| Compound Tujingpi tincture is a commonly used compound preparation of traditional Chinese medicine and western medicine. It’s made of the extract of cortex Pseudolaricis and benzoic acid, salicylic acid. For the anti-epidermal fungi and antipruritic effect that compound Tujingpi tincture is commonly used in clinical to treat tinea manus, tinea pedis and tinea corporis, etc. Currently, there is only research on the traits, the identification, the inspection(ethanol content and microbial limit test) and the determination of content of total acids in its quality standard, and yet the content limitation of Tujingpi’s main ingredients has not been clearly defined, which cannot reflect its internal quality effectively. To improve and enhance the quality standard system of compound Tujingpi tincture, four steps are included in this study:firstly, A HPLC method was established to determine the Tujingpi’s multi-index components and main chemical composition of the preparation; secondly, the quantitative analysis of multi-components with singe marker (QAMS) was established for simultaneous determination of four diterpenoids in the preparation; thirdly, the fingerprint of the preparation was established; and finally, a verification of the microbial limit test was developed for the preparation.In the first part of this study, the contents of the main ingredient of Tujingpi, pseudolaric acid B(PAB), and two main chemical components in compound Tujingpi tincture, benzoic acid(BA) and salicylic acid(SA) were determined by HPLC. The results showed that the calibration curves were linear in the ranges of1.29～25.78μg·ml-1(r=0.9998) for pseudolaric acid B,0.022～0.266mg·ml-1(r=0.9999) for benzoic acid and0.0110.110mg·ml-1(r=0.9998) for salicylic acid. The average recoveries (n=6) were100.4%,100.9%and100.5%, with RSD were1.00%,0.84%and 0.75%, respectively. This method is convenient, accurate, and with good reproducibility that can be applied for the quality control of compound Tujingpi tincture.In the second part, the content determination of pseudolaric acid A, pseudolaric acid B, pseudolaric acid B-O-β-D-glucoside and pseudolaric acid C in Compound Tujingpi tincture were determined by HPLC. The quantitative analysis of multi-components with singe marker(QAMS) was adopted for the quality evaluation of compound Tujingpi tincture. The results displayed that the calibration curves have good linear relationship with the peak area in the ranges of11.598～82.845μg·ml-1for pseudolaric acid B-O-β-D-glucoside,1.206～27.009μg·ml-1for pseudolaric acid C,14.746～126.392μg·ml-1for pseudolaric acid B and0.159～2.274μg·ml-1for pseudolaric acid A. The average recoveries (n=6) of four diterpenoids were99.0%(RSD=0.29%),99.5%(RSD=0.15%),101.5%(RSD=0.17%),99.1%(RSD=0.26%), respectively. Using pseudolaric acid B as the internal reference substance, the relative correction factors(RCF) of the other three diterpenoids were0.6814,1.2253and1.0650, respectively. The content of other three diterpenoids was simultaneously calculated according to their RCFs. The contents of17samples were also determined by external standard method, comparing the difference of the two methods to validate the accuracy and feasibility of the QAMS. The results indicated that there no significant differences between the calculated values and the measured values. The QAMS method is feasible and accurate to evaluate the contents of four diterpenoids in compound Tujingpi tincture, which can be used for the quality control of it.In the third part, the fingerprint of compound Tujingpi Tincture was established by HPLC. The fingerprint of compound Tujingpi Tincture contains13common peaks with reference to the peak of pseudolaric acid B. The standard chromatographic fingerprints were established and the similarity of14batches was calculated. The establishment method has good precision, reproducibility and stability, which provides necessary supplement to the study on quality control of compound Tujingpi Tincture.In the last part, a verification of the microbial limit test for compound Tujingpi tincture was established according to the Chinese Pharmacopoeia2010edition. The results showed that the membrane filtration method can be used for bacterial count, and the medium dilution method can be used for molds count, the membrane filtration method can be used for specified mirco-organisms. The establishment method can effectively remove the bacteristatic activity, and make the compound Tujingpi tincture detectable accurately, so it can be used for the quality control of compound Tujingpi Tincture. |
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