Quantitative determination of free and esterified cholesterol concentrations in cholesterol-fed rabbit aorta using near-infrared- Fourier transform-Raman spectroscopy

Abstract

Near-infrared-Fourier transform-Raman spectroscopy is a rapid and non-destructive technique that can provide reliable qualitative in situ information about the chemistry of biological samples. We have combined this technique with partial least squares (PLS) regression to perform a quantitative determination of free cholesterol and cholesterol esters in two synthetic sample series, in order to evaluate the performance and accuracy of the Raman-PLS method. In 65 ternary mixtures containing various proportions of cholesterol, cholesterol linoleate and oleate, the standard errors of prediction (SEP) are 1.3%, 1.2% and 0.9%, respectively. In the case of these ternary mixtures, it is also possible to extract quantitative information for a single component independently. For the second series of experiments, concerning the delicate problem of quantitative analysis of cholesterol palmitate and stearate mixtures, the standard error of prediction for 49 samples is 3.0%. After this preliminary study, the technique was used for the quantitative analysis of free and esterified cholesterol in rabbit arterial wall. Comparing the Raman-PLS results with the biochemical results, the SEP are 0.24 mg g −1 of tissue in 31 rabbit aorta samples for free cholesterol and 0.35 mg g −1 of tissue in 30 samples for esterified cholesterol. Using the Raman-PLS method, quantitative analysis is performed in approximately 10 min without destruction of the samples.