Reverse Phase High Pressure Liquid Chromatography of Vitamin A in Margarine, Infant Formula, and Fortified Milk

Abstract

Retinol was determined in margarine (50 mg), infant formula (1 ml), and fortified milk (1 ml) by saponification in centrifuge tubes, extraction of the unsaponifiable lipid with hexane, and high pressure liquid chromatography (HPLC) in 90% methanol on a 25 cm × 3.2 mm column containing 10 μm LiChrosorb reverse phase. β-Carotene was determined using the same column and 99% methanol as eluant. The vitamins in the eluate were identified and measured from their absorption at 325 (retinol) and 453 nm (β-carotene), using a computing integrator. Retinol and carotene were prominent peaks in the chromatograms from properly fortified samples and were satisfactorily separated from other materials. In a survey of 12 different margarines for retinol, the results from liquid chromatography agreed with those of a variety of Spectrophotometric and fluorometric procedures but they were obtained with greater ease and they could be interpreted more confidently. The recovery of retinol from oil was better than 99%. The coefficients of variation were 6.8% for 12 replicate analyses of a margarine for retinol and 5.4% for 10 replicate analyses of a margarine for β-carotene.

Storage stability evaluation of some packed vegetable oil blends

“…The results presented on the stability of minor components, viz., oryzanol and tocopherols, show that under the conditions used in the study, packaged oils and their blends may retain appreciable amounts of these natural antioxidants and therefore may provide adequate nutrition. Various methods are available for the determination of βcarotene in vegetable oils and related products based on the analysis of unsaponifiable matter using reversed phase C-18 columns and methanol/acetonitrile/methylene chloride (16), methylene chloride/acetonitrile (17), and 90% methanol (18) as the mobile phases. In this study, the β-carotene content was determined by injecting the oil sample solution directly into the mobile phase.…”

Consumer Acceptability and p‐Carotene Content of Beef as Related to Cattle Finishing Diets

“…Liver, ribeye steak, and ground beef samples were prepared by protocols adapted from Thompson and Maxwell (1977). After saponification, samples were extracted with hexane and evaporated to dryness.…”

“…Concentration, purity and stability of ␤-carotene (Fluka, Ronkonkom, NY) and retinol (Fluka, Ronkonkom, NY) standards were monitored spectrophotometrically with a diode array spectrophotometer (Model 8451A, Hewlett Packard, Palo Alto, CA). Concentrations of standards were determined using absorptivities of 2,480 at 450 nm for ␤-carotene (in methylene chloride) [Davies, 1976] and 1832 at 325 nm for retinol (Thompson and Maxwell, 1977). Peak identity was confirmed by comparison of retention times to ␤-carotene and retinol standards and by standard addition prior to saponification.…”

Isocratic nonaqueous reversed-phase liquid chromatography of carotenoids

“…Although modern liquid chromatography (HPLC) is gradually emerging as a method of choice for carotenoid separation, no systematic study of its merits and shortcomings has yet been conducted. Rather, a diversity of HPLC systems has been devised for specific applications, often of limited scope, such as the determination of /3-carotene and/or some selected derivatives in pizza (4), oil, margarine, and milk (5)(6)(7)(8)(9), orange and citrus juice (10)(11)(12)(13)(14), tomatoes (15), and tobacco leaves (16,17). HPLC has proven particularly useful for photosynthetic pigment profiling in algae and higher plants (18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28).…”