By
Stevan Teji 11505005
BACHELOR’S DEGREE in
FOOD TECHNOLOGY
FACULTY OF LIFE SCIENCES AND TECHNOLOGY
SWISS GERMAN UNIVERSITY The Prominence Tower
Jalan Jalur Sutera Barat No. 15 Alam Sutera Tangerang, Banten 15143 - Indonesia
July 2019
ANALYSIS OF ALPHA-GLUCOSIDASE INHIBITOR ACTIVITY IN WILD Stenochlaena palustris FROM KALIMANTAN ORIGIN
By
Stevan Teji 11505005
BACHELOR’S DEGREE in
FOOD TECHNOLOGY
FACULTY OF LIFE SCIENCES AND TECHNOLOGY
SWISS GERMAN UNIVERSITY The Prominence Tower
Jalan Jalur Sutera Barat No. 15 Alam Sutera Tangerang, Banten 15143 - Indonesia
July 2019
[Revision after thesis defense on July 19th 2019]
STATEMENT BY THE AUTHOR
I hereby declare that this submission is my own work and to the best of my knowledge, it contains no material previously published or written by another person, nor material which to a substantial extent has been accepted for the award of any other degree or diploma at any educational institution, except where due acknowledgement is made in the thesis.
Stevan Teji
_____________________________________________
Student Date
Approved by:
Maria D.P.T. Gunawan Puteri, S.T.P., M.Sc., Ph.D.
_____________________________________________
Thesis Advisor Date
Eisuke Kato, Ph.D.
_____________________________________________
Thesis Co-Advisor Date
Dr. Dipl. –Ing. Samuel P. Kusumocahyo
_____________________________________________
Dean Date
ABSTRACT
ANALYSIS OF ALPHA-GLUCOSIDASE INHIBITOR ACTIVITY IN WILD S.
palustris FROM KALIMANTAN ORIGIN
By
Stevan Teji
Maria D.P.T. Gunawan Puteri, S.T.P. M.Sc, Ph.d., Advisor Eisuke Kato, Ph.D., Co-Advisor
SWISS GERMAN UNIVERSITY
S. palustris is a fern commonly occurs in Malaysia, Indonesia, India, and some other Asian countries, the fronds of the fern are consumed for dietary and medicinal purposes.
S. palustris was mentioned as a potent alpha-glucosidase inhibitor source on par with the current commercial synthetic alpha-glucosidase inhibitors. Considering the quality of herbal medicines is determined by which cultivation technologies were applied to and the ecological conditions in which the plants grow. With a thoroughly controlled excavation process, the effect of the pre-treatments of sample can be analysed on whether or not it affects the biochemical activity. This study found that S. palustris from Kalimantan origin have active glucosidase inhibitory activity on sucrase and maltase also anti-amylase activity. Methanol (50.15% maltase inhibition at 10 mg/ml, 65.87% Alpha-amylase inhibition at 1 mg/ml) and 50% aq. methanol (54.84% maltase inhibition at 10 mg/ml, 61.34 ± 0.45 % alpha-amylase inhibition at 1 mg/ml) are statistically better solvent compared to water for both alpha-glucosidase and alpha- amylase inhibitor. Kaempferol 3-O-β-glucopyranoside or astragalin is present I S.
palustris and highly possible to be the major compound responsible on S. palustris maltase inhibitory activity.
Keywords: S. palustris, Alpha-glucosidase inhibitors, Alpha-amylase inhibitors, Column chromatography, LC-MS, NMR
© Copyright 2019 by Stevan Teji All rights reserved
DEDICATION
I dedicate this works for the sake of health improvement of the diabetics.
ACKNOWLEDGEMENTS
First and foremost, thank God for all His blessing and guidance, I can continue working up to this point.
I am very grateful of the support of my family, especially my parents for their never ending support and love.
Biggest appreciation to my thesis advisor Mrs. Maria D.P.T. Gunawan Puteri, S.T.P., M.Sc., Ph.D., my thesis co-advisor Mr. Eisuke Kato, Ph.D., my mentor Mrs. Della Rahmawati, S.Si., M.Si., for guiding me throughout the duration of this study, and enabling me undergone the exchange program during this period.
Special thanks to Elissa Florentina, Evelyn Antoinette, James Emmanuel, and Imelda Adhitama for all the support during the completion of this thesis work.
I would also thank my friends, classmates, laboratory assistants, fellow researchers and everyone who contributed directly or indirectly for the completion of this work.
TABLE OF CONTENTS
STATEMENT BY THE AUTHOR ... 3
ABSTRACT ... 4
DEDICATION ... 6
ACKNOWLEDGEMENTS ... 7
TABLE OF CONTENTS ... 8
LIST OF FIGURES ... 10
LIST OF TABLES ... 11
Chapter 1 - INTRODUCTION ... 12
1.1 Background... 12
1.2 Research Objectives ... 13
1.3 Significance of the Study ... 14
1.4 Research Questions ... 14
1.5 Hypotheses ... 14
Chapter 2 - LITERATURE REVIEW ... 15
2.1 Diabetes mellitus... 15
2.2 Alpha-Glucosidase Inhibitor (AGI) ... 16
2.3 Alpha-Amylase Inhibitor (AAI) ... 16
2.4 S. palustris ... 17
2.5 Extraction by Maceration ... 19
2.6 Purification through Multiple Stage Fractionation ... 19
2.7 Identification with LC-MS ... 20
Chapter 3 - RESEARCH METHOD ... 21
3.1 Venue and Time ... 21
3.2 Material and Equipment ... 21
3.2.1 Sample materials ... 21
3.2.2 Chemicals... 21
3.2.3 Equipments ... 22
3.3 Design of Experiments ... 22
3.4 Experimental Procedure ... 24
3.4.1 Sample preparation ... 24
3.4.3 Liquid-liquid Fractionation ... 25
3.4.4 Thin Layer Chromatography... 25
3.4.5 Silica Column Fractionation ... 25
3.5 Analytical procedure ... 27
3.5.1 Alpha-glucosidase Inhibitory Activity ... 27
3.5.2 Alpha-Amylase inhibition assay ... 28
3.5.3 LC-MS ... 29
3.5.4 1H-NMR ... 29
3.5.5 Statistical analysis ... 29
Chapter 4 – RESULT AND DISCUSSION ... 30
4.1 Solvent Selection ... 30
4.2 Fractionation of Methanol Extract ... 34
4.2.1 First Stage Fractionation by Liquid-liquid Fractionation ... 34
4.2.2 Second Stage Fractionation by Column Chromatography ... 37
4.3 Active Compound Prediction by LC-MS and NMR... 42
Chapter 5 – CONCLUSIONS AND RECOMMENDATIONS ... 46
5.1 Conclusions ... 46
5.2 Recommendations ... 46
REFERENCES ... 47
APPENDICES ... 52
CURRICULUM VITAE ... 105
