Studies On Chromatogram-effect Relation Of Zanthoxylum Nitidum (Roxb.) DC. On Tumor Cell Lines HeLa, HepG-2

The curative effect of traditional Chinese medicine is results of combination effects of many compositions, which cooperation, complementation or restrict with each other. At present, one or few markers are employed for evaluation the quality of traditional Chinese medicine. It is thus not useful or feasible to analyze the quality evaluation of traditional Chinese medicine. Fingerprint technology was introduced for the identification and quality evaluation of traditional Chinese medicine by SDA. Since 2000 SDA promulgating《Fingerprint technical requirements fo TCM injections (temporary)》, fingerprint has been accepted by research and industrial groups, and become to a hot topic. But the fingerprint research at the present stage is confined to "chemical composition group" analysis, which makes it difficult to determine the main therapeutic activity and quality of TCM. Information resulted from fingerprint should be corresponded or highly correlated with pharmacodynatics. Establishing a quality evaluation systerm, which can reflect the main therapeutic activity and quality of TCM , is the issue that be considered and resolved in the study.In this paper, Zanthoxylum nitidum was studied. chosen tumor cell lines HeLa, HepG-2 as efficacy model, and established the fingerprint. On the basis of that, multiple linear regression was applied to find the correlation between Zanthoxylum nitidum fingerprint and pharmacodynamics.Methods(1) Screening the anticancer capacity site of Zanthoxylum nitidumUsing reflux method, and four kinds of solvents extraction: petroleum ether, chloroform, ethyl acetate and water on Zanthoxylum nitidum, get the solvent extracts; Chosen tumor cell lines HeLa,HepG-2 as efficacy model, the anticancer capacity of four solvent extracts were determined using MTT method; Simultaneously, these extracts were chromatographed with fast HPLC methods. The anticancer capacity site of Zanthoxylum nitidum was determined by the results of MTT method and fast HPLC methods.(2) The establishment of Zanthoxylum nitidum fingerprintAccording to the chemical fingerprint of the requirements, the chromatographic fingerprint similarity evaluation system software, which was promulgated by the State Pharmacopoeia Commission, was used to establish and evaluate the HPLC fingerprint of anti-tumor activity site.(3) Detected the anticancer capacity of different sources of Zanthoxylum nitidumChosen tumor cell lines HeLa, HepG-2 as efficacy model, the anticancer capacity of 43 sources of Zanthoxylum nitidum extracts were determined using MTT method.(4) Construct"spectrum-effect"relationships modelMultiple linear regression was applied to construct a multivariate regression model,which could predict the anticancer capacity from the fast chromatograms.Results(1) Chloroform extract was the anti-tumor active site of Zanthoxylum nitidum (Roxb.)DC. , because this site contains alkaloids which have anticancer capacity, such as nitidinechloride, liriodenine, cheletythrine, ethoxychelerythrine.(2) The HPLC-fingerprint of Zanthoxylum nitidum was established, and the similarity of fingerprint had reach to 0.7. All date were done with the chromatographic fingerprint similarity evaluation system software, choiced out 24 fingerprints peaks, including nitidinechloride, ethoxychelerythrine.( 3 ) There was huge difference on anticancer capacity between Zanthoxylum nitidum samples. Follow sourses of Zanthoxylum nitidum had better anticancer capacity: Guangxi Napo county, Datang town, Daxin county, Rongshui county, Fusui county.(4) Variables were selected by multiple linear regression, 17 fingerprints peaks related to anticancer capacity to HeLa, and the spectrum-effect relationships:Y=28.071+18.451X1-13.693X2-4.670X4+9.028X6+6.292X7-1.817X8-3.428X11+10.852X14-5.395X15+6.936X17+9.263X18+8.904X19+4.567X20-9.226X21 -8.954X22+3.619X23-5.357X24 (R2=0.957);17 fingerprints peaks related to anticancer capacity to HepG-2, and the spectrum-effect relationships:Y=70.255-16.695X1-2.756X3+11.519X5+3.107X6-4.491X7-1.737X8 -1.052X9+13.224X10-11.944X11-9.035X12-25.106X15+9.911X16+10.796X17 +10.489X18+3.432X20-12.643X21+6.152X23 (R2=0.977).Get 10 Zanthoxylum nitidum samples for verification, the deviation of predictive value and true value was not exceed 10%. The result indicated that the multiple linear regression could predict the anticancer capacity to HeLa, HepG-2 from the fast chromatograms.Conclusion(1) Different sourses of Zanthoxylum nitidum have different compositions, which leads to different curative effect. So we can not detect one or few markers for evaluation the quality of Zanthoxylum nitidum. (2) HPLC-fingerprint of Zanthoxylum nitidum makes a better reflect on pharmacologically active components, such as nitidinechloride, ethoxychelerythrine and other composition, so we can evaluate the quality of Zanthoxylum nitidum by this way.(3) the spectrum-effect relationships on HeLa, HepG-2 has good predictive and describing abilities to evaluate the quality and therapeutic activity of Zanthoxylum nitidum. It is more reasonable to evaluate the quality of TCM by the bioactivity- fingerprint.