基于荧光定量PCR方法检测金龙胶囊配方中守宫投料量的初步研究

Preliminary study of determination of Shougong in Jinlong Capsules based on fluorescence quantitative PCR method

基于COⅠ序列设计中国壁虎特异性引物对GZG1/GZG2,采用荧光定量PCR技术,对金龙胶囊配方中的中国壁虎组分进行定性及定量分析。所建立的中国壁虎定量分析的标准曲线为y=-3.012 7x+34.501,其中y为Ct值,x为COⅠ片段拷贝数以10为底的对数值。对10份自制供试样品的测定中,含有中国壁虎的供试品均出现扩增,不含中国壁虎的全伪品组及空白对照组未见扩增,其中全正品组、正伪品1∶1组及正伪品1∶4组所得中国壁虎COⅠ片段拷贝数依次为11.511×10^6,6.416×10^6,2.553×10^6拷贝/μL,大致符合投料比,可为含有动物药成分的中成药提供质量控制的新思路。

A pair of species-specific primer(GZG1/GZG2) based on COⅠ sequence regions for identification of Gekko chinensis were designed. A fluorescent quantitative PCR method was established to identify and quantify G. chinensis from Jinlong Capsules Formula. A standard curve for quantitative analysis of G. chinensis was established(the standard curve equation: y=-3.012 7x + 34.501, y is Ct value, x is lg N, N is the copies of COⅠ fragment from G. chinensis). Samples included G. chinensis appeared amplification, while falsify group(not included G. chinensis) and negative control did not have amplification products. The copy number of COⅠ region of G. chinensis was respectively 11.511×106, 6.416×10^6,2.553×10^6 copies/μL in all quality goods, quality goods-adulterants 1∶1, quality goods-adulterants 1∶4. The results accorded with proportion of adding amount roughly. This study can provide a new strategy for quality control of Chinese patent medicine containing animal drug ingredients.