High-throughput mass spectrometry for screening microbial culture supernatants for aromatic amino acids and selected derivatives.

Sustainable production of aromatic amino acids (AAAs) and their derivatives are central to developing renewable routes for aroma- and flavour-related chemicals with myriads of applications in pharmaceutical industry, food and nutrition, cosmetics and personal care, polymer synthesis, and biotechnology. The aim of the present study was to establish and validate a high-throughput mass spectrometry (HT-MS) workflow for rapid screening of AAAs and selected aromatic derivatives, i.e., kynurenic acid, indole-3-acetic acid, and p-coumaric acid, using an automated solid-phase extraction system coupled to time-of-flight detection. A method based on fast cycle time, 15 s per sample, and minimal sample handling was elaborated to enable high-throughput screening of large culture collections. Validation using pure standards confirmed excellent linearity (R² ≥ 0.99), accuracy, precision, and on-instrument stability at 4 °C. Although strong matrix effects were observed in microbial culture supernatants, analyte detection remained selective and reproducible, supporting reliable semi-quantitative screening. The results demonstrate that the proposed workflow supports semi-quantitative, comparative analysis of AAAs and their derivatives across large sample sets. Overall, the developed HT-MS method provides a robust and efficient platform for rapid pre-screening of microbial collections accelerating the discovery and optimization of aromatic metabolite production.