In situ product removal during enzymatic cephalexin synthesis by complexation

Abstract

In this paper, ‘complexation’ indicates the formation of clathrate type inclusion compounds of cephalexin with naphthalene derivatives. These inclusion compounds readily crystallise in solution, resulting in specific co-crystals of complexing agent and cephalexin with a set ratio between both components. Complexation is used for in situ product removal during enzymatic kinetic cephalexin synthesis to prevent undesired hydrolysis. In order to achieve this, beneficial reaction conditions have to be matched with conditions that are beneficial for complexation. In the work described here, a pH of 7.5 and a temperature of 293 K meet these requirements best. The results were compared to predictions obtained with a model originally developed for cephalexin synthesis and which is now extended with complexation. For 1,5-dihydroxy-naphthalene, the course of the reaction was predicted accurately. For 2-naphthol, this was not the case; synthesis was enhanced and hydrolysis reduced compared to the model predictions for immobilised enzyme. On the other hand, the course of reactions could be predicted accurately by the model for liquid enzyme. Apparently, the reduced reaction rate (∼30% residual activity) is such that mass transfer can keep up with it and diffusion limitation was lifted resulting in higher cephalexin concentrations. The effect of in situ complexation on productivity is discussed. It was found that complexation has a beneficial effect on overall cephalexin productivity and in most cases, hydrolysis is suppressed. The effects were most pronounced for liquid enzyme in combination with complexation with 1,5-dihydroxy-naphthalene for which, also experimentally, the highest cephalexin concentrations were measured.