In an effort to enhance the sensitivity, the colorimetric assays of glucose employ bienzyme reaction. In the presence of oxygen, the GOD oxidizes the glucose to gluconic acid and H2O2. The H2O2 is then utilized to oxidize a chromogenic substrate in a secondary
reaction with HRP. As shown in Fig. 3a, the resulting color changed from almost colorless to red for glucose detection and the calibration curves were obtained. The relative intensity of color was proportional to the concentration of glucose within the linear range, and the mean intensity was shown in the calibration curves in Fig. 3b. The equation of curve was y = 8.28 x + 92.28, R2 = 0.997.Under the optimized conditions, the color intensity is linear with
the concentration over the range of 1.0 × 10−3 to 11.0 × 10−3M with a limit of detection (LOD) of 3×10−4M for glucose. The LOD is given by 3S0/S, where 3 is the factor at the 99% confidence level, S0 is the standard deviation of the blank measurements (n = 10), and S is the slope of the calibration curve.