NADPH-mediated seedless in situ formation of gold or gold-platinum nanoparticles for the enzymatic determination of atropine

Abstract

The without-seeds in situ formation of gold nanoparticles from NADPH and its application to the colorimetric determination of atropine (a tropane alkaloid) in cereals is reported. The method is based on a chemical hydrolysis, followed by an enzymatic oxidation by NADP catalyzed by tropinone reductase in the presence of Au(III) or Au(III)/Pt(II). During this reaction, the formed NADPH reduces the metal ion precursor to AuNPs (or AuPtNPs) and the absorption due to the plasmon band (550 nm or 575 nm) is measured. The method (AuPtNPs) allows the determination of the analyte in the concentration range 0.025 to 0.09 mM with an RSD of 3% (n = 5) and is applied to its determination in spiked buckwheat samples using the standard addition method, with 96.9 ± 2.0% recovery. It is also demonstrated that NAD(P)H and NADH have different kinetics for AuNP generation, which could be used to discriminate between these two cofactors.

Keywords: Gold nanoparticles; Gold-platinum nanoparticles; NADH; Tropane alkaloid; Tropinone reductase.

Fig. 1

Scheme of the reactions studied in this work

Fig. 2

A) Molecular absorption spectra of AuNPs and AuPtNPs synthetised in the presence of NADPH and Au/III) or Au(III)/Pt(II), respectively. B) Variation with time at Abs_540nm of the corresponding nanoparticles. In both cases, a concentration of NADPH of 1 × 10^–4 M was used

Fig. 3

A) TEM image of the AuNPs. B) TEM image of the AuPtNPs

Fig. 4

Calibration curve for the reaction using the following concentrations of metals: [Au(III)] = 1 × 10^–4 M; Pt(II) = 4 × 10^–4 M. A) Abs = f([NADH] representation; B) fitting to Eq. (4)

Fig. 5

Comparison of the kinetics of the reaction between NADPH and NADH (1,5 × 10^–4 M for both compounds in A; 1.5 × 10^–5 M for both compounds in B). A) [Au(III)] = 1 × 10^–4 M.; B) [Au(III)] = 1 × 10^–4 M. [Pt(II)] = 4 × 10^–4 M

Fig. 6

Calibration lines obtained for atropine determination with TRase, Au(III) and Pt(II). A) Calibration line; B) logistic curve for the reaction using the following concentrations of metals: [Au(III)] = 4 × 10^–4 M; Pt(II) = 1 × 6·10^–3 M