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Title
The effects of (-)-epigallocatechin-3-O-gallate (EGCG), a green tea catechin, on blood cholesterol
Author/Creator
Naumovski, Nenad
Institution
University of Newcastle. Faculty of Science and Information Technology, School of Environmental and Life Sciences
Description
Research Doctorate - Doctor of Philosophy (PhD)
Description
Background: The catechin, (-)-epigallocatechin-3-O-gallate (EGCG), the most abundant compound in green tea, has been linked to numerous beneficial health effects, including a reduction in blood cholesterol levels and protection against cardiovascular disease. Previous studies, mostly with extracts of green tea containing mixtures of catechins and other compounds, in animal models of hypercholesterolaemia including the cholesterol-fed rabbit, have shown that these preparations can lower blood cholesterol. Three plausible mechanisms by which they could lower cholesterol have been postulated: 1) an increase in the LDL-receptor and 2) a reduction in cholesterol synthesis postulated from studies in the hypercholesterolaemic rabbit model and in cultured human HepG2 liver cells and 3) an inhibition of intestinal cholesterol absorption postulated from studies in mice, rats and hamsters. However, it is not known whether EGCG, as a pure compound, can lower cholesterol in the rabbit model and in humans and whether it works through the three postulated mechanisms of action. Hypotheses and Aims: The working hypothesis for this thesis was that ‘pure EGCG will lower cholesterol in hypercholesterolaemic rabbits and humans’. Therefore, the aim was to determine the effect of pure EGCG on cholesterol in hypercholesterolaemic rabbits and in humans with moderately elevated cholesterol. For an experiment with the hypercholesterolaemic rabbits, it was hypothesised that ‘pure EGCG will lower cholesterol by up-regulating the LDL-receptor, reducing cholesterol synthesis and inhibiting the intestinal cholesterol absorption’. Therefore, the aims were to determine whether pure EGCG could lower cholesterol in this animal model and by which mechanisms of action. An absorption study was then conducted in humans for which it was hypothesised that ‘pure EGCG will be absorbed better when given in capsule form without food compared to given in capsule form with a breakfast. It was also hypothesised that ‘incorporating the EGCG in a strawberry sorbet will improve the EGCG absorption compared to taking EGCG in capsule form with a breakfast’. Therefore, the aim was to determine, which of the three EGCG delivery formats, was the best for maximising the systemic absorption of the catechins in humans. Finally, a pilot intervention study in humans was conducted for which it was hypothesised that ‘pure EGCG will lower cholesterol in mildly hypercholesterolaemic humans’. Therefore, the aim was to determine whether pure EGCG, given by the best of the three methods of delivery tested in the absorption study, could lower cholesterol in humans with moderately elevated cholesterol. Methodology: For the animal model study, 12 New Zealand white rabbits were made hypercholesterolaemic by feeding with 0.25% (w/w) cholesterol for two weeks. Then, for four weeks, one group (6) was fed 0.25% (w/w) cholesterol and 2% (w/w) EGCG and the control group (6) was fed 0.25% (w/w) cholesterol only. Blood and faecal samples were collected prior to and at the end of the treatment period. Liver samples were also collected at the end of the study. Among other measurements, blood cholesterol, lathosterol, and squalene, LDL cholesterol, hepatic LDL receptor and CD36 protein and faecal neutral sterols and bile acids were determined. For the absorption of EGCG study, 4 human subjects ingested on three separate occasions after fasting overnight and in random order, 500mg of EGCG taken either in capsule form with 1) water only or 2) a breakfast cereal and milk or 3) incorporated in 200g of a strawberry sorbet. Venous blood samples were taken before ingestion and after 0.5, 1, 2, 3, 5 and 8 hours. The plasma concentration of EGCG was analysed by HPLC-MS and the area under the concentration-time curve (AUC) and other pharmacokinetic parameters were determined. For the human pilot EGCG intervention study, 10 volunteers (6 males and 4 females) with moderate hypercholesterolaemia (5.5-7.5 mmol/l) were recruited for a placebo-controlled, double-blind, parallel design study. After a 2-week baseline period, the subjects were given EGCG or gelatine (placebo) for 4 weeks. Venous blood samples were collected on day 0 and day 28. Plasma cholesterol and triglycerides and HDL cholesterol were analysed using commercially available kits and LDL cholesterol was calculated using the Friedewald equation. Outcomes: In the animal study, the hypotheses were mostly supported in that there was a 85% reduction in serum cholesterol and a 92% reduction in LDL cholesterol in the EGCG group at the end of the treatment period compared to control (p<0.05). The lathosterol to squalene ratio, an index of cholesterol synthesis, was also significantly lower (p=0.03) in the EGCG group (0.20±0.02) compared to control (0.62±0.16). After the 4-week treatment period, the hepatic LDL- receptor was significantly increased (+59%, p<0.001) as well as the hepatic CD36 protein (+62%, p=0.002) in the 2%EGCG group. However, there were no significant differences in faecal sterol excretion between the two groups. In the EGCG absorption study, the first hypothesis was supported in that the plasma EGCG concentrations were markedly higher over the 8-hour period (AUC) when taken without food (174±68) than when EGCG capsules were taken with a breakfast cereal and milk (64±53, p=0.044) or when the EGCG was taken incorporated in a strawberry sorbet (44±23, p=0.019). However, the second hypothesis was not supported in that there was no significant difference between the EGCG capsules taken with a breakfast or taken incorporated within a strawberry sorbet. Therefore, for the pilot study, 1g/day EGCG was given in two doses of 500mg EGCG, one at least 30min before the morning meal and the other 30min before the evening meal. In this pilot study, the hypothesis was partially supported in that the LDL cholesterol was 25% lower (p=0.026) in the EGCG group than in the control group at the end of the 4-week treatment period, although the plasma total cholesterol was not different between the two groups. Conclusions: Pure EGCG exhibited strong cholesterol lowering properties in the cholesterol-fed rabbit model of hypercholesterolaemia by increasing the hepatic LDL-receptor and possibly by reducing cholesterol synthesis but not by increasing the faecal excretion of neutral or acidic sterols. The systemic absorption of EGCG in healthy human subjects was highest when the catechin was taken in capsule form without food on an empty stomach. Finally, in a 4-week pilot study in 10 subjects with moderate hypercholesterolaemia, 1g/day EGCG, given in capsule form without food, resulted in a 25% lower LDL cholesterol concentration compared to control. The human pilot study also showed that amount of EGCG given over the 4-week period was well tolerated, as no serious adverse effects were noted.
Relation
University of Newcastle Research Higher Degree Thesis
Date
2012
Keyword(s)
EGCG;  cholesterol;  green tea catechin;  human studies;  animal studies;  EGCG absorption in humans
Resource Type
thesis
Rights
Copyright 2012 Nenad Naumovski
Identifier
http://hdl.handle.net/1959.13/930758
Language
eng
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"The effects of (-)-epigallocatechin-3-O-gallate (EGCG), a green tea catechin, on blood cholesterol"
 
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