α-Amylase Monitoring by a Novel Amperometric Biosensor Based on Au Electrode: Its Optimization, Characterization, and Application

Figures & data

Scheme 1. Schematic representation of enzymatic reactions in the reaction medium.

Table 1. The effects of glucose oxidase amount, gelatin amount, and glutaraldehyde percentage on the biosensor response

	R^2	Slopes (y)	Linear Ranges (U/ml)
Glucose oxidase amount (mg)^a
2.5	0.988	73.641x − 12.024	0.819–13.110
5.0	0.957	71.110x + 144.26	4.097–12.291
7.5	0.981	65.667x + 127.86	0.819–13.110
10.0	0.970	78.269x − 20.893	0.819–9.833
Gelatin amount (mg)^b
2.5	0.983	76.926x − 46.672	0.819–13.110
5.0	0.973	53.773x + 22.090	1.639–13.110
7.5	0.988	77.264x − 43.727	1.639–13.110
Glutaraldehyde percentage (%)^c
1.25	0.990	54.584x − 11.838	0.819–13.110
2.50	0.998	62.829x + 7.846	0.819–16.388
5.00	0.985	75.916 − 30.534	0.819–13.110

^aThe amount of gelatin and the percentage of glutaraldehyde were kept constant at 7.5 mg and 5.0%, respectively. Working conditions: phosphate buffer; ph 7.5, 50 mm; t = 35°c; 5 mg/ml starch solution and 65.55 u/mg α-amylase.

^bThe amount of glucose oxidase and the percentage of glutaraldehyde were kept constant at 2.5 mg and 5.0 %, respectively. Working conditions were the same as mentioned above.

^cThe amount of glucose oxidase and the amount of gelatin were kept constant at 2.5 and 7.5 mg, respectively. Working conditions were the same as mentioned above.

Figure 1. The effect of pH on the biosensor response. [Working conditions: phosphate buffers, 50 mM and pH 6.0, 6.2, 6.4, 6.5, 6.6, 6.8, 7.0. T = 35°C. 5 mg/ml starch solution and 6.555 U/ml standard solution of α-amylase was used. The amount of glucose oxidase and gelatin was kept constant at 5 mg and glutaraldehyde percentage was %2.5.]

Figure 2. The effect of temperature on the biosensor response. [Working conditions: Phosphate buffer; pH 6.5, 50 mM; 5 mg/ml starch solution and 6.555 U/ml standard solution of α-amylase was used. Amount of glucose oxidase, amount of gelatin and glutaraldehyde percentage was kept constant as 2.5 mg, 7.5 mg, and 5%, respectively.]

Figure 3. Linear range of the biosensor. [Working conditions: Phosphate buffer; pH 6.5, 50 mM; T = 35°C; 5 mg/ml starch solution and α-amylase activity between 0.819-13.110 U/ml. Amount of glucose oxidase, amount of gelatin and glutaraldehyde percentage was kept constant as 2.5 mg, 7.5 mg, and 5%, respectively.]

Table 2. Results for determinations of α-amylase activity in baker's yeast samples

Sample	Found by the biosensor* (U/ml)	Found by the reference method* (U/ml)	Relative difference (%)
Baker's Yeasts	3.241 ± 0.104	3.160 ± 0.032	2.563

*Results were given as average ± standard deviation (n = 3).