Evaluating a spectroscopic assay for detection of aquatic one-carbon substrates methanol and formaldehyde

A sensitive assay consisting of enzymatic oxidation of methanol to a formaldehyde intermediate, followed by formation of a spectroscopically active chromophore, 3,5-diacetyl-1,4-dihydrolutidine (DDL), was evaluated for detection of methanol and formaldehyde in environmental and bacterial culture samples. It was hypothesized DDL could be detected at low concentrations via absorbance or fluorescence spectroscopy and subsequently used to quantify formaldehyde in aquatic media in the environment and in culture. The sensitivity of the chromophore synthesis was tested in both absorbance and fluorescence spectroscopy; ultimately, the limit of detection for the fluorescent method was superior to the absorbance-based method. The efficacy of the enzymatic reaction was then tested to determine the limits of detection, sensitivity, and linear range. Ultimately, quenching effects at low concentrations, relatively low initial reaction velocities for the enzymatic oxidation, and potential reaction reversibility merit further investigation of the fluorometric method and necessitate improvement of the method. Finally, formaldehyde quantification was evaluated in both culture and environmental samples, demonstrating success in the former and inefficiency in the latter. Future directions of this project will involve refinement of the fluorometric quantification and examining formaldehyde concentrations at a more resolved scale in the growth phase of methylotroph culture.