A Preliminary Study On The Analysis Of Secondary Metabolites Of Taxol - Producing Bacteria

Taxol is a tetracyclic diterpenoid compounds, which is recognized as one of broad-spectrum and competent anticancer drugs. The main source of taxol is from plant tissue extraction, chemical synthesis, yew cell culture and microbial fermentation. Currently, paclitaxel production mainly depends on the extraction in the yew bark, because of its very low content in Taxus, we should search for its substitution and precursor substances instead of looking for itself. With the increasing of its source, taxane and precursor substances are also continued to deepen. Endophytic fungi biosynthesis of paclitaxel has its own obvious advantages, so more and more domestic and foreign researchers pay attention to the its investigation and development.In my study, genome shullfing of high taxol with engineering strains HDFS4-26was acted as the original strain, we fermentated it by using bioreactors. After fermentation, with downstream engineering, fermentation products were seprated, purified and identified. The fermentation broth and mycelium were extracted by chloroform and mixed solvent of ethyl acetate:methanol (V:V=1:1), then discarded the aqueous phase and vacuumed distillation, at last we got its extraction; After that we extracted the products by using normal phase chromatography, reversed phase chromatography, semi-preparative HPLC, Sephadex chromatography and recrystallizating extraction to isolate and purify the product. In this experiment,five compounds were isolated from the fermentation broth of HDFS4-26by using1H-NMR,13C-NMR and HPLC-MS. We identificated three taxane diterpenoids, which named paclitaxel, cephalomannine, baccatinⅢ; Two non-taxane compounds, which named β-sitosterol and flavonoids of Isovitexin.Purification technologies applicating in this experiment are simple, the separation result is very obvious, and it also has a small amount of solvent using. Therefore, the result provides a useful reference for the separation and purification of intermediates in the production of paclitaxel.