Long-term performance monitoring of a secondary electrospray ionisation high-resolution mass spectrometry platform using a vapor generator for routine quality control.

Secondary electrospray ionisation high-resolution mass spectrometry (SESI-HRMS) is a powerful, non-invasive tool for real-time breath analysis. However, instrumental drift, capillary degradation and other technical variations compromise clinical applications, especially for long-term studies, which require robust quality control (QC) methodologies. In this work, we present the use of an evaporation chamber-based reference gas system to monitor SESI-MS system performance over almost two years. Targeted QC measurements with deuterated butyric acid were recorded three times on each measurement day. Analytical stability was systematically quantified across both positive and negative ion modes. In a standalone continuous characterization, the reference gas generator delivered a stable signal over several hours. A Linear Mixed-Effects Model (LMM) was constructed to evaluate the influence of time-of-day on instrument performance. This revealed significant effects: positive mode signal intensity progressively increased throughout the day, whereas for negative mode, an initial drop in signal intensity and subsequent stabilisation was observed. Furthermore, the longitudinal QC framework successfully tracked capillary performance over time, providing objective performance metrics to retrospectively evaluate the impact of targeted hardware maintenance on system performance. The implementation of a structured QC protocol provides a valid methodology for long-term SESI-HRMS performance monitoring, ensuring data integrity and supporting the translation of SESI-HRMS into large-scale clinical applications.