摘要
中药的化学成分是药物发挥药效作用的物质基础,是中药现代化的关键所在。本文利用多种现代分析方法,对中药山茱萸的炮制工艺、山茱萸炮制前后化学成分的变化规律、山茱萸苷类化合物的质谱特征、山茱萸与五种临床常用配伍药材配伍前后的化学变化等一系列问题进行了系统研究,从而为阐明山茱萸炮制原理、建立炮制前后药材的组分库及质谱指纹图谱、规范山茱萸炮制方法和诠释配伍机制提供了理论依据和科学方法。
本文首先首次采用电喷雾多级串联质谱技术和高分辨质谱FTICR-MS及SORI-CID技术对山茱萸现有标准品化合物马钱苷、熊果酸、齐墩果酸和没食子酸进行了质谱研究。总结了这些化合物的特征质谱行为,并且利用超高分辨率、准确质量确证了中性碎片丢失,进一步证明了对其碎裂机理的推断。同时实验表明通过氢氘转换和人为加入金属离子可以提高山茱萸苷类化合物分析灵敏度,从而为其提供较多结构信息。并且在串联质谱中表现不同的碎裂行为,为山茱萸苷类化合物结构分析提供补充信息,能够有效的鉴定山茱萸皂苷的结构。利用这些特征可以简便快捷地区分出山茱萸中结构类似的皂苷类化合物、有机酸化合物及其同分异构体。
其次,利用高效液相色谱法(HPLC)和薄层扫描法(TLC),采用正交设计对山茱萸有效成分超声提取工艺和山茱萸炮制工艺进行了优化。并对其水分含量、灰分含量、主要有效成分含量进行了分析测定,建立了相应质量标准,对生药的鉴别和制剂的质控起到重要的作用。
之后,利用液质联机(HPLC- ESI)法、电喷雾质谱法(ESI-MS)和分光光度法(UV)法,对山茱萸炮制前后有效成分的变化规律进行了研究,发现山茱萸炮制前后有效物质的质和量都发生了较大的变化,与山茱萸生用和制用效用不同相符。同时根据苷类化合物的电喷雾质谱规律以及色谱保留时间在线分析鉴定了山茱萸中的七种苷类化合物。建立了一种有效检测山茱萸中苷类化合物的快速灵敏的分析方法,为建立山茱萸药材的质量控制标准提供了借鉴。
最后,以中医药理论为指导,利用高效液相色谱、电喷雾质谱等方法对山茱萸与临床常用配伍药材白芍、生晒参、红参、五味子和熟地配伍前后的变化进行研究,结果发现山茱萸中有效成分在酸性和中性溶液中比较稳定,但对其他药材中有效成分有所影响,部分化学变化与煎煮过程有较大的关系。为组方的合理性和科学性提供了一定的化学物质基础依据。
The chemical components in traditional Chinese medicine(TCM)are responsible for their therapeutic effects and very important in the modernization of TCM. A serious of systematic studies-the processing procedure,variety regulations before and after the processing,mass spectrometry characteristics of glycosides,chemical changes happened in the combinations of Fructus Corni with other five TCM common used in clinic, was carried on by using various modern analytical methods to provide theoretical basis and scientific methods for clarifing the processing principle,establishing the components database and mass spectrometry fingerprints,gauging the processing method and annotating the combination mechanisms.
Standard substances of Fructus Corni in existence were firstly analyzed by using electrospray ionization tandem mass spectrometry (ESI-MSn) and Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) using sustained off-resonance irradation/collision-induced dissociation (SORI-CID) method at high mass resolution. Mass spectrometry character behavior has been summed up and product ion mass spectra have been examined at high mass resolution allowed unambiguous determination of the elemental composition of fragment ions. Both the hydrogen and deuterium exchange and the alkali metal cationization on fragmentation pathways in mass experiments can provide complementary useful structural information. These characteristics can be used to distinguish similar structure in glycoside compounds, organic acid compounds and their isomers simply and efficiently.
By using HPLC and UV analysis methods, L9 (34) orthogonal designs were carried on for the optimizations of the ultrasonic extraction Procedure and the processing procedure of Fructus Corni. And then the moisture content, ash content, and the main effective constituents were determined and the corresponding quality standards were established which played an important role in the identification and quality control.
The variety regulation of effective components in crude and processed Fructus Corni was investigated by using HPLC- ESI, ESI-MS and UV. The results show that there is an obvious change in the procedure of Fructus Corni processing, which agreed with the effect dissimilarity of crude Fructus Corni with processed Fructus Corni. Then the extracts of Frutus Corni have been analyzed by HPLC-ESI-MSn. According to these mass spectrometry characteristic regulations and the retention time (tR) of HPLC, the structures of seven kinds of glycosides have been identified. The characteristic fragment ions of CID of glycosides have been discussed, which are specific and useful for the identification of some glycosides in Fructus Corni.
Guided by the Chinese medicine theories, the changes before and after the combinations Fructus Corni with different medicine herbs in different proportion were studied by using HPLC, ESI, etc. The experiment results show that the effective components in Frucus Corni are stable in acid and neutral solutions, but they could influence the constituents in other medicine materials. A part of changes occurred in the decoction. It provides chemical basis for studying traditional Chinese medicine prescription.
引文
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