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[Pages:290]GREEN TEA EXTRACT LOADED LIPOSOMES: FORMATION, CHARACTERIZATION AND STABILITY

A THESIS SUBMITTED TO THE GRADUATE SCHOOL OF NATURAL AND APPLIED SCIENCES

OF MIDDLE EAST TECHNICAL UNIVERSITY

BY DAMLA DA

IN PARTIAL FULLFILLMENT OF THE REQUIREMENTS FOR

THE DEGREE OF MASTER OF SCIENCE IN

FOOD ENGINEERING

JUNE 2017

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Approval of the thesis:

GREEN TEA EXTRACT LOADED LIPOSOMES: FORMATION, CHARACTERIZATION AND STABILITY

Submitted by DAMLA DA in partial fulfillment of the requirements for the degree of Master of Science in Food Engineering Department, Middle East Technical University by,

Prof. Dr. G?lbin Dural ?nver Dean, Graduate School of Natural and Applied Sciences

Prof. Dr. Serpil ahin Head of Department, Food Engineering

Asst. Prof. Dr. Mecit Halil ?ztop Supervisor, Food Engineering Dept., METU

_______________ _______________ _______________

Examining Committee Members:

Prof. Dr. Esra Yener Food Engineering Dept., METU

Asst. Prof. Dr. Mecit Halil ?ztop Food Engineering Dept., METU

Prof. Dr. Serpil ahin Food Engineering Dept., METU

Prof. Dr. Servet G?l?m umnu Food Engineering Dept., METU

Asst. Prof. Dr. Elif Yola?aner Food Engineering Dept., Hacettepe University

_______________ _______________ _______________ _______________ _______________

Date: 13.06.2017

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I hereby declare that all information in this document has been obtained and presented in accordance with academic rules and ethical conduct. I also declare that, as required by these rules and conduct, I have fully cited and referenced all material and results that are not original to this work.

Name, Last name : Damla Da

Signature

:

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ABSTRACT

GREEN TEA EXTRACT LOADED LIPOSOMES: FORMATION, CHARACTERIZATION AND STABILITY

Da, Damla M.S., Department of Food Engineering Supervisor: Asst. Prof. Dr. Mecit Halil ?ztop

June 2017, 268 pages

Polyphenol-rich green tea extract was encapsulated into liposomes using microfluidization and ultrasonication at two different mediums (acetate buffer and distilled water) to overcome the instability towards oxygen, light, temperature and alkaline conditions. The liposomes loaded with green tea extract by microfluidization were further coated with anionic biopolymers (gum arabic, whey protein) and cationic biopolymer (lysozyme, chitosan) to provide a protective layer over the liposomal surface. The stability of both uncoated and coated liposomes was explored by particle size, zeta potential, transmission electron microscopy, total phenolic content, antioxidant activity and NMR Relaxometry experiments during 28-days storage at 4?C. Moreover, in vitro digestion in the simulated gastric and intestinal juice was performed for uncoated liposomes. The results indicated the biopolymer coated liposomes showed better stability compared to uncoated liposomes during storage. Addition of lysozyme, gum arabic and whey protein to uncoated liposomes increased the particle size from 35 to 43 nm while the increase was recorded as 38 nm to 356 nm after chitosan addition. The zeta potential measurements of uncoated liposomes prepared in distilled water decreased from -30.2 to -23.2 mV at the end of 28th day. This decrease in zeta potential was eliminated by coating of liposomes with biopolymers. The biopolymer layer around the liposomes was also investigated through transmission electron microscope images. Results indicated that lysozyme, gum arabic, whey protein could provide increased stability to liposome possessing fragile structure.

Keywords: Green tea extract, liposome, encapsulation, biopolymer coating, stability

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?Z

YEL ?AY ?Z?T? LE DOLDURULMU LPOZOM SSTEMLERNN OLUTURULMASI, KARAKTERZASYONU VE DAYANIKLILII

Da, Damla Y?ksek Lisans, Gida M?hendislii B?l?m? Tez Y?neticisi:Yrd. Do?. Dr. Mecit Halil ?ztop

Haziran 2017, 268 sayfa

Polifenol bakimindan zengin yeil ?ayin oksijen, iik, sicaklik ve ortamin asiditesine kari dayaniksizliinin ?stesinden gelmek i?in, yeil ?ay ?z?t? iki farkli ortamda (asetat tampon ??zeltisi ve saf su) mikroakikanlatirma ve ultrasonikasyon y?ntemleri kullanilarak lipozomlara enkaps?le edilmitir. Ek olarak, mikroakikanlatirma y?ntemi ile hazirlanan yeil ?ay ?z?t? ile y?kl? lipozomlarin lipozomal y?zeyleri lipozomlara koruyucu bir katman salamak i?in anyonik biyopolimerler (arap zamki, peynir alti suyu proteini) ve katyonik biyopolimer (lizozom, kitozan) ile kaplanmitir. Kaplanmami ve kaplanmi lipozomlarin dayaniklilii, par?acik boyutu, zeta potansiyeli, transmisyon elektron mikroskobu, toplam fenolik madde miktari, antioksidan aktivite ve NMR Relaksometre deneyleri ile 4? C'de 28 g?nl?k depolama s?resi boyunca incelenmitir. Ayrica, kaplanmami lipozomlar i?in sim?le edilmi gastrik ve bairsak suyundaki in vitro sindirim deneyleri yapilmitir. Elde edilen sonu?lar dorultusunda, biyopolimer kapli lipozomlarin kaplanmami lipozomlara g?re depolama s?resi boyunca daha dayanikli olduu g?sterilmitir. Lipozomlarin lizozom, arap zamki ve peynir alti suyu proteini ile kaplanmasi, lipozomlarin par?acik boyutunu 35 nm'den 43 nm'ye arttirirken, kitozan ile kaplanmasi, lipozomlarin par?acik boyutunu 38 nm'den 356 nm'ye arttii g?zlemlenmitir. Saf suda hazirlanan kaplanmami lipozomlarin zeta potansiyeli ?l??mleri 14. g?n?n sonunda -30.2 mV'den -23.2 mV'ye d?m?t?r. Zeta potansiyelindeki bu azalma, lipozomlarin biyopolimerlerle kaplanmasiyla ortadan kaldirilmitir. Lipozomlarin etrafindaki biyopolimer katmani transmisyon elektron mikroskop g?r?nt?leri ile de incelenmitir. Sonu?lar, lizozom, arap sakizi, peynir alti suyu protein polimerlerinin kirilgan yapiya sahip lipozomlarin dayanikliliini arttiini g?stermektedir.

Anahtar Kelimeler: Yeil ?ay ?z?t?, lipozom, enkaps?lasyon, biyopolimer kaplama, dayaniklilik

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To my beloved family, vii

ACKNOWLEDGEMENTS

I would first like to express my sincere gratitude to my advisor Asst. Prof. Mecit Halil ?ztop for the continuous support of my thesis study and related research, his patience, motivation, and immense knowledge. The door to Asst. Prof. ?ztop office was always open whenever I ran into a trouble spot or had a question about my research or writing. He has been helpful in providing critical advice during my master which enables to pursue a career in research. He was and remains my best role model for a scientist, mentor and teacher. It has been an honor and a privilege being involved in his project under the guidance of him.

Every result described in this thesis was accomplished with the help and support of fellow labmates. I would like to offer my special thank to irvan Sultan Uuz, Pelin Po?an, Elif Akba, Mete Kilerciolu, Simge Tutku Yildirim, Selen G?ner, rem Ala?ik, ?ari Helin Kara?am and the rest of ?ztop Lab members for their endless supports. I would like to thank G?zde Nezihe Tuncer, Cansu Deirmenci, Ayenur Yaylaci and B?ra Akdeniz who as good friends, were always willing to help and give their best suggestions. I would like to thank Emrah Kirtil for his contribution to the discussion of the part of my thesis.

This research has been financially supported by The Scientific and Technological Research Council of Turkey (TUBITAK) with proposal number 113O442. I am deeply grateful to them for giving me this opportunity for my research. Additionally, I would like to thank METU Central Laboratory for the experiments performed there.

Finally, it is a pleasure to express my deepest thanks and gratitude to Tayfun Efe Ertop for his continuous support and encouragement throughout my years of study. I owe a deep sense of gratitude to my mother, Huriye Da, father, aban Da, and brother, Uur Da, for providing me with unfailing support and continuous encouragement throughout my life with their endless love.

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