Yeong et al., 1983), and 65% (Mustafa et al., 2004). A similar variation was also observed for the composition of amino acids. Lysine appears to be the first limiting amino acid for animals, followed by the sulfur-contain amino acids (methionine and cysteine) and

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tryptophan (Agunbiade et al., 1999; Alimon, 2004). PKC has been reported to be lacking in methionine and lysine (Siew, 1989; Rhule, 1996; Ng, 2004; Sundu and Dingle, 2005), deficient in lysine and threonine (Carvalho et al., 2006) and deficient in lysine, leucine, threonine and phenylalanine compared to the other protein concentrates used in feed for farm animals (Babatunde et al., 1975). In contrast, other studies have stated that PKC is a rich source of protein with methionine (Lyman et al., 1956; Agunbiade et al., 1999; Carvalho et al., 2006). A summary of the studies on the amino acid contents of PKC is presented in table 2.5. Researches on PKC have been mainly focused on animal feeds, and included analysis and amino acid contents as related to animal nutrition. Various studies on the addition of PKC to feed for ruminants and non-ruminants have produced belie findings. One study concluded that PKC is not a good feed source for pigs (Babatunde et al., 1975).

Relatively low levels of essential amino acids (lysine and methionine), high dietary fiber and grittiness in PKC have reduced its level of inclusion in broiler diets (Sekoni et al., 2008). It has also been suggested that PKC is not a good protein source in compound feed for ruminants and non-ruminants (Adesehinwa, 2007). Another study noted that higher inclusion levels of PKC showed a decreasing trend in lysine contents (Rhule, 1996; Adesehinwa, 2007). It has been proposed that the addition of sulfur-contain amino acids and lysine would enhance the nutritive value of PKC used for poultry feed, compensating for its lack of essential amino acids and a high amount of arginine (Sundu and Dingle, 2005).

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Table 2.5: A summary of amino acid contents in PKC (mg/g protein) Amino acid Contents Authors

Alanine 35.55 - 42.26 Babatunde et al., 1975; Cheah et al., 1989; Perez et al., 1999; Alimon, 2004; Illuyemi et al., 2006 Aspartic acid 36.30 - 102.29

Glutamic acid 157.73 - 218.20 Proline 26.25 - 37.80

Valine 28.44 - 54.00 Lyman et al., 1956, 1958; Babatunde et al., 1975;

Cheah et al., 1989; Siew, 1989; Sreedhara and Kurup, 1998; Perez et al., 1999; Alimon, 2004;

Illuyemi et al., 2006 Threonine 21.33 - 37.88

Histidine 4.37 - 23.40 Isoleucine 32.20 - 50.87 Leucine 60.70 - 125.82 Lysine 9.84 - 37.11 Methionine 16.95 - 21.39 Phenylalanine 26.80 - 43.00 Arginine 48.68 - 149.00

Serine 35.05 - 53.10 Babatunde et al., 1975; Siew, 1989; Perez et al., 1999; Alimon, 2004; Illuyemi et al., 2006 Glycine 38.84 - 50.31

Tyrosine 18.59 - 29.80

Tryptophan 7.21 - 12.42 Lyman et al., 1956, 1958; Babatunde et al., 1975;

Siew, 1989; Sreedhara and Kurup, 1998; Perez et al., 1999

Cysteine 7.65 - 17.39 Babatunde et al., 1975; Cheah et al., 1989; Siew, 1989; Perez et al., 1999; Alimon, 2004; Illuyemi et al., 2006

In contrast to the above findings, Nwokolo et al. (1976) reported that all the essential amino acids are present in PKC; they also indicated that amino acid availability is higher than 85%, except for valine which is available at 68.4%. Nevertheless, PKC has been named as a high-energy diet due to its high quality protein contents and high fiber contents (Siew, 1989). A few authors have also suggested that PKC contains high quality amino acids (Cornelius, 1966; Umunna, et al., 1980; Omoregie, 2001; Atill, 2004), and is also a good source of protein (Alimon, 2004).

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The availability of amino acid contents is affected by the palm kernel oil extraction process and the antagonistic effect of some amino acids. The availability of protein is reduced due to the heat that generates from the processing of palm kernel oil (Sundu and Dingle, 2005). Further, browning (which can be caused by the heat generated in the screw press) in commercial PKC can seriously affect its nutritional value, particularly with regard to amino acids such as lysine, cysteine, and methionine (Cornelius, 1966). Browning may be the result of a Maillard reaction that takes place when the sugar and amino acids interact at high temperatures and at relatively low water activity.

Antagonistic effects are mainly seen between arginine and lysine (Sundu and Dingle, 2005), especially when low levels (0.5 - 2.0%) of lysine are present (James et al., 1967). When the arginine contents are high in a feed, lysine has to be supplied to keep the correct ratio between arginine and lysine, or the availability of lysine will be reduced. The authors (James et al., 1967) also mentioned that PKC has a high amount of arginine; as a result the lysine in PKC may be affected by the arginine contents. Therefore, to make PKC a high quality protein feed or food, browning and antagonistic effects should be minimized.

2.1.3.4 Mineral Contents

PKC contains 0.6 - 28.9 mg/kg of copper (Table 2.6). The excess usage (90%) of PKC in sheep feed causes copper toxicity because the dietary requirement of copper for sheep is only 4 - 6 mg/kg (Hair-Bejo and Alimon, 1995). The authors speculated that high contents of copper in PKC are probably due to the contamination that

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occurred during the oil extraction process. The most probable step for contamination will be at the pressing of palm kernel where a screw press is used.

Table 2.6: A summary of mineral contents in PKC Mineral Quantity (mg/kg) Authors

Ca 2000 - 3400 Babatunde et al., 1975; Jaafar and Hamali, 1989;

Siew, 1989; Mustaffa et al., 1991; Akpanabiatu et al., 2001; Awaludin, 2001; Chin, 2001; Alimon, 2004

Cu 0.60 - 28.90 Jaafar and Hamali, 1989; Siew, 1989; Hair-Bejo and Alimon, 1995; Akpanabiatu et al., 2001; Chin, 2001;

Alimon, 2004; Akpan et al., 2005; Gill and Hill, 2008

Fe 4.05 - 6130 Siew, 1989; Akpanabiatu et al., 2001; Alimon, 2004 K 1900 - 9300 Akpanabiatu et al., 2001; Alimon, 2004; Kolade et

al., 2005

Mg 100 - 5000 Jaafar and Hamali, 1989; Siew, 1989; Akpanabiatu et al., 2001; Alimon, 2004; Gill and Hill, 2008 Mn 17.10 - 520 Jaafar and Hamali, 1989; Siew, 1989; Akpanabiatu

et al., 2001; Alimon, 2004 Mo 7000 - 7900 Alimon, 2004

Na 1200 Akpanabiatu et al., 2001

P 4100 - 7900 Babatunde et al., 1975; Jaafar and Hamali, 1989;

Siew, 1989; Mustaffa et al., 1991; Akpanabiatu et al., 2001; Awaludin, 2001; Chin, 2001; Alimon, 2004; Kolade et al., 2005

S 1900 - 2300 Akpanabiatu et al., 2001; Alimon, 2004 Se 0.23 - 0.30 Alimon, 2004; Gill and Hill, 2008

Zn 3.70 - 340 Jaafar and Hamali, 1989; Siew, 1989; Akpanabiatu et al., 2001; Alimon, 2004; Akpan et al., 2005; Gill and Hill, 2008

Another important mineral-related parameter in PKC is the Ca: P ratio. The Ca: P ratio in PKC is adequate for beef cattle fattening, but not enough for high-producing dairy cattle. Therefore, the use of PKC as a whole feed requires a supplementation with minerals (Awaludin, 2001). Further, special attention needs to be given to

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