Heart-Cut Two-Dimensional Countercurrent Chromatography for the Isolation of a Furan Fatty Acid Triacylglycerol from Latex Gloves and Identification of Further Lipid Compounds

Abstract

Hevea brasiliensis latex from specific genotypes such as RRIM501 or PB235 is one of the richest natural sources of furan fatty acid 9-(3-methyl-5-pentylfuran-2-yl)-nonanoic acid (9M5), which is mainly present in the form of triacylglycerols. In this study, we successfully isolated a triacylglycerol esterified with three 9M5 molecules (tri-9M5) with a purity of 97% from the lipid extract of latex gloves by means of countercurrent chromatography (CCC). The gas chromatography with a mass spectrometry (GC/MS) spectrum of tri-9M5 not only featured the diagnostic fragment ion [M-RCOO]+ of triacylglycerols but also a fragment ion shifted by 16 Da to higher mass which corresponded with [M-RCO]+. [M-RCO]+ was only detected in triacylglycerols substituted with at least one furan fatty acid. Additionally, five γ-tocotrienyl fatty acid esters (γ-T3-FA esters), namely, γ-T3-16:0, γ-T3-18:0, γ-T3-18:1n-9, γ-T3-18:2n-6 and γ-T3-20:0, were detected in the sample. Contributions of γ-T3-FA esters with 18:1n-9 and 18:2n-6 which co-eluted in GC/MS could be calculated after mathematical correction for contributions of the 13C2 isotope peak of γ-T3-18:2n-6 to γ-T3-18:1n-9. This was necessary for quantitation of these two γ-T3-FA esters. Improved separation of the γ-T3-FA esters could be achieved by the novel heart-cut recycling CCC mode (four cycles). Sterols detected in disposable latex gloves were β-sitosterol, Δ5-avenasterol, and stigmasterol along with small quantities of 24-methylene-cycloartenol and Δ5-citrostadienol.