Title Page
Contents
ABSTRACT 15
Ⅰ. Introduction 16
1.1. Background 16
1.1.1. Introduction to succinoglycan (SG) 16
1.1.2. Isolation of SG 17
1.1.3. Structural properties of SG 17
1.1.4. Physical properties of SG 18
1.1.5. Physiological properties of SG 19
1.1.6. Applications of SG 20
1.2. Summary 22
Ⅱ. Rheological, antibacterial, antioxidant properties of D-mannitol-induced highly viscous succinoglycans produced by Sinorhizobium meliloti Rm 1021 24
2.1. Introduction 24
2.2. Materials and Methods 27
2.2.1. Materials 27
2.2.2. Growth conditions and production of succinoglycans 27
2.2.3. Nuclear magnetic resonance (NMR) spectroscopy 28
2.2.4. Differential scanning calorimetry (DSC) 28
2.2.5. Thermal gravimetric analysis (TGA) 28
2.2.6. Viscosity measurements 28
2.2.7. Rheological measurements 28
2.2.8. Field emission scanning electron microscopy (FE-SEM) 29
2.2.9. Antibacterial test 29
2.2.10. DPPH radical scavenging activity 29
2.2.11. Hydroxyl radical scavenging activity 30
2.3. Results and Discussion 31
2.3.1. Characterization of SG 31
2.3.2. Rheological properties of SG 36
2.3.3. Morphologies of SG 40
2.3.4. Antibacterial effect of SG 42
2.3.5. Antioxidant activity of SG 43
2.4. Conclusions 45
Ⅲ. Rheological, thermal, and physicochemical properties of carboxyethyl-succinoglycan 46
3.1. Introduction 46
3.2. Experimental 48
3.2.1. Materials 48
3.2.2. Production of succinoglycans 49
3.2.3. Synthesis of carboxyethyl-succinoglycan 49
3.2.4. Nuclear magnetic resonance (NMR) spectroscopy 50
3.2.5. Fourier transform infrared (FTIR) spectroscopy 50
3.2.6. Differential scanning calorimetry (DSC) 50
3.2.7. Thermal gravimetric analysis (TGA) 50
3.2.8. Viscosity measurements 50
3.2.9. Rheological measurements 51
3.2.10. Field emission scanning electron microscopy (FE-SEM) 51
3.2.11. Antibacterial test 51
3.2.12. DPPH radical scavenging activity 52
3.2.13. Hydroxyl radical scavenging activity 52
3.3. Results and Discussion 53
3.3.1. Characterization of CE-SG 53
3.3.2. Rheological properties of CE-SG 57
3.3.3. Morphologies of CE-SG 60
3.3.4. Antibacterial effect of CE-SG 60
3.3.5. Antioxidant activity of CE-SG 61
3.4. Conclusion 63
Ⅳ. Injectable, self-healable and adhesive hydrogels using oxidized succinoglycan/chitosan for pH-responsive drug delivery 64
4.1. Introduction 64
4.2. Materials and Methods 66
4.2.1. Materials 66
4.2.2. Preparation of OSG 67
4.2.3. Synthesis of OSG/CS hydrogels (OSG/CS) 67
4.2.4. Nuclear magnetic resonance (NMR) spectroscopy 68
4.2.5. Differential scanning calorimetry (DSC) 68
4.2.6. Fourier transform infrared (FTIR) spectroscopy 68
4.2.7. Thermal gravimetric analysis (TGA) 68
4.2.8. Rheological measurements 68
4.2.9. Compressive, tensile, and adhesive strength tests 69
4.2.10. Antibacterial test 69
4.2.11. Field emission scanning electron microscopy (FE-SEM) 70
4.2.12. Equilibrium swelling ratio 70
4.2.13. Controlled drug release tests 71
4.2.14. In vitro degradation of hydrogels 71
4.2.15. In vitro cytotoxicity tests for cell proliferation. 72
4.3. Results and Discussion 72
4.3.1. Characterization of OSG and OSG/CS 72
4.3.2. Rheological properties of OSG/CS 78
4.3.3. Tensile tests and self-healing property of OSG for injectability 79
4.3.4. Adhesion ability and antibacterial effect of OSG/CS 83
4.3.5. Morphologies of OSG/CS 87
4.3.6. OSG/CS swelling behavior 88
4.3.7. Controlled drug release from OSG/CS 90
4.3.8. In vitro cytotoxicity and cell proliferation of OSG/CS 92
4.4. Conclusions 95
Ⅴ. Conclusion 96
References 97
Abstract (in Korean) 111
〈Table 1-1〉 Recent patents related succinoglycan for commercial purpose 21
〈Table 2-1〉 Degree of non-carbohydrate substitution of SG 33
〈Table 3-1〉 Degree of substitution of carboxyethyl-succinoglycan according to molar ratio of reactants 55
〈Table 4-1〉 The experimental conditions of the hydrogel components 73
〈Figure 1-1〉 Isolation and structural analysis of succinoglycan and its rheological and physiological properties 18
〈Figure 2-1〉 (a) Repeating unit of succinoglycan (SG). The possible substitution for succinyl group is shown with R. (b) ¹H NMR spectra for SG... 32
〈Figure 2-2〉 Viscosity of SG (1.0 wt%) according to the D-mannitol concentration of SG_M5, SG_M25, SG_M50, SG_M75, and SG_M100 versus... 33
〈Figure 2-3〉 (a) DSC spectra, and (b) TGA spectra of SG according to the D-mannitol concentration of SG_M5, SG_M10, SG_M20, SG_M30, SG_M40,... 34
〈Figure 2-4〉 Viscosity of SG (1.0 wt%) according to the D-mannitol concentration of SG_M5, SG_M10, SG_M20, SG_M30, SG_M40, and... 35
〈Figure 2-5〉 Viscosity of SG (1.0 wt%) in the presence (0, 100 mM, and 1 M) of (a) NaCl, (b) CaCl₂, and (c) FeCl₃ according to the D-mannitol concentration... 38
〈Figure 2-6〉 FE-SEM images of the surface of SG according to the D- mannitol concentration of (a) SG_M5, (b) SG_M10, (c) SG_M20, (d) SG_M30,... 40
〈Figure 2-7〉 Evaluation of the antibacterial effect (%) of SG according to the D-mannitol concentration of SG_M5, SG_M10, SG_M20, SG_M30, SG_M40,... 41
〈Figure 2-8〉 ¹H NMR spectra for succinoglycan (SG) and an acetate- and succinate-free succinoglycan (ASF-SG) 43
〈Figure 2-9〉 Scavenging activity of SG according to the D-mannitol concentration of SG_M5, SG_M10, SG_M20, SG_M30, SG_M40, and... 44
〈Figure 3-1〉 Structure of succinoglycan (SG) and possible reaction mechanism of carboxyethylation 53
〈Figure 3-2〉 (a) ¹H NMR spectra and (b) FTIR spectra for SG and CE-SG (SG:CPA_1:10, SG:CPA_1:50, and SG:CPA_1:100). Abbreviations for... 54
〈Figure 3-3〉 (a) DSC spectra, and (b) TGA spectra of CE-SG according to molar ratio of reactants 56
〈Figure 3-4〉 Viscosity of CE-SG (1.0 wt%) according to molar ratio of reactants versus (a) shear rate (0.01 to 1000 s⁻¹) and (b) different... 58
〈Figure 3-5〉 FE-SEM images of the surface of SG and CE-SG according to molar ratio of reactants. The images were the magnified images of a-f... 59
〈Figure 3-6〉 Evaluation of the antibacterial effect (%) of CE-SG according to molar ratio of reactants against (a) E. coli. and (b) S. aureus. Comparison... 61
〈Figure 3-7〉 Scavenging activity of CE-SG according to the molar ratio of reactants on (a) DPPH radical and (b) hydroxyl radical 62
〈Figure 4-1〉 OSG/CS synthetic route 73
〈Figure 4-2〉 ¹H NMR spectra for SG and OSG 74
〈Figure 4-3〉 FTIR spectra of (a) SG and OSG, (b) OSG/CS according to the molar ratio, (c) DSC spectra, and (d) TGA spectra of CS and OSG/CS... 75
〈Figure 4-4〉 Rheological measurement of OSG/CS. Variation of the storage modulus (G') and loss modulus (G") versus (a) angular frequency (0.1 to 100... 77
〈Figure 4-5〉 Self-healing behavior and tensile properties of OSG/CS. Hydrogels cut and healed for 12 h: (a) uncut OSG/CS, (b) healed OSG/CS.... 80
〈Figure 4-6〉 The cyclic continuous step strain measurements of OSG/CS according to molar ratio, in which the strain was switched from 0.5% strain for... 81
〈Figure 4-7〉 The self-healing ability of OSG/CS through two cut specimens integration with different colors 82
〈Figure 4-8〉 Adhesion processes of OSG/CS (a) Fingertips and joints with hydrogel attached, (b) Adhesive strength of OSG/CS according to the ratio of... 83
〈Figure 4-9〉 Evaluation of the resistance of OSG/CS to bacterial clones. Images of surviving E. coli. and S. aureus. on agar against (a) OSG:CS_1:3,... 85
〈Figure 4-10〉 Evaluation of the resistance of CS, SG, OSG, and OSG/CS to bacterial clones. Images of surviving E. coli. and S. aureus. on agar for (a)... 86
〈Figure 4-11〉 FE-SEM images of the fractured surface of OSG/CS according to the ratio of (a) OSG:CS_1:3, (b) OSG:CS_1:2, (c) OSG:CS_1:1, (d)... 87
〈Figure 4-12〉 Swelling ratio profiles for OSG/CS in PBS (a) pH 7.4 and (b) pH 2.0 at 37 °C. (c) Cumulative amount (%) of 5-fluorouracil (5-FU) released... 89
〈Figure 4-13〉 (a) UV absorption spectra according to 5-FU concentration. (b) Calibration curve at 266 nm for 5-FU release from OSG/CS 89
〈Figure 4-14〉 Comparison of UV absorption spectra of solvent containing 5-FU loaded OSG/CS and non-drug loaded OSG/CS that dissolves over time in... 90
〈Figure 4-15〉 Time-dependent remaining weight (%) of OSG/CS in PBS pH 7.4, pH 4.7, and pH 2.0 at 37 °C 92
〈Figure 4-16〉 WST-8 assays for (a) cell viability and (b) cell proliferation of HEK 293 cells cultured on OSG 93
〈Figure 4-17〉 The Calcein AM (live cells in green)/Propidium iodide (PI, dead cells in red) staining of (a) HEK293 cells and (b) HeLa cell cultured on... 94