仿刺参雌性生殖腺皂苷提取分离工艺优化及体外抗氧化活性研究

Optimizations of extraction and separation process of saponins and in vitro antioxidant activity from the female gonad of Apostichopus japonicus

目的:优化超声辅助提取和高速逆流色谱(HSCCC)分离仿刺参雌性生殖腺皂苷工艺并探究其抗氧化活性。方法:以提取时间、乙醇体积分数、超声时间、料液比为考察因素,采用响应面试验优化皂苷提取工艺;针对高速逆流色谱分离工艺的关键参数溶剂体系配比、流动相体积流量、转速进行优化,并通过DPPH自由基、ABTS自由基清除试验评价粗提物及分离组分的抗氧化活性。结果:粗皂苷的最佳提取条件为提取时间7 d,乙醇体积分数83%,超声时间33 min,料液比1∶20(g/mL),此条件下皂苷提取率为(0.66±0.02)%。在HSCCC溶剂体系配比为V_(乙酸乙酯)∶V_(正丁醇)∶V_(甲醇)∶V_(水)为2.0∶3.0∶0.2∶4.8,流动相体积流量为2 mL/min,转速为800 r/min的条件下分离粗皂苷提取物得到组分1和组分2。粗皂苷提取物、组分1和组分2的DPPH自由基清除率分别可达(36.23±0.55)%,(35.07±0.20)%和(38.40±0.14)%,ABTS自由基清除率分别可达为(17.55±0.42)%,(24.49±0.50)%和(33.65±0.34)%。结论:经工艺优化获得了较高的仿刺参雌性生殖腺粗皂苷得率及较好的分离效果,且相较于粗皂苷,经HSCCC分离后得到的组分2的抗氧化活性有所提高。

Objective:This study aimed to optimize the ultrasonic-assisted extraction technology and HSCCC separation process of saponins from the female gonad of Apostichopus japonicus and to explore their antioxidant activities.Methods:Extraction time,ethanol concentration,ultrasonic time,and solid-to-solvent ratio as the influencing factors were considered in this study,and the extraction process of saponin was optimized using the response surface methodology.The ratio of solvent systems,the volume flow of mobile phases,and the rotation speeds of HSCCC were optimized.The DPPH radical and ABTS radical scavenging experiments were conducted to evaluate the antioxidant activities of the crude extracts and the separated fractions.Results:The optimal extraction condition of the crude saponin were extraction time of 7 d,ethanol concentration of 83%,ultrasound time of 33 min,and solid-to-solvent ratio of 1∶20(g/mL).Under these conditions,the extraction rate of saponins was(0.66±0.02)%.Components 1 and 2 were separated from the crude saponin by HSCCC with V_(acetate)∶V_(butanol)∶V_(methanol)∶V_(water)=2.0∶3.0∶0.2∶4.8,2 mL/min volume flow,and 800 r/min rotation speed.The DPPH radical scavenging rate of the crude saponin,component 1,and component 2 were(36.23±0.55)%,(35.07±0.20)%,and(38.40±0.14)%,respectively,while the ABTS radical scavenging rate were(17.55±0.42)%,(24.49±0.50)%and(33.65±0.34)%,respectively.Conclusion:A higher yield of crude saponin from the female gonad of Apostichopus japonicus with better separation was obtained after process optimization.Compared with the crude saponin,the antioxidant activity of component 2 was improved after HSCCC separation.