Differentiation of wild-type and PAP1-overexpressing tobacco by volatile organic compound profiling using TP-SESI mass spectrometry.

Real-time profiling of plant volatiles provides insight into metabolic regulation, yet rapid and high-throughput characterization remains analytically challenging. Here, we apply temperature programming secondary electrospray ionization (TP-SESI) mass spectrometry to compare volatile organic compound (VOC) profiles from wild-type and PAP1-overexpressing tobacco (Nicotiana tabacum). This study extends the application of TP-SESI, which was originally developed for plant volatile analysis, to evaluate genotype-specific metabolic differences. TP-SESI enabled sensitive detection of VOCs and semi-volatile compounds spanning three major plant volatile classes: terpenoids, phenylpropanoids/benzenoids, and fatty acid derivatives, across both genotypes. While the two genotypes shared many detected features, PAP1-overexpressing plants were associated with upregulation of these indicated VOCs, suggesting broad metabolic reprogramming associated with PAP1 activation. Statistical analyses revealed clear separation between genotypes, demonstrating TP-SESI captures robust, reproducible metabolic differences in intact plant tissue. These results suggest that TP-SESI is capable of detecting genotype-associated variations in plant volatile emissions and may be useful for examining transcription factor-related changes in plant metabolism.