Received on December 15, 2004
Accepted on May 24, 2005

Lipids, Lipoproteins, and Cardiovascular Risk Factors

Quantification of Lipoprotein Subclasses by Proton Nuclear Magnetic Resonance-Based Partial Least-Squares Regression Models

¹ Institute of Human Nutrition, Centre for Advanced Food Research, The Royal Veterinary and Agricultural University, Frederiksberg, Denmark
² Umetrics AB, Malmö, Sweden
³ Centre for Advanced Food Studies, Department of Food Science, Quality and Technology, The Royal Veterinary and Agricultural University, Frederiksberg, Denmark
⁴ Novo Nordisk A/S, Virology & Molecular Toxicology, Måløv, Denmark

^* To whom correspondence should be addressed. E-mail: mpe{at}kvl.dk.

Background: Cardiovascular disease risk can be estimated in part on the basis of the plasma lipoprotein profile. Analysis of lipoprotein subclasses improves the risk evaluation, but the traditional methods are very time consuming. Novel, rapid, and productive methods are therefore needed.

Methods: We obtained plasma samples from 103 fasting people and determined the plasma lipoprotein subclass profiles by an established ultracentrifugation-based method. Proton nuclear magnetic resonance (NMR) spectra were obtained from replicate samples on a 600 MHz NMR spectrometer. From the ultracentrifugation-based reference data and the NMR spectra, we developed partial least-squares (PLS) regression models to predict cholesterol and triglyceride (TG) concentrations in plasma as well as in VLDL, intermediate-density lipoprotein (IDL), LDL, 3 LDL fractions, HDL, and 3 HDL subclasses.

Results: The correlation coefficients (r) between the plasma TG and cholesterol concentrations measured by the 2 methods were 0.98 and 0.91, respectively. For LDL- and HDL-cholesterol concentrations, r = 0.90 and 0.94, respectively. For the cholesterol concentrations in the LDL-1, LDL-2, and LDL-3 fractions, r = 0.74, 0.78, and 0.69, respectively, and for HDL subclasses HDL_2b, HDL_2a, and HDL₃, cholesterol concentrations were predicted with r = 0.92, 0.94, and 0.75, respectively. The TG concentrations in VLDL, IDL, LDL, and HDL were predicted with correlations of 0.98, 0.85, 0.77, and 0.74, respectively. The cholesterol and TG concentrations in the main lipoprotein fractions and in LDL fractions and HDL subclasses predicted by the PLS models were 94%-100% of the concentrations obtained by ultracentrifugation.

Conclusion: NMR-based PLS regression models are appropriate for use in research in which analyses of the plasma lipoprotein profile, including LDL and HDL subclasses, are required in large numbers of samples.