The synthesis of 4-deoxy-4-fluoro- d -[U- 14 C]glucose from d -[U- 14 C]galactose is reported. A 24-h incubation of Pseudomonas putida with 4-deoxy-4-fluoro- d -[U- 14 C]glucose gives 95 ± 5% release of fluoride and 4.8 ± 0.2% of the initial radioactivity as 14 CO 2 . After centrifugation, Dowex-1 [borate 2 − ] column chromatography of the cell supernatant, which amounts for 52.4 ± 1.3% of the initial radioactivity, allows the isolation of a major radioactive metabolite. By 13 C- and 1 H-n.m.r. spectroscopy and by mass spectrometric analysis, this metabolite is identified as 2,3-dideoxy- d - glycero -pentonic acid. Extensive dialysis of the remaining cell pellet, followed by sonication and appropriate centrifugation, allows isolation of a cell envelope fraction with 0.4 ± 0.05% of the initial radioactivity. Gel filtration of this sodium dodecylsulphate solubilised fraction shows all the radioactivity to be in a large molecular weight peptidoglycan-protein complex (>400 000 daltons). Following lysozyme treatment, this complex now elutes from the same column with a lower molecular weight (