Stereo-selective conversion of mandelonitrile to (R)-(−)-mandelic acid using immobilized cells of recombinant Escherichia coli

Abstract

Immobilized cells of a recombinant Escherichia coli expressing nitrilase from Pseudomonas putida were used to catalyze the hydrolysis of mandelonitrile (2-hydroxy-2-phenylacetonitrile) to ( R )-(−)-mandelic acid. The cells had been immobilized by entrapment in an alginate matrix. Conditions for the hydrolysis reaction were optimized in shake flasks and in a packed bed reactor. In shake flasks the best conditions for the reaction were a temperature of 40 °C, pH 8, biocatalyst bead diameter of 4.3 mm, sodium alginate concentration in the gel matrix of 2 % (w/v, g/100 mL), a cell dry mass concentration in the bead matrix of 20 mg/mL, an initial substrate concentration of 50 mM and a reaction time of 60 min. Under these conditions, the conversion of mandelonitrile was nearly 95 %. In the packed bed reactor, a feed flow rate of 20 mL/h at a substrate concentration of 200 mM proved to be the best at 40 °C, pH 8, using 4.3 mm beads (2 % w/v sodium alginate in the gel matrix, 20 mg dry cell concentration per mL of gel matrix). This feed flow rate corresponded to a residence time of 0.975 h in the packed bed.

Characterization of a Nitrilase and a Nitrile Hydratase from Pseudomonas sp. Strain UW4 That Converts Indole-3-Acetonitrile to Indole-3-Acetic Acid

“…2 and 3). These data agree well with other reported pseudomonad nitrilases, which have temperature optima from 40 to 55°C (27,28,(33)(34)(35). The UW4 enzyme retained Ͼ50% activity at 55°C; however, Ͻ20% activity was retained at 60°C.…”

“…The UW4 enzyme retained Ͼ50% activity at 55°C; however, Ͻ20% activity was retained at 60°C. The pH optima of the UW4 nitrilase is slightly lower than that of other pseudomonad nitrilases, which display maximum activity at pHs between 7 and 9 (27,28,33,34). Rather, it is similar to that of R. rhodochrous K22 (pH 5.5) and Arthrobacter nitroguajacolicus ZJUTB06-99 (pH 6.5) (36,37).…”

Kinetics and mechanism of regioselective monoacetylation of 3-aryloxy-1,2-propandiols using immobilized Candida antarctica lipase

“…The prevalent application of biocatalysis in non-aqueous 10 media has included stereo-selective synthesis of enantiopure 11 drugs over the past two decades [1][2][3][4]. The production of 12 enantiomerically pure compounds has gained significant impor- 13 tance in chemical and pharmaceutical industry.…”

Synthesis of (R)-(-)-mandelic Acid by Hydrolysis of (R)-(-)-mandelic Acid Ethyl Ester by Bacillus Cereus CGMCC No.12336

“…, cephalosporin [2], antineoplastic drugs [3] and (+)-crassalactone A [4]. Therefore, (R)-(-)-mandelic acid become an important chiral chemical intermediate [5][6].…”