고성능박층크로마토그래프 (HPTLC : high performance thin layer chromatography)와 고성능액체크로마토그래프 (HPLC : high performance liquid chromatography)를 이용하여 상엽 (Morus alba Linné의 잎) 중의 생리활성 물질인 rutin을 확인, 정량하는 방법을 연구하였다. ...
고성능박층크로마토그래프 (HPTLC : high performance thin layer chromatography)와 고성능액체크로마토그래프 (HPLC : high performance liquid chromatography)를 이용하여 상엽 (Morus alba Linné의 잎) 중의 생리활성 물질인 rutin을 확인, 정량하는 방법을 연구하였다. 정량법은 국제적으로 통용되는 ICH (international conference on harmonisation of technical requirements for registration of pharmaceuticals for human use [의약품 규제 조화 국제 회의])의 기준에 따라서 validation을 수행하였다. HPTLC 분석은 HPTLC pre-coated silica gel 60 F254 plate을 이용하여 ethlyacetate : water : formic acid : acetic acid (100 : 15 : 7 : 7)로 8cm로 전개하여 UV 254nm에서 검출하였다. 검량선은 62.5 - 2000μg/mL의 농도범위에서 R2 = 0.9992 ± 0.000125 인 직선성을 나타내었고, 검출한계 (LOD)는 31.25μg/mL로 나타났다. rutin의 회수율은 85.04 - 90.23 %였으며, 시료의 반복실험 결과로 Rf값이 0.17 ± 0.005, 함량은 0.1385 ± 0.00228mg/g으로 나타났다. HPLC 분석은 C-18 칼럼 (5um, 4.6×250mm)을 사용하였고, 이동상으로는 메탄올-0.5 % 인산용액의 혼합액으로 단계적인 비율로 하여 유속 1mL/min, 검출기는 PDA 검출기를 사용하여 254nm에서 검출하였다. 검량선은 15.625 - 2000μg/mL의 농도범위에서 R2 = 0.9998 ± 0.000037 인 직선성을 나타내었다. rutin의 회수율은 100.14 - 104.10 %였으며, 시료의 반복실험 결과로는 머무름시간이 19.0101 ± 0.00763분, 함량은 0.8589 ± 0.00215mg/g으로 나타났다. 시중에 판매되고 있는 10종의 상엽의 함량을 분석한 결과는 HPTLC 분석법에 따르면 0.0955 - 0.3819mg/g으로 나타났으며, HPLC 분석법에 따르면 0.1201 - 1.5288mg/g으로 나타났다. 단일 품목을 HPTLC로 분석한 경우 분석 시간은 chamber에서 전개할 때 40분이 소요되며 전개용매로 사용되는 용매의 양은 35mL이었고, HPLC로 분석한 경우 분석시간은 225분이 소요되었고 이동상으로 사용되는 용매의 양은 225mL이었다.
고성능박층크로마토그래프 (HPTLC : high performance thin layer chromatography)와 고성능액체크로마토그래프 (HPLC : high performance liquid chromatography)를 이용하여 상엽 (Morus alba Linné의 잎) 중의 생리활성 물질인 rutin을 확인, 정량하는 방법을 연구하였다. 정량법은 국제적으로 통용되는 ICH (international conference on harmonisation of technical requirements for registration of pharmaceuticals for human use [의약품 규제 조화 국제 회의])의 기준에 따라서 validation을 수행하였다. HPTLC 분석은 HPTLC pre-coated silica gel 60 F254 plate을 이용하여 ethlyacetate : water : formic acid : acetic acid (100 : 15 : 7 : 7)로 8cm로 전개하여 UV 254nm에서 검출하였다. 검량선은 62.5 - 2000μg/mL의 농도범위에서 R2 = 0.9992 ± 0.000125 인 직선성을 나타내었고, 검출한계 (LOD)는 31.25μg/mL로 나타났다. rutin의 회수율은 85.04 - 90.23 %였으며, 시료의 반복실험 결과로 Rf값이 0.17 ± 0.005, 함량은 0.1385 ± 0.00228mg/g으로 나타났다. HPLC 분석은 C-18 칼럼 (5um, 4.6×250mm)을 사용하였고, 이동상으로는 메탄올-0.5 % 인산용액의 혼합액으로 단계적인 비율로 하여 유속 1mL/min, 검출기는 PDA 검출기를 사용하여 254nm에서 검출하였다. 검량선은 15.625 - 2000μg/mL의 농도범위에서 R2 = 0.9998 ± 0.000037 인 직선성을 나타내었다. rutin의 회수율은 100.14 - 104.10 %였으며, 시료의 반복실험 결과로는 머무름시간이 19.0101 ± 0.00763분, 함량은 0.8589 ± 0.00215mg/g으로 나타났다. 시중에 판매되고 있는 10종의 상엽의 함량을 분석한 결과는 HPTLC 분석법에 따르면 0.0955 - 0.3819mg/g으로 나타났으며, HPLC 분석법에 따르면 0.1201 - 1.5288mg/g으로 나타났다. 단일 품목을 HPTLC로 분석한 경우 분석 시간은 chamber에서 전개할 때 40분이 소요되며 전개용매로 사용되는 용매의 양은 35mL이었고, HPLC로 분석한 경우 분석시간은 225분이 소요되었고 이동상으로 사용되는 용매의 양은 225mL이었다.
A method was developed for the identification and quantitative determination of rutin in Mori Folium (the leaves of Morus alba Linné) by HPTLC and HPLC. The HPTLC analysis of rutin was performed on TLC plates pre-coated with silica gel 60F254 as the stationary phase and ethylacetate : water :...
A method was developed for the identification and quantitative determination of rutin in Mori Folium (the leaves of Morus alba Linné) by HPTLC and HPLC. The HPTLC analysis of rutin was performed on TLC plates pre-coated with silica gel 60F254 as the stationary phase and ethylacetate : water : formic acid : acetic acid (100:15:7:7) as the mobile phase. Rutin was determined by densitometry at 254 nm. The system was found to give compact spot for rutin (Rf value of 0.17 ± 0.05). The linear regression analysis data for the calibration plots showed good linear relationship with R2=0.9913 in the concentration range 62.5-2000μg/mL with respect to peak area. The amount of rutin in Mori Foilum measured by the HPTLC was 0.0955 - 0.3819mg/g. The HPLC analysis of rutin was performed on C18 column as the stationary phase and methanol and 0.5% phosphoric acid by gradient condition as the mobile phase. Rutin was determined by PDA detector at 254 nm. The system was found to give peak for rutin (RT value of 19.0101 ± 0.00763 min). The linear regression analysis data for the calibration plots showed good linear relationship with R2=0.9998 in the concentration range 15.625 - 2000μg/mL with respect to peak area. The amount of rutin in Mori Foilum measured by the HPLC was 0.8589mg/g. Statistical analysis of the data showed that the method is reproducible and selective for the estimation of rutin in Mori Folium. In the result of content analysis for 10 circulated Mori Folium, the content of rutin is 0.1201 - 1.5288mg/g. In this study, analytical method of HPTLC and HPLC for rutin of Mori Folium was established. But, it showed the difference of rutin content by HPTLC between HPLC and this difference would need further study.
A method was developed for the identification and quantitative determination of rutin in Mori Folium (the leaves of Morus alba Linné) by HPTLC and HPLC. The HPTLC analysis of rutin was performed on TLC plates pre-coated with silica gel 60F254 as the stationary phase and ethylacetate : water : formic acid : acetic acid (100:15:7:7) as the mobile phase. Rutin was determined by densitometry at 254 nm. The system was found to give compact spot for rutin (Rf value of 0.17 ± 0.05). The linear regression analysis data for the calibration plots showed good linear relationship with R2=0.9913 in the concentration range 62.5-2000μg/mL with respect to peak area. The amount of rutin in Mori Foilum measured by the HPTLC was 0.0955 - 0.3819mg/g. The HPLC analysis of rutin was performed on C18 column as the stationary phase and methanol and 0.5% phosphoric acid by gradient condition as the mobile phase. Rutin was determined by PDA detector at 254 nm. The system was found to give peak for rutin (RT value of 19.0101 ± 0.00763 min). The linear regression analysis data for the calibration plots showed good linear relationship with R2=0.9998 in the concentration range 15.625 - 2000μg/mL with respect to peak area. The amount of rutin in Mori Foilum measured by the HPLC was 0.8589mg/g. Statistical analysis of the data showed that the method is reproducible and selective for the estimation of rutin in Mori Folium. In the result of content analysis for 10 circulated Mori Folium, the content of rutin is 0.1201 - 1.5288mg/g. In this study, analytical method of HPTLC and HPLC for rutin of Mori Folium was established. But, it showed the difference of rutin content by HPTLC between HPLC and this difference would need further study.
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