Study On Antibacterial "Chromatogram-Effect" Relationship Of Zanthoxylum Nitidum (Roxb.) DC

Chinese herbals have a long history and significant effects, current common mode of quality evaluation of Chinese medicine in China is to detect the main known components as indicators.This model only considers the individual component of many complex components that Chinese medicine contained, ignores the synergies and the antagonistic effect between components.Therefore,it is difficult to evaluate the intrinsic quality of traditional Chinese medicine objectively and effectively,also to take advantage of Chinese medicine mixed with fake. As a result,fingerprint of Chinese medicine come into being as a quality evaluation model that can fully reflect the existing state of chemical substances contained,which has became most effective and admited internationally as means to control quality of Chinese medicine and natural medicine,widely accepted home and abroad as a model for traditional Chinese medicine's assessment.However,current fingerprinting technology also display the existing state of chemical composition groups of Chinese medicine only,still can not explain the correlation of composition with the efficacy.To truly realize the objective and effective quality evaluation of traditional Chinese medicine,in addition to chemical components which are to be expressed using fingerprint technology,further researchs on associativity between fingerprint and the efficacy are needed to establish a new model of quality assessment that can predict the efficacy of Chinese medicine by determine its fingerprints, that is "chromatogram-effect "relationship model.ObjectiveRutaceae plant Zanthoxylum nitidum was used to construct chemical fingerprint, Staphylococcus aureus, Escherichia coli, Bacillus subtilis, Salmonella typhimurium and Klebsiella pneumoniae were used as pharmacodynamic models to obtain quantitative characteristics of chemical fingerprint and pharmacodynamic data. On this basis, back technique was used to find the corresponding relationship between chemical chromatogram and effectiveness of Zanthoxylum nitidum (Roxb.) DC.Methods1. Antibacterial active position and process for extraction of Zanthoxylum nitidum (Roxb.) DC. were selected by antibacterial test.Zanthoxylum nitidum (Roxb.) DC.was extracted by petroleum ether, chloroform, ethyl acetate and water separately, got extracts of different polar solvent; the effective part extracted by soxhlet extraction, ultrasonic extraction, reflux extraction and so on, extracts by different processes were obtained. Microdilution method was used to detect antibacterial activity of extracts by different solvent and different processes to Staphylococcus aureus, Escherichia coli, Bacillus subtilis, Salmonella typhimurium and Klebsiella pneumoniae. Effective antibacterial part and good extraction process of Zanthoxylum nitidum (Roxb.) DC.were definitely.In addition, antibacterial activity of 43 Zanthoxylum nitidum (Roxb.) DC. samples was determined by agar diffusion method,got inhibition zone of all the samples to the five kinds of bacteria.2. HPLC chemical fingerprint of Zanthoxylum nitidum (Roxb.) DC.was Established.HPLC conditions were optimized, according to the requirements of chemical fingerprint, chromatographic fingerprint evaluation system software of Chinese medicine promulgated by state pharmacopoeia commission was used to establish and evaluat HPLC fingerprint of Zanthoxylum nitidum (Roxb.) DC. antibacterial activity part.3. "Chromatogramm-activity" relationship model of Zanthoxylum nitidum (Roxb.) DC.was constructed.The HPLC fingerprint peak areas of Zanthoxylum nitidum (Roxb.) DC. were quantified and combined with antibacterial activity measured by disc agar diffusion method,used back technique to construct antibacterial "chemical chromatogram"-" efficacy" relationship of Zanthoxylum nitidum (Roxb.) DC.Results1. Extract of Zanthoxylum nitidum (Roxb.) DC. by chloroform had better antibacterial activity,which contained Nitidine chloride and other alkaloids. Technology of sample solution preparation was directly refluxed by chloroform. Antibacterial activity of samples from the highest to the lowest:Guangxi Fusui 2> Guangxi Daxin Shuolong> Guangxi Napo> Guangxi Qinzhou Nali town> control medicine 1> Guangxi Chongzuo.2. Most similarity of HPLC fingerprint established were about 0.9, there were 24 peaks shared which include Nitidine chloride, Chelerythrine and other ingredients.The established HPLC fingerprint of Zanthoxylum nitidum (Roxb.) DC.could be used to distinguish non-medicinal parts and pseudo commodeties, which was conducive to regulate the quality standards of Zanthoxylum nitidum (Roxb.) DC.3. Back technique showed that:In the 24 fingerprint peaks of HPLC fingerprints, there were 17 peaks contributed to the antibacterial action on Staphylococcus aureus,"chromatogram-effect" relational expression of Staphylococcus aureus was:Y=11.562-9.542X1+3.149X3-3.720X4+7.903X5-3.265X7-1.422X8+1.929X10+1.919X11-1.614X12-1.143X13-3.969X14-1.438X15-3 .036X16+3.412X20+5.826X21+1.882X22+3.836X24(R2=0.921);17 peaks contribu-ted to the antibacterial effect on Escherichia coli,"chromatogram-effect" relational expression of Escherichia coli was:Y= 11.074-5.969X1+2.102X2+ 2.809X3+2.051X4+3.522X5-1.638X6-0.840X7-0.789X8-0.844X10+1.070X11-0.57 5X13-1.150X15-2.031X16-1.795X18+1.302X21-0.576X23+3.952X2(R2=0.914);13 peaks contributed to the antibacterial effect on Bacillus subtilis,"chromatogram-effect"relational expression Of Bacillus subtilis was:Y=12.397-8.362X1+2.211 X3-2.200X4+2.656X5-2.516X8+1.421X9+1.347X11-2.552X16+2.946X18+1.366X1 9+2.311X20+3.835X21+3.119X24 (R2=0.908);13 peaks contributed to the antibacterial effect on Salmonella typhi,"chromatogram-effect" relational expression of Salmonella typhi was:Y=7.505-4.742X1-2.478X4+2.999X5+1.969 X6+1.958X11-1.170X12-1.108X13-1.273X15+4.023X19+1.740X20+2.953X21+1.90 2X22+0.412X23 (R2=0.936);17 peaks Contributed to the Antibacterial action on Klebsiellapneumoniae,"chromatogram-effect" relational expression of Klebsie-llapneumoniae was:Y=11.103-8.016X1+1.177X3-1.717X4+3.849X5-1.183X7-1.355X8+1.525X10+1.206X11-1.217X12-0.623X13-1.435X14-1.612X16+1.556X19 +2.235X20+4.794X21+1.162X22+2.910X24 (R2=0.919).We found that the calculated value derived from "chromatogram-effect" relationship is very close to the actual value By verified 10 samples.Conclusion1. Zanthoxylum nitidum (Roxb.) DC.of different places, different sources have different chemical compositions which cause the difference of efficacy. Therefore, the traditional quality control of Zanthoxylum nitidum (Roxb.) DC. that use contents of single or several chemical components can not reflect the quality of medicines well.2. By analyze HPLC fingerprint of efficacy part, not only better reflect active ingredients such as Chelerythrine, and also identify the authenticity of the herb better, in order to control the quality and efficacy of Zanthoxylum nitidum (Roxb.) DC.3. The "chromatogram-effect" relationship in this study can evaluate the internal quality of Zanthoxylum nitidum (Roxb.) DC. objectively and effectively, provide the basis for the use of herbs.