An enzymatic assay with formate oxidase for point-of-care diagnosis of methanol poisoning

Abstract

Gas chromatographic analysis for quantification of plasma methanol requires laboratory equipment and personnel, and it is typically unavailable in short time notice, especially in low- and middle-income countries. Detection of formate with the enzyme formate oxidase (FOX) is a promising method that can make the diagnosis of methanol poisoning simple and fast. The aims of this study were to test the sensitivity and specificity of a modified FOX-enzyme and to test the specificity of a point-of-care (POC)-model containing FOX-enzyme with samples from patients with metabolic acidosis. The sensitivity and specificity of FOX-enzyme in aqueous solution were evaluated with a spectrometer and by visual detection for colour change. Formate concentrations between 1 and 20 mmol/L were used to test sensitivity, and 18 potentially interfering substances were tested for specificity. The sensitivity of the FOX-enzyme was 100% and the specificity 97%. When specificity of the POC-model was tested, no false positives were detected. As such, the sensitivity and specificity of this modified FOX-enzyme for detection of formate were high. The results with this enzyme confirm the potential for its use in formate analysis as a fast diagnosis of methanol poisoning.

Keywords: bedside testing; diagnosis; formate; methanol poisoning; point-of-care-testing.

FIGURE 1

Illustration of principles for point‐of‐care model

FIGURE 2

Illustration of point‐of‐care model with three level scales for semiquantitative detection of formate; negative <1 mmol/L, low positive 1–10 mmol/L and high positive >0 mmol/L. Note. Adapted from “Formate test for bedside diagnosis of methanol poisoning,” by KE Hovda et al., 2021, Basic Clin Pharmacol Toxicol., 129, p. 87

FIGURE 3

Sensitivity of formate oxidase (FOX)‐enzyme in aqueous solutions evaluated with a spectrometer for nine formate concentrations between 1 and 20 mmol/L. Y‐axis is the ratio between the measured value at a given time and start (T = 0). A ratio ≥0.8 after 5 min was defined as a negative sample and a ratio ≤0.7 as a positive sample.

FIGURE 4

Sensitivity of formate oxidase (FOX)‐enzyme in aqueous solutions evaluated with a spectrometer for low concentrations of formate. Y‐axis is the ratio between the average measured value for each concentration at a given time and start (T = 0). A ratio ≥ 0.8 after 5 min was defined as a negative sample and a ratio ≤0.7 as a positive sample.

FIGURE 5

Illustration of the reaction between formate and formate oxidase (FOX)‐enzyme that gives a colour change to blue after 5 min. From left; formate 10 mmol/L; 5 mmol/L; 2 mmol/L and control (without formate)