High-Throughput Quantitation of Plasma Trimethylamine N-oxide Using Desorption Electrospray Ionization Mass Spectrometry for Rapid Cardiovascular Disease Screening

Trimethylamine N-oxide (TMAO) is an emerging biomarker of cardiovascular disease (CVD) risk, but current detection methods are limited by low throughput and lengthy workflows. To address this, we developed a high-throughput desorption electrospray ionization-mass spectrometry (DESI-MS) platform for rapid and accurate quantitation of TMAO in plasma. The method involves protein removal, spot deposition, and DESI-MS analysis using isotope-labeled internal standards for calibration. Validation showed strong linearity (R² > 0.97), precision (CV < 20%), minimal matrix effects, and low carry-over (<5%). In a cohort of 197 patients from National Taiwan University Hospital, DESI-MS demonstrated high correlation with LC-MS/MS (R = 0.96), 92.9% concordance in risk classification, and a 10-fold reduction in processing time. Risk stratification revealed a 1.55-fold higher prevalence of coronary stenosis in the high-risk group. Capable of processing up to 2,000 samples per day, this DESI-MS platform shows strong potential for large-scale clinical screening and personalized cardiovascular risk assessment.