Rapid Enzyme-Independent Catechol Reactions In Biologically-Relevant Aqueous Media

Catecholic compounds are widely used in biological and bioanalytical systems, yet their chemical stability under routine experimental conditions remains incompletely characterized. Here, we demonstrate that catechol undergoes rapid, enzyme- and cell-free transformations in biologically-relevant aqueous media, producing spectrally-active chromophoric species (“browning”) under conditions commonly used in in vitro studies. In neutral to mildly alkaline phosphate buffer (pH 7.0-8.0) alone, catechol browning occurred within hours in a temperature-dependent manner. The presence of amine-containing components – including amino acids (glycine, lysine, proline), the biocompatible buffer tris(hydroxymethyl)aminomethane (Tris), and bovine serum albumin (BSA) – markedly accelerated browning, with detectable optical changes within minutes and strong pH dependence. BSA-enhanced browning was accompanied by reproducible alterations in electrophoretic mobility under denaturing conditions. These findings indicate that catechol can undergo rapid, non-enzymatic chemistry in commonly used buffers and protein-containing systems, potentially confounding spectrophotometric and protein-based measurements. The results underscore the need to account for enzyme- and cell-free catechol reactivity in experimental design and data interpretation.