In-a-Day Electrochemical Detection of Coliforms in Drinking Water Using a Tyrosinase Composite Biosensor

摘要

A rapid method for the detection of fecal contamination in water based on the use of a tyrosinase composite biosensor for improved amperometric detection of beta-galactosidase activity is reported. The method relies on the detection of phenol released after the hydrolysis of phenyl beta-D-galactopyranoside (PG) by beta-galactosidase. Under the optimized PG concentration and pH (4.0) values, a detection limit of 1.2 X 10~(-3) unit of beta-galactosidase/mL~(-1) was obtained. The capability of the sensor for the detection of Escherichia coli was evaluated using polymyxin B sulfate to allow permeabilization of the bacteria membrane. A detection limit of 1 X 10~(6) cfu of E. coli mL~(-1) was obtained with no preconcentration or pre-enrichment steps. To improve the analytical characteristics for bacteria detection, the processes involving galactosidase induction during incubation and membrane permeabilization were optimized. Using 0.25 mM isopropyl beta-D-thiogalactopyranoside for the enzyme activity induction, and 10 (mu)g mL~(-1) polymyxin B sulfate as permeabilizer agent, it was possible to detect bacteria concentrations as low as 10 cfu mL~(-1) after 5 h of enrichment. The possibility of detecting E. coli at the required levels for drinking water quality assessment (1 cfu/100 mL) is demonstrated, the time of analysis being shorter than 6.5 h and involving a simple methodology.