Gossypetin Prevents the Progression of Nonalcoholic Steatohepatitis by Regulating Oxidative Stress and AMP-Activated Protein Kinase.

Nonalcoholic steatohepatitis (NASH) is a severe liver metabolic disorder, however, there are still no effective and safe drugs for its treatment. Previous clinical trials used various therapeutic approaches to target individual pathologic mechanisms, but these approaches were unsuccessful because of the complex pathologic causes of NASH. Combinatory therapy in which two or more drugs are administered simultaneously to patients with NASH, however, carries the risk of side effects associated with each individual drug. To solve this problem, we identified gossypetin as an effective dual-targeting agent that activates AMP-activated protein kinase (AMPK) and decreases oxidative stress. Administration of gossypetin decreased hepatic steatosis, lobular inflammation and liver fibrosis in the liver tissue of mice with choline-deficient high-fat diet and methionine-choline deficient diet (MCD) diet-induced NASH. Gossypetin functioned directly as an antioxidant agent, decreasing hydrogen peroxide and palmitate-induced oxidative stress in the AML12 cells and liver tissue of MCD diet-fed mice without regulating the antioxidant response factors. In addition, gossypetin acted as a novel AMPK activator by binding to the allosteric drug and metabolite site, which stabilizes the activated structure of AMPK. Our findings demonstrate that gossypetin has the potential to serve as a novel therapeutic agent for nonalcoholic fatty liver disease /NASH. SIGNIFICANCE STATEMENT: This study demonstrates that gossypetin has preventive effect to progression of nonalcoholic steatohepatitis (NASH) as a novel AMP-activated protein kinase (AMPK) activator and antioxidants. Our findings indicate that simultaneous activation of AMPK and oxidative stress using gossypetin has the potential to serve as a novel therapeutic approach for nonalcoholic fatty liver disease /NASH patients.

Enhanced bioavailability and hepatoprotective effect of silymarin by preparing silymarin-loaded solid dispersion formulation using freeze-drying method.

This study aimed to develop a solid dispersion formulation of silymarin (Silymarin-SD) using freeze-drying method to enhance its oral bioavailability (BA) by inhibiting the intestinal first-pass effect and increasing its solubility and permeability. Silymarin-SD formulation (i.e., silymarin:tween 80:hydroxypropyl cellulose (HPC) = 1:1:3 (w/w/w) significantly increased silymarin permeability in the duodenum, jejunum, and ileum by decreasing the efflux ratio of silymarin and by inhibiting silymarin-glucuronidation activity, in which tween 80 played a crucial role. As a result, orally administered Silymarin-SD formulation increased plasma silymarin concentrations and decreased silymarin-glucuronide in rats compared with silymarin alone and silymmarin:D-α-tocopherol polyethylene glycol 1000 succinate (1:1, w/w) formulation. In addition to modulating intestinal first-pass effect, Silymarin-SD formulation showed a significantly higher cumulative dissolution for 120 min compared with that of silymarin from the physical mixture (PM) of the same composition as Silymarin-SD and silymarin alone; the relative BA of silymarin-SD increased to 215% and 589% compared with silymarin-PM and silymarin alone, respectively. This could be attributed to the amorphous status of the Silymarin-SD formulation without chemical interaction with excipients, such as tween 80 and HPC. Moreover, the hepatoprotective effect of Silymarin-SD in acetaminophen-induced acute hepatotoxicity, as estimated from the alanine aminotransferase and aspartate aminotransferase values, was superior to that of silymarin. In conclusion, the increase in the dissolution rate and intestinal permeability of silymarin, and the inhibition of silymarin-glucuronidation by the Silymarin-SD formulation, prepared using tween 80 and HPC, increased its plasma concentration and resulted in a superior hepatoprotective effect compared to silymarin.

Concomitant Administration of Red Ginseng Extract with Lactic Acid Bacteria Increases the Plasma Concentration of Deglycosylated Ginsenosides in Healthy Human Subjects.

With the increased frequency of red ginseng extract (RGE) and lactic acid bacteria (LAB) co-administration, we aimed to investigate the interactions between RGE and LAB with regard to in vitro and in vivo deglycosylation metabolism and the pharmacokinetics of ginsenosides. As a proof-of-concept study, five healthy humans were administered RGE (104.1 mg of total ginsenosides/day) with or without co-administration of LAB (2 g, 1 billion CFU/day) for 2 weeks, and the plasma concentrations of ginsenosides in human plasma were monitored. The plasma exposure to compound K (CK), ginsenoside Rh2 (GRh2), protopanaxadiol (PPD), and protopanaxatriol (PPT) in the concomitant administration RGE and LAB groups increased by 2.7-, 2.1-, 1.6-, and 3.5-fold, respectively, compared to those in the RGE administration group, without a significant change in Tmax. The plasma concentrations of GRb1, GRb2, and GRc remained unchanged, whereas the AUC values of GRd and GRg3 significantly decreased in the concomitant administration RGE and LAB groups. To understand the underlying mechanism, the in vitro metabolic activity of ginsenosides was measured during the fermentation of RGE or individual ginsenosides in the presence of LAB for 1 week. Consistent with the in vivo results, co-incubation with RGE and LAB significantly increased the formation rate of GRh2, CK, PPD, and PPT. These results may be attributed to the facilitated deglycosylation of GRd and GRg3 and the increased production of GRh2, CK, PPD, and PPT by the co-administration of LAB and RGE. In conclusion, LAB supplementation increased the plasma concentrations of deglycosylated ginsenosides, such as GRh2, CK, PPD, and PPT, through facilitated deglycosylation metabolism of ginsenosides in the intestine.

Simultaneous Analysis of a Combination of Anti-Hypertensive Drugs, Fimasartan, Amlodipine, and Hydrochlorothiazide, in Rats Using LC-MS/MS and Subsequent Application to Pharmacokinetic Drug Interaction with Red Ginseng Extract.

Fimasartan, amlodipine, and hydrochlorothiazide are commonly used in combination therapies as antihypertensive drugs. This study aimed to develop and validate an analytical method for fimasartan, its active and major metabolite fimasartan-amide, amlodipine, and hydrochlorothiazide in rat plasma using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The standard calibration curves for fimasartan (1−500 ng/mL), its active and major metabolite fimasartan-amide (0.3−100 ng/mL), amlodipine (0.5−200 ng/mL), and hydrochlorothiazide (5−5000 ng/mL) were linear with R2 > 0.9964, and the inter- and intra-day accuracy and precision and stability were within the acceptable criteria. Using this validated analytical method, the pharmacokinetic interaction of these triple combination drugs between single administration and concomitant administration of the triple combination was investigated; the results did not reveal a significant difference in any of the pharmacokinetic parameters. Based on these results, we investigated the effects of red ginseng extract (RGE) on the pharmacokinetics of fimasartan, fimasartan-amide, amlodipine, and hydrochlorothiazide after oral administration of the combination in rats. No significant difference was observed in the pharmacokinetic parameters of fimasartan, fimasartan-amide, amlodipine, and hydrochlorothiazide, except for the Tmax values of amlodipine. The delayed Tmax value of amlodipine was attributed to its decreased intestinal permeability after repeated RGE treatments. In conclusion, using a combination of antihypertensive drugs and simultaneous analytical methods, we established efficient drug interaction and toxicokinetic studies using a small number of animals.

Effect of Lactic Acid Bacteria on the Pharmacokinetics and Metabolism of Ginsenosides in Mice.

This study aims to investigate the effect of lactic acid bacteria (LAB) on in vitro and in vivo metabolism and the pharmacokinetics of ginsenosides in mice. When the in vitro fermentation test of RGE with LAB was carried out, protopanaxadiol (PPD) and protopanaxadiol (PPD), which are final metabolites of ginsenosides but not contained in RGE, were greatly increased. Compound K (CK), ginsenoside Rh1 (GRh1), and GRg3 also increased by about 30%. Other ginsenosides with a sugar number of more than 2 showed a gradual decrease by fermentation with LAB for 7 days, suggesting the involvement of LAB in the deglycosylation of ginsenosides. Incubation of single ginsenoside with LAB produced GRg3, CK, and PPD with the highest formation rate and GRd, GRh2, and GF with the lower rate among PPD-type ginsenosides. Among PPT-type ginsenosides, GRh1 and PPT had the highest formation rate. The amoxicillin pretreatment (20 mg/kg/day, twice a day for 3 days) resulted in a significant decrease in the fecal recovery of CK, PPD, and PPT through the blockade of deglycosylation of ginsenosides after single oral administrations of RGE (2 g/kg) in mice. The plasma concentrations of CK, PPD, and PPT were not detectable without change in GRb1, GRb2, and GRc in this group. LAB supplementation (1 billion CFU/2 g/kg/day for 1 week) after the amoxicillin treatment in mice restored the ginsenoside metabolism and the plasma concentrations of ginsenosides to the control level. In conclusion, the alterations in the gut microbiota environment could change the ginsenoside metabolism and plasma concentrations of ginsenosides. Therefore, the supplementation of LAB with oral administrations of RGE would help increase plasma concentrations of deglycosylated ginsenosides such as CK, PPD, and PPT.