Simultaneous determination of molecular species of monoacylglycerols, diacylglycerols and triacylglycerols in human very-low-density lipoproteins by reversed-phase liquid chromatography

Abstract

The aim of the present study was to investigate the applicability of a previously developed method for the analysis of triacylglycerol molecular species to the simultaneous determination of triacylglycerols, diacylglycerols and monoacylglycerols of human very-low-density lipoproteins (VLDL). Ten elderly women were recruited for the study. Blood was obtained in fasting conditions and VLDL were isolated by ultracentrifugation. Neutral lipids were separated by solid-phase extraction and were subsequently injected on a reversed-phase HPLC system, with an elution system composed of acetone in acetonitrile. The method allowed the separation of four monoacylglycerols, 18 diacylglycerols and 24 triacylglycerols, including the resolution of positional isomers of diacylglycerols. Monoacylglycerols were composed of oleic, linoleic, palmitic and stearic acids. The major diacylglycerols were 1,2-dilinoleoyl-glycerol and 1,3-dilinoleoyl-glycerol (14.24+/-1.02 and 17.93+/-1.42%, respectively). The main triacylglycerols quantified were dioleoyl-stearoyl-glycerol (OOS), oleoyl-dipalmitoyl-glycerol (OPP), trilinoleoyl-glycerol (LLL) and linoleoyl-distearoyl-glycerol (LSS), accounting for 11.25+/-2.15, 10.14+/-2.05, 9.35+/-2.30 and 8.56+/-1.56%, respectively. An inverse relationship between polarity and fatty acid disappearance from triacylglycerols (r(2)=0.82, P<0.05) and from diacylglycerols (r(2)=0.93, P<0.01) was discovered. In conclusion, the method allowed, for the first time, the easy, rapid and simultaneous determination in a single chromatogram of triacylglycerol, diacylglycerol and monoacylglycerol molecular species of human VLDL by reversed-phase HPLC.

Quantification of major lipid classes in human triacylglycerol‐rich lipoproteins by high‐performance liquid chromatography with evaporative light‐scattering detection

“…Homan and Anderson [19] did not succeed in finding MG in rat tissues. In contrast with these results, we showed the presence of this lipid class in VLDL in a recent study published by our group, achieving the quantification of monoleoyl-glycerol, monolinoleoyl-glycerol, monopalmitoyl-glycerol, and monostearoyl-glycerol [22]. Due to the non-linear response of the ELSD, it is relatively difficult to detect at the same time peaks of very low and very high areas (i. e. TG and MG), since the detector can be easily saturated.…”

“…This is true for a narrow range of amount injected. When dealing with a wide variety of standard amounts or concentrations, the responses are better fitted to polynomial curves [22,29]. In the present study curves adjusted to linear equations showed r 2 coefficients lower than 0.98, whereas secondorder polynomial curves yielded r 2 values very close to 1.…”

“…SeppanenLaakso et al [14] optimized a HPLC-ELSD method for the quantification of lipid classes in human plasma. However, to our knowledge, no method has been developed for analysis of the lipid classes in TRL by HPLC, which are characterized for an extremely high concentration of TG [21,22]. In the present study, we optimized a HPLC method, coupled with ELSD, for the quantitative analysis of CE, TG, free cholesterol (FC), monoacylglycerols (MG), and phospholipids (PL) in human CM and VLDL.…”

Determination of Omega Fatty Acid Profiles in Egg Yolk by HILIC-LC-MS and GC-MS

“…Elution order of PLs in reversed phase depends on the differences in the chain lengths and the number of double bonds of acyl residues. The elution order of lipids in reversed phase is presented using the coefficient equivalent carbon number (ECN) for fatty acid (Perona and Ruiz-Gutierrez 2003). ECN is defined as ECN = CN − 2n (where CN is the sum of the total carbon atom number of fatty acyl groups and n is the number of the sum of double bonds).…”

A review of chromatographic methods for the assessment of phospholipids in biological samples

“…RP-HPLC is used to separate individual molecular species of a particular phospholipid class (Bunger and Pison, 1995). The sample will be separated based on a partition mechanism and will elute based on lipophilicity owing to the combined chain length and number of double bonds present in the fatty acids side chains (Perona and Ruiz-Gutierrez, 2003). Shorter chains will have longer retention times .…”