1H NMR spectroscopy quantifies visibility of lipoproteins, subclasses, and lipids at varied temperatures and pressures

Abstract

Supplementary key words metabolomics • serum lipids • intermediate density lipoprotein • very low density lipoprotein • low density lipoprotein • high density lipoprotein • high pressure nuclear magnetic resonanceLipoprotein particles of different size and composition mediate lipid metabolism by transporting them to their place of destination via the blood or the lymphatic system (1, 2). The outer layer of all lipoprotein classes is covered by hydrophilic moieties of apolipoproteins and of amphiphilic lipids, such as free unesterified cholesterol, phospholipids (PLs), and SM, in order to guarantee water solubility of the assembly. The core of the particles is mainly composed of cholesteryl esters (CEs), triacylglycerides (TGs), and also free cholesterols. The lipoprotein composition directly affects the molecular dynamics of the lipids and may alter the conformation of apolipoproteins, which has direct influence on metabolic processes like receptor binding or cholesterol exchange (3-5).With rising temperature, lipid assemblies usually are subjected to a phase transition from a solid-like gel state to a liquid-crystalline-like state with higher internal mobility. The core lipids of LDLs undergo a broad phase transition Abstract NMR-based quantification of human lipoprotein (sub)classes is a powerful high-throughput method for medical diagnostics. We evaluated select proton NMR signals of serum lipoproteins for elucidating the physicochemical features and the absolute NMR visibility of their lipids. We separated human lipoproteins of different subclasses by ultracentrifugation and analyzed them by 1 H NMR spectroscopy at different temperatures (283-323 K) and pressures (0.1-200 MPa). In parallel, we determined the total lipid content by extraction with chloroform/methanol. The visibility of different lipids in the 1 H NMR spectra strongly depends on temperature and pressure: it increases with increasing temperatures but decreases with increasing pressures. Even at 313 K, only part of the lipoprotein is detected quantitatively. In LDL and in HDL subclasses HDL2 and HDL3, only 39%, 62%, and 90% of the total cholesterol and only 73%, 70%, and 87% of the FAs are detected, respectively. The choline head groups show visibilities of 43%, 75%, and 87% for LDL, HDL2, and HDL3, respectively. The description of the NMR visibility of lipid signals requires a minimum model of three different compartments, A, B, and C. The thermodynamic analysis of compartment B leads to melting temperatures between 282 K and 308 K and to enthalpy differences that vary for the different lipoproteins as well as for the reporter groups selected. In summary, we describe differences in NMR visibility of lipoproteins and variations in biophysical responses of functional groups that are crucial for the accuracy of absolute NMR quantification.

Chronic Oleoylethanolamide Treatment Decreases Hepatic Triacylglycerol Level in Rat Liver by a PPARγ/SREBP-Mediated Suppression of Fatty Acid and Triacylglycerol Synthesis

“…In particular, the signals at 3.35-3.32 and 3.17 ppm of methyl groups (-N + (CH 3 ) 3 ) were ascribable to PC/PE and SM, respectively. Moreover, the presence of PC and PE were confirmed by resonances at 3.60 and 4.07 ppm, respectively, while the multiplets at 5.74 ppm were diagnostic of the characteristic sphingenine moiety vinyl protons (HO-CH-CH=CH-) and, therefore, suggested the presence of sphingolipids, including sphingomyelin, as also reported in the literature for specific systems [40,51,52]. All diacylglycerophospholipids (DAGP) are represented by the backbone glycerol sn-2 proton multiplet at~5.…”

“…By NMR spectroscopy we identified specific characteristic sphingenine moieties, suggesting the possible presence of sphingomyelin [ 40 , 51 , 52 ]. On the basis of the NMR result, we investigated on the different sphingolipid species by TLC and LC-MS/MS analysis.…”

NMR-Based Metabolomics in Investigation of the Radiation Induced Changes in Blood Serum of Head and Neck Cancer Patients and Its Correlation with the Tissue Volumes Exposed to the Particulate Doses

“…The initial (week-1) response to the treatment is manifested by the simultaneously increased levels of the branched-chain amino acids (BCAAs: leucine, valine and isoleucine), N-acetyl-glycoprotein (NAG), Nacetylcysteine, glutamine, creatinine and tyrosine as well as the decreased lipid signals at 3.2 ppm and the deformations (decrease) of the downfield part of the resonance at 0.9 ppm. The lipid band at 0.9 ppm derives from the terminal CH 3 groups of fatty acids, triglycerides and phospholipids as well as from the cholesterol methyls [19,20]. Thus, the changes in its shape and intensity indicate the variation in the mutual contributions of the individual components.…”

Human blood lipoprotein predictions from1H NMR spectra: protocol, model performances and cage of covariance

“…A few studies have characterised signature signals of the LP main fractions and subfractions. This has been done mathematically using either curve fitting 16 or by calculating selectivity ratio from PLS models developed to predict the LP 18,19 , and experimentally by measuring pure fractions after UC 33 . The concentrations of the three main populations of protons, cholesterol (0.75-0.6 ppm), methyl (0.95-0.80 ppm) and methylene (1.4-1.2 ppm), differ significantly among the 1 H NMR spectra of the four main fractions.…”