贝莱斯芽孢杆菌发酵对乳康方剂中药有效成分提取效果的影响

doi:10.16431/j.cnki.1671-7236.2023.06.038

摘要/Abstract

摘要： 【目的】探究贝莱斯芽孢杆菌发酵对自拟乳康方剂中主要药物活性成分王不留行黄酮苷、咖啡酸和芦丁含量变化的影响。【方法】试验建立测定乳康方剂中药中王不留行黄酮苷、咖啡酸和芦丁含量的高效液相色谱（HPLC）检测方法，考察该方法的线性范围、精密度、稳定性、重复性、加样回收率；以乳康方剂中药王不留行、蒲公英、黄芪、当归、甘草药粉为主要成分占30%，黄豆粉占10%、CaCO₃占0.2%、超纯水占59.8%，配制发酵中药培养基，发酵组接种2%（v/w）贝莱斯芽孢杆菌种子液混合均匀后发酵，发酵后加入2倍量水，反复文火煎煮，过滤为待测样；HPLC测定发酵与非发酵乳康方剂中药水煎提取液中的王不留行黄酮苷、咖啡酸和芦丁含量。【结果】建立的测定乳康方剂中王不留行黄酮苷、咖啡酸和芦丁含量的HPLC方法分别在0.01~0.5、5~120和10~100 μg/mL范围内线性关系良好，建立的方法经考察精密度、稳定性、重复性、加样回收率均良好。在相同物理提取条件下，经贝莱斯芽孢杆菌发酵的乳康方剂中药提取液中王不留行黄酮苷、咖啡酸和芦丁含量分别为4.99、6.86和5.81 mg/mL，非发酵的乳康方剂中药提取液中王不留行黄酮苷、咖啡酸和芦丁含量分别为3.70、3.01和3.27 mg/mL，二者相比差异极显著（P<0.01），说明贝莱斯芽孢杆菌适用于发酵乳康方剂中药，与非发酵水煎提取相比提取效果更佳，有利于王不留行黄酮苷、咖啡酸和芦丁药效活性成分释放。【结论】贝莱斯芽孢杆菌发酵乳康方剂中药能显著提高主要有效成分含量，为乳康方剂中药成分高效提取提供了技术参考。

关键词: 贝莱斯芽孢杆菌; 泌乳; 中药; 发酵; 有效成分

Abstract: 【Objective】 The experiment was conducted to explore the effect of fermentation by Bacillus velezensis on the content changes of the main active components of the self-prepared Rukang prescription, including the flavonoid glycosides from Vaccaria segetalis, caffeic acid and rutin.【Method】 The study established high performance liquid chromatography(HPLC) methods and investigation method linear range, precision, stability, repeatability, sample recovery for the determination of flavonoid glycosides from Vaccaria segetalis, caffeic acid and rutin in self-prepared Rukang prescription.The fermentation culture medium was prepared with the main ingredients of self-prepared Rukang prescription, such as the powder of Vaccaria segetalis, Taraxacum mongolicum Hand.-Mazz, Astragalus membranaceus, Angelica sinensis, Glycyrrhiza uralensis Fisch., accounting for 30%, soybean powder accounting for 10%, CaCO₃ accounting for 0.2%, and ultra-pure water accounting for 59.8%.The fermentation group was inoculated with 2% (v/w) of Bacillus velezensis seed liquid, mixed evenly, and then fermented.After fermentation, double amount of water was added, repeatedly simmered, and filtered as samples to be tested.The contents of flavonoids glycosides from Vaccaria segetalis, caffeic acid and rutin in the fermented and non-fermented Rukang prescription were determined by HPLC.【Result】 The established HPLC methods for the determination of flavonoid glycosides from Vaccaria segetalis, caffeic acid and rutin in Rukang prescription, the linear relationship was good within the range of 0.01-0.5, 5-120 and 10-100 μg/mL, respectively, and the methodological investigations were good, the established method had good precision, stability, repeatability and recovery rate.Under the same physical conditions, the contents of flavonoids glycosides from Vaccaria segetalis, caffeic acid and rutin in the extract of self-prepared Rukang prescription fermented by Bacillus velezensis were 4.99, 6.86 and 5.81 mg/mL, respectively.The contents of flavonoids glycosides from Vaccaria segetalis, caffeic acid and rutin in the non-fermented self-prepared Rukang prescription were 3.70, 3.01 and 3.27 mg/mL, respectively.The difference between the two groups was extremely significant (P<0.01).The results showed that Bacillus velezensis was suitable for fermenting self-prepared Rukang prescription, and the extraction effect was better than that of non-fermented group.The fermentation of Bacillus velezensis was beneficial to the release of flavonoids glycosides from Vaccaria segetalis, caffeic acid and rutin in self-prepared Rukang prescription.【Conclusion】 The fermentation of self-prepared Rukang prescription by Bacillus velezensis could significantly increase the content of main effective ingredients, which provided a technical reference for the efficient extraction of self-prepared Rukang prescription.

Key words: Bacillus velezensis; lactate; Chinese medicine; ferment; active component

中图分类号:

S859.5

薛沾枚, 刘雪松, 张艳, 张备, 孟维珊, 张国华, 王岩, 钟鹏, 邱景会, 汤继龙, 史同瑞. 贝莱斯芽孢杆菌发酵对乳康方剂中药有效成分提取效果的影响[J]. 中国畜牧兽医, 2023, 50(6): 2540-2549.

XUE Zhanmei, LIU Xuesong, ZHANG Yan, ZHANG Bei, MENG Weishan, ZHANG Guohua, WANG Yan, ZHONG Peng, QIU Jinghui, TANG Jilong, SHI Tongrui. Effect of Bacillus velezensis Fermentation on Extraction of Active Components from Traditional Chinese Medicine of Rukang Prescription[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(6): 2540-2549.

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