LIST OF CONTENTS - prr.hec.gov.pkprr.hec.gov.pk/jspui/bitstream/123456789/1386/2/2071S-0.pdfiii LIST...
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iii LIST OF CONTENTS Acknowledgement…………………………………………………………….. i Contents……………………………………………………………………….. ii List of contents……………………………………………………………….. iii List of tables………………………………………………………………….. vii List of figures………………………………………………………………… x List of Appendices……………………………………………………………. xi Abstract……………………………………………………………………….. xii 1. INTRODUCTION………………………………………………………... 1 2. REVIEW OF LITERATURE…………………………………………… 6 2.1. Vitamin K; a preamble ………………………………………… 7 2.2. Dietary sources, requirements and deficiencies………………… 8 2.2.1. Spinach: source of vitamin K 1 ..……………………… 9 2.2.2. Natto: a source of vitamin K 2 ………………………… 10 2.2.3 Vitamin K requirements …….………………………… 11 2.2.4. Deficiency of vitamin K…….………………………… 14 2.3. Absorption and metabolic route ………………………………… 15 2.4. Coagulation activities ……..……….…………………………… 17 2.4.1. Vitamin K and anticoagulants………………………… 18 2.5. Vitamin K-dependent proteins ………………….………………. 19 2.6. Vitamin K and bone health ……………………....……………… 22 2.6.1. Socio-demographic studies and vitamin K…………….. 24 2.7. Product development …….……………………………………… 25 2.7.1. Hedonic response……………………………………… 26 2.8. Efficacy study …………………………………………………... 27 3. MATERIALS AND METHODS………………………………..….......... 31 3.1. Materials…………………………………………………………. 31 3.2 Characterization of spinach and soybean..………………………... 31 3.2.1. Moisture content………………………………………... 31 3.2.2. Crude protein …………………………………………... 31 3.2.3. Crude fat ……….……………………………………..... 32 3.2.4. Crude fiber …………………………………………….. 32 3.2.5. Ash …………………………………………………….. 32
Transcript of LIST OF CONTENTS - prr.hec.gov.pkprr.hec.gov.pk/jspui/bitstream/123456789/1386/2/2071S-0.pdfiii LIST...
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LIST OF CONTENTS
Acknowledgement…………………………………………………………….. i
Contents……………………………………………………………………….. ii
List of contents……………………………………………………………….. iii
List of tables………………………………………………………………….. vii
List of figures………………………………………………………………… x
List of Appendices……………………………………………………………. xi
Abstract……………………………………………………………………….. xii
1. INTRODUCTION………………………………………………………... 1
2. REVIEW OF LITERATURE…………………………………………… 6
2.1. Vitamin K; a preamble ………………………………………… 7
2.2. Dietary sources, requirements and deficiencies………………… 8
2.2.1. Spinach: source of vitamin K1..……………………… 9
2.2.2. Natto: a source of vitamin K2………………………… 10
2.2.3 Vitamin K requirements …….………………………… 11
2.2.4. Deficiency of vitamin K…….………………………… 14
2.3. Absorption and metabolic route ………………………………… 15
2.4. Coagulation activities ……..……….…………………………… 17
2.4.1. Vitamin K and anticoagulants………………………… 18
2.5. Vitamin K-dependent proteins ………………….………………. 19
2.6. Vitamin K and bone health ……………………....……………… 22
2.6.1. Socio-demographic studies and vitamin K…………….. 24
2.7. Product development …….……………………………………… 25
2.7.1. Hedonic response……………………………………… 26
2.8. Efficacy study …………………………………………………... 27
3. MATERIALS AND METHODS………………………………..….......... 31
3.1. Materials…………………………………………………………. 31
3.2 Characterization of spinach and soybean..………………………... 31
3.2.1. Moisture content………………………………………... 31
3.2.2. Crude protein …………………………………………... 31
3.2.3. Crude fat ……….……………………………………..... 32
3.2.4. Crude fiber …………………………………………….. 32
3.2.5. Ash …………………………………………………….. 32
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3.2.6. Nitrogen free extract (NFE)……………………………. 32
3.3. Minerals profile ………………………………………………….. 32
3.4. Extraction of vitamin K ……………………………….…………. 33
3.4.1. Preparation of sample………………………………..…. 33
3.4.2. Quantification of vitamin K ….……………………..…. 33
3.5. In vitro studies…………………………………………………… 33
3.5.1. Total phenolics….……..………..……………………... 33
3.5.2. Antioxidant activity …………......…………………….. 34
3.5.3. Free radical scavenging ability ………………………... 34
3.5.4. Ferric reducing antioxidant power (FRAP)……………. 35
3.6. Product development ……………………………………………. 35
3.6.1. Preparation of cooked and reconstituted spinach……… 35
3.6.1.1. Cooked spinach……………………………… 35
3.6.1.2. Reconstituted spinach……………………….. 35
3.6.2. Preparation of Natto…………………………………… 35
3.7. Analysis of vitamin K enriched dietary sources..……………….. 36
3.8. Sensory evaluation ……………………………………………… 36
3.9. Selection of best products ……….…….………………………... 37
3.10. Bioevaluation studies ………………………………………….. 37
3.10.1. Physical parameters…………………………………... 38
3.10.1.1. Feed and water intakes……………………... 38
3.10.1.2. Body weight gain…………………………... 40
3.10.2. Coagulation assays…………………………………… 40
3.10.2.1. Bleeding & clotting time…………………… 40
3.10.2.2. Blood prothrombin time (PT)………………. 40
3.10.2.3. Activated partial thromoplastin time (APTT). 40
3.10.2.4. Plasma fibrinogen level…………………….. 40
3.10.2.5. International normalized ratio (INR)……….. 41
3.10.3. Bone mineral density…………………………………. 41
3.10.4. Vitamin K-dependent proteins………………………... 41
3.10.4.1. Osteocalcin (OC)…………………………… 41
3.10.4.2. Undercarboxylated osteocalcin (ucOC)…….. 41
3.10.4.3. Protein induced by vitamin K absence or antagonist
-II (PIVKA-II)………………………………… 42
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3.10.5. Serum vitamin K contents…………………………….. 42
3.10.6. Serum TBARS………………………………………… 42
3.10.7. Serum lipid profile…………………………………….. 43
3.10.7.1. Cholesterol…………………………………… 43
3.10.7.2. Low density & high density lipoproteins……. 43
3.10.7.3. Triglycerides…………………………………. 43
3.10.8. Liver and kidney functioning tests……………………... 43
3.10.9. Hematological aspects………………………………….. 43
3.10.10. Electrolytes balance…………………………………… 44
3.10.11. Statistical Analysis…………………………………….. 44
4. RESULTS AND DISCUSSION …………………………………………… 45
4.1. Nutritional analysis of raw materials………………………………. 45
4.1.1. Proximate Composition………………………………........ 45
4.1.2. Mineral profile…………………………………………….. 46
4.1.3. Vitamin K content…………………………………………. 49
4.1.4. Antioxidant potential………………………………………. 51
4.2. Nutritional analysis of vitamin K enriched products ……..………… 54
4.2.1. Proximate composition ………………………………….... 54
4.2.2. Mineral profile ……………………………………………. 57
4.2.3. Phylloquinone & menaquinone-7…………………………. 61
4.2.4. Antioxidant potential……………………………………… 64
4.3. Sensory evaluation ………….………………………………….…… 68
4.4. Efficacy study ………………………………………………………. 71
4.4.1. Physical parameters ………………..……………………… 72
4.4.1.1. Feed intake………………………………………. 72
4.4.1.2. Water intake……………………………………... 73
4.4.1.3. Body weight gain………………………………... 76
4.4.2. Blood coagulation ……..………………………………….. 82
4.4.2.1. Bleeding time……………………………………. 82
4.4.2.2. Clotting time (CT)……………………………….. 84
4.4.2.3. Prothrombin time (PT)…………………………... 88
4.4.2.4. Activated partial thromboplastin time (APTT)….. 91
4.4.2.5. Plasma fibrinogen………………………………... 95
4.4.2.6. International normalized ratio (INR)…………….. 97
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4.4.3. Bone mineral density (BMD) ………….……..…………… 99
4.4.4. Vitamin K-dependent proteins …………………………….. 102
4.4.4.1. Carboxylated osteocalcin (cOC)…………………. 102
4.4.4.2. Undercarboxylated osteocalcin (ucOC)………….. 106
4.4.4.3. Protein induced by vitamin K absence or antagonist
-II (PIVKA-II)……………………………………. 111
4.4.5. Serum vitamin K contents …………………………………. 115
4.4.5.1. Serum phylloquinone…………………………….. 115
4.4.5.2. Serum menaquinone-4 (MK-4)…………………... 119
4.4.5.3. Serum menaquinone-7 (MK-7)…………………... 122
4.4.6. Thiobarbituric acid reactive substances (TBARS)…………. 126
4.4.7. Serum lipid profile………………………………………….. 129
4.4.8. Liver functioning tests……………………………………… 132
4.4.9. Kidney functioning tests……………………………………. 134
4.4.10. Hematological analysis……………………………………. 137
4.4.10.1. Red blood cells indices………………………….. 137
4.4.10.1.1. Red blood cells (RBCs)………………... 137
4.4.10.1.2. Hemoglobin (Hb)……………………… 137
4.4.10.1.3. Hematocrit (Hct)……………………….. 137
4.4.10.1.4. Mean corpuscular volume (MCV)……. 139
4.4.10.2. White blood cells indices……………………….. 139
4.4.10.2.1. White blood cells (WBCs)……………. 139
4.4.10.2.2. Lymphocytes………………………….. 139
4.4.10.2.3. Monocytes…………………………….. 139
4.4.10.2.4. Neutrophils……………………………. 139
4.4.11. Serum electrolytes………………………………………… 142
5. SUMMARY……...…………………………………………………………… 146
RECOMMENDATIONS………………………………………………………. 153
LITERATURE CITED………………………………………………………… 154
APPENDICES………………………………………………………………….. 193
