目的:基于成分含量和体外降糖活性优选麦冬、山药、五味子配伍比例及提取工艺。方法:以配伍比例、甲醇浓度、溶剂倍数、提取时间为因素,以麦冬皂苷D(OPD)、麦冬皂苷D'(OPD'),麦冬甲基二氢高异黄酮A(MA)、麦冬甲基二氢高异黄酮B(MB),五味子醇甲(SA),尿囊素(DA)含量及体外HepG2细胞糖代谢改善、STZ损伤NS-1细胞修复等为指标,运用星点设计-效应面法结合G1-熵权法优化麦冬山药五味子配伍比例及提取工艺。结果:麦冬、山药、五味子体外降糖最佳配伍比例为麦冬20～28 g、山药5～15 g、五味子10～14 g;最佳提取工艺参数范围为甲醇浓度>85%、溶剂倍数>8、提取时间>90 min。其中,麦冬15 g、山药7 g、五味子6 g配伍后,用6倍91%甲醇提取60 min,所得提取物对HepG2细胞试验组上清葡萄糖消耗量较少,平均为2.13 mmol·L-1;对INS-1细胞活力增殖较低,吸收度A平均为1.12。麦冬25 g、山药25 g、五味子6 g配伍后,用10倍91%甲醇提取120 min,所得提取物对HepG2细胞试验组上清葡萄糖含量消耗较多,平均为3.24 mmol·L-1,对INS-1细胞活力增殖较高,吸收度A平均为1.97。配伍比例优化试验拟合方程为Y＝0.155+0.14X1+0.013X2+0.096X3+0.12X1X1+0.10X2X2+0.082X3X3+0.054X1X2-0.15 X1X3-0.20X2X3(R-Sq=96.59%);提取工艺优化实验拟合方程为Y'＝0.63-0.010X'1+0.11X'2+0.12X'3-0.099X'1X'1-0.075X'2X'2-0.069X'3X'3+0.054X'1X'2+0.12X'1X'3+0.011X'2X'3(R-Sq=94.86%)。在此条件下,分别进行3次验证试验,试验数据组间无明显差异。结论:基于网络药理学数据,从麦冬、山药、五味子主要降糖成分和体外降糖活性情况考虑,明确配伍比例及提取工艺范围,为麦冬的“酸甘化阴”配伍中降糖活性组分研究提供参考。麦冬、山药、五味子配伍可促进细胞对上清液中葡萄糖的摄取,同时,可不同程度增加STZ致INS-1细胞损伤增殖的活力,促进细胞修复。
Objective: To explore the compatibility ratio and extraction process of Ophiopogonis Radix based on composition changes and hypoglycemic activity in vitro. Methods: The Star-point design and G1-entropy weight method were used to explore the the compatibility ratio and extraction process of Ophiopogonis Radix which were measured against the contents of ophiopogonin D (OPD), ophiopogonin D'(OPD '), methylophiopogonanone A(MA) methylophiopogonanone B (MB), schisandrin A (SA), allantoin (DA), improvement of glucose metabolism of HepG2 cells and repair of NS-1 cells damaged by STZ in vitro as indexes and were influenced by the factors as follows:the compatibility ratio, methanol concentration, solvent multiple and extraction time. Results: The best compatibility ratios of Ophiopogonis Radix were 20-28 g, 5-15 g and 10-14 g. The best extraction process parameters were methanol concentration > 85%, volume multiple > 8 times and extraction time > 90 min. Ophiopogonis Radix 15 g, Dioscoreae Rhizoma 7 g and Schisandrae Chinensis Fructus 6 g were extracted with 6 times 91% methanol for 60 min. The glucose consumption of the supernatant of HepG2 cell test group was less by this extract substance, with an average of 2.13 mmol·L-1. The proliferation of INS-1 cells was low, with an average absorbance value of 1.12. Ophiopogonis Radix 25 g, Dioscoreae Rhizoma 25 g and Schisandrae Chinensis Fructus 6 g were extracted with 10 times 91% methanol for 120 min. The extract substance consumed more glucose in the supernatant of HepG2 cell test group, with an average of 3.24 mmol·L-1. It had higher activity and proliferation of INS-1 cells, with an average absorbance value of 1.97.The final equation were that as follows:compatibility proportion optimization test:Y＝0.155+0.14X1+0.013X2+0.096X3+0.12X1X1+0.10X2X2+0.082X3X3+0.054X1X2-0.15 X1X3-0.20X2X3(R-Sq=96.59%) and extraction process optimization test:Y'＝0.63-0.010X'1+0.11X'2+0.12X'3-0.099X'1X'1-0.075X'2X'2-0.069X'3X'3+0.054X'1X'2+0.12X'1X'3+0.011X'2X'3(R-Sq=94.86%). In this condition, three validation tests were carried out, and there were no significant difference between the test data groups. Conclusion: Based on the network pharmacology data, considering the main hypoglycemic components and in vitro hypoglycemic activity of Ophiopogonis Radix, Dioscoreae Rhizoma and Schisandrae Chinensis Fructus, the compatibility ratio and extraction process range were determined, which laid a foundation for the study of the hypoglycemic active components in the compatibility of Ophiopogonis Radix.The combination of Ophiopogonis Radix, Dioscoreae Rhizoma and Schisandrae Chinensis Fructus can promote the uptake of glucose in the supernatant, increase the proliferation activity of INS-1 cells injured by STZ in varying degrees, and promote cell repair.
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