Quantification of the Molecular Species of TAG and DAG in Lesquerella (Physaria fendleri) Oil by HPLC and MS

Abstract

Ten diacylglycerols (DAG) and 74 triacylglycerols (TAG) in the seed oil of Physaria fendleri were recently identified by high-performance liquid chromatography (HPLC) and mass spectrometry (MS). These acylglycerols (AG) were quantified by HPLC with evaporative light scattering detector and electrospray ionization mass spectrometry of the lithium adducts of the AG in the HPLC fractions of lesquerella oil. The MS 1 ion signal intensities of molecular ions [M ? Li] ? in HPLC fractions of an HPLC peak were used to estimate the ratios of AG in the HPLC peak. The ratios of TAG with the same mass in HPLC fractions were estimated by the ratios of the sums of MS 2 ion signal intensities from the neutral loss of the three fatty acids [M ? Li -FA] ? . The ratio of DAG with the same mass were estimated by the ratio of the sums of two MS 2 ion signal intensities [M ? Li -FA] ? and [FA ? Li] ? from the two different FA of a DAG. We have estimated the contents of ten molecular species of DAG and 74 molecular species of TAG in P. fendleri oil using this new method. The content of ten DAG combined was about 1 % and 74 TAG was about 98 %. The contents of DAG in decreasing order were: LsLs (0.25 %), LsLn (0.25 %), LsO (0.24 %), and LsL (0.11 %); and the contents of TAG in decreasing order were: LsLsO (31.3 %), LsLsLn (24.9 %), LsLsL (15.8 %), LsL-OH20:2 (4.3 %), LsO-OH20:2 (2.8 %), and LsLn-OH20:2 (2.5 %).

Identification of multiple lipid genes with modifications in expression and sequence associated with the evolution of hydroxy fatty acid accumulation in Physaria fendleri

“…Castor bean LPAAT2 shows enhanced HFA activity (Arroyo‐Caro et al ., ), which helps explain in part why castor bean TAGs accommodate HFAs at all three positions. However, unlike castor bean, positional analysis and HPLC measurement of TAG species in Physaria fendleri demonstrated that only low quantities of HFAs are present at the sn ‐2 position of TAGs (Hayes et al ., ; Lin and Chen, ), although there is some variability in other Paysonia and Physaria species, which show higher proportions of HFAs in the sn ‐2 position. These chemical analyses, together with the lack of divergence of the LPAAT2 sequence in Physaria (Figure ), suggest that Physaria LPAAT2 is unlikely to be specialized for HFA incorporation.…”

Ratios of Regioisomers of the Molecular Species of Triacylglycerols in Lesquerella (Physaria fendleri) Oil Estimated by Mass Spectrometry

“…Elongation seems more effective than hydroxylation to incorporate hydroxy FA into TAG at the sn-1/3 positions in lesquerella as the order of OH20 > diOH18 > OH18. The contents of TAG containing diOH18 and diOH20 were very low [6]. The block of elongation (from C 18 to C 20 ) may be used to increase the content of hydroxy FA, e.g., ricinoleate, at the sn-2 position of TAG in lesquerella.…”

“…We have recently reported the contents (%) of all of these 74 molecular species of TAG in lesquerella oil [6]. From these contents and the ratios of regioisomers (Table 1) of the molecular species of TAG, the contents of individual FA at the sn-2 and sn-1/3 positions were estimated as shown in Table 2.…”

“…The content of ten DAG combined was about 1 %, 74 TAG was about 98 % and 13 tetraacylglycerols was about 1 % [6,7]. The contents of the molecular species of TAG were: LsLsO (31.3 %), LsLsLn (24.9 %), LsLsL (15.8 %), LsL-OH20:2 (4.3 %), LsO-OH20:2 (2.8 %), and LsLn-OH20:2 (2.5 %) [6]. Minor monohydroxy and dihydroxy FA were identified in lesquerella oil and their structures were different from those of minor FA in castor oil as OH 12 18:2 9,14 (OH 12 18:2 9,13 in castor oil) and diOH 12,13 18:2 9,14 (diOH 11,12 18:2 9,13 in castor oil) [4].…”

“…We have recently identify and quantify ten molecular species of diacylglycerols (DAG), 74 molecular species of triacylglycerols (TAG) and 13 molecular species of tetraacylglycerols in lesquerella oil using HPLC and electrospray ionization mass spectrometry (ESI-MS) [4][5][6][7]. The content of ten DAG combined was about 1 %, 74 TAG was about 98 % and 13 tetraacylglycerols was about 1 % [6,7]. The contents of the molecular species of TAG were: LsLsO (31.3 %), LsLsLn (24.9 %), LsLsL (15.8 %), LsL-OH20:2 (4.3 %), LsO-OH20:2 (2.8 %), and LsLn-OH20:2 (2.5 %) [6].…”