Debre Berhan University College of Natural and Computational Sciences Department of Chemistry Evaluation of Antimicrobial Activities, Antioxidant Properties and Phytochemical Components of Aloe ankoberensis By: Minda Getaneh Advisor: Minbale Gashu (PhD) February, 2021 Debre Berhan, Ethiopia LETTER OF APPROVAL SHEET Debre Berhan University College of Graduate studies Department of Chemistry This is to certify that the thesis prepared by Minda Getaneh, entitled: Evaluation of Antibacterial Activities, Antioxidant Properties and Phytochemical Components of Aloe ankoberensis, submitted in partial fulfillment of the requirement of the Degree of Masters of Science in Chemistry complies with the regulation of the University and meets the accepted standards with respect to originality and quality. Approved by the Examining Board: Name Signature Date External examiner-------------------------------------- ---------------- Internal examiner ---------------------------------------------------- ------------- Advisor ----------------------------------------------------- -------------- Head of the Department ------------------------------------------------------- -------------- Declaration I declare that this thesis work entitled “Evaluation of Antibacterial Activities, Antioxidant Properties and Phytochemical Components of Aloe ankoberensis” is based on my work done under the esteemed supervisor of Dr. Minbale Gashu and the data presented in this work are original and authentic .They have not been published before in any journal .Beside that, I have properly acknowledged the reference materials I used in this thesis work. Name: ------------------------------------------------------------ Signature: -------------------------------------------------------- Date: -------------------------------------------------------------- Acknowledgements First and for most I would like to thank the Almighty God and St.Mary who gave me health, moral and strength throughout my life. I would like to express my sincere appreciation and gratitude to my advisor Dr. Minbale Gashu for availing his time, encouragement and guidance throughout the experiment and his advice, comments and support in getting samples and chemicals. I am gratefull to express my thanks to Debre Brehan University for instrumental support. I would also like to thanks to Wegere preparatory school leaders and teachers for their encouragement and helped me a lot in all way to complete this research work success fully. i Abstract In Ethiopia there are many species of medicinal plants which may contain disease- curing bioactive compounds which are distributed all over the country. The traditional medicine still plays an important role in the primary health care in Ethiopia. Aloe ankoberensis is one of the most common endemic plants in Ethiopia used in traditional medicines. In this study the leaves of Aloe ankoberensis were extracted using hexane, chloroform, ethyl acetate and ethanol using maceration. The antibacterial activities of the extracts were investigated against Gram positive, and Gram negative bacteriausing Muller Hinton Agar medium by disc diffusion method. Ethanol extract of aloe ankoberensis indicated a good antibacterial activity against a Staphyloccus aures, Listerial monocytogenes Escherichia coli and, candida albicans with zone of inhibition 11, 11, 13 and 10, respectively. Chloroform extract of Aloe ankoberensis also showed a medium antibacterial activity against a Staphyloccus aureus and listerial monocytogenes and Escherichia coli and, candida albicans with zone of inhibition 9, 7, 8, and 8, respectively. The antioxidant activity of the extracts of Aloe ankobernsis was determined using 2,2-Diphenyl-1-picrylhydrazyl (DPPH) assay. DPPH radical scavenging activity of hexane, chloroform, ethyl acetate, and ethanol extracts were 83.73%-96.04%, 72.14%- 95.43%, 67.57%-91.64%, 42.61%- 90.47%, respectively. Generally ethanol crude extract had good antibacterial and antioxidant activities than ethyl acetate, chloroform and hexane extracts due to the presence of saponins and phenolics compounds. Keywords: Aloe ankoberensis, antibacterial activities, Antioxidant activity ii Table of content Acknowledgements .......................................................................................................... i Abstract ............................................................................................................................ ii Table of content .............................................................................................................. iii List of table……………………………………………………………………………..………...v List of figure ..................................................................................................................... vi List of acronyms ............................................................................................................. vii 1. Introduction ................................................................................................................. 1 1.1 Background of the study ............................................................................................ 1 1.2. Statement of the problem………………………………………………………………… 2 1.3. Significance of the study .......................................................................................... 2 1.4. Objectives of the study ............................................................................................. 3 1.4.1 General objectives of the study ........................................................................... 3 1.4.2. Specific objectives ............................................................................................. 3 2. Literature review ............................................................................................................ 4 2.1. The family Asphodelaceae ....................................................................................... 4 2.2. The genus aloe......................................................................................................... 5 2.3 Traditional use of aloe species .................................................................................. 5 2.4. Antioxidant activity .................................................................................................... 6 2.5. Active phytochemicals in medicinal plants ............................................................... 8 2.5.1 Phenols ............................................................................................................... 8 2.5.2. Terpenoids ......................................................................................................... 8 2.5.3 Alkaloids……………………………………………………………………………...…8 2.5.4 Tannins .................................................................................................................. 9 2.8. Phytochemical screening ......................................................................................... 9 2.9. Test microganismsfor antibacterial activity ........................................................... 9 2.9.1. Escherichia coli .................................................................................................. 9 2.9.2 Salmonella typhi ................................................................................................... 10 2.9.3 Staphylococcus aureus ..................................................................................... 10 2.9.4 Candida albicans .............................................................................................. 10 2.9. The role of UV-Vis spectrometer in analysis .......................................................... 11 iii 3. Materials and methods .............................................................................................. 12 3.1. Plant materials........................................................................................................ 12 3.2. Chemicals and reagents ......................................................................................... 12 3.3. Instruments and apparatus ..................................................................................... 12 3.4. Preparation and Extraction of the plant materials ................................................... 12 3.5 Anti-microbial activity by Disc Diffusion Method ...................................................... 13 3.6. Preparations of stock solutions, reagents and extracts .......................................... 14 3.7 Antioxidant capacity assay (DPPH free radical scavenging activity) ....................... 15 3.8. Phytochemical screening test ................................................................................. 15 3.8.1. Test of saponins .............................................................................................. 16 3.8.2. Test for flavonoids ........................................................................................... 16 3.8.3 Test for terpenoids ............................................................................................ 16 3.8.4. Tests for steroids ............................................................................................. 16 3.8.5. Test for alkaloid (Wagner’s test) ...................................................................... 16 3.8.6. Test for phenolic compound (ferric chloride test) ............................................. 16 3.9. Analytical methods ................................................................................................