刺糖总黄酮的提取工艺优化及其成分的快速分析

Optimization of Extraction Process and Rapid Analysis of Total Flavonoids from Saccharum alhagi

目的优化刺糖总黄酮的提取工艺,并用超高效液相色谱-四极杆串联时间飞行质谱(UPLC-Q-TOF-MS)法分析其黄酮类化学成分。方法对乙醇浓度、液料比、提取时间、提取温度等单因素考察,并进行4因素3水平的响应面研究。以芦丁标准品为对照,采用紫外分光光度法测定刺糖醇提液中总黄酮的含量;采用CORTECS UPLC-C18柱(2.1 mm×100 mm,1.6μm),以0.1%甲酸水(A)-乙腈(B)为流动相进行梯度洗脱,流速为0.3 mL·min^-1,柱温为25℃。应用电喷雾离子源分别在正、负离子模式下采集数据,建立骆驼刺属植物化合物的质谱数据库,并结合在线数据库和Masslynx 4.1工作站对使用AB-8型大孔树脂富集纯化后的刺糖提取物进行定性分析。结果通过响应面法优化后的刺糖醇提取最佳工艺为:乙醇浓度67%,料液比为1∶25,提取时间75 min,提取温度75℃。刺糖提取物中共鉴定出40个化学成分,包括7个黄酮及黄酮苷类,20个黄酮醇及黄酮醇苷类,4个异黄酮类,2个苯丙素及苷类,2个二氢黄酮类,2个甾体类,2个酚类,1个其他成分。结论 Box-Benhnken响应面优化刺糖总黄酮的提取工艺稳定、可行。首次应用UPLC-Q-TOF-MS技术对刺糖中黄酮类成分进行定性分析,为刺糖药效物质研究及质量标准的建立提供了科学参考。

Objective The extraction process of total flavonoids from Saccharum alhagi were optimized and the chemical constituents of flavonoids were analyzed by UPLC-Q-TOF-MS.Methods After the investigation of single factors with ethanol concentration,ratio of liquid to material,extraction time and extraction temperature,the response surface of 4 factors and 3 levels was designed.With the rutin reference substance as control,the content of total flavonoids in alcohol extract of Saccharum alhagi was determined by ultraviolet spectrophotometry.Using CORTECS UPLC-C18 column(2.1 mm×100 mm,1.6 μm) with gradient elution consisting of 0.1% formic acid in water(A) and acetonitrile(B) at a flow rate of 0.3 mL·min^-1,and a column temperature 25℃.Mass spectrometric data were obtained under source of ESI in negative and positive modes.The database of related compounds of Saccharum alhagi was established for screening,and the extract of Saccharum alhagi enriched and purified by AB-8 macroporous resin was qualitatively analyzed by database and Masslynx 4.1 workstation.Results The best extraction process of Saccharum alhagi alcohol optimized by response surface method was as follows:ethanol concentration67%,ratio of liquid-solid is 25 times,extraction time 75 min,extraction temperature 75℃.A total of 40 kinds of components were identified from the alcohol extract of Saccharum alhagi,including seven kinds of flavonoids and glycosides,twenty kinds of flavonols and glycosides,four kinds of isoflavones,two kinds of phenylpropanoid and glycosides,two kinds of dihydroflavonoids,two kinds of steroids,two kinds of phenols and one kind of others.Conclusion The extraction process of total flavonoids from Saccharum alhagi optimized by Box-Benhnken response surface is stable and feasible.The rapid qualitative analysis of flavonoids in Saccharum alhagi was carried out by UPLC-Q-TOF-MS for the first time,which can provide a scientific reference for the study of effective substances and the establishment of quality standard.