基于叶绿体基因matK及UPLC对市售大黄的种质资源鉴定和质量分析

Molecular identification and quality analysis of Rhei Rhizoma based on chloroplast matK gene and UPLC

大黄是多基原大宗常用药材,不同基原大黄其药理活性存在一定差异。本研究基于叶绿体基因matK序列对市售大黄饮片的种质资源进行鉴定,通过UPLC对其总蒽醌和游离蒽醌含量进行测定,以期确定市售大黄的主要种质及质量。本研究从27省40市收集89份市售大黄样品并提取DNA,通过PCR扩增matK基因,扩增的matK基因序列的分析结果表明,收集的市售大黄样品都是同一基原,即掌叶大黄,共鉴定了其8个基因型:Rp1、Rp2、Rp3、Rp4、Rp5、Rp6、Rp10和Rp12,其中来自甘肃、四川和云南省的Rp4和Rp6是主要流通基因型,分别占总样品量40.45%和37.08%。系统发育树结果表明,8个基因型主要分成2大支,其中主要基因型Rp4和Rp6聚在同一大支,遗传距离分析结果表明8个基因型遗传距离在0.001~0.010之间。利用UPLC检测市场样品中的总蒽醌和游离蒽醌含量结果表明,93.26%样品符合药典标准。样品之间总蒽醌含量和游离蒽醌的含量差异显著,其中游离蒽醌相差1.01%;总蒽醌含量相差3.79%,说明市售大黄样品质量存在参差不齐的现象,但是不同采集省份之间市售大黄总蒽醌和游离蒽醌含量没有显著性差异,各基因型之间的大黄样品中总蒽醌和游离蒽醌含量没有显著性差异。本文对大黄市场样品的种质资源、产地来源和药材质量进行研究,有利于指导大黄市场样品流通及临床科学合理用药,为其他中药材市场样品种质及质量鉴定提供参考。

Rhei Rhizoma is commonly used as a traditional Chinese medicine with multiple botanical origins.Different botanical sources may have different pharmacological activities.The germplasm resources of commercial Rhei Rhizoma were determined based on the chloroplast gene matK,and the anthraquinone and free anthraquinone content was determined by UPLC to analyze quality of commercial Rhei Rhizoma.Eighty-nine commercial Rhei Rhizoma samples were collected from 40 cities in 27 provinces.DNA was extracted and the matK gene was amplified by PCR.Results indicated that the collected samples were from the same botanical origin,Rheum palmatum,and 8 genotypes were identified,including Rp1,Rp2,Rp3,Rp4,Rp5,Rp6,Rp10 and Rp12.Rp4 and Rp6,cultivated in Gansu,Sichuan and Yunnan provinces were the main circulating genotypes,representing 40.45%and 37.08%of the total samples,respectively.Phylogenetic tree analysis showed that the eight genotypes were mainly divided into two branches,of which the main genotypes Rp4 and Rp6 were in one branch.Genetic distance analysis indicated that the genetic separation of the eight genotypes was between 0.001 and 0.010.UPLC analysis indicated that 93.26%of the samples met the Pharmacopoeia standards.There were significant differences in the content of total anthraquinone and free anthraquinone among the samples,in which the difference in free anthraquinone was 1.01%and the difference in total anthraquinone content was 3.79%,indicating that the quality of commercial Rhei Rhizoma samples varies considerably.There was no significant difference in the content of total anthraquinone and free anthraquinone in commercial Rhei Rhizoma among different collection provinces and genotypes.This study will help guide the circulation of Rhei Rhizoma in the market and provides valuable insights for molecular identification and quality analysis of other traditional Chinese medicines.