New spin coated multilayer lactate biosensor for acidosis monitoring in continuous flow assisted with an electrochemical pH probe

Abstract

A LOx-based electrochemical biosensor for high-level lactate determination was developed. For the construction of the biosensor, chitosan and Nafion layers were integrated by using a spin coating procedure, leading to less porous surfaces in comparison with those recorded after a drop casting procedure. The analytical performance of the resulting biosensor for lactate determination was evaluated in batch and flow regime, displaying satisfactory results in both modes ranging from 0.5 to 20 mM concentration range for assessing the lactic acidosis. Finally, the lactate levels in raw serum samples were estimated using the biosensor developed and verified with a blood gas analyzer. Based on these results, the biosensor developed is promising for its use in healthcare environment, after its proper miniaturization. A pH probe based on common polyaniline-based electrochemical sensor was also developed to assist the biosensor for the lactic acidosis monitoring, leading to excellent results in stock solutions ranging from 6.0 to 8.0 mM and raw plasma samples. The results were confirmed by using two different approaches, blood gas analyzer and pH-meter. Consequently, the lactic acidosis monitoring could be achieved in continuous flow regime using both (bio)sensors.

Keywords: Continuous monitoring; Diffusion limiting membrane; Healthcare assessment; Lactate amperometric biosensor; Spin coating procedure.

Fig. 1

Conceptual scheme of the proposed mechanism of lactate detection with the SCPC-PB-LOxAuNPs-Chit-PVC-Nafion biosensor. a Oxygen and lactate reaching the enzymatic layer through the diffusion limiting layer and resulting into pyruvate and oxygen peroxide. b Later transformation of hydrogen peroxide into water due to the catalytic effect of PB layer

Fig. 2

Fabrication scheme of the (bio)sensors developed. a Fabrication of SCPC-PB-LOxAuNPs-Chit-PVC-Nafion® biosensor. b Fabrication of PANI-based pH probe

Fig. 3

Analytical performance of the SCPC-PB-LOxAuNPs-Chit-PVC-Nafion® lactate biosensor in batch regime in PBS 0.1 M pH 7.4. a Chonoamperogram (E = 0.05 V) corresponding to the calibration plot of LA ranging from 0.5 to 20 mM; (inset: lactate calibration plot). b Interferent assay performed by using ascorbic acid (AA), uric acid (UA), dopamine (DA), and glucose (GL) as interfering species at 0.2, 0.2, 0.006, and 5 mM, respectively, prior to the addition of 0.5, 5, and 5 mM of LA

Fig. 4

Chonoamperogram recorded in presence of LA ranging from 0.5 to 20 mM in 0.1 M PBS at pH 7.4 with the biosensor developed in flow regime at 0.05 V. The inset shows the lactate calibration plot

Fig. 5

Dynamic response and average calibration graph for the carry-over test (five consecutive calibrations by alternatively increasing and decreasing the pH in buffer solutions) performed with the PANI-prepared pH sensor

Fig. 6

a Correlation between the concentration values of LA determined with the blood analyzer in serum samples and those determined with the biosensor developed. b Results of pH obtained by the blood analyzer, the PANI pH sensor developed, and a micro pH-meter