A novel and simple methodology for co-obtaining of enantiomerically pure alpha-hydroxyacids and alpha-ketoacids was developed by enantioselective oxidation of alpha-hydroxyacids bearing a substituent with an aryl group using alpha-hydroxyacid dehydrogenase (alpha-HADH). A high-throughput method was firstly established for screening of enantioselective alpha-HADHs. Sinorhizobium sp. ZJB1101 with high activity and excellent enantioselectivity of alpha-HADH for oxidation of alpha-hydroxyacids bearing a substituent with an aryl group was isolated and identified. This strain has potential for co-production of (R)-alpha-hydroxyacids and alpha-ketoacids in near theoretical yields, while no consecutive oxidation of alpha-ketoacids was observed. The green conversion system appears promising for potential applications in industry. (C) 2012 Elsevier Ltd. All rights reserved.