Microbial Transformation Of Curcumin To Its Derivatives

Curcumin, a poly-phenolic compound, has shown a wide range of pharmacological activities. However, the clinical use of curcumin is limited to some extent because of its poor water solubility and low bioavailability. To overcome these problems, many approaches have been attempted and structural modification of curcumin by microbial transformation has been proven to be an alternative. Microbial cells can produce a variety of different enzymes, which can catalyze substrate to generate a series of derivatives and lay the foundation for finding new curcumin derivatives. In this paper, a strain capable of transforming curcumin to its derivatives was isolated from a soil sample and subsequently identified. The separation and purification of transformed products, and the optimization of fermentation medium and biotransformation conditions were also investigated. Firstly, a novel yeast strain ZJPH0802, capable of transforming curcumin to two major products of Ⅰ and Ⅱ, was isolated from a soil sample. The isolate was identified as Pichia kudriavzevii ZJPH0802according to its physiological and biochemical characteristics and26S rDNA D1/D2region sequence analysis as well. TLC analysis showed that product Ⅰ had a Rf value of0.424and product Ⅱ had a Rf value of0.712. HPLC analysis showed that the retention times of above two compounds were26min and62min, respectively.Subsequently, gel column chromatography and preparative thin layer chromatography were carried out for the separation and purification of transformed products by Pichia kudriavzevii ZJPH0802. Through above methods we get0.005g of product Ⅰ and0.008g of product Ⅱ, respectively. The purity of both above products could reach above95%by HPLC analysis. UV analysis showed that the maximum absorption wavelength of product Ⅰ and Ⅱ were the same of280nm. LC-MS analysis showed that the product Ⅰ had a molecular weight of374and372for product Ⅱ. According to the fragments ions information, product Ⅰ was tentatively identified as hexahydrocurcumin and product Ⅱ was tetrahydrocurcumin.Then, the optimization of fermentation medium, in order to increase the yield of hexahydrocurcumin, was carried out by single factor experiments. The optimal fermentation medium was determined as follows (g/L):glycerol50, peptone5, KH2PO40.2, K2HPO40.2, MgSO4·7H2O0.4. The biotransformation process was then optimized by investigating some key reaction parameters including curcumin concentration, cell concentration, buffer pH, and reaction time. Under the optimal conditions (curcumin concentration50mg/L, wet cells50g/L, pH6.5, reaction for40h), the yield of hexahydrocurcumin was47.89%, corresponding to a4-fold of original medium.Finally, the optimization of fermentation medium, in order to increase the yield of tetrahydrocurcumin, was also carried out by orthogonal experiment. The optimal fermentation medium was determined as follows (g/L):glucose25, yeast extract2.5, NH4C16.5, KH2PO41, K2HPO41, MgSO4·7H2O0.5. The biotransformation process was subsequently optimized by investigating some key reaction parameters including the solvent, substrate concentration, cell concentration, buffer pH, and reaction time. Under the optimal conditions (dehydrated alcohol as substrate solvent, curcumin concentration50mg/L, wet cells80g/L, pH6.5, reaction for24h), the yield of tetrahydrocurcumin was77.43%, an increase of1.2-fold compared with original medium.