A major concern in San Joaquin Valley, CA, is that high levels of Se in agricultural drainage water are concentrating to hazardous levels in evaporation ponds. The primary objective of this study was to determine factors that affect Se biomethylation and thus permanently remove Se directly from evaporation pond water. Pond water samples (14-2000 μg Se L-1) were set up as laboratory mesocosms. The natural formation of dimethylselenide (DMSe) in unamended water was less than 1% of the total Se inventory after 40 d of incubation. No Se methylation took place in autoclaved unamended pond water. L-Methionine (10 μM) stimulated DMSe production in nonsterile pond water, and in autoclaved water (1 μM). Increasing the temperature to 35°C and the addition of 1% (wt/wt) glucose with a fungal inoculum, Alternaria alternata, doubled DMSe production over the controls after 25 d of incubation. Carbon sources such as glucose, maltose, sucrose, and galacturonic acid at 2 g C L-1 under ambient conditions slightly enhanced indigenous Se methylation (1.5-fold). Of the amino acids tested, L-methionine (0.02 g C L-1) stimulated DMSe evolution from pond water more so than L-cysteine, L-cystine, and L-serine. The proteins, egg albumen, casein, and gluten (2 g C L-1) dramatically increased Se biomethylation causing a 23, 41, and 10% Se loss from the inventory, respectively, after 43 d of incubation. The stimulation of Se volatilization from evaporation pond water through specific amendments could prove to be promising as a field detoxification technique.