Development of an enzymatic biosensor to determine eugenol in dental samples.

A GCE/CRGO-ßCD's/ADA-SPE/AuNPs biosensor was successfully developed to determine eugenol in dental samples. The optimal conditions to construct the biosensor were obtained from an experimental design based on the response surfaces methodology. The GCE/CRGO-ßCD/ADA-SPE/AuNPs biosensor exhibited a very good analytical performance for the quantification of eugenol. Thus, it shows a linear range between 1.3 × 10-8 and 1 × 10-5 mol L-1, with a sensitivity of (5.3 ± 0.3) x 10-3 A mol-1 L. The limits of detection and quantification were 4 × 10-9 mol L-1 and 1.3 × 10-8 mol L-1, respectively. Biosensors had an intraday and inter day reproducibility of 5% and 8%, respectively. The repeatability was of 3%, and the stability was 21 days (a decrease of 30% in current responses was observed after the fourth week). Recovery studies were performed in order to validate the proposed method. Recovery percentages were between 94 and 108%. A value of the apparent Michaellis-Menten constant, KMapp, of 3.1 × 10-6 mol L-1 was obtained using both Lineweaver-Burk and Eadi-Hofstee methods.

Determination of kinetic parameters of the enzymatic reaction between soybean peroxidase and natural antioxidants using chemometric tools.

The oxidation of eugenol, isoeugenol and vanillin natural antioxidants catalyzed by the soybean peroxidase enzyme was studied using uv-vis spectroscopy. An experimental design was used to optimize the different variables. The multivariate curve resolution method was used to obtain the profiles of antioxidant absorbance's as a function of time due to uv-vis absorption bands of both antioxidants and the enzymatic reaction product/s show a strong overlap. From these results, apparent Michaelis-Menten constants as well as the kinetic parameters k1 and k3 involved in the catalytic cycle of peroxidases were calculated. The antioxidant apparent acidity constants were also determined at different pH's from uv-vis spectrophotometric measurements. Values of k1 were (0.6 ±â€¯0.1) × 105 M-1 s-1, (2.0 ±â€¯0.2) × 105 M-1 s-1 and (7.0 ±â€¯0.5) × 106 M-1 s-1 and k3 (4.0 ±â€¯0.2) × 105 M-1 s-1, (6.0 ±â€¯0.6) × 105 M-1 s-1 and (6.0 ±â€¯0.9) × 106 M-1 s-1 for eugenol, isoeugenol and vanillin, respectively.