All glassware used in media preparation and culturing was cleaned according to the USGS National Water-Quality Assessment (NAWQA) protocols to minimize contamination from trace organic and inorganic constituents [36 ].
Metformin standards (as Metformin hydrochloride) for the analysis of concentrations in growth media (0–1000 mg L-1) were prepared by dilutions of a primary Metformin stock solution (10 mg mL-1 dissolved in mineral water; Toronto Research Chemicals). Mobile phase solvents included 0.1% UPLC-grade formic acid in HPLC-grade water (Sigma-Aldrich; solvent A), UPLC-grade 0.1% formic acid in HPLC-grade acetonitrile (Sigma-Aldrich; solvent B). Aliquots of 0.2 μm-filtered water were dispensed into 1.5 mL (12 x 32 mm) amber borosilicate glass SUN-SRI standard opening auto-sampler vials (Thermo Scientific). Internal standard (IS) solutions of deuterated Metformin (Toronto Research Chemicals) were prepared from frozen stock solutions (1 mg mL-1 in Milli-Q water) diluted to yield a working solution (of 998 ng mL-1). Internal standard (10 μg L-1) was added to each sample and standard.
Metformin standards (as Metformin hydrochloride) for the analysis of concentrations in growth media (0–1000 mg L-1) were prepared by dilutions of a primary Metformin stock solution (10 mg mL-1 dissolved in mineral water; Toronto Research Chemicals). Mobile phase solvents included 0.1% UPLC-grade formic acid in HPLC-grade water (Sigma-Aldrich; solvent A), UPLC-grade 0.1% formic acid in HPLC-grade acetonitrile (Sigma-Aldrich; solvent B). Aliquots of 0.2 μm-filtered water were dispensed into 1.5 mL (12 x 32 mm) amber borosilicate glass SUN-SRI standard opening auto-sampler vials (Thermo Scientific). Internal standard (IS) solutions of deuterated Metformin (Toronto Research Chemicals) were prepared from frozen stock solutions (1 mg mL-1 in Milli-Q water) diluted to yield a working solution (of 998 ng mL-1). Internal standard (10 μg L-1) was added to each sample and standard.