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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 To separate HolotoxinA1 from sea cucumber and obtain HolotoxinA1 reference substance. To establish a HPLC-ELSD quantitative analysis method for HolotoxinA1 in Apostichopus ja-ponicus, and to evaluate the quality of the sea cucumber ginseng The basis. Methods HolotoxinA1 was separated and purified by macroporous resin and HPLC. The structure of Holotoxi-nA1 was determined by mass spectrometry and nuclear magnetic resonance. The HPLC-ELSD method was developed for the determination of HolotoxinA1 by Hypersil Gold C18 column (150 mm × 2.1 mm, 3 μm) and gradient elution with 5 mmol·L-1 ammonium acetate-acetonitrile. Results According to the spectral data, it was determined that HolotoxinA1 was separated and prepared from the sea cucumber ginsenoside. HPLC-ELSD was used to determine its purity as reference substance. The established HPLC-ELSD method showed a good linear relationship (r = 0.999) at 0.2-25μg · mL-1 with a detection limit of 0.05μg · mL-1. Five kinds of sea cucumber ginseng from different sea areas were identified with Holotoxin A1 Quantitative analysis showed that the content of HolotoxinA1 was the highest (588.20μg · g-1) in the fossilized sea cucumber of East China Sea, and the content of HolotoxinA1 was the lowest (21.33μg · g-1).