基于多指标综合评分和正交设计优化雷公藤悬浮细胞总萜类物质的提取工艺

Optimization of extraction process of total terpenoids from Tripterygium wilfordii suspension cells based on orthognal design and multi-index comprehensive evaluation method

目的优化雷公藤悬浮细胞总萜类物质的提取工艺。方法采用UPLC-QQQ-MS测定雷公藤悬浮细胞中雷公藤甲素、雷酚内酯和雷公藤红素的含量。色谱条件为:Waters HSS T3色谱柱(2.1 mm×100 mm,1.8μm);0.1%甲酸水溶液-乙腈梯度洗脱;柱温30℃;流速0.3 mL/min;进样量5μL。以3种萜类成分含量的综合评分为指标,在单因素试验的基础上,设计L9(34)正交试验优化甲醇浓度、提取时间和溶剂用量3个参数,并通过验证试验判断该工艺的可行性。结果雷公藤悬浮细胞萜类成分最佳提取工艺为:加入50倍量80%甲醇,浸渍12 h后,超声提取0.5 h,在此条件下,雷公藤甲素、雷酚内酯和雷公藤红素的提取量分别为(52.36±0.60)、(170.73±1.57)、(388.14±5.89)μg/g,综合评分稳定。结论基于多指标综合评分法和正交设计优化的雷公藤悬浮细胞总萜类物质提取工艺合理可行、重复性好,具有一定的实用性,适用于雷公藤悬浮细胞总萜类物质的大规模提取,可为雷公藤药用资源的开发利用提供参考依据。

Objective To optimize the extraction process of total terpenoids from Tripterygium wilfordii suspension cells. Methods The contents of triptolide, triptophenolide and celastrol were determinated by UPLC-QQQ-MS. The separation and analysis were performed on Waters HSS T3 chromatographic column （2.1 mm ×100 mm, 1.8 μm）, with 0.1% acetic acid solution-acetonitrile as the mobile phase for gradient elution. The column temperature was set at 30℃ with flow rate of 0.3 mL/min, and the sample volume was 5 μL. The composite score of 3 kinds of terpenes contents was used as index, based on single factor tests, three parameters （methanol concentration, extraction time and solvent volume） were evaluated with an L9 （34） orthogonal design. The feasibility was checked by validation tests. Results The optimum extracting conditions of total terpenoids from Tripterygium wilfordii suspension cells were 50 times volumes of 80% methanol as the solvent and ultrasonic extracting 0.5 h after dipping 12 h. In this condition, the average yields of triptolide, triptophenolide and celastrol were （52.36±0.60）, （170.73±1.57）, （388.14±5.89） μg/g, respectively. The comprehensive score was stable. Conclusion The extraction process of total terpenoids from Tripterygium wilfordii suspension cells based on orthognal design and multi-index comprehensive evaluation is reasonable, repeatable and suitable for massive extraction of total terpenoids from Tripterygium wilfordii suspension cells.