仿刺参中海参皂苷HolotoxinA_1的分离制备及其含量的分析

Isolation,preparation and analysis of triterpene glycoside Holotoxin A_1 in Apostichopus japonicus

目的分离制备海参皂苷HolotoxinA1单体,获得HolotoxinA1对照品;建立仿刺参(Apostichopus ja-ponicus)中HolotoxinA1的HPLC-ELSD定量分析方法,以此作为评价来源于不同海域的仿刺参中海参皂苷质量的依据。方法应用大孔树脂、HPLC对仿刺参中的HolotoxinA1进行分离纯化、制备海参皂苷Holotoxi-nA1单体,通过质谱、核磁确定其结构。采用Hypersil Gold C18(150mm×2.1mm,3μm)色谱柱,5mmol·L-1醋酸铵—乙腈溶液梯度洗脱建立海参皂苷HolotoxinA1的HPLC-ELSD方法。结果根据波谱数据确定分离制备的海参皂苷单体为三萜皂苷HolotoxinA1,HPLC-ELSD分析其纯度可作为对照品。建立的HPLC-ELSD方法在0.2～25μg·mL-1线性关系良好(r=0.999),检测限为0.05μg·mL-1;5种来源于不同海域的仿刺参中的海参皂苷均以HolotoxinA1含量最多,定量分析可知韩国东海岸产仿刺参中HolotoxinA1含量最多(588.20μg·g-1)福建产仿刺参中HolotoxinA1含量最少(21.33μg·g-1)。

Objective Prepareing HolotoxinA1 as the reference substance and establishing HPLC-ELSD method to evaluate the quality of triterpene glycosides in Apostichopus japonicus from different areas. Methods The HolotoxinA1 was separated and purified by macroporous resin and HPLC; The structure of HolotoxinA1 was determined by MS and NMR. The HPLC-ELSD method was performed on a Hy-persil Gold C18（150 mm×2. 1 mm,3 μm）, using acetonitrilewater （ containing 5 mmol · L^-11 ammonium acetate ） as the mobile phase with gradient elution. Results The HolotoxinA1 was characterized by the MS and NMR and could be used as reference substance by the HPLC-ELSD analysis. The established method of HPLC-ELSD indicated that the calibration curve showed good linearity from 0. 2μg·mL^-1 to 25 μg·mL^-1 （r=0. 999） and the lower limit of detection was 0. 05 μg·mL^-1 at a signal to noise ratio of 3. Conclusion This HolotoxinA1 is one of the most abundant triterpene glycosides in Apostichopus japonicus. The Apostichopus japonicus from east coast of South Korea has the most HolotoxinA1 （588. 20 μg·mL^-1）and the Apostichopus japonicus from Fujian province has the least HolotoxinAl（21.33μg·mL^-1） according to the analysis.