A modified and sensitive spectrophotometric coupling diazotization method for the analysis of Organophosphate insecticides i. e., Chlorpyrifos, in sub-microgram levels is described. During alkaline hydrolysis of Chlorpyrifos to produce 1, 2, 4-trichloropyridine (TCP) in alkaline medium, which coupled with diazotized p-aminoacetophenone to form orange-red color dye at measure 505 nm. The color system obeys Beer’s law in the following working range of 1.2 to 10.5 µg (0.048 -0.42 ppm) in a final solution of 25 ml. The Molar Absorptivity, Sandell’s Sensitivity, Correlation coefficient, Standard deviation and Relative standard deviation have been determined The method is highly reproducible and have been successfully applied for determination of Chlorpyrifos insecticides in environmental and biological samples.