Optimization of culture conditions for transformation of vitamin D3 to calcitriol by Actinomyces hyovaginalis isolate A11-2

Abstract

Background: Actinomyces hyovaginalis A11-2, a soil isolate obtained throughout a previous screening program able to transform vitamin D3 to its biologically active form, calcitriol (1α, 25-dihydroxy vitamin D3). Methods and Findings: Various culture conditions were studied to optimize the biotransformation process by the respective isolate. Product analysis was determined using TLC/HPLC coupled assay and the results revealed that the maximum calcitriol production was achieved using nutrient broth as a medium for preculture. The optimal timing of substrate (vitamin D3) addition was found to be two days after the beginning of the main culture and at concentration of 20 mg% w/v. Main culture medium components of 15 g fructose, 15 g skim milk, 1 g dipotassium hydrogen phosphate, 0.2 g sodium fluoride per liter and initial pH of 7.8 as well as continuing the main culture for 96 hrs after addition of vitamin D3 proved to be optimum for the biotransformation process. Testing vitamin D3 biotransformation by the cell lysate of the tested isolate in comparison to the intact cells revealed that 118 μg calcitriol was obtained using the lysate of cells obtained from 1 L culture in a time not exceeding 6 hrs. Conclusion: Physiological optimization of vitamin D3 biotransformation gave calcitriol production of 220 μg/L. Further genetic and biochemical studies will be conducted in the future for the characterization of the enzymes involved and to improve the vitamin D3 biotransformation process. © Copyright iMedPub.