Optimisation of conditions for the resolution of 1,2-epoxyoctane in a bioreactor

Abstract

Due to recent legislation requiring the determining of the pharmacokinetic effect of both enantiomers separately, of any new racemic drug before commercialisation, much research is done to improve and optimise methods for obtaining chirally pure compounds important for the pharmaceutical industry, for example epoxide precursors. To date most experiments regarding the biocatalytic chiral separation of 1,2-epoxyoctane has been done in batch processes. The aim of this study was to optimise the enantioselective hydrolysis of 1,2-epoxyoctane by Rhodosporidiurn tondoides in both a batch and continuous process. The batch process was optimised in terms of stir speed, biomass (cell) concentration and reaction time, while the flow-through reactor (continuous process) was optimised with regards to the flow rate as a function of the pressure and the amount of chitosan and biomass in the reactor. Initial inconsistencies of epoxide concentrations in preliminary studies were found to be due to adsorption by reaction and sampling vessels, and the lower than expected solubility of 1,2-epoxyoctane (3.85 mM instead of 6 mM as reported by previous investigators). The results from the batch process suggest that as the reaction time increases, the % ee-epox increases initially, but decreases after 40 minutes. Optimum yield in terms of % ee-epox were obtained at medium stir speed (400 rpm) and biomass (cell) concentration (13 %). Below these values the % ee-epox increases with an increase in stir speed and/or biomass concentration. Above these values however, the increased stir speed and/or biomass concentration causes abrasion between cells, which negatively affects the % ee-epox. The % ee-diol reached a steady state after 10 minutes, and the effect of the different operating conditions on % ee-diol was negligible. In the flow-through reactor chitosan was used as a spacer material (antifouling agent) to help decrease the fouling due to biomass deposition. The use of chitosan as a spacer ensured higher and stabilised flow rates for extended periods of time. In initial studies 0.5 g chitosan increased the flow rate by 34 % with a resistance removal of 25 %. For 1 g chitosan these values were 130 % flow increase and 57 % resistance removal. The flow rate was optimised in relation to the chitosan amount, biomass (cell) amount and pressure. The maximum flow rate was obtained at a pressure of 40 kPa, using the minimum amount of cells (0.4 g) and a maximum amount of chitosan (1.6 g)