title: Preparation of (R)- and (S)-N-protected 3-hydroxypyrrolidines by hydroxylation with Sphingomonas sp. HXN-200, a highly active, regio- and stereoselective, and easy to handle biocatalyst.
abstract: Hydroxylation of N-benzylpyrrolidine 8 with resting cells of Sphingomonas sp. HXN-200 gave N-benzyl-3-hydroxypyrrolidine 15 in 53% ee (S) with an activity of 5.8 U/g CDW. By changing the "docking/protecting group" in pyrrolidines, hydroxylation activity and enantioselectivity were further improved and the enantiocomplementary formation of 3-hydroxypyrrolidines was achieved: hydroxylation of N-benzoyl-, N-benzyloxycarbonyl-, N-phenoxycarbonyl-, and N-tert-butoxycarbonyl-pyrrolidines 9-12 gave the corresponding 3-hydroxypyrrolidines 16-19 in ee of 52% (R), 75% (R), 39% (S), and 23% (R), respectively, with an activity of 2.2, 16, 14, and 24 U/g CDW, respectively. Simple crystallizations increased the ee of 16-18 to 95% (R), 98% (R), and 96% (S), respectively. Hydroxylation of pyrrolidines 8-12 with soluble cell-free extracts of Sphingomonas sp. HXN-200 and equimolar NADH gave 3-hydroxypyrrolidines 15-19 in nearly the same ee as the products generated by whole cell transformation, suggesting that this strain possesses a novel soluble alkane monooxygenase. Cells of Sphingomonas sp. HXN-200 were produced in large amounts and could be stored at -80 degrees C for 2 years without significant loss of activity. The frozen cells can be thawed and resuspended for biohydroxylation, providing a highly active and easy to handle biocatalyst for the regio- and stereoselective hydroxylation of nonactivated carbon atoms. These cells were used to prepare 1.0-3.2 g (66.4-93.5% yield) of 3-hydroxypyrrolidines 16-19 by hydroxylation of pyrrolidines 9-12 on 0.9-2 L scale. Preparative hydroxylation was also achieved with growing cells as biocatalysts; hydroxylation of pyrrolidine 11 on 1 L scale gave 1.970 g (79.7% yield) of 3-hydroxypyrrolidine 18.
authors.affiliation: Institute of Biotechnology, ETH Zurich, Hoenggerberg, CH-8093 Zurich, Switzerland. zhi@biotech.biol.ethz.ch
authors.forename: Z
authors.initials: Z
authors.lastname: Li
authors.forename: H J
authors.initials: HJ
authors.lastname: Feiten
authors.forename: D
authors.initials: D
authors.lastname: Chang
authors.forename: W A
authors.initials: WA
authors.lastname: Duetz
authors.forename: J B
authors.initials: JB
authors.lastname: van Beilen
authors.forename: B
authors.initials: B
authors.lastname: Witholt
mesh_terms: D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D006900:Hydroxylation; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D011760:Pyrrolidinones / Q000737:chemistry*; D013055:Spectrophotometry, Infrared; D020578:Sphingomonas / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D013237:Stereoisomerism
chemical_list: D011760:Pyrrolidinones