Rapid serial diluting biomicrofluidic provides EC50 in minutes

Abstract

Complex water effluents pose a toxicity risk to biological wastewater treatments and environmental discharge. Dynamic sampling and risk assessment of effluents would mitigate downstream hazards, but few methods are available to assess microorganism toxicity on the minute timescale. To rapidly evaluate unknown aqueous effluents, a segmented-flow microfluidic device is refined for real-time gross toxicity detection and quantification. A microfluidic chip integrating a magnetic stirrer for serial dilution is demonstrated to rapidly determine EC50 values of known and unknown toxicants, where 100 nL samples are suspended into a continuous oil phase with no interfering surfactants. A five logarithmic dilution sequence is evaluated in zinc > copper > nickel cations. Resazurin concentration of 10 μM is optimal for Enterococcus faecalis at 0.1 optical density. The adaptable method is transferable to other microorganisms, such as common baker's yeast, Saccharomyces cerevisiae, where the resazurin reduction rate is 30% of Enterococcus faecalis (4 nM/s vs. 13 nM/s per 100 nL droplet). Zinc and nickel cations are observed to increase the base resazurin reduction rate of baker's yeast by 25%, whereas copper is found to be more cytotoxic than mercury cations. Keywords: Microfluidic, Droplet, EC50, Heavy metals, Enzyme kinetics, Resazurin