Integrated L-phenylalanine separation in an E. coli fed-batch process: from laboratory to pilot scale.

Pilot-scale reactive-extraction technology for fully integrated L-phenylalanine (L-Phe) separation in Escherichia coli fed-batch fermentations was investigated in order to prevent an inhibition of microbial L-Phe production by-product accumulation. An optimal reactive-extraction system, consisting of an organic kerosene phase with the cation-selective carrier DEHPA (di-2-ethylhexyl phosphonic acid) and an aqueous stripping phase including sulphuric acid, was found particularly efficient. Using this system with two membrane contactors, mass-transfer coefficients of up to 288 x 10(-7) cm s(-1) for the aqueous/organic and 77 x 10(-7) cm s(-1) for the organic/stripping phase were derived from experimental data using a simple modelling approach. Concentration factors higher than 4 were achieved in the stripping phase as compared to the aqueous donor phase. Reactive extraction enabled a 98% cation portion of L-Phe in the stripping phase, leading to final product purity higher than 99% after L-Phe precipitation. A doubling of L-Phe/glucose yield was observed when kerosene/DEHPA was added to the fermentation solution in the bioreactor to experimentally simulate a fully integrated L-Phe separation process.

Enhanced pilot-scale fed-batch L-phenylalanine production with recombinant Escherichia coli by fully integrated reactive extraction.

A fully integrated process for the microbial production and recovery of the aromatic amino acid L-phenylalanine is presented. Using a recombinant L-tyrosine (L-Tyr) auxotrophic Escherichia coli production strain, a fed-batch fermentation process was developed in a 20-l-scale bioreactor. Concentrations of glucose and L-Tyr were closed-loop-controlled in a fed-batch process. After achieving final L-phenylalanine (L-Phe) titres >30 g/l the process strategy was scaled up to 300-l pilot scale. In technical scale fermentation L-phenylalanine was continuously recovered via a fully integrated reactive extraction system achieving a maximum extraction rate of 110 g/h (final purity >99%). It was thus possible to increase L-Phe/glucose selectivity from 15 mol% without to 20.3 mol% with integrated product separation.