Title Page

Contents

Abstract 12

Ⅰ. INTRODUCTION 14

Ⅱ. MATERIALS AND METHODS 17

1. Materials 17

2. Fabrication of Gelatin/alginate coacervates with or without cryoprotectant 18

3. Freeze-drying process of complex coacervates 18

4. Characteristics of rehydrated coacervates 19

4-1. Appearance of rehydrated coacervates and DLS 19

4-2. Fluorescence microscope 19

5. Characteristics of freeze-dried coacervates 20

5-1. Scanning electron microscope (SEM) analysis 20

5-2. ATR-FTIR 20

6. In vitro profiles 21

6-1. bFGF stability measurement 21

6-2. In vitro release test 21

7. In vitro cell activity study 22

7-1. In vitro HDFs viability test 22

7-2. In vitro HDFs Procollagen synthesis 23

7-3. In vitro HDFs migration test 23

8. Statistical Analysis 24

Ⅲ. RESULTS 25

1. Characterization of Gelatin/alginate coacervates 25

2. Effects of cryoprotectants on Characteristics of rehydrated coacervates 27

2-1. Effect on aggregation and turbidity changes upon rehydraion after freeze-drying 27

2-2. Effects on particle size and PDI value 29

2-3. Fluorescence microscope images 31

3. Effects of cryoprotectants on characteristics of freeze-dried coacervates 34

3-1. Morphology and texture of freeze-dried coacervates 34

3-2. Effect of trehalose during freeze-drying process confirmed by SEM images 35

3-3. ATR-FTIR spectra of GA/SA Coacervate with trehalose 37

4. In vitro profiles 39

4-1. bFGF stability after freeze-drying process 39

4-2. Controlled release rate of Freeze-dried coacervates with trehalose 40

5. In vitro cell activity test 42

5-1. In vitro HDFs viability assay 42

5-2. In vitro HDF procollagen synthesis 44

5-3. In vitro HDF migration test 45

Ⅳ. DISCUSSION 47

Ⅴ. CONCLUSION 50

REFERENCES 51

ABSTRACT IN KOREAN 56

Table. 1. Characteristics of liquid state coacervates encapsulating bFGF (n＝3). 25

Table. 2. Cryoprotectants and their final concentrations. 25

Table. 3. Sample name abbreviations and their compositions. 26

Table. 4. ATR-FTIR spectra of trehalose powder, GA/SA/trehalose mixture and CAT-F/D 38

Table. 5. Statistical difference in release rate of various coacervate relative to bFGF solution.... 41

Table. 6. Cell culture conditions for each test 43

Figure. 1. Appearance of coacervates(CA) before freeze-drying and after rehydration of freeze-dried coacervates. (a) without cryoprotectant (b)... 28

Figure. 2. (a) Size and (b) PDI values of acetic acid or citric acid coacervates before freeze-drying (AA, CA) and after rehydaration of... 29

Figure. 3. Effects of cryoprotectants on (a) Size and (b) PDI value of rehydrated coacervates, (CA-F/D, CAT-F/D, CAM-F/D, CAW-F/D) 30

Figure. 4. Fluorescence microscope images of Citric acid coacervates with or without cryoprotectant. (a) before freeze-drying (b) without... 32

Figure. 5. Fluorescence microscope images of Acetic acid coacervates with or without cryoprotectant. (a) before freeze-drying. (b) without... 33

Figure. 6. Morphology and texture of freeze-dried coacervates (a) with various concentration of cryoprotectants. (b) texture of freeze-dried... 34

Figure. 7. SEM images of freeze-dried coacervates with or without trehalose 2%(w/v) (a) without trehalose (b) with trehalose... 36

Figure. 8. ATR-FTIR spectra of trehalose, GA/SA/trehalose powder mixture and CAT-F/D 38

Figure. 9. bFGF stability after F/D 39

Figure. 10. In vitro release profiles using Transwell method. comparison of AA-F/D, CA-F/D, AAT-F/D and CAT-F/D 41

Figure. 11. In vitro activity assay of coacervates at 5 days (n＝3). (a) HDF viability assay before and after freeze-drying without trehalose (b)... 42

Figure. 12. PICP ELISA assay of supernatants collected at day 5 44

Figure. 13. (a) Representative bright field images of the HDF scratch wound assay (b) Wound closure of various conditions relative to the... 46