β-galactosidase Determination by an Electrochemical Biosensor Mediated with Ferrocene

Figures & data

Figure 1. The effect of mediator, ferrocene concentration on the biosensor performance [Ferrocene concentrations used: -•-•-: 1 mM, -▪-▪- : 2.5 mM, -▴-▴-: 5 mM, -♦-♦-: 7.5 mM. Working conditions: The amount of glucose oxidase immobilized on the electrode was 45 U, 50 mM citrate buffer (containing 100 mM lactose, and of course containing ferrocene as indicated above, pH 4.8), T = 35 °C. Chronoamperometric medium: at a constant potential: 250 mV, t.puls:40 ms, t.meas:20 ms].

Figure 2. The effect of lactose concentration on the biosensor performance [Lactose concentration levels added to the working buffer:-♦-♦- :10 mM, -▪-▪-:25 mM -▴-▴-: 50 mM, GGG GGGGGGGGGGGGGG: 75 mM, -•-•-:100 mM. Working conditions: The amount of glucose oxidase immobilized on the electrode was 45 U, 50 mM citrate buffer (containing 1 mM ferrocene, and of course containing lactose as indicated above, pH 4.8), T = 35 °C. Chronoamperometric medium: at a constant potential:250 mV, t.puls:40 ms, t.meas:20 ms].

Figure 3. The effect of temperature on the biosensor performance. [The amount of glucose oxidase immobilized on the electrode was 45 U. β-galactosidase standard used in the experiments: 0.188 U/mL. Working conditions: 50 mM citrate buffer (containing 100 mM lactose, 1 mM ferrocene, pH 4.8). Chronoamperometric medium: at a constant potential: 250 mV, t.puls:40 ms, t.meas:20 ms.]

Figure 4. The effect of pH and buffer concentration on the biosensor performance. [For both parameters the amount of glucose oxidase immobilized on the electrode and β-galactosidase standard used in the experiments were same as 45 U and 0.188 U/mL, respectively. For optimum pH studies; Working conditions: 50 mM citrate buffers with differing pHs (containing 100 mM lactose, 1 mM ferrocene), T = 35 °C. For investigation of buffer concentration (inner graph); Working conditions: citrate buffers with differing concentrations (containing 100 mM lactose, 1 mM ferrocene), T = 35 °C. Chronoamperometric medium was same for two tests as follows; a constant potential: 250 mV, t.puls:40 ms, t.meas:20 ms.]

Figure 5. Calibration graph for β-galactosidase activity. [The amount of glucose oxidase immobilized on the electrode was 45 U. Working conditions: 50 mM citrate buffer (containing 100 mM lactose, 1 mM ferrocene, pH 4.8), T = 35 °C. Chrono-amperometric medium: at a constant potential: 250 mV, t.puls: 40 ms, t.meas:20 ms.]

Table 1. Results obtained from the reproducibility studies of the biosensor based on glucoseoxidase.

Biosensor	Linear range (μM)	R^2	y
1	4,7×10^−3–3,29×10^−1	0.9885	3620.2×−131.87
2	4,7×10^−3–3,29×10^−1	0.9933	3549.2×−151.65
3	4,7×10^−3–3,29×10^−1	0.9894	3629×−131.9

Table 2. β-galactosidase activity determination in artificial intestinal juice by the biosensor and by a spectrophotometric reference method.

β-galactosidase (U/ml) Added	Found by the biosensor	Found by the reference method	% Recovery	% Relative difference
0.188	0.18	0.195	96	8