基于灰色关联分析方法对鹿茸“补肾健骨”作用与品质相关性研究

Nowadays, due to the lack of research on the correlation between CERVI CORNU PANTOTRICHU effect and its quality, a new method, based on the grey correlation analysis, is put forward for the first time to study the correlation between the CERVI CORNU PANTOTRICHU quality and its effect of tonifying the kidney and strengthening the bone. The research can be divided into three sections. Firstly, different qualities of CERVI CORNU PANTOTRICHU are chosen. The effects of CERVI CORNU PANTOTRICHU on the proliferation and differentiation of both MC3T3-E1 and BMSCs are studied. Also the effect of CERVI CORNU PANTOTRICHU on ovariectomized rats as a pharmacological model of osteoporosis is studied. Secondly, calculate the relative correlation degree (ri) with the grey correlation analysis.The correlation between CERVI CORNU PANTOTRICHU effect and its quality is established according to the result. The main compositions from different qualities of CERVI CORNU PANTOTRICHU have been determined at the same time. Then, Stepwise regression analysis is used the main compositions content as the variables and the"composite effect index"as the dependent variables. The compositions which have the greatest affect upon tonifying the kidney and strengthening the bone are determined as the "chemical characteristic". Finally,the samples of CERVI CORNU PANTOTRICHU are added, and the characteristic compositions are determined and cluster analysis is processed. Macroscopic identification of the sub-classification are described as the "macroscopic characteristic". A new quality evaluation pattern and indicator system about CERVI CORNU PANTOTRICHU （Chinese crude drugs）can be established on the base of its clinical effect.

针对鹿茸功效与品质相关性研究的空白，本研究提出基于灰色关联度分析方法对鹿茸"补肾健骨"作用与品质相关性进行研究。拟选择具代表性的不同品质的鹿茸药材为研究对象，通过其对小鼠成骨细胞增殖与分化、大鼠骨髓间充质干细胞增殖及向成骨分化、以及对去卵巢大鼠所致骨质疏松防治作用研究，以灰色关联度分析方法的相对关联度ri，建立鹿茸"补肾健骨"作用与品质的相关性及关联程度；同时，测定不同品质的鹿茸中的各主成分含量，并分别以此为自变量、鹿茸"补肾健骨"的"综合效应指数"为因变量，进行逐步回归分析，确定对鹿茸"补肾健骨"作用影响最显著的化学成分或成分类群，即"化学表征"（含量测定指标）；在此基础上，对放大鹿茸药材的样本，进行"表征成分"含量测定与聚类分析，将得到的"各类"鹿茸药材性状特征进行描述，即"性状表征"（性状鉴别指标）。以期建立以"临床功效"为核心与导向的新型鹿茸及中药材品质评价模式与指标体系。

本研究针对鹿茸功效与品质相关性研究的空白，首次对8个不同品质鹿茸药材，对小鼠成骨细胞增殖与大鼠骨髓间充质干细胞成骨分化及对大鼠去卵巢所致肾虚骨质疏松的防治作用进行实验研究；首次引入灰色关联度分析模型，建立了8个鹿茸样品的“补肾健骨”作用与其品质相关性；进而对鹿茸“补肾健骨”作用影响最显著的化学成分类群进行统计学筛选；同时对放大样本的主要成分测定结果进行了判别与主成分分析；首次以透光率（T%）与总色度（E*ab、三维色度L*、a*、b*），对其主要特征进行了量化表征。重要显示：① 鹿茸通过促进成骨细胞分化与骨髓间充质干细胞成骨分化，对去卵巢大鼠所致的骨质疏松产生防治作用；其作用机制可能是与上调BMP-2/Smad1、Smad5/Runx2信号传导通路相关；不同品质鹿茸药材药效结果的灰关联度分析评价与质量的灰关联度方法评价结果具有一致性，并且与传统药材规格的划分基本吻合。② 在鹿茸的8类主要化学成分中，对其“补肾健骨”药效影响最显著的是氨基酸类成分；在马鹿茸多肽7个不同分子量肽段中，分子量在0.2K~3K的游离氨基酸总量最高（初步体内外药效实验结果表明其“补肾健骨”作用最强）；对鹿茸药材放大样本的成分分析中，以对氨基酸类成分的判别分析和对8类主要成分含量的主成分分析，可有效地区别梅花鹿茸与马鹿茸。③ 对传统评价鹿茸药材品质的量化表征结果，其透光率依“蜡粉片”、“蜡片”、“血片”、“白片”等依次增加；总色度值E*ab（无论内色、外色）均依“血片”、“蜡片”、“蜡粉片”、“白片”递增，不同规格、等级梅花鹿茸饮片的L*差异性最显著。本研究不仅发现了鹿茸药效作用的主要途径与主要药效物质类群；而且揭示了鹿茸“补肾健骨”作用与其药材品质的相关性；并对鹿茸药材传统品质评价特征进行了量化表征；进而建立了以“临床功效”为核心与导向的新型鹿茸品质评价模式，也为建立新型中药材品质评价模式提供了示范作用。