PHYTOCHEMICAL AND INHIBITION STUDIES OF GARCINIA KOLA HECKEL (GUTTIFERAE) SEED EXTRACTS ON SOME KEY ENZYMES INVOLVED WITH DIABETES


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TABLE OF CONTENTS

TITTLE
ABSTRACT
TABLE OF CONTENT
ABBREVIATIONS AND ACRONYMS

CHAPTER ONE
1.0 INTRODUCTION
1.1 Medicinal Plants
1.1.1 Alkaloids
1.1.2 Phenolics
1.1.3 Terpenoids
1.2 Plants in Traditional Medicine
1.3 Plants in Modern Medicine
1.4 Impact of Technological Advancement in Medicinal Plant Research
1.5 Challenges in Medicinal Plant Research
1.6 Statement of Research Problem
1.7 Justification
1.8 Aim and Objectives of Research
1.8.1 Aim of research
1.8.2 Objectives of research
1.9 Hypothesis

CHAPTER TWO
2.0 LITERATURE REVIEW
2.1 The Taxon Guttiferae/Clusiaceae
2.1.1 Chemistry of Guttiferae
2.1.2 The genus Garcinia
2.1.3 Ethnobotany and pharmacological activities of Garcinia spp
2.1.4 Isolated compounds from Garcinia spp
2.2 Garcinia kola
2.2.1 Description and Distribution
2.2.2 Ethnobotany and pharmacological activities of Garcinia kola
2.2.3 Isolated compounds from Garcinia kola
2.3 Diabetes
2.4 Glycosidase
2.5 α-Amylase
2.5.1 Substrate
2.5.2 Structure
2.5.3 Medicinal applications of α-amylase
2.6 α-Glucosidase
2.6.1 Substrate
2.6.2 Structure
2.6.3 Medicinal application of α-glucosidase
2.7 Glycosidase Mechanisms
2.7.1 Inverting mechanism
2.7.2 Retaining mechanism

CHAPTER THREE
3.0 MATERIALS AND METHODS
3.1 Materials
3.1.1 Solvents
3.1.2 Chromatographic adsorbents
3.1.3 TLC solvents
3.1.4 TLC spray reagent
3.1.5 Extraction/Assay buffer
3.1.6 Components of buffer
3.1.7 Reagents
3.1.8 Substrates
3.1.9 Machines
3.2 Methods
3.2.1 Collection and identification of plant materials
3.2.2 Preparation of plant material
3.2.3 Preparation of extract from Garcinia kola powder
3.2.4 Preliminary phytochemical screening
3.2.5 TLC profile of crude extracts
3.3 Biological Activity
3.3.1 Extraction of crude intestinal α-glucosidase
3.3.2 Extraction of crude pancreatic α-amylase
3.3.3 Inhibitors
3.3.4 Enzymes
3.3.5 Enzyme Assay
3.4. Column Chromatography
3.4.1 Development of solvent system
3.4.2 Silica-gel column chromatography of ethyl acetate extract
3.4.3 Isolation of compound ZAAK
3.5 Physico-Chemical Studies of Compound ZAAK
3.5.1 Melting point determination for ZAAK
3.5.2 Chemical test
3.6 Structural Elucidation of ZAAK
3.6.1 Fourier transformed infrared spectroscopy of ZAAK
3.6.2 Gas chromatography-mass spectrophotometry of ZAAK
3.7 Statistical Analysis


CHAPTER FOUR
4.0 RESULTS
4.1 Percentage Yield
4.2 Preliminary Phytochemical Screening
4.3 TLC profiles of crude extracts
4.3.1 TLC profiles of Garcinia kola hexane extract
4.3.2 TLC profiles of Garcinia kola ethyl acetate extract
4.3.2 TLC profiles of Garcinia kola methanol extract
4.4 Crude Enzyme Yield
4.5 Crude Enzyme Activity
4.6 Inhibitory effect of G. kola Extracts on α-Amylase
4.7 Inhibitory effect of G. kola Extracts on α-Glucosidase
4.8 Column Chromatography of Garcinia kola Ethyl acetate Extract
4.9 Physico-Chemical Studies on ZAAK
4.10 Fourier Transformed Infrared Spectroscopy of ZAAK
4.11 Gas chromatography-mass spectrophotometry of ZAAK

CHAPTER FIVE
5.0 DISCUSSION

CHAPTER SIX
6.0 SUMMARY, CONCLUSION AND RECOMMENDATIONS
6.1 Summary
6.2 Conclusion
6.3 Recommendations
REFERENCES
APPENDICES



ABSTRACT

Garcinia kola is an Angiosperm belonging to the family Guttiferae. It is known in commerce as bitter kola. The plant seeds have been used in the treatment of a wide range of diseases including diabetes, and its importance in folkloric medicine as a purgative, mastcatory, aphrodisiac etc. is eminent. Diabetes mellitus is a metabolic disorder of multiple etiologies characterized by chronic hyperglycemia leading to severe complications such as neuropathy, nephropathy, retinopathy and foot ulcer. n-Hexane, ethyl acetate and methanol extracts were prepared successively in a soxhlet apparatus at 50ºC. Qualitative phytochemical screening was carried out. Column chromatographic analysis was carried out on the ethyl acetate extract and the structure of the isolated compound was elucidated via Gas Chromatography-Mass Spectrophotometry and Fourier Transformed-Infra Red spectroscopy. Pancreatic α-amylase and intestinal α-glucosidase were extracted from porcine pancrease and rat small intestine under specified conditions. Steroids/triterpenes, phenolics, flavonoids, cardiac glycosides, alkaloids, coumarins and phlobatannins were detected. Methanol, ethyl acetate and n-hexane extracts inhibited α-amylase with IC50 = 0.78 ± 0.32 mg/ml, 3.44 ± 3.46 mg/ml, 4.89 ± 4.62 and α-glucosidase IC50 = 2.67 ± 0.74 mg/ml, 1.68 ± 1.27 mg/ml, 10.29 ± 4.08 mg/ml respectively. The compound ZAAK was isolated from ethyl acetate extract. Fourier Transformed-Infrared spectra revealed the presence of carboxylic acid and an ester in ZAAK. Total ion chromatogram of ZAAK revealed three major peaks corresponding to ZAAK1 ZAAK2 and ZAAK3. The mass spectra identified ZAAK1 ZAAK2 and ZAAK3 as 1-pentadecanecarboxylic acid, (Z)-11-Octadecenoic acid and octadecanoic acid, 2-(2-hydroxyethoxy) ethyl ester respectively.


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