Studies On The Anti-lung Cancer Material Basis Of Traditional Chinese Medicine Compound Formulae SanCaoFang

The subject mainly studied the anti-lung cancer material basis of Traditional Chinese Medicine（TCM） compound formulae SanCaoFang. This subject also reviewed the current investigative methods and the advanced studies of TCM compound formulae SanCaoFang and its component medicines.The subject was abided by the hypothesis of "three layer and multi-dimension structure" of TCM material basis which been proposed by our laboratory to explain the material basis of SCF from macroscopy to microcoscopy.TCM compound formulae SanCaoFang which be composed of Brunellae cum fructu,et al,was used to treat non-small cellular lung cancer in Jiangsu hospital integrated traditional and western medicine.The patients could be improved symptoms and life quality,reduced recurrence and prolonged survival time by this proved recipe.For research of consequence and development of preparation,the subject studied the anti-lung cancer material basis of this compound,main content as follow:（1） The different solvent parts of SCF were obtained by reflux extraction;these were the 95%alcohol extraction（solvent partⅠ）,the 60% alcohol extraction（solvent partⅡ）,the 30%alcohol extraction（solvent partⅢ） and the water extraction（solvent partⅣ）.The chemical constituents of the solvent parts were compared by extraction rate,preliminary chemical analysis,assay of contents of effective and HPLC.（2） The first,the anti-lung cancer material basis of SanCaoFang was explored initially by the subject.In vitro experiments were taken to the different solvent parts and compatibility of SCF.Two models of human lung cancer cell,SPC-A-1 and A549,were established,and MTT was used for in vitro cytostasis experiment.Experimental result indicated that,solvent partⅠhave strong inhibition activity to the two human lung cancer cell,and IC₅₀ was respective 8.34 and 12.21 mg crude drug/mL;solvent partⅡalso had inhibition activity,and IC₅₀ was respective 3.01 and 2.77 mg crude drug/mL; inhibition activity of solvent partⅢandⅣwas comparative weak; furthermore,compatible inhibition activity of partⅠandⅡwas significantly strengthened,and IC₅₀ was respective 0.45 and 0.54 mg crude drug/mL.In vivo experiments were also taken to the different solvent parts and compatibility of SCF.C57BL/6J small black rats injected with Lewis lung cancer cell were used for in vitro experiment;anti-tumor ability and anamnestic reaction were used as evaluation index.Results showed that solvent partⅠandⅡhave strong anti-tumor ability,tumor inhibition rate of high-dose were respective 47.99%and 49.99%,while low-dose group were respective 4.46%and 18.41%;the compatible tumor inhibition rate of high-dose and low-dose were respective 64.12%and 30.27%;compared with the tumor weight of negative control group,tumor weight of high-dose group and compatibility of partⅠandⅡwere significantly decreased（p＜0.05）.（3） For the next experiment of researching the anti-lung cancer material basis of SanCaoFang,and on account of the previous results,identification of the effect solvent partsⅠandⅡof SCF,macroporous adsorptive resins were used to depurate constituents from the solvent parts.Total flavonoids and total terpenes were taken as the index,SP825 and HP20 series which were screened from five different macroporous adsorptive resins had the best depurate effect,and after screening,optimum technology was selected: wet method upper sample,first washed with mass water and 10%alcohol, then respectively washed with 50%and 95%alcohol,finally eluent was collected.After that over-column fluid（component A_Ⅰ）,water eluent（component B_Ⅰ）,10%alcohol eluent（component C_Ⅰ）,50%alcohol eluent（component D_Ⅰ） and 95%alcohol eluent（component E_Ⅰ） of partⅠwere obtained;over-column fluid（component A_Ⅱ）,eluent（component B_Ⅱ）, 10%alcohol eluent（component C_Ⅱ）,50%alcohol eluent（component D_Ⅱ） and 95%alcohol eluent（component E_Ⅱ） of partⅠwere also obtained. Through HPLC spectral analysis and assaying of the component,flavonoid was concluded as the main content of component D_Ⅰand D_Ⅱand terpenes was the main content of component E_Ⅰand E_Ⅱ.（4） On account of the previous results,the subject researched the anti-lung cancer material basis of SanCaoFang in the next step.In vivo and in vitro experiments were taken to the different composition and compatibility of SCF.Results showed that,the component E_Ⅰ、D_Ⅱand E_Ⅱhad good anti-proliferative effect to SPC-A-1 cell in all components,their IC₅₀ were 6.35 mg crude drug/mL、7.71 mg crude drug/mL and 6.75 mg crude drug/mL respectively,the component E_Ⅰ,D_Ⅱand E_Ⅱalso had good anti-proliferative effect to A549 cell in all components,their IC₅₀ were 8.05 mg crude drug/mL,6.79 mg crude drug/mL and 6.88 mg crude drug/mL respectively;the anti-proliferative effect to SPC-A-1 and A549 cells was elevated significantly（p＜0.05） after compatibility the component D and E, IC₅₀ were 0.71 mg crude drug/mL,0.68 mg crude drug/mL respectively.C57BL/6J small black rats injected with Lewis lung cancer cell were used for in vitro experiment;anti-tumor ability and immunifaction were used as evaluation index.Results showed that the component D and E have strong anti-tumor ability in all components,tumor inhibition rate of their high dose groups were 31.89%and 51.02%respectively,and tumor inhibition rate of their low dose groups were 23.09%and 31.37% respectively;tumor inhibition rate of the compatibility of the two components were 55.08%and 34.13%in high low dose groups;the high dose groups of the component D and E and the high low dose groups of the compatibility,their tumor weight were reduced significantly（p＜0.05） with negative control group;With negative control group,thymus index of the component E was no significant difference（p＞0.05）,thymus index of the high dose group of the component D significantly increased（p＜0.05）;With negative control group,spleen index of the high dose group of the component E significantly decreased（p＜0.05）,spleen index of the component D was no significant difference（p＞0.05）;With negative control group,spleen index of the compatibility was no significant difference（p＞0.05）,thymus index of the high dose group of the compatibility was significantly increased（p＜0.05）.The compatibility of the two components had attenuate and synergetic effection.The material basis of SCF was elucidates by the results of in vivo and in vitro experiments,it had compatibility between the component D and E. The third layer（The material basis of TCM constructed of difference components,the components have compatibility relation） of the hypothesis of "three layer and multi-dimension structure" of TCM material basis was evidenced by the results.（5） Ascription was identified though chromatograph fingerprint peak, and combined with pharmacectic experimental data of each components and their compatibility,the possible existed compatibility relation of monomer elements from the components constructed by the elements of the same chemistry sort was initially prompted,and it was also the initial research of the second layer（components part constructed of elements of the same chemistry sort,the elements from the components have compatibility relation）of the hypothesis of "three layer and multi-dimension structure" of TCM material basis.