Green synthesis approach: extraction of chitosan from fungus mycelia.
Chitosan, a natural product derived from chitin, possesses hypocholesterolemic properties similar to those of cholestyramine, but there has been no report concerning its effects on the equilibrium between dietary cholesterol and de novo cholesterol synthesis in the liver. In this work, we studied the effects of chitosan on plasma and liver cholesterol levels, liver weight, and the key regulatory enzyme of cholesterogenesis 3-hydroxy-3-methylglutaryl coenzyme-A (HMG-CoA) reductase in rats fed a sterol diet containing 1% cholesterol and 0.2% cholic acid. The animals given the sterol diet showed increases in plasma and liver cholesterol, which were lowered by 54% in plasma and 64% in liver by 5% chitosan, while cholestyramine completely blocked such increases. HMG-CoA reductase activity was considerably increased in the sterol-cholestyramine group, but was greatly decreased in both sterol and sterol-chitosan groups. There was no change in liver weight or appearance after treatment with chitosan, but cholestyramine-treated animals manifested secondary effects from the treatment, including smaller yellowish livers. High mol wt chitosans [> 750 kilodaltons (kDa)] were found to be less effective as hypocholesterolemia than a 70-kDa preparation. Also, when the 70-kDa chitosan was used at 2.5%, 5%, and 7.5% of the total diet, its effectiveness was greatest at the higher concentrations; indeed, incorporation of 7.5% chitosan in the sterol diet for 3 weeks completely prevented any decrease in plasma high density lipoprotein cholesterol or increase in the plasma cholesterol level and liver weight. This formula greatly reduced the increase in liver cholesterol content due to the sterol diet, with values of 8.8 +/- 1.3 for the sterol-chitosan diet vs. 18.2 +/- 0.8 mg/g tissue for the sterol diet. The increased intake of sterols considerably lowered both HMG-CoA reductase activity (33-fold) and HMG-CoA reductase mRNA levels (3-fold) in rat liver, but in the sterol-chitosan group, HMG-CoA reductase activity was 7.7 times more elevated than in the sterol group, although it was still lower than the control value, whereas HMG-CoA reductase mRNA levels were normal. The results obtained did not differ significantly when rats were studied for 1, 3, or 6 weeks. These results taken collectively indicate that the 7.5% chitosan formula maintained adequate cholesterol homeostasis in rats, despite a greatly increased intake of cholesterol.