Pharmacological Studies of Methanolic Extract of Glycosmis pentaphylla This Thesis Paper is submitted in Partial Fulfillment of the Requirement for the Degree of Bachelors of Pharmacy, East West University Submitted by Tahiya Islam ID: 2013-1-70-002 Supervised by Dr. Shamsun Nahar Khan Ph. D, Postdoc, Harvard University Chairperson and Associate Professor Department of Pharmacy East West University Submission Date: 19 December, 2016 This Research paper is dedicated to my beloved Parents DECLARATION BY THE CANDIDATE I hereby declare that this dissertation, entitled “Pharmacological studies of Methanolic Extract of Glycosmis pentaphylla” is an authentic and genuine research work carried out by me under the guidance of Dr. Shamsun Nahar Khan, Chairperson and Associate Professor, Department of Pharmacy, East West University, Dhaka. ______________________ Tahiya Islam ID # 2013-1-70-002 Department of Pharmacy East West University, Dhaka. ENDORSEMENT BY HEAD OF THE DEPARTMENT This is to certify that the dissertation entitled “Pharmacological studies of Methanolic Extract of Glycosmis pentaphylla” is a genuine research work carried out by Tahiya Islam, under the supervision of Shamsun Nahar Khan (Ph. D, Postdoc, Harvard University, Chairperson, Department of Pharmacy, East West University, Dhaka). I further certify that no part of the thesis has been submitted for any other degree and all the resources of the information in thus connection are duly acknowledged. Dr. Shamsun Nahar Khan Ph. D, Postdoc, Harvard University Chairperson and Associate Professor Department of Pharmacy East West University CERTIFICATE This is to certify that, the thesis on “Pharmacological studies of Methanolic Extract of Glycosmis pentaphylla” submitted to Department of Pharmacy, East West University, Aftabnagar, Dhaka, in partial fulfillment of the requirements for the degree of Bachelors of Pharmacy (B. Pharm), was carried out by Tahiya Islam (ID # 2013-1-70-002) under my guidance and supervision and that no part of the thesis has been submitted for any other degree. We further certify that all the sources of information of in this connection are duly acknowledged. ___________________________ Dr. Shamsun Nahar Khan Ph. D, Postdoc, Harvard University Chairperson and Associate Professor Department of Pharmacy East West University Aftabnagar, Dhaka ACKNOWLEDGEMENT Above all, I express my gratitude to Almighty Allah for giving me the strength, energy and patience to carry out this research work. I would like to express my gratitude and admiration to Dr. Shamsun Nahar Khan, Associate Professor and Chairperson, Department of Pharmacy, East West University, for her suggestion, careful guidance, sincere help, constructive criticism and valuable time without which I would not have been able to complete this work. She had been very enthusiastic and supportive in my research. I would like to convey my deepest love and obedience to my parents for their support, inspiration and guidance towards me throughout my life until today, which keeps me strong and firm to do the things I needed to do. I would also like to take the opportunity to express my whole hearted gratitude to my fellow researcher friends specially Nadia Akther, Mohammad Ali and near and dear ones who offered encouragement, information, inspiration and assistance during the period of constructing the research report . I am thankful to the laboratory instructors for their kind support during the laboratory work. ABSTRACT Purpose: The research work was carried out to determine the pharmacological activities of methanolic extract of Glycosmis pentaphylla. Method: Methanolic extract was administered orally to the animal model (Swiss albino) and the effects were determined by comparing with respect to control group which were treated with 5% CMC. For every experiment, positive control was used. Different experiments were used to determine the pharmacological profile which was collected from the internationally published publications and journals. Result: The CNS activity was evaluated by open field method and hole board test. In the open field method and hole board experiment, the crude extract of Glycosmis pentaphylla (200mg/kg, 400mg/kg and 800mg/kg) dose dependently reduces the number of peripheral locomotion, central locomotion and leaning in the open field test and reduces the number of head dipping and head poking in the hole board test. The reduction is significant when it is compared to the standard drug. The aim of the study was also to investigate the possible toxicity of the plant Glycosmis pentaphylla and especially to establish the safety of the methanolic extract of this plant by focusing on its chronic toxicity in mice. For finding chronic toxicity several tests were done such as CBC (Cell Blood count) test and Liver Function test. All data were analyzed by using SPSS analytical method. Conclusion: After summarizing all the results, it can be said that Glycosmis pentaphylla may have several pharmacological activities but to prove the hypothesis, further higher studies are needed to be conducted. Keywords: Glycosmis pentaphylla, Neuro pharmacological effect and Toxicity test. Table of Contents Chapter 1 Introduction 1-35 1.1 Medicinal Plants 1 1.1.1 Medicinal plants all around the world 1 1.1.2 History of medicinal plants 2-3 1.1.3 Traditional and alternative medicine 3-4 1.1.4 Future of medicinal plants 4 1.1.5 Characteristics of medicinal plants 4 1.1.6 Medicinal plants in Bangladesh 5 1.1.7 Practice of traditional medicine in Bangladesh 5 1.2 Herbal products and their toxicity profiles 6 1.3 Nervous system 6-7 1.3.1 Central nervous system 7-8 1.3.2 8-9 Peripheral nervous system 1.3.3 Somatic nervous system 9 1.3.4 Autonomic nervous system 9 1.3.5 Nerve cells 10 1.3.6 Cell body or soma 10 1.3.7 Axon 10 1.3.8 Dendrite 11 1.3.9 Synapse 11 1.3.10 Different central nervous system disorders 11-12 1.4 Toxicology 12 1.4.1 Toxicity 13 1.4.2 Exposure 13 1.4.3 Route of exposure 13-14 1.4.4 Acute toxicity 14 1.4.5 Chronic toxicity 14 1.4.6 Evaluation of herbal toxicity 14-15 1.5 Hematology 15 1.5.1 Cellular elements of blood 15-16 1.5.2 Plasma 16 1.5.3 Red Blood Cell 16-17 1.5.4 Count of Red Blood Cell 17-25 1.5.5 White Blood Cell 25-26 1.5.6 Platelets 27-28 1.5.7 Hepatotoxicity 28-29 1.5.8 Liver 29-31 1.5.9 Liver Function tests 31-35 Chapter 2 Plant Introduction 36 2.1 Introduction of the Plant 37 2.1.1 Description of Glycosmis pentaphylla 37-38 2.2 Description 38-39 2.3 Geographical distribution 39 2.4 Chemical composition 39-41 2.5 Medicinal uses 41 Chapter 3 Materials and methods 42 3.1 Preparation of plant extraction 43-44 3.1.1 Crystal formation 44 3.2 Experimental Animals 44-45 3.3 45 CNS Activity Test 3.3.1 45 Materials for CNS Activity Test 3.3.2 45 Chemical Agents Used in Test 3.3.3 45-46 Doses Used in CNS Activity Test of the Extract: 3.3.4 Methods for CNS Activity Test: 46 3.3.5.1 46-47 Open Field Test: 3.3.5.2 47-48 Hole Board Test 3.4 Toxicity Test 48 3.4.1 Materials for Toxicity Test 48 3.4.2 Chemical Agents Used Toxicity Test 49 3.4.3 Doses Used for Toxicological Activity of the Extract 49 3.5 Chronic Toxicity Test 49 3.6 Hematological parameters 49 3.7 Serum biochemical parameters 49 3.8 Histopathological studies 49 3.9 Statistical Analysis 50 Chapter 4 Results and Discussion 51 4.1 Consequences of dosing: 52 4.2 CNS Activity Test of Methanolic Extract of Glycosmis 53 pentaphylla 4.2.1 Open Field Test 53 Total Number of Peripheral locomotion, Central 53- 57 4.2.1.1 locomotion, Leaning, Rearing 4.2.1.4 Hole Board Test: 58-59 4.3 Chronic Toxicity Test: 60-67 Conclusion 68-69 Chapter 5 Reference 70-73 List of Figure Serial Title Page Figure 1 Classification of Nervous system 7 Figure 2 Different parts of brain 8 Figure 3 Peripheral Nervous System 9 Figure 4 10 Neuron Figure 5 Cellular elements of blood 15 Figure 6 Blood plasma 16 Figure 7 Red Blood Cells 17 Figure 8 Structure of hemoglobin 18 Figure 9 Hematocrit Determination 20 Figure 10 Equation to determine the percent of hematocrit 22 Figure 11 24 Equation to determine MCV, MCH and MCHC Figure 12 25 White Blood Cell Figure 13 Various kinds of White Blood Cells 26 Figure 14 Platelet 27 Figure 15 Platelet aggregation to form a blood clot 28 Figure 16 Human Liver 30 Figure 17 31 Mouse Liver Figure 18 3D Structure of Alkaline Phosphatase 33 Figure 19 34 3D structure of Aspartate Transaminase Figure 20 39 Glycosmis pentaphylla Figure 21 43 Herbarium Sheet of Glycosmis pentaphylla Figure 22 44 Rotary filter for crystal formation Figure 23 45 Swiss albino mice Figure 24 47 Open field test Figure 25 48 Hole board test Figure 26 Consequences of dosing 52 Figure 27 54 Graphical representation of number of central locomotion Figure 28 55 Graphical representation of number of peripheral locomotion Figure 29 56 Graphical representation of number of leaning Figure 30 58 Graphical representation of number of head poking and dipping Figure 31 59 Sacrifice of a Swiss albino mice Figure 32 Graphical representation of different counts of WBC 61 Figure 33 62 Graphical representation of different counts of RBC Figure 34 63 Graphical representation of platelet count Figure 35 64 Graphical representation of Liver Function test Figure 36 Graphical representation of the initial and final body weight of 65 female mice.