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Study on Biological Activities and Essential Oil of Cinnamomum Tamala

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Study on Biological Activities and Essential Oil of Cinnamomum Tamala STUDY ON BIOLOGICAL ACTIVITIES AND ESSENTIAL OIL OF CINNAMOMUM TAMALA A DISSERTATION SUBMITTED FOR THE PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE MASTERS DEGREE OF SCIENCE IN CHEMISTRY BY Dhan Bahadur G.C. Exam Symbol No.: Chem.426/072 T.U. Regd. No.: 5-2-37-111-2011 CENTRAL DEPARTMENT OF CHEMISTRY INSTITUTE OF SCIENCE AND TECHNOLOGY TRIBHUVAN UNIVERSITY, KIRTIPUR KIRTIPUR,KATHMANDU NEPAL May, 2019 BOARD OF EXAMINER AND CERTIFICATE OF APPROVAL This dissertation entitled “STUDY ON BIOLOGICAL ACTIVITIES AND ESSENTIAL OIL OF Cinnamomum tamala” by Dhan Bahadur G.C., under the supervision of Prof. Ram Chandra Basnyat, PhD, Central Department of Chemistry, Tribhuvan University, Nepal, is hereby submitted for the partial fulfillment of the Master of Science (M.Sc.) Degree in Chemistry. This dissertation has been accepted for the award of a degree. _________________________ Supervisor Prof. Ram Chandra Basnyat, PhD Central Department of Chemistry Tribhuvan University Kirtipur,Kathmandu, Nepal Internal Examiner External Examiner Prof. Niranjan Parajuli, PhD Prof. Daman Raj Gautam, PhD Central Department of Chemistry Amrit Science College Tribhuvan University Tribhuvan University Kirtipur,Kathmandu, Nepal Lainchaur, Kathmandu Head of the Department Prof. Ram Chandra Basnyat, PhD Central Department of Chemistry Tribhuvan University Kritipur, Kathmandu, Nepal ii RECOMMENDATION LETTER This is to certify that the dissertation work entitled “STUDY ON BIOLOGICAL ACTIVITIES AND ESSENTIAL OIL OF Cinnamomum tamala” has been carried out by Dhan Bahadur G.C. as partial fulfillment for the requirement of Master Degree in Chemistry under my Supervision. To the best of my knowledge, this work has not been submitted to any other degree in this institute. ________________________________ Supervisor Prof. Ram Chandra Basnyat, PhD Central Department of Chemistry Tribhuvan, University Kirtipur, Kathmandu, Nepal iii DECLARATION I, Dhan Bahadur G.C., hereby declare that the work presented herein is genuine work done originally by me and has not been published or submitted elsewhere for the requirement of a degree program. Any literature, data or works done by others, presented in this dissertation are cited, has been given due acknowledgment and listed in the reference section. _____________________________ Dhan Bahadur G.C. Central Department of Chemistry Tribhuvan, University Kirtipur, Kathmandu, Nepal iv ACKNOWLEDGEMENTS The successful completion of this study was possible with the support, assistance, motivation and advice of various people. I would like to express my wholehearted gratitude and appreciation to the following: I would like to express my deepest gratitude and sincere appreciation to my supervisor and Head of the Central Department of Chemistry Prof. Ram Chandra Basnyat, Ph.D., Central Department of Chemistry, for his constant support, inspirable guidance, invaluable suggestions, and regular feedback at all stages of my dissertation work. My deepest appreciation goes to all my respected teachers of CDC. I would also like to convey my thanks to all the administration staff as well as laboratory staffs of the department for their continuous assistance throughout the dissertation work. I would especially like to thank National Herbarium and Plant Laboratories (KATH), for the identification of plants. My immense thanks go to Department of Plant Resources, Thapathali, Kathmandu, for providing GC/MS analysis. I am thankful to the authors of books and journals which I have used as a reference in this work. My special thanks go to all my colleagues whom I have been surrounded and enjoyed, for their co-operation, support and encouragement during the work. I would also like to convey my thanks to my seniors and my juniors for their help and suggestion during this work. Last but not least, my sincere obligation goes to my dearest family. Thank you so much your love, encouragement, inspiration and cooperation that has made this dissertation possible. Thank you all. Dhan Bahadur G.C. Email: [email protected] v LIST OF TABLES Table 1 : Structures of some major compounds found in Cinnamomum tamala. 19 Table 2 : Composition of artificial sea water. 25 Table 3 : Sample collection and identification. 35 Table 4 : Table showing % yield of methanol and hexane extract. 35 Table 5 : Micro-chemical analysis of phytochemicals of leaves. 36 Table 6 : Micro-chemical analysis of phytochemicals of Bark. 36 Table 7 : Total phenolic content of Cinnamomum tamala extracts. 42 Table 8 : Total flavonoid content of Cinnamomum tamala extracts. 44 Table 9 : Calculation of LC50 value of methanol extract of leaves and bark of Cinnamomum tamala. 45 Table 10 : Antibacterial activity of methanol extract of C. tamala leaves and bark. 47 Table 11 : Antifungal activity of methanol extract of C. tamala bark and leaves. 47 Table 12 : Organoleptic properties of essential oil from C. tamala leaves. 48 Table 13 : Chemical constituents of essential oil from Cinnamomum tamala leaves of Bhadgaun, Gulmi. 50 Table 14 : Structures of some major compounds of the Essential Oil from C. tamala leaves. 52 Table 15 : Antioxidant activity of Ascorbic acid. 61 Table 16 : Antioxidant activity of Methanol extract of Bark. 61 Table 17 : Antioxidant activity of Methanol extract (cold 61 percolation) of Leaf. vi Table 18 : Antioxidant activity of Methanol extract of Leaf. 62 Table 19 : Antioxidant activity of Hexane extract of Bark. 62 Table 20 : Antioxidant activity of Hexane extract of Leaf. 62 Table 21 : Total phenolic content in methanol extract of Bark. 63 Table 22 : Total phenolic content in methanol extract of Leaf 63 Table 23 : Total phenolic the content in hexane extract of Bark. 63 Table 24 : Total phenolic content in hexane extract of Leaf. 64 Table 25 : Total flavonoid content in methanol extract of Bark. 64 Table 26 : Total flavonoid content in hexane extract of Leaf. 64 Table 27 : Total flavonoid content in methanol extract of Leaf. 65 Table 28 : Total flavonoid content in hexane extract of Bark. 65 vii LIST OF FIGURES Figure 1 : Industrial uses of medicinal plants. 3 Figure 2 : Cinnamomum tamala leaves. 4 Figure 3 : Cinnamomum tamala bark. 4 Figure 4 : Mechanism of DPPH free radical scavenging by 8 ascorbic acid. Figure 5 : General chemical representation of a polyphenol. 10 Figure 6 : Generic structure of flavonoid. 10 Figure 7 : Flow chart for the extraction, fractionation and analysis of Cinnamomum tamala leaf and bark using Soxhlet apparatus. 22 Figure 8 : Antioxidant activity of standard ascorbic acid. 38 Figure 9 : Comparision of % radical scavenging between ascorbic acid and methanol extract of C. tamala bark. 38 Figure 10 : Comparision of % radical scavenging between ascorbic acid and hexane extract of C. tamala bark. 38 Figure 11 : Comparision of % radical scavenging between ascorbic acid and methanol extract (cold percolation) of C. tamala leaves. 39 Figure 12 : Comparision of % radical scavenging between ascorbic acid and methanol extract of C. tamala leaves. 39 Figure 13 : Comparision of % radical scavenging between ascorbic acid and hexane extract of C. tamala leaves. 40 Figure 14 : IC50 values of different extract of the plant along with ascorbic acid. 40 Figure 15: Variation of absorbance with concentration for standard Gallic acid. 41 Figure 16: Total phenolic content in different Cinnamomum tamala plant extract. 42 viii Figure 17: Variation of absorbance with concentration for Quercetin. 43 Figure 18: Total flavonoid content in different C. tamala plant extracts. 44 ix LIST OF SPECTRA Spectra 1: GC Chromatogram of the essential oil from C.tamala leaves of Bhadgaun, Gulmi. 49 x LIST OF ACRONYMS AND ABBREVIATIONS C. tamala : Cinnamomum tamala DMSO : Dimethyl Sulphoxide DPPH : 1,1-diphenyl-2-picryl hydrazyl FCR : Folin-Ciocalteau Reagent GAE : Gallic Acid Equivalent GC/MS : Gas Chromatography/Mass Spectrometry IC50 : Inhibitory Concentration for 50% Inhibition IR : Infra-Red LC50 : Lethal Concentration for 50% Mortality MHA : Mueller Hinton Agar MHB : Mueller Hinton Broth ppm : Part per million QE : Quercetin Equivalent ROS : Reactive Oxygen Species RT : Retention Time TAC : Total Antioxidant Capacity TFC : Total Flavonoid Content TPC : Total Phenolic Content UV : Ultraviolet ZOI : Zone of Inhibition xi ABSTRACT The present work is focused on the study of antioxidant, antimicrobial and toxicity of the leaves and bark extract of the Cinnamomum tamala and quality assessments of essential oil of Cinnamomum tamala leaves collected from Gulmi district. The phytochemical screening showed that the plant extracts possess volatile oil, quinones, glycosides, flavonoids, terpenoids, etc. Antioxidant activity studied by DPPH radical scavenging assay showed the inhibitory concentration (IC50) value of 90.35 μg/mL and 204.31 μg/mL for methanolic and hexane bark extracts respectively as compared to standard ascorbic acid having IC50 value 55.40 μg/mL. The total phenolic content calculated showed more value for methanolic bark extract 196.5 mg GAE/gm than the methanolic leaf extract 153.41 mg GAE/gm. Similarly, the highest flavonoid content was found in methanol extract of bark 167.82 mg QE/gm. Methanolic extract of bark and leaf demonstrated significant toxicity to A. salina with LC50 value of 275.42 μg/mL and 251.18 μg/mL respectively. The antimicrobial activity was carried out by well diffusion method which showed the highest ZOI value 15 mm for Staphylococcus aureus by methanolic extract of bark followed by methanolic extract of the leaf (9 mm) as compared to standard neomycin
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