Seung-Shi Ham

Antimutagenic effects of subfractions of Chaga mushroom (Inonotus obliquus) extract

Inonotus obliquus is a mushroom commonly known as Chaga that is widely used in folk medicine in Siberia, North America, and North Europe. Here, we evaluated the antimutagenic and antioxidant capacities of subfractions of Inonotus obliquus extract. The ethyl acetate extract was separated by vacuum chromatography into three fractions, and the fraction bearing the highest antimutagenic activity was subsequently separated into four fractions by reversed phase (ODS-C18) column chromatography. The most antimutagenic fraction was then separated into two subfractions (subfractions 1 and 2) by normal phase silica gel column chromatography. Ames test analysis revealed that the subfractions were not mutagenic. At 50 μg/plate, subfractions 1 and 2 strongly inhibited the mutagenesis induced in Salmonella typhimurium strain TA100 by the directly acting mutagen MNNG (0.4 μg/plate) by 80.0% and 77.3%, respectively. They also inhibited 0.15 μg/plate 4NQO-induced mutagenesis in TA98 and TA100 by 52.6-62.0%. The mutagenesis in TA98 induced by the indirectly acting mutagens Trp-P-1 (0.15 μg/plate) and B(α)P (10 μg/plate) was reduced by 47.0-68.2% by the subfractions, while the mutagenesis in TA100 by Trp-P-1 and B(α)P was reduced by 70.5-87.2%. Subfraction 1 was more inhibitory than subfraction 2 with regard to the mutagenic effects of 4NQO, Trp-P-1, and B(α)P. Subfractions 1 and 2 also had a strong antioxidant activity against DPPH radicals and were identified by MS, 1H NMR and 13C NMR analyses as 3β-hydroxy-lanosta-8, 24-dien-21-al and inotodiol, respectively. Thus, we show that the 3beta-hydroxy-lanosta-8, 24-dien-21-al and inotodiol components of Inonotus obliquus bear antimutagenic and antioxidative activities.

Anticancer activity of subfractions containing pure compounds of Chaga mushroom (Inonotus obliquus) extract in human cancer cells and in Balbc/c mice bearing Sarcoma-180 cells

The Chaga mushroom (Inonotus obliquus) has been used in folk medicine to treat cancers. However, limited information exists on the underlying anticancer effects of the major component of I. obliquus in vivo. We hypothesize that the pure compounds (3β-hydroxy-lanosta-8,24-dien-21-al, inotodiol and lanosterol, respectively) separated from I. obliquus would inhibit tumor growth in Balbc/c mice bearing Sarcoma-180 cells (S-180) in vivo and growth of human carcinoma cells in vitro. To test this hypothesis, the growth inhibition of each subfraction isolated from I. obliquus on human carcinoma cell lines (lung carcinoma A-549 cells, stomach adenocarcinoma AGS cells, breast adenocarcinoma MCF-7 cells, and cervical adenocarcinoma HeLa cells) was tested in vitro. Then, after S-180 implantation, the mice were fed a normal chow supplemented with 0, 0.1 or 0.2 mg of subfraction 1, 2 or 3 per mouse per day. All of the subfractions isolated from I. obliquus showed significant cytotoxic activity against the selected cancer cell lines in vitro. Subfraction 1 was more active than subfraction 2 and subfraction 3 against the A549, AGS and MCF-7 cancer cell lines in vitro. In in vivo results, subfraction 1 isolated from I. obliquus at concentrations of 0.1 and 0.2 mg/mouse per day significantly decreased tumor volume by 23.96% and 33.71%, respectively, as compared with the control. Subfractions 2 and 3 also significantly inhibited tumor growth in mice bearing S-180 as compared with the control mouse tumor. Subfraction 1 isolated from I. obliquus showed greater inhibition of tumor growth than subfractions 2 and 3, which agrees well with the in vitro results. The results suggest that I. obliquus and its compounds in these subfractions isolated from I. obliquus could be used as natural anticancer ingredients in the food and/or pharmaceutical industry.

Masou salmon (Oncorhynchus masou) ethanol extract decreases 3-hydroxy-3-methylglutaryl coenzyme A reductase expression in diet-induced obese mice.

This study was designed to evaluate the hypocholesterolemic effects of masou salmon 70% ethanol extract (MSE) and to determine the molecular mechanism by which MSE exerts its effects in high-fat (HF) diet-induced obese mice. We hypothesize that the MSE may contain abundant n-3 fatty acids, so a diet containing MSE may also have hypolipidemic effects by assessing several key gene expressions in cholesterol metabolism such as the low-density lipoprotein (LDL) receptor, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, and cholesterol 7alpha-hydroxylase (CYP7A1). To test this hypothesis, C57BL/6J mice were fed a 40% HF diet for 5 weeks, after which time the animals were fed an HF diet containing 0 mg/kg, 75 mg/kg, or 150 mg/kg MSE (HF, HF + MSE 1, and HF + MSE 2 groups, respectively) for an additional 4 weeks (n = 8 in each group, for a total of 24 mice). We found that feeding MSE with an HF diet prevented hypercholesterolemia in diet-induced obese mice; daily MSE feeding reduced total cholesterol levels in plasma and liver by 12.3% and 16.2%, respectively. Furthermore, we examined the expression of key cholesterol metabolism genes by reverse transcription-polymerase chain reaction and found that messenger RNA levels of HMG-CoA reductase were decreased by up to 5-fold, but the expression of both LDL receptor and CYP7A1 did not change. Thus, MSE may exert its hypocholesterolemic effect by altering the expression of HMG-CoA reductase.

Platycodon grandiflorum root attenuates vascular endothelial cell injury by oxidized low-density lipoprotein and prevents high-fat diet–induced dyslipidemia in mice by up-regulating antioxidant proteins

We hypothesized that a Platycodon grandiflorum root (PG) ethyl acetate extract (PGEA) would help reduce the vascular cell injury caused by oxidized low-density lipoprotein (oxLDL) and prevent high-fat (HF) diet-induced dyslipidemia and oxidative stress by up-regulating antioxidant proteins. We investigated the protective effects of PGEA against vascular endothelial cell injury induced by oxLDL and dyslipidemia induced by an HF diet, and the mechanisms underlying these effects were studied. The protective effects of PGEA were investigated with respect to calf pulmonary arterial endothelial (CPAE) cell viability and the lactate dehydrogenase release during oxLDL treatment. The in vivo effects of PGEA were examined using C57BL/6 mice, which were fed an HF diet for 9 weeks. The HF diet was supplemented with 0, 25, or 75 mg/kg PGEA during the last 4 weeks of the experimental period. Histologic analyses of hepatic lipid accumulation were performed. The changes in antioxidant protein levels induced by PGEA, which protects against HF diet-induced oxidative stress, were measured using a proteomics approach. We found that PGEA exhibited antioxidant activity. In CPAE cells, PGEA inhibited both oxLDL-induced cell death and lactate dehydrogenase release. In the HF diet-induced obese mice that received PGEA, we observed significantly reduced plasma and hepatic lipid levels, demonstrating that PGEA has beneficial effects on hyperlipidemia. In addition, we found that PGEA caused the up-regulation of antioxidant proteins. These findings suggest that the antioxidant effects of PGEA may protect against oxidative stress-related diseases.

Antigenotoxicity of Enzymatic Browning Reaction Products of Potatoes in the Micronucleus Test

This study investigated the antigenotoxic effects of enzymatic browning reaction products (PEBRPs) obtained by reaction of polyphenol compounds with oxidase extracted from potato. Each of the PEBRPs by themselves at 100 mg/kg did not induce an increased frequency of micronucleated polychromatic erythrocytes (MNPCEs) irrespective of the sampling time (up to 72 h), while the treatment with benzo[a]pyrene (B[a]P) significantly increased the incidence of MNPCEs (P < 0.05). Significant reductions were observed in the frequencies of MNPCEs (P < 0.05) when all PEBRPs were given to the mice 12 h before they were exposed to 100 mg/kg of B[a]P and inhibitory effects were 60%, 70%, and 60% in the catechol (Ca)-PEBRPs, hydroxyhydroquinone (HHQ)-PEBRPs, and pyrogallol (Py)-PEBRPs, respectively. When three kinds of PEBRPs were fed to mice 12 h before injecting 100 mg/kg of B[a]P, the most significant decrease (P < 0.05) in the frequencies of MNPCEs induced by B[a]P were observed and the relative frequency inhibitions by Ca-PEBRPs, HHQ-PEBRPs, and Py-PEBRPs were 70%, 70%, and 60%, respectively. Also, when each type of PEBRP was given to mice one time every day for 5 days, significant reductions were observed in the frequencies of MNPCEs induced by B[a]P (P < 0.05). The strongest relative frequency inhibitions were 60% and 70%, respectively, at 200 mg/kg for Ca-PEBRPs and HHQ-PEBRPs, but Py-PEBRPs had their strongest inhibitory effect at a concentration of 100 mg/kg. These results indicates that enzymatic browning reaction products of potatoes have a strong modulatory effect on B[a]P-induced MNPCEs.