山豆根指纹图谱、含量测定及成分分析研究

发布时间：2018-02-04 11:29

本文关键词： 成分分析含量测定山豆根生物碱指纹图谱　出处：《北京中医药大学》2017年硕士论文　论文类型：学位论文

【摘要】：研究目的:山豆根为临床常用药,近年来关于山豆根在临床使用中出现中毒的现象屡有报道,原因之一是将毒性大的山豆根与毒性小的北豆根混用,导致患者中毒。而目前关于山豆根尚缺乏系统性的质量标准方面的研究。本研究通过山豆根进行指纹图谱、有效成分含量测定,并结合高分辨质谱进行成分分析,为完善补充现有关于山豆根的质量标准提供参考依据和思路。研究方法:(1)研究采用UPLC-PDA对山豆根70%甲醇部位的提取物进行指纹图谱分析,所得cdf格式文件导入到国家药典委员会出版的《中药色谱指纹图谱相似度评价系统A版》软件进行分析,生成山豆根的指纹图谱共有模式、共有峰以及相似度。并且指定氧化苦参碱作为参照峰,通过计算其他共有峰的相对保留时间和相对峰面积对不同产地的山豆根的差异进行评价。(2)研究通过对山豆根的提取方法进行单因素考察初步确定山豆根的最佳提取条件(包括不同方法、不同溶剂、不同时间),再通过HPLC-PDA对样品进行色谱条件考察,实现对苦参碱、氧化苦参碱、槐果碱、氧化槐果碱等四种生物碱的分离,最终进行定量,最后结果再采用SAS 8.2统计分析软件进行聚类分析,将山豆根按产地分为三类,实现对不同产地山豆根的差异评价。(3)研究通过UPLC-Q-TOF/MS对道地产区山豆根进行成分而分析,所得数据通过Waters公司的配套软件Unify进行智能匹配分析,最终确定山豆根中可能含有的成分。研究结果:(1)13批山豆根含有27个共有色谱峰,指认了其中2个分别是苦参碱和氧化苦参碱;不同批次山豆根药材共有峰相对保留时间在3.925%～4.588%之间,而相对峰面积的RSD较大,且13批山豆根药材的相似度大部分大于0.9,但其中2批北豆根的相似度则较差。(2)13批山豆根中有12批苦参碱与氧化苦参碱的总含量达到药典标准(≥0.7%),有1批含量低于药典标准,且13个产地按照苦参碱、氧化苦参碱、槐果碱、氧化槐果碱的含量大概可以分为三类。(3)通过Unify软件智能匹配,并结合文献中的特征碎片离子比对,推断出山豆根中含有的56种成分。研究结论:(1)本研究中的不同产地的山豆根在化学成分上相似度良好,该方法精密度、稳定性及重复性较好,特征性及专属性较强,可以为山豆根和北豆根的鉴别提供思路,从而避免因临床混用导致的药物中毒事件的发生。(2)结果表明,不同产地四种生物碱含量差异显著,道地产区有效成分含量相对较高。苦参碱和氧化苦参碱总量大于1.00%的产地大部分为广西,与广西为山豆根的道地产区这一事实相符合,且苦参碱与氧苦参碱的含量呈现"此消彼长";聚类分析结果表明,不同产地对山豆根生物碱的含量影响较大,不同分组的产地区分不够明显,可能是由于产地数量有限的原因导致,提示应该增加样本的产地、批次以获取更为准确客观的研究结果。(3)最终推断结果为进一步分析山豆根化学成分提供了参考思路,但是推断结果需要进一步验证。
[Abstract]:Research purposes: tonkinensis is commonly used in clinical medicine, in recent years on the Rrst in clinical use in poisoning have repeatedly reported that one of the reasons is Shandougen and toxicity big small Beidougen mix, resulting in poisoning. At present, the quality standard of radix sophorae tonkinensis is still a lack of systematic aspects of the study. The study of fingerprint by Shandougen, determination of active ingredients, combined with component analysis of high resolution mass spectrometry, to provide reference and ideas for the improvement of sophoratonkinensis supplement the existing quality standards. Research methods: (1) research by UPLC-PDA of Shandougen 70% methanol extracts were part of the fingerprint analysis, the CDF the format of the file into the A software version > < chromatographic fingerprint evaluation system published by the State Pharmacopoeia Commission analysis, fingerprint generation sophoratonkinensis common mode, there are The peak and similarity. And specify Oxymatrine as the reference peak, through the calculation of other common peak difference of relative retention time and relative peak area of different origin of sophoratonkinensis evaluation. (2) to study the optimum extraction condition was investigated by single factor extraction method of radix sophorae tonkinensis initially identified Shandougen (including the different methods, different solvents, different time), and then investigated by HPLC-PDA chromatographic conditions for samples of matrine, Oxymatrine, sophocarpine, Oxysophocarpine and separation of four kinds of alkaloids, the final quantity, and then the final results using SAS 8.2 statistical analysis software for cluster analysis, the mountain according to the origin of root is divided into three categories, realize the evaluation of differences of different origin Shandougen. (3) through the study of UPLC-Q-TOF/MS on real estate areas of Rrst composition and analysis, the data through the Waters company supporting software Matching analysis of intelligent Unify, and ultimately determine the Rrst may contain ingredients in. Results: (1) the 13 batch of Rrst containing 27 common peaks were identified, of which 2 are respectively of matrine and Oxymatrine; different batches of Shandougen common peak relative retention time in 3.925% ~ 4.588%, while the RSD of the relative peak area is larger, and the 13 batch of similarity Shandougen most of more than 0.9, but the 2 batch of rhizomamenispermi similarity is poor. (2) the total content of the 13 batch of Rrst in 12 batches of matrine and oxymatrine to reach the standard of Chinese Pharmacopoeia (more than 0.7%), 1 group was lower than the standard of Pharmacopoeia, and 13 samples in accordance with matrine, Oxymatrine, sophocarpine, Oxysophocarpine contents can be divided into three categories. (3) by Unify software, intelligent, and combined with the characteristic fragment ions in comparingliterature, infer 56 contain tonkinensis in . conclusions: (1) in the study of different origin Shandougen good similarity in chemical composition, the precision, stability and repeatability, characteristic and specificity is strong, can provide ideas for the identification of Rrst and Beidougen, so as to avoid the drug caused by clinical mix poisoning (2). The results show that the difference of four kinds of alkaloid content in different areas significantly, the content of effective components of real estate areas is relatively high. Matrine and Oxymatrine is greater than 1% of the total area most of the Guangxi, and Guangxi for the fact that the real estate areas of Rrst accord and the content of matrine and oxygen matrine showed a "shift"; cluster analysis showed that the effect of alkaloid content in different areas of Rrst larger, different groups of origin differentiation is not obvious, which may be due to the limited number of places,. Should increase the sample from batch to obtain more accurate and objective results. (3) the final inference results provide a reference for the further analysis of Rrst chemical composition, but the inference results need to be further verified.

【学位授予单位】：北京中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R284.1

【参考文献】