목차

I. INTRODUCTION 1
1. Chiral separation by HPLC using immobilized stationary phase 1
1.1 Chirality 1
1.2 Techniques for chiral switching 3
1.2.1 Capillary electrophoresis (CE) 4
1.2.2 Gas chromatography (GC) 4
1.2.3 Supercritical fluid chromatography (SFC) 4
1.2.4 Nuclear magnetic resonance spectroscopy (NMR) 5
1.2.5 Thin layer chromatography (TLC) 5
1.2.6 Mass spectrometry (MS) 6
1.2.7 High performance liquid chromatography (HPLC) 6
1.3 HPLC for chiral separation 7
1.3.1 Principle of separation by HPLC 7
1.3.2 Methodologies for chiral separation by HPLC 9
1.3.3 Chromatography parameters in separation by HPLC 12
1.4 Chiral stationary phase for enantioseparation by HPLC 14
1.4.1 Polysaccharide 15
1.4.2 Cyclodextrin 17
1.4.3 Cyclofructan 19
1.4.4 Macrocyclic antibiotics 19
1.4.5 Protein 19
1.4.6 Pirkle 20
1.4.7 Ligand exchange 20
1.4.8 Other chiral stationary phase 21
2. Design of experiment in analytical method development 22
2.1 Screening designs 25
2.2 Response surface methods 26
2.2.1 Symmetrical designs 26
2.2.2 Asymmetrical designs 27
2.2.3 Mixture design 28
2.2.4 Split-plot design 28
3. Introduction about Alpha ? Lipoic acid 30
3.1 Chemical properties 30
3.2 Pharmacology properties 30
3.2.1 Biological activities 31
3.2.2 Diseases treatment 32
3.2.3 RALA versus racemic mixture 35
4. Studies about enantioseparation of Alpha ? lipoic acid 36
4.1 Chiral separation of ALA by CE 36
4.2 Chiral separation of ALA by LC-MS/MS 36
4.3 Chiral separation of ALA by HPLC 38
5. Research objectives 39
II. EXPERIMENTAL 40
1. Materials and Apparatus 40
1.1 Materials 40
1.2 Apparatus 40
2. HPLC operation conditions 41
3. Preparation of test solutions 41
4. Method development 42
4.1 Preliminary experiments 42
4.1.1 Selection of mobile phase 42
4.1.2 Influence of guard column on the chiral separation of ALA 43
4.2 Screening design ? DoE 43
4.3 Optimization design ? DoE 44
4.4 Confirmation for optimal condition 45
5. Method validation 46
5.1 Specificity 46
5.2 System suitability 46
5.3 Linearity and Sensitivity 46
5.4 Accuracy 46
5.5 Precision 46
5.6 Robustness 47
5.7 Stability of solution 47
6. Application of the proposed method for determination S-(-)-alpha lipoic acid in raw materials and formulations 48
III. RESULTS 49
1. Method development 49
1.1 Preliminary experiments 49
1.1.1 Selection of mobile phase 49
1.1.2 Influence of guard column on the chiral separation of ALA 52
1.2 Screening design ? DoE 53
1.3 Optimization design ? DoE 55
1.4 Confirmation 60
2. Method validation 61
2.1 Specificity 61
2.2 System suitability 63
2.3 Linearity and Sensitivity 64
2.4 Accuracy 65
2.5 Precision 65
2.6 Robustness 66
2.7 Stability of solution 67
3. Application of the proposed method for determination S-(-)-alpha lipoic acid in raw materials and formulations 68
4. Comparison with conventional methods 69
IV. CONCLUSION 71
V. REFERENCES 72
VI. 국문초록 80