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GRAPES SEAWEEDS), ROOTS of Tamarindus Indica (TAMARIND), Eichhornia Crassipes (WATER HYACINTH) and Dracaena Surculosa (SPOTTED DRACAENA)

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GRAPES SEAWEEDS), ROOTS of Tamarindus Indica (TAMARIND), Eichhornia Crassipes (WATER HYACINTH) and Dracaena Surculosa (SPOTTED DRACAENA) PH0500004 PHILIPPINE NUCLEAR RESEARCH INSTITUTE - SClENTiFiC LIBRARY AND DOCUMENTATION CENTER POLYTECHNIC UNIVERSITY OF TEE PHILIPPINES Sta. Mesa, Manila CC MPARATIVE STUDY ON THE PEROXIDASE ACTIVITY FROM TH^JLOATS OF Caulerpa lentillifera (GRAPES SEAWEEDS), ROOTS OF Tamarindus indica (TAMARIND), Eichhornia crassipes (WATER HYACINTH) AND Dracaena surculosa (SPOTTED DRACAENA) An UNDERGRADUATE THESIS Presented to the Faculty of the Department of Natural Sciences College of Science In Partial Fulfillment of the Requirements for the Degree of BACHELOR OF SCIENCE IN CHEMISTRY By Maan Dyann N. Ber-6sil Cherrie Joy C. Magtibay February 2005 POLYTECHNIC UNIVERSITY OF THE PHILIPPINES Sta. Mesa, Manila APPROVAL SHEET This is to certify that this thesis entitled "COMPARATIVE STUDY ON THE PEROODASE ACTIVITY FROM THE FLOATS OF Caiderpa lentillifera (GRAPES SEAWEEDS), ROOTS OF Tamarindus indica (TAMARIND), Eichhornia crassipes (WATER HYACINTH) AND Dracaena surculosa (SPOTTED DRACAENA)" prepared and si bmitted by Maan Dyann N. Berosil and Cherrie Joy C. Magtibay to fulfill the requirements for the degree of Bachelor of Science in Chemistry has been examined by the Committee on Oral Examination, successfully defended and approved with a grade of PASSKD on February 21, 2005. ai>i ? Abigail P. Cid Adviser Lorna T.(yEnerva, Ed. D. Examiner IU Joy]$. Htofilefia, Ph. D. Erigr. Liduvina C. Examiner Examiner The Department of Natural Sciences endorses acceptance of this thesis as a partial fulfill snent of the requirements for the degree of Bachelor of Science in Chemistry. laida R. parmiento, Ed. D. Chaij", Department of Natural Sciences College of Science This thesis is hereby officially accepted as partial fulfillment of the requirements for .he degree of Bachelor of Science in Chemistry. Normita B: Geirospe, Ed. D. Dean, College of Science li ACKNOWLEDGEMENT We would like to give credit to Christian Jay Cambiador and Ms. Maria Celestina Honrado that have a big part in our thesis. Christian Jay for sharing his God-given talent with us from the very beginning our thesis and for jeing a.k.a "student thesis adviser. Ma'am Cel for being appioachable that made us feel comfortable to ask whenever we had problems during our laboratory experiment, for guiding us throughout our thesis even though she was just assigned to supervised us and for staying with us even it is already an overtime. We would also like to thanks the PNR1 employees and staff, for imdoubtfully letting use their facilities and resources. To Ms. Elvira Z. Sombrito for giving her recommendations for us to avail a laboratory room. To Ms. Merrian Tangonan or Ma'am Yen for assisting us at times Ma'am Cel is not around. To Ms. Xena for sparing us some of the reagents we needed. To Sir Chito for trusting us to used almost everyday the UV-Vis spectrophotometer. To Biboy for providing and lending us the apparatus we needed. To Kuya Glenn for cutting some parts of the roots of tamarind tree. And to all the chemists and biologists in the ARD building for helping us iround. To our frrnds namely Arvie, Jheng, Nico, Bamba, Marco, and Mamu for providing us some of the reagents we needed. Ill To the bangkero, who helped us get our water hyacimh in the Pasig river. To Ms. Abigail P. Cid, our thesis adviser, for her unfailing help, support and guidance throughout our thesis. And for her time and patience in correcting our mistakes in our manuscript in order to finish and have a satisfying results. To our panelist namely Engr. Liduvina Nery, Dr. Lonia Enerva, and Dr. Joy Hofileiia, for their time even though they are very busy people amd for iheir questions and advise which made us improve our thesis. To our parents, family and friends for their love and support. To my Mom for accompanying in Navotas port to get our seaweeds sample and for willingly get the burden we have just to lighten ours (Cj). To Mhark and Jet for being our inspiration. For their encouraging words and their smiles that cheered up us to continue our thesis. To all those people we have forgotten to mention, for their love, support and prayers. And most especially to our Almighty God for His blessings and guidance vvhenever we cross the very dangerous Commonwealth Ave., just to able to reach ihe PNRI and for giving us the people we have right now that helped and inspired us to do our best. IV TABLE OF CONTENTS Page TITLE PAGE ' i APPROVAL SHEET ii ACKNOWLEDGMENT Hi LISTOFFIGURES . vii LIST OF TABLES ' viii LIST OF GRAPHS xi ABSTRACT x CHAPTER I: INTRODUCTION 1 1.1 Objectives 2 1.2 Significance of the Study 3 1.3 Scope and Limitation 3 1.4 Definition of Terms 4 CHAPTER II: REVIEW OF RELATED LITERATURE 5 2.1 Raw Materials 5 2.2 Peroxidase 12 2.3 Foreign Studies on Peroxidase 14 2.4 Local Studies on Peroxidase :... 15 CHAPTER III: METHODOLOGY ...; 16 3.1 General Outline 16 3.2Reagents 17 v 3.3 Materials 17 3.4 Sampling of Plants 18 3.5 Preparation of Buffer Solutions • 18 3.6 Preparation of Crude Enzyme Extract 19 3.7 Selection of Optimum pH 20 3.8 Spectrophotometric Peroxidase Determination 21 3.9 Protein Determination 22 CHA1TER IV: RESULTS AND DISCUSSION 23 CHA111'ER V: CONCLUSION AND RECOMMENDATION 28 REFERENCES 29 APPENDICES Appendix A: Calculations 32 Appendix B: Pictures 34 Appendix C: Cc.tification 39 CURRICULUM VITAE 41 VI LIST OF FIGURES Page Figi. -J2. 1 Different Parts of Tamarind 5 Figi;-; 2.2 Water Hyacintn Plant 7 Figui-'. 2.3 Spotted Dracaena Plant 9 Figure 2.4 Grapes Seaweeds 10 Figu! c 3.1 General Procedures in Determining Peroxidase Activity 16 of the Four Different Varieties of Plants vu LIST OF TABLES Page Table 4.1 pH Range Usrti for the Different Plant Samples 23 Table 4.2_Optimum pH of Plant Samples 24 Table 4.3 Peroxidase Activity and Absorbance of Different Varieties of Plants .... 25 Table 4.4 Total Protein Concentration and Specific Activity of the Four Plant 27 Samples Tab'c 4.5 Percent Degradation of Peroxidase Activity of the Four Plant Samples... 27 vui LIST OF GRAPHS Page Grapl 4.1 Calibration Curve of Standard Horseradish Peroxidase 24 Graph. 4.2 Calibration Curve of Standard Bovine Serum Albumin 26 IX ABSTRACT Peroxidase activities from four different varieties of plant roots were investigated through the use of UV-Vis Spectrophotometer. Hydrogen, peroxide was used as the substrate and phosphate buffer at a pH that have been determined to be the optimal pH for peroxidase activity for the specific sample type. The four plant root extracts showed an assay pH optimum of 7.5 for the Tamarindus indica (tamarind) and Eichhornia crassipes (water hyacinth), pH 5.5 for Dracaena snrculosa (spotted dracaena) and pH7.0 for Canlerpa leniillifera (grapes seaweeds) using Maehly and Chance method. Determination of peroxidase at 510 nm of the four extracts indicated that, spotted dracaena gave the highest peroxidase activity with 361.07U ml"1, followed by tamarind with 57.11U ml'1, then water hyacinth with 29.39 U ml'1 and lastly, grapes seaweeds with 7.55 U ml"1. The specific peroxidase activities of the spotted dracaena, water hyacinth, tamarind and grapes seaweeds are 0.3224, 0.2048, 0.0721 and 0.0341 U mg'1 respectively. The peroxidase of the four plant tissues that were kept at ultra low personal freezer for almost a week was degraded. CHAPTER I INTRODUCTION Peroxidase has been widely spread in various plants, animal tissues, bacteria and fungi [I]. There are peroxidases that have been obtained in the cystalline form such as the verdoperoxidase (myeloperoxidase) from leukocytes, lactoperoxidase from milk, cytochrome C peroxidase from yeast and horeseradish perosidase from horseradish [2]. Peroxidase from horseradish or HRP has been used in clinical analysis which is combined with other enzymes especially oxidase to determine many clinical important chemicals by spectrometry. Enzymes from peroxidase is conjugated to antibodies for ELISA [3,4].Soybean peroxidase has been proved to be a good catalyst, for phenol resins [4,5'j It has also widely used in the contamination of water polluted with phenols [6,7]. Mos* of this phenols and anilines are toxic chemicals [8]. Removal of these chemicals (Iron1, wastewater by peroxidase instead of the chemical and physical methods has some advantages such as low cost, no formation of by-product and efficiency. It has beea found that these phenols can be removed by enzyme-catalyzed polymerization using horseradish peroxidase that has little substrate specificity. Phenols from waste water are oxidized to phenoxy radicals, which undergo further reaction yielding various polymers and oligomers that are less toxic [9]. The polymers and oligomers products from the read ion can be removed from waste water since they have low solubility properties in water [6,10]. Peroxidase replaces the potential carcinogen formaldehyde in glues arid varnishes [7]. The baking industry in Europe has started using soybean peroxidase instead of potassium bromate in bread making and dough conditioning [7], Other functions of peroxidase are used to improved digestibility of wood and straw, used to bleach pulp, fiber in paper or textile industries [4, 6]. 1.1 OBJECTIVES The general objective of this study is to be able to determine peroxidase activity from the floats of Caulerpa lentillifera, roots of Tamarindus indica, Eichhornia crassipes, and Dracaena surculosa, by using UV-Vis spectrophometer. Specific objectives of this study are the following: 1 to be able to determine the optimum pH of the floats of Caulerpa lentillifera, roots of Tamarindus indica,FJchhornia crassipes, and Dracaena surculosa; 2 to determine the total protein concentration using Bicinchoninic Acid Protein Assay of the floats of Caulerpa lentillifera, roots of Tamarindus Indica, Eichhornia crassipes, and Dracaena surculosa; 3.
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