This document details an analytical method developed for the identification and quantitation of eight plant phytoestrogens (biochanin A, coumestrol, daidzein, equol, formononetin, glycitein, genistein, and prunetin) in soy milk. The method utilizes gas chromatography tandem mass spectrometry (GC/MS/MS) with an Agilent 7000 Series Triple Quadrupole GC/MS instrument.
The procedure involves extracting phytoestrogens from soy milk using liquid-liquid extraction with ethyl acetate, followed by derivatization to trimethylsilyl ethers using trimethylchlorosilane (TMCS) and N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA). The analysis employs specific Multiple Reaction Monitoring (MRM) transitions optimized for each analyte to ensure high selectivity and sensitivity.
| Column Type | Agilent J&W HP-5 (5% phenyl, 95% methylpolysiloxane) |
|---|---|
| Column Dimensions | 30 m x 0.25 mm id |
| Carrier Gas | Helium |
| Flow Rate | 1.2 mL/min |
| Injector Temperature | 280 °C |
| Ion Source Temperature | 300 °C |
| Electron Energy | -70 eV |
| Dwell Time | 50 msec |
The study analyzed eight phytoestrogens: biochanin A, coumestrol, daidzein, equol, formononetin, glycitein, genistein, and prunetin.
Ethyl acetate was used as the extracting solvent for the liquid-liquid extraction of phytoestrogens from soy milk.
The analytes were derivatized using trimethylchlorosilane (TMCS) and N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA).
MassHunter software was used for instrument control and data analysis.