Quantitative Profiling of Metabolites, Sugars, and Lipids in Pollen Tubes by Mass Spectrometry.

Pollen tube germination and elongation are energy-demanding, metabolically active processes essential for plant reproduction. Recent research combining metabolomics, imaging, and molecular genetics has highlighted the crucial roles of sucrose synthase activity, starch breakdown, and lipid signaling in maintaining pollen tube health and promoting precise growth, particularly under stress conditions. This chapter presents a mass spectrometry-based workflow for high-resolution profiling of metabolites, sugars, and lipids in pollen and pollen tubes. The workflow includes metabolite extraction, chemical derivatization, and chromatographic separation steps compatible with both gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS). These techniques enable simultaneous quantification of polar metabolites, lipids, and fatty acids, capturing the metabolic complexity that supports pollen tube germination, elongation, and fertilization. It also covers the application of internal standards and isotopic normalization to ensure precise quantification and reproducibility across experiments. These analytical approaches collectively provide a foundation for comparative metabolite profiling in reproductive biology and can be adapted to various plant species, lipid signaling, and membrane biogenesis, thereby offering a mechanistic understanding of how pollen tubes sustain polarized growth under both developmental and stress conditions.