Glycolic Acid → Glyoxylic Acid

Participants

Reaction Steps

Demonstration of the enzymatic synthesis of glyoxylic acid on a large scale was performed in an Amicon Model 2000 High-Output Stirred Cell, where a 1.6 mm thick Teflon® sheet was substituted for the filtration membrane, and which had a magnetically driven paddle stirrer. Glycolate oxidase (2000 IU, isolated from spinach) was added to 2.0 L of solution containing glycolic acid (113 g, 1.49 moles), ethylenediamine (95 g, 1.58 moles), FMN (9.7 mg, 0.02 mmoles), and catalase [2.8×106IU, from Aspergillus niger (Sigma)], and the resulting mixture (final pH =8.9) was stirred at 15° C. under 6 atmospheres of oxygen. Aliquots were removed at regular intervals for analysis by HPLC to monitor the progress of the reaction. After 77 hours, HPLC analysis of the reaction mixture indicated that glyoxylic acid (110 g, 99.6% yield), formic acid (0.2%), and oxalic acid (0.2% yield) were the only reaction products; complete conversion of glycolic acid was attained. Glycolate oxidase and catalase activities were 74% and 87% of their initial activities, respectively The reaction was terminated by sparging the solution with nitrogen, then heating the reaction mixture to 70° C. for 5 minutes under a nitrogen blanket. Precipitated protein was removed by centrifugation, and FMN removed by filtration of the reaction mixture through activated carbon. Any remaining soluble protein was removed by filtration using a Millipore Minitan Filtration System with a 10,000 mw cutoff filter, then glyoxylate and ethylenediamine (EDA) were separated by ion exchange chromatography.

Provenance & Evidence