Ultra‐High‐Performance Liquid Chromatography‐Mass Spectrometry‐based Chemical Profiling of <i>Glycine max</i> Seeds, Preparative Isolation of Its Major Monoglycerides, and Their Spectroscopic Characterization

ABSTRACT Soybean ( Glycine max L. seeds) constitutes a staple in many parts of the world and is associated with several health benefits. It contains oil and proteins besides bioactive isoflavones such as daidzein and genistein. Saturated and unsaturated fatty acids in soybean oil are chemically converted to monoglycerides. Monoglycerides are used in the pharmaceutical, cosmetic, and food industries. There is an increasing demand for high‐quality monoglycerides, especially from natural sources. The aim of this study was the chemical profiling of G. max seeds and the development of a preparative high‐performance liquid chromatography (prep‐HPLC) protocol for the purification of its monoglycerides. Methanol extract of soybeans was subjected to liquid‐liquid partitioning with water‐saturated butanol to prepare an enriched fraction. Its chromatographic profiling was carried out on an ultra HPLC‐mass spectrometry system using a C18 column (2.1 × 50 mm, 1.7 µm) that revealed the presence of monoglycerides and fatty acids. Monolaurin (18.6%), monotridecanoin (13.0%), and monomyristin (9.8%) were the major monoglycerides. α‐Linolenic acid (10.9%) and tetracosa‐15,18,21‐trienoic acid (9.4%) were the major fatty acids. A protocol for the preparative‐scale isolation of monolaurin and monomyristin was developed. It involved a gradient elution with acetonitrile‐water with 0.1% formic acid on a C18 column (10 × 250 mm, 5 µm) at a flow rate of 4 mL/min. The injection volume was 200 µL, and the run time was 20 min. In conclusion, the current study established the presence of monolaurin and monomyristin along with other monoglycerides in G. max seeds. An efficient protocol for the rapid isolation of key monoglycerides was also developed.