| For several decades, the quality of Chinese patent medicine has been controled only with one marker. The characteristic that Chinese medicine exerts its effect on human body through multi-components and multi-target determines the necessity of monitoring multi-components in Chinese medicines. Therefore, the quality control mode that simultaneously determines multiple components in Chinese medicines was called for quality evaluation. However, the supply of reference substances used for quantification cannot meet its demands. Apart from this, it will raise the detection cost while using traditional external standard method to simultaneously quantify the multi-compounds. These arising problems hinder applying quality control mode of simultaneously determination of multiple components to the fields of production, research, supervision and administration. The quality evaluation mode that is so-called QAMS, which is short for quantitative analysis of multi-components by single marker offers a new solution.The action of Zhizi Jinhua pill is to clear Heat and purge Fire, and cool Blood and relieve toxicity. Zhizi Jinhua pill is commonly used to cure the mouth and tongue sores, swelling and aching of gum and so on due to extremely abundant Heat in the Lung and Stomach. The assay of Zhizi Jinhua pill in China Pharmacopoeia (2010edition) is only to determine the content of geniposide. Now QAMS is applied to the quality evaluation of Zhizi Jinhua pill, to probe the feasibility of applying the method to Chinese patent medicines, raise the quality control level, and decrease the detection cost.Part1The quantification of three different structural types of ingredients in Zhizi Jinhua pill by using external standard methodA high performance liquid chromatography method was developed to simultaneously determine the contents of berberine, baicalin, wogonoside, baicalein, aloe-emodin, wogonin, rhein, emodin, chrysophanol and physcion in Zhizi Jinhua pill. The analysis was performed on a Syncronis C18column (4.6mm x250mm,5μm) by gradient elution with mobile phrases consisted of methanol, acetonitrile and0.2%phosphoric acid solution. The flow rate was1mL· min-1. The column temperature was set at30℃and the detection wavelength was set at228nm,274nm,254nm. The validation of method was carried out at two detection wavelength modes. One was single detection wavelength of254nm, which meant the peak areas of ten compounds were wholly determined at254nm. The other was multiple detection wavelengths of228nm,274nm and254nm due to the different structures of the ten compounds. In this mode, the peak areas of alkaloids, flavonoids and free anthraquinones were determined at228nm,274nm and254nm respectively. The precision, linearity, stability, repeatability and accuracy experiments met the requirements of quantitative analysis of Chinese medicines. The results showed that the analysis method achieved good resolution and no interference of impurities. The method was simple and accurate and therefore can be used for the quantitative analysis of the ten components mentioned above in Zhizi Jinhua pill.Part2The establishment of quality evaluation method of QAMS used for simultaneously determining three different structural types of ingredients in Zhizi Jinhua pill1. Emodin was chosen as the internal reference substance. The relative correction factors (RCF) of berberine, baicalin, wogonoside, baicalein, aloe-emodin, wogonin, rhein, chrysophanol and physcion to emodin were calculated through slope correction with the peak areas determined at two different detection modes mentioned in part1. Through repeatability, ruggedness tests, the RCF values at two detection modes were as follows: RCF at single detection wavelength of254nm: RCF at multiple detection wavelengths of228nm,274nm,254nm: The letters from A to J represent berberine, baicalin, wogonoside, baicalein, aloe-emodin, wogonin, rhein, emodin, chrysophanol and physcion respectively.The results showed that the RCF can be used as a constant for the determination of alkaloids, flavonoids and free anthraquinones in Zhizi Jinhua pill.2. Compared the difference retention times (△tR) and the relative retention value (RTR) of the ten components in ruggedness test, the difference in retention times was chosen for locating the nine target peaks. In order to locate peaks more accurately, it was necessary to combine the difference in retention times with the ultraviolet absorption spectra of the determined compounds and the specific chromatogram of Zhizi Jinhua pill. The identified differences in retention time was as following: J/H=16.23min.3. The experiment explored the effect of another location method named linear regression. When using this method to locate the target peaks, the internal reference substance must be chosen as one of the correction point (n≥2) due to the test components’different structures. In practical application, it was flexible according to the reference substances in stock. It was found that the location of target peaks was much more accurate when the emodin and berberine were selected as the correction points. The accuracy of location was not improved when more correction points were used. With the ultraviolet absorption spectra of the determined compounds and the reference chromatogram, it was much more accurate to locate the components while using this method.4. The established QAMS method for the quantification of alkaloids, flavonoids and anthraquinones in Zhizi Jinhua pill was directly applied to evaluate the quality of Zhizi Jinhua pill. The contents of these ten ingredients in all samples were also determined by the external standard method. No significant differences were observed between the quantitative results of the two methods with the relative error less than5%. The results indicated that the QAMS method was feasible to simultaneously quantify three different structural types of ingredients in Zhizi Jinhua pill... |
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