Quantitative Raman Analysis of Carotenoid Protein Complexes in Aqueous Solution

Carotenoids are naturally abundant fat-soluble pigmented compounds, with dietary, antioxidant and vision protection advantages. The dietary carotenoids, Beta Carotene, Lutein and Zeaxanthin, complexed with in bovine serum albumin (BSA) in aqueous solution, were explored using Raman spectroscopy to differentiate and quantify their spectral signatures. UV visible absorption spectroscopy was employed to confirm the linearity of responses over the concentration range employed (0.05-1mg/ml) and, of the 4 source wavelengths, 785nm, 660nm, 532nm, 473nm, 532nm was chosen to provide the optimal response. After preprocessing to remove water and BSA contributions, and correct for self-absorption, a partial least squares model with R2 of 0.9995, resulted in a accuracy of Root Mean Squared Error of Prediction for Beta Carotene of 0.0032 mg/ml and Limit of Detection 0.0106mg/ml. Principal Components Analysis clearly differentiated solutions of the three carotenoids, based primarily on small shifts of the main peak at ~1520cm-1. Least squares fitting analysis of the spectra of admixtures of the carotenoid:protein complexes showed reasonable correlation between norminal% and fitted%, yielding 100% contribution when fitted with individual carotenoid complexes and variable contributions with multiple ratios of admixtures. The results indicate the technique can potentially be used to quantify carotenoid content of human serum and to identify their differential contributions for application in clinical analysis.