A 96-well headspace iodometric assay with a starch-KI paper for high-throughput screening of formaldehyde in fresh-food contacting liquids.

The misuse of formaldehyde in fresh foods remains a food-safety concern, necessitating high-throughput screening methods suitable for food-associated aqueous samples. This study presents a 96-well microplate headspace iodometric assay, coupled with a starch-KI paper sensor and digital image analysis, for measuring formaldehyde in sample-contacting liquids. Classical hypoiodite oxidation chemistry was adapted as an in-situ headspace iodine-generation system: formaldehyde was oxidized under alkaline conditions, and following acidification, iodine formed in solution partitioned into the headspace and was captured on a starch-KI-impregnated paper sheet covering the wells. The captured iodine produced a brown iodine/polyiodide-associated color response in the dry starch-KI-cellulose matrix, which was quantified by flatbed scanner-based image analysis. The paper sensor provides uniform color responses, signal stability for at least 60 min, and excellent within-plate well-to-well repeatability (3% RSD, n = 96 at 0.6 mM). Under the optimized conditions, the assay exhibited a linear working range of 0.2-1.2 mM, with a detection limit of 0.06 mM. Interference testing revealed compound-dependent tolerance toward representative carbonyl compounds. Spike recoveries of 100-118% (n = 5) were obtained across seven seafood, offal, vegetable, and mushroom matrices. Application to real samples revealed millimolar levels of releasable formaldehyde in selected storage liquids and rinsates. A preliminary comparison of three representative samples covering low, intermediate, and high concentration levels showed no statistically significant difference between the proposed assay and conventional sulfite titration (paired t-test, P > 0.05). An AGREE greenness score of 0.86 indicated a favorable analytical greenness profile. Compared with reported fluorescence- and acetylacetone-based colorimetric assays, the proposed platform integrates simple aqueous reagents, headspace-based matrix separation, patterning-free paper sensing, and 96-well throughput, providing a practical tool for routine screening of releasable formaldehyde in fresh-food-contacting liquids. The method targets formaldehyde released into sample-contacting liquids rather than total formaldehyde in edible tissues. Elevated responses indicate screening findings needing confirmatory analysis.