| Abstract
| - Phytosterols have been shown to reduce cholesterol absorption in humans. Supplementingphytosterols in fat-free formulations, however, has yielded controversial results. In the present study,we investigated the effect of supplementing test meals with different fat-free phytosterol products oncholesterol incorporation into mixed micelles during simulated digestion and accumulation of micellarcholesterol by Caco-2 cells: control orange juice (OJ), orange juice supplemented with eithermultivitamin/multimineral tablets (MVT), multivitamin/multimineral tablets containing phytosterols(MVT+P), and phytosterol powder (PP). These combinations were added to Ensure-based test mealsand spiked with cholesterol of natural isotopic composition or 13C2-cholesterol to differentiate externalfrom endogenous cholesterol. After simulated gastric/small intestinal digestion, micelle fractions wereanalyzed for cholesterol enzymatically (n = 6−20/product) and by high-performance liquid chromatography−tandem mass spectrometry (n = 12/product) and added to Caco-2 cells to determine theaccumulation of 13C2-cholesterol (n = 10−24/product). As compared to OJ, PP and MVT+Psignificantly decreased cholesterol micellarization (determined enzymatically) by 70 ± 39 (mean ±SD) and 70 ± 39%, respectively (P< 0.001, Bonferroni). The stable isotope experiments revealedthat both PP and MVT+P reduced cholesterol micellarization [by 25 ± 12 (P = 0.055) and 21 ± 8%(P = 0.020), respectively, Fisher's protected LSD test] and Caco-2 cell accumulation (by 28 ± 8 and10 ± 8%, respectively; P< 0.010, Bonferroni). OJ+P did not inhibit micellarization or accumulationof cholesterol by Caco-2 cells. This study shows that fat-free phytosterol-containing products cansignificantly inhibit cholesterol micellarization and Caco-2 cell bioaccessibility, albeit to different extentsdepending on individual formulations. This is most likely explained by inhibition of cholesterolmicellarization. Keywords: Phytosterols; food matrix; cholesterol absorption; micellarization; Caco-2 cells; isotopiclabel; mass spectrometry
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