Simultaneous Measurement of Carotenoids and Vitamins A and E in Human Serum by High Performance Liquid Chromatography (HLPC) and the Effect of Consuming Eggs on Serum Carotenoid and Lipid Concentrations and Macular Degeneration

摘要

The aims of this project were to: (1) develop a simultaneous measurement and identification of carotenoids, tocopherols, and retinol concentrations in human serum using a new C30 ProntoSlL Column by High Performance Liquid Chromatography (HPLC); (2) determine if the consumption of 12 store-bought eggs per week for 1 year will raise serum lutein and zeaxanthin levels and MPOD while preventing the progression of AMD in individuals with early to intermediate stages of Dry AMD; (3) determine if the consumption of 12 store-bought eggs per week for 1 year will raise serum lipid, lipoprotein cholesterol, and other cardiovascular disease (CVD) related biochemical concentrations in these individuals; and (4) determine if the consumption of a meal containing high amounts of lutein and zeaxanthin, which may occur with the consumption of 12 eggs per 1 week, alters the serum concentration of other important carotenoids as well as vitamins A and E.;Previously, HPLC instruments utilizing a C18 300 x 4.6 mm column was needed to separate and identify the various isomers of carotenoids from human serum. In the first experiment, an HPLC method utilizing a new C30 ProntoSIL 250 x 4.6 mm column, at 22°C, a flow rate of 0.5 mL/min, and an injection volume of 30 microL with an 80 minute running time was optimized for the determination of the various carotenoid isomers, tocopherols, and retinol simultaneously in human serum. A diode array detector (DAD) was used to measure serum carotenoids (set at a wavelength of 450 nm) and retinol (set at a wavelength of 325 rim) while a fluorescent detector was used to measure tocol (internal standard) and various tocopherols (set at an excitation wavelength of 290 nm and an emission wavelength of 330 nm).;The next phase of the project involved identifying if the consumption of 12 eggs per week for 1 year produces changes in serum lutein and zeaxanthin concentrations and on MPOD and the progression of AMD in patients previously diagnosed with early AMD. Forty-five adults were randomized to the intervention (n=27) or control (n=18) group. Serum lutein, zeaxanthin and MPOD for both eyes were measured at 0 and 12 months. Results showed greater increases in lutein (52% vs. 6%) and zeaxanthin (83% vs. 0%) in the intervention group compared to the control group, however only serum zeaxanthin concentrations changed significantly.;Since recent studies have suggested that eating 1 or more eggs daily may increase risk of death from cardiovascular disease nearly as much as smoking, the third experiment examined the effects of consuming 12 eggs per week for 1 year on serum concentration of lipids, lipoprotein cholesterol, C-reactive protein (CRP), apolipoprotein (Apo)-A1 and B, and glucose. Results showed greater increases in HDL and LDL, 6.25% and 3.3% respectively, in the intervention group. The intervention group's apolipoprotein Al, C-reactive protein, total cholesterol, and triglycerides levels increased, while apolipoprotein ratio (B/A1) and lipoprotein (a) levels decreased.;Lastly, it is possible that eating a meal or a supplement containing a high concentration of certain individual carotenoids (polar vs nonpolar) may cause a reduction in the other carotenoids in the serum simultaneously. This experiment investigated how a diet of 12 eggs per week for 1 year influenced the serum alpha-cryptoxanthin, beta-cryptoxanthin, lycopene, alpha-carotene, beta-carotene, alpha-tocopherol, and retinol concentrations in the subjects. A group of 45 adults were randomized to intervention (n=27) and control (n=18) groups. Serum carotenoid and vitamin levels were obtained at the beginning and at the end of the 12-months trial period. Results showed a statistically significant 18% increase in alpha-carotene level in the control group, and no significant increase in serum levels of any other carotenoid, tocopherol, or retinol in both control and intervention groups. In addition, beta-carotene levels decreased in control and intervention groups, - 9% and -18% respectively.