Highly Sensitive and Selective Detection of L-Tryptophan by ECL Using Boron-Doped Diamond Electrodes

Abstract

L-tryptophan is an amino acid that is essential to the metabolism of humans. Therefore, there is a high interest for its detection in biological fluids including blood, urine, and saliva for medical studies, but also in food products. Towards this goal, we report on a new electrochemiluminescence (ECL) method for L-tryptophan detection involving the in situ production of hydrogen peroxide at the surface of boron-doped diamond (BDD) electrodes. We demonstrate that the ECL response efficiency is directly related to H₂O₂ production at the electrode surface and propose a mechanism for the ECL emission of L-tryptophan. After optimizing the analytical conditions, we show that the ECL response to L-tryptophan is directly linear with concentration in the range of 0.005 to 1 µM. We achieved a limit of detection of 0.4 nM and limit of quantification of 1.4 nM in phosphate buffer saline (PBS, pH 7.4). Good selectivity against other indolic compounds (serotonin, 3-methylindole, tryptamine, indole) potentially found in biological fluids was observed, thus making this approach highly promising for quantifying L-tryptophan in a broad range of aqueous matrices of interest.

Keywords: L-tryptophan; boron-doped diamond; electrochemiluminescence.

Figure 1

Lower applied voltage plotted against the corresponding oxidation ECL peak emission during the oxidation of luminol in log10 scale. Three electrodes, Au (short dash line) GC (long dash line), and BDD (continuous line), were tested, starting at 0 V before then sweeping to the lower potential and finally sweeping to 1 V, at a scan rate of 0.1 V.s⁻¹ in a 0.1 M PBS solution with 1 µM of luminol (n = 3).

Figure 2

(a) CV (dashed line) and ECL (solid line) emission from 500 nM L-tryptophan in 0.1 M PBS solution. CV scan was 0 V → −3 V → 1.5 V → 0 V, at a scan rate 0.25 V.s⁻¹. (b) Resulting calibration curve for L-tryptophan obtained in the same experimental conditions (inset: ECL peak emission for each concentration of tryptophan measured (0, 5, 10, 25, 50, 100, 250, 500, 1000 nM) in 0.1 M PBS (pH 7.4) on BDD electrodes after successive reduction and oxidation).

Figure 3

(a) ECL signal intensity versus wavelength from BDD electrode and (b) ECL emission plotted against time for four electrodes (yellow: Au, red: Pt, black: GC and turquoise: BDD) recorded with the PMT. In both cases a solution containing 0.1 M PBS (pH 7.4) and 500 nM tryptophan was used; CV scan was 0 V → −3 V → 1.5 V → 0 V, at a scan rate 0.1 V.s⁻¹.

Figure 4

Proposed mechanism for the ECL of L-tryptophan at BDD electrode using successive electro-reduction and electro-oxidation.

Figure 5

ECL maximum peak emission from 10 µM of five different indolic compounds (tryptophan, tryptamine, indole, skatole (3-methylindole) and serotonin) on BDD electrodes after successive reduction and oxidation. CV scan was 0 V → −3 V → 1.5 V → 0 V, at a scan rate 0.1 V.s⁻¹.