Using The Uncertainty Derivation And Enrichment Of Minor Components To Separate And Identify The Potential New Active Ingredients In Isoniazid

Objectives: Develop a new active ingredients by screening,separating and purifying technology through uncertainty derivation methods,applying thin layer chromatography enrichment and high performance liquid chromatography(HPLC)to find potential new active ingredients from isoniazid(INH).Methods: INH was derived by employing the uncertainty derivation method,and enriching the active substances in new ingredients.Isoniazid were formulated into 10% methanol solution.The derivatives were sampled application and derived,then expanded in one dimension.According to the different partition coefficients,the derivatives were separated and different color bands were generated on thin layer plate.Then the developed thin layer plate were turned in 90 angle and developed a second dimension by focusing the line-shaped bands into spots on edge of the thin layer plate.In this study,thin layer plate control,INH control and INH derivative were set up.Bacteriostatic test of H37 Rv was done on the culture media,which were put in 37?incubator for 4 weeks.And then active INH derivatives were screened in order to be separated and purified by HPLC,to be identified whether or not it is a new anti-tuberculosis active ingredient.Bacteriostatic TestResults: In this experiment,we obtained 12 bands of INH derivative by using uncertainty derivation method and thin layer chromatography enrichment.Bacteriostatic test of MTB demonstrated that there were no MTB found in the band 5 and band 6 of INH derivatives,or INH control band.While there were MTB found in the other INH derivative band and thin layer plate control band.This suggests that thin layer plate control band had no effect on our study and INH control had anti-tuberculosis effect.This also suggests that band 5 and band 6 of INH derivatives have anti-tuberculosis active ingredients to inhibit the growth of H37 Rv.We separated and purified the active ingredients by using HPLC,and proved the active ingredients were potential new,which were different from the existing isoniazid.Further purification of band 5 showed that the band was unstable to purify and easy to break down.The improvement of purification methods are under progress and final structural identification is proposed.Conclusions: Through using the uncertainty derivation method,thin layer chromatography enrichment and HPLC,the potentially new active components of INH were screened,separated and purified,which laid the foundation for further structural analysis and ingredient identification.