Low Cost Sensor for E. Coli/ Colliform Detection in Water

Fabrication of Low Cost Optical Detection System For E. coli 0157 Based On

4-Methyl Umbelliferyl- ß -D-Galactoside– ß-Galactosidase Reaction.

Joseph Michael D. Racho, Erba V. Del Rosario, Karlen C. Dawal,

Dan R. Laurente, and Raymond J. Sucgang*1,

In cooperation with the

Chemists’ & Educators’ Support for the

Advancement of Relevant Science Education, Inc., CESARSED, Inc.

1R.J. Sucgang Center for Research in the Natural Sciences

Napti, Batan, Aklan

Keywords: Sensor, Optical detection, E.coli

This paper reports the fabrication of a handheld, simple to use sensor which detects contamination by deadly strains of E coli in drinking water. E. coli 0157, predominately 0157:H7, has been implicated as the causative organism of hemolytic uremic syndrome and hemorrhagic colitis. The sensor works based on irreversible color changes that occur subsequent to hydrolysis of a chromogenic substrate by a ß-galactosidase enzyme of E.coli. The recognition material, 4-methyl umbelliferyl- ß -D-galactoside has two components: a fluorescent dye and a “quencher” which is sensitive to cleavage by ß-galactosidase. In the absence of E.coli, the quencher binds to the dye and prevents fluorescence, while the interaction of ß-galactosidase with the quencher results to the dissociation of the complex ,7-hydroxy-4-methylcoumarin, giving off a blue fluorescence.

The sensor uses an LED light source shining on a photodetector. When the colorimetric reaction has taken place, light shines through a beaker containing the sample, and onto a light-sensitive meter. A clear tube of water or other clear solution is the BLANK and has zero absorbance. A fraction of the incident light that is blocked by the sample, can be used for quantitative determination of the bacterial population in the water. The E. coli population is related to the absorbance reading, following Beer’s law. The amount of incident and transmitted light is expressed in foot candle units, by a LX 104 light meter. The sensor shows outstanding response over the range of 0.8 x107 to 1.35 x108 CFUs E. coli concentration. The bacterial numbers in each test solution has been confirmed using standard plate count method. Excellent sensitivity and linearity (R2=0.984) has been achieved using the sensor, working with 6 replicate measurements per particular concentration. The specificity of the sensing system to E. coli, was tested by spiking other bacterial strains, ( Staphyloccus aureus) during the 4th to 6th determination. The sensor is highly selective and specific in its responses to E. coli. The cost of the sensor is very low in contrast to the cost of acquiring a laboratory spectrophotometer.

This work demonstrated that assembly of a sensitive, portable, low cost devices from an ordinary light meter, an LED light source, and traditional biochemical chromogenic reagents for the detection and quantization of bacterial populations is practical.

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