基于psb A-trn H序列的秦艽分子鉴定及其药效分析

Molecular Identification and Pharmacodynamic Analysis of Gentiana Using psb A-trn H Sequence

秦艽相近相似混伪品较多,且药效差异显著,为准确区分西北地区秦艽混伪品及药效,试验利用psb Atrn H基因作为条形码鉴定5种来自西藏、四川、青海地区相近秦艽的药材,基因测序后,在GenBank数据库中通过序列比对选取药材同源序列,利用MEGA 7.0软件计算双参数(K2P)遗传距离,并基于邻近法构建系统进化树,分析物种系统发育关系;另外,利用紫外分光光度法检测试验样本龙胆苦苷含量及DPPH清除率,评估不同地区药材龙胆苦苷含量及抗氧化能力。结果表明,结合K2P遗传距离、系统发育关系和psb A-trn H基因相似性确定试验样品中3种为粗茎秦艽(Gentiana crasicaulis),另外2种分别为麻花秦艽(G. straminea)和黄管秦艽(G.officinalis)。所有样品龙胆苦苷含量在8.56%~12.76%,其中大小表现为黄管秦艽>麻花秦艽>粗茎秦艽;DPPH清除率范围为66.49%~90.68%,其中麻花秦艽>黄管秦艽,而3个不同地区的粗茎秦艽DPPH清除率差异较大。综上,psb A-trn H基因序列在秦艽植物鉴定和系统分类研究上十分有效,不同地区试验样品中龙胆苦苷含量均较高,抗氧化能力较强。

There are many similar counterfeit varieties in the medicinal materials of Gentiana and significant differences in their pharmacodynamics content. To accurately distinguish the Gentiana samples in northwestern China and identify its pharmacodynamics, in this study, 5 similar medicinal materials of Gentiana from Tibetan, Sichuan, Qinghai were identified using psb A-trn H sequences as the bar code. After gene sequencing, the homologous sequences of the medicinal materials were selected by sequence alignment in GenBank database, the Kimura 2-parameter genetic distance was calculated using MEGA 7.0software, and the phylogenetic tree was constructed based on the neighbor-joining method to analyze the phylogenetic relationship of species. Additionally, gentiopicroside content and DPPH radical scavenging rate in the test samples were detected using ultraviolet spectrophotometry to evaluate gentiopicroside content and antioxidant capacity of medicinal materials in different regions. The results showed that in all five samples, three belonged to Gentiana crasicaulis, the other two belonged to G.straminea and G. officinalis, respectively, according to K2P distance, phylogenetic relationship, and psb A-trn H gene similarity.Gentiopicroside contents in the samples ranged from 8.56% to 12.76%(G. officinalis>G. straminea>G. crasicaulis), DPPH radical scavenging rates were 66.49%-90.68%(G. straminea>G. officinalis), and the DPPH radical scavenging rates in the samples of G. crasicaulis from three different regions were of difference. Based on research above, these results indicated that the gene psb A-trn H could be very effective on identification of Gentiana and phylogenetic study, the content of gentiopicroside in the test samples from different regions was high and the antioxidant capacity was strong.