Citrus limonoids and flavonoids: extraction, antioxidant activity and effects on hamster plasma cholesterol distribution

Four in vitro models were used to measure the antioxidant activity of 11 citrus phytochemicals. The citrus limonoids and bergapten showed very weak antioxidant activity. The flavonoids demonstrated mild, to moderate, to strong antioxidant activity. In addition to some other commonly accepted structural features our data indicated that the hydroxyl group in position 6 of ring A could also increase the antioxidant activity of flavonoids. Compared with the active flavonoids, limonoids are highly oxygenated triterpenoids, with fewer hydroxyl groups to stabilize unpaired electrons (or scavenge free radicals). Bergapten lacks a hydroxyl group. This is the first report on the antioxidant activity of limonoids and neoeriocitrin.

   A feeding study using Syrian hamsters was followed to determine the effect of citrus limonoids and flavonoids on plasma cholesterol. Hamsters fed with limonin, limonin 17-Beta-D-glucopyranoside and grapefruit pulp significantly inhibited the increase of LDL/HDL-cholesterol (36.6%, 52.9% and 57% respectively) compared with the basal control (65.8%) and the pectin control (70%). Furthermore, hamsters fed with limonin had significantly larger LDL particle size (21.21 nm) compared with the control group (19.96 nm). Further studies demonstrated that LDLs from hamsters fed with limonin and limonin 17-Beta-D-glucopyranoside were less susceptible to oxidation. These data suggest that limonin, limonin 17-Beta-D-glucopyranoside and grapefruit pulp have potential inhibitory effects against atherogenesis.

    Supercritical CO2 (SC-CO2) was attempted to extract limonoids from grapefruit seeds and molasses. Limonin aglycone was successfully extracted with SC-CO2 directly from grapefruit seeds with the yield of 6.3 mg/g seeds at 48.3 MPa, 50˚C and 60 min with CO2 top feeding; and the limonin glucoside was extracted using SC-CO2 and ethanol as co-solvent from the defatted seeds with the yield of 0.73 mg/g seeds at 42 MPa, 52˚C, 45% ethanol (XEth=0.45) and 40 min with CO2 top feeding; and limonin glucoside also was extracted using SC-CO2 and ethanol with the yield of 0.61mg/g grapefruit molasses at 48.3 MPa, 50˚C and 10% ethanol (XEth=0.1), 40 min with CO2 top feeding. CO2 flow rate was around~5 l/min in experiments. The results demonstrated SC-CO2 extraction of limonoids from citrus juice industry byproducts has practical significance for future commercial production.