갯방풍의 페놀화합물 동시 분석법 개발 및 On-Line HPLC-ABTS 분석법에 의한 항산화능 측정
Method Development for Simultaneous Determination of Phenolic Compounds and Antioxidant Activity Test by On-Line HPLC-ABTS Assay from Glehnia littoralis
- 주제(키워드) 도움말 HPLC , Glehnia litoralis , simultaneous determination , method validation , On-Line HPLC-ABTS assay
- 발행기관 강릉원주대학교 일반대학원
- 지도교수 도움말 전중균, 김상민
- 발행년도 2017
- 학위수여년월 2017. 2
- 학위명 석사
- 학과 및 전공 도움말 일반대학원 KIST강릉분원학.연협동과정
- 실제URI http://www.dcollection.net/handler/kangnung/000000009298
- 본문언어 한국어
초록/요약 도움말
Glehnia littoralis is a perennial plant that lives in coastal sand. It has been used as a substitute for “bangpung” plant with similar efficacy to prevent stroke, and studied to be effective to inflammation and cardiovascular disease. However, most studies have focused only on the secondary metabolites of roots as traditional medicines, and there has been no report of aerial part which is consumed as a vegetable. In this study, new analytical method was developed for 16 phenolic compounds by using HPLC-DAD and validated by according to the verification guidelines. Based on this method, quantification analysis of major phenolic compounds was carried out after various extraction and processing process. In addition, antioxidant activity of G. littoralis was analyzed by on-line HPLC-ABTS antioxidant analysis. In results, new HPLC-DAD method was developed by using switching solvent system from water-acetonitrile to water-methanol and Hydrosphere C18 column and a detection at 254 and 330 nm. This method could separate ten coumarins, four flavonoids, and two hydrocinnamic acids well in a 65 min running time. The validity of the developed method has been verified according to the verification guidelines with linearity, accuracy and precision (intra and inter day precision), detection limit (LOD), quantitation limit (LOQ) and recovery. Quantification result showed that total phenolic content was the highest in flower among three parts of G. littoralis (flower, aerial part and root). Most abundant compounds were chlorogenic acid and rutin. 95% Ethanol solvent was more effective to extract phenolic compounds than water. Heat treatment with various conditions decreased total amount of phenolic compounds slightly. Among 16 phenolic compounds, only 5 compounds of chlorogenic acid, caffeic acid, rutin, hyperoside and isoquercetin were identified to show ABTS radical scavenging activity by on-line HPLC-ABTS assay. The antioxidant capacity of chlorogenic acid and rutin were over 80% of total antioxidant acitivity. Antioxidant activity of individual compounds evaluated by TEAC value was in the following order : caffeic acid 1.37, chlorogenic acid 1.43, isoquercetin 1.56, hyperoside 2.19, rutin 2.34. The quantification results by new developed HPLC-DAD method and antioxidant activity result in this study can be used a basic data in the development of processed foods and quality standardization for G. littoralis.
more목차 도움말
I. 서 론 1
Ⅱ. 재료 및 방법 14
1. 시료 및 시약 14
1.1. 시료 14
1.2. 시약 14
2. 표준액과 시험용액 제조 15
2.1. 표준액의 제조 15
2.2. 추출방법 최적화 15
2.3. 시료의 제조 15
3. LC-MS 분석에 의한 화합물 동정 16
3.1. LC-MS 분석조건 16
4. HPLC-DAD 분석법 개발 17
4.1. 최적 파장의 선택 17
4.2. 컬럼의 선택 17
4.3. 이동상의 선택과 용리 조건 최적화 17
5. HPLC-DAD 분석법의 검증 18
5.1. 특이성 18
5.2. 직선성 18
5.3. 정밀성 및 정확성 18
5.4. 검출한계 및 정량한계 19
5.5. 회수율 19
6. 추출물 제조 및 페놀화합물 정량 분석 20
6.1. 부위별 페놀화합물 정량 20
6.2. 주정 및 열수 추출물의 정량 20
6.3. 가공조건 별 주요 화합물 변화 22
6.3.1. 건조 처리 22
6.3.2. 덖음 처리 22
6.3.3. 증숙 처리 22
6.3.4. 가공식품의 페놀화합물 정량 22
7. On-line HPLC-ABTS assay 이용한 항산화능 측정 23
7.1. On-line HPLC-ABTS assay 분석조건 23
7.2. 각 화합물의 항산화도 측정 23
7.3. 개별 페놀화합물의 항산화능 정량 23
III. 결과 및 고찰 24
1. LC-MS 분석에 의한 화합물 동정 24
2. 추출 방법 최적화 24
3. HPLC-DAD 분석법 개발 28
3.1. 최적 파장의 선택 28
3.2. 컬럼의 선택 28
3.3. 이동상의 선택 용리 조건 최적화 32
4. HPLC-DAD 분석법의 검증 35
4.1. 특이성 35
4.2. 직선성, 검출한계와 정량한계 35
4.3. 정밀성 및 정확성 35
4.4. 회수율 39
5. 다양한 조건 별 페놀화합물 정량 분석 39
5.1. 부위별 정량 39
5.2. 용매 별 추출물 정량 45
5.3. 열처리에 의한 화합물 변화 45
5.4. 가공식품의 페놀화합물 정량 49
6. On-line HPLC-ABTS 분석법을 이용한 갯방풍의 항산화능 분석 54
IV. 결론 59
Ⅴ. 참고논문 61
Abstract 69
