Colorimetric Fluoride Screen Reveals Defluorination Capacity in Growing Microbial Cultures

Colorimetric measurements of inorganic fluoride (F–) offer a rapid screening of biodefluorination, but existing methods fall short when they are applied to growing cultures. By systematically replacing mineral salt medium components, we demonstrate that a modified HEPES/Cl–-rich medium allows colorimetric F– quantification (20–200 μM) during biodefluorination of 1-fluorodecane (FD), 4,5,5-trifluoropent-4-enoic acid (TFEA), and 4,4,4-trifluoro-3-(trifluoromethyl) crotonic acid (SFCA). Applied in a deep 96-well microtiter plate format, this approach explored the biodefluorination of these compounds in growing axenic cultures, resting cell incubations, and mixed environmental cultures. Sphingopyxis sp. strain NJF-3 and Rhodococcus sp. strain NJF-7 exhibited differential defluorination of TFEA and SFCA. Resting cells exhibited limited defluorination activity to TFEA and SFCA, but strain NJF-7 displayed higher TFEA defluorination (219 μM F–) than strain NJF-3 (39.1 μM F–), indicating a greater growth-independent defluorination capacity. Defluorination by environmentally sourced microbial communities varied, potentially reflecting differences in sample origin (water vs soil). Correlation analysis of intermediates measured in different SFCA-defluorinating cultures indicated that abiotic and biotic reactions contribute to SFCA transformation, with a β-oxidation-type pathway inferred for the biotransformation. The refined approach expands the tool box for investigating biodefluorination in microbial enrichment and pure culture studies.