Monitoring 12 isoflavones during soybean seed germination using LC-MS/MS

Soybean isoflavones have been linked to potential cardiovascular protective effects. This study investigated changes in isoflavone content during germination in five Taiwanese soybean cultivars. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was established to quantify 12 individual isoflavones within 15 min, using a Kinetex F5 column and Agilent 6470 mass spectrometer in MRM mode. The method showed good linearity (3–800 ng/mL, r² > 0.99), sensitivity (LLOQ = 3 ng/mL), and precision (RSD < 13.6 %). Total isoflavone concentrations increased progressively during the germination period and peaked on day 4. Malonyl-genistin was the dominant component, contributing 55–70 % of the total isoflavone content, and exhibited a 1.8- to 2.5-fold increase from day 1 to day 4. Glycoside forms also increased during germination, while aglycones and acetyl-glucosides remained relatively constant. These results indicate active isoflavone transformation during germination and suggest that day 4 may be a favorable time point for harvesting to maximize bioactive isoflavone yield. This analytical approach can support future studies on soybean sprout quality and processing optimization aimed at enhancing functional compound accumulation.