Possible mechanisms for antioxidant activity of glycyrrhizinic acid

Possible mechanisms of antioxidant activity of glycyrrhizinic acid (GA) were studied. GA did not exhibit antiradical properties in the experiments with stable radical 1,1-diphenyl-2-picrylhydrazyl at the concentration range of 1–100 μM. These data were confirmed by the study of GA effect on luminol chemiluminescence in a cell-free system in the presence of hydrogen peroxide. At the same time, GA decreases (in a dose-dependent manner) the generation of reactive oxygen species by neutrophils activated with both phorbol 12-myristate 13-acetate (PMA) and the chemotactic peptide, N-formyl-Met-Leu-Phe (FMLP). Using dichlorodihydrofluorescein (DCF) fluorescence it has been demonstrated that direct addition of GA to neuron culture did not decrease the level of free radical formation. However, preincubation of cells with GA resulted in the decrease in free radical production rate and increase in reduced intracellular glutathione level.     

Effect of mevinolin and glycyrrhizinic acid on cholesterol and bile acid metabolism in cultured rabbit hepatocytes]

A comparison of effects of two hypocholesterolemic drugs--mevinolin and glycyrrhizinic acid, on cholesterol and bile acid metabolism in cultured rabbit hepatocytes has been carried out. The following parameters have been determined: i) cholesterol synthesis from [2-14C]acetate; ii) bile acid production from newly synthesized and [4-14C]-labeled HDL2 cholesterol, and, iii) total cholesterol efflux into the incubation medium Mevinolin (0.5 microgram/ml) inhibited [2-14C] acetate incorporation into cholesterol by more than 90%. Conversely, glycyrrhizinic acid did not influence cholesterol synthesis even when used at high (100 micrograms/ml) concentrations but stimulated the conversion of endogenous (by 37%) and exogenous (by 18%) cholesterol into bile acids and increased, in addition, the proportion of bile acids in the total sterol pool released from hepatocytes into the incubation medium. At the same time, mevinolin used at 0.5 microgram/ml decreased the bile acid production by endogenous (by 27%) and exogenous (by 40%) cholesterol. The data obtained suggest that glycyrrhizinic acid exerts hypocholesterolemic action by stimulation of cholesterol conversion into bile acids without any effect on cholesterol synthesis. As for mevinolin, it has a cholesterol-suppressing effect via a mechanism of cholesterol synthesis inhibition only.     

Interaction of glycyrrhizin and glycyrrhetinic acid with DNA

Glycyrrhizin (GL), a molecule of glycyrrhetinic acid (GA), is an aqueous extract from licorice root. These compounds are well known for their anti-inflammatory, hepatocarcinogenesis, antiviral, and interferon-inducing activities. This study is the first attempt to investigate the binding of GL and GA with DNA. The effect of ligand complexation on DNA aggregation and condensation was investigated in aqueous solution at physiological conditions, using constant DNA concentration (6.25?mM) and various ligands/polynucleotide (phosphate) ratios of 1/240, 1/120, 1/80, 1/40, 1/20, 1/10, 1/5, 1/2, and 1/1. Fourier transform infrared and ultraviolet (UV)-visible spectroscopic methods were used to determine the ligand binding modes, the binding constants, and the stability of ligand-DNA complexes in aqueous solution. Spectroscopic evidence showed that GL and GA bind DNA via major and minor grooves as well as the backbone phosphate group with overall binding constants of K(GL-DNA)=5.7×10(3) M(-1), K(GA-DNA)=5.1×10(3) M(-1). The affinity of ligand-DNA binding is in the order of GL>GA. DNA remained in the B-family structure, whereas biopolymer aggregation occurred at high triterpenoid concentrations.     

Determination of uric acid in serum using isotachophoresis

An operational system is described for the isotachophoretic determination of uric acid in serum, making use of column coupling. The method has been compared with a standard enzymatic procedure. With the present technique small amounts of serum (ca. 3 μl) can be applied without any pretreatment. Urate recovery was 99.0–100.5%. Under the non-physiological measuring conditions used, 12–28% of control serum uric acid was bound to macromolecules of molecular weight exceeding 25,000. The day-to-day variations of the isotachophoretic procedure were smaller than those of the enzymatic method, whereas standard deviations were comparable. The isotachophoretic procedure is less influenced by certain metabolites.     

Synthesis and immunomodulating activity of new diglycopeptides of of glycyrrhizinic acid and it's 30-methyl ester

New amino acid derivatives of glycyrrhizic acid and its methyl ester were selectively synthesized using active N-succinimide esters. The compounds with residues of glycine ethyl ester and alanine methyl and butyl esters increased the level of agglutinins and hemolysins in blood serum of mice two- to threefold in comparison with the control upon parenteral administration at a dose of 2 mg/kg for 14 days. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2004, vol. 30, no. 2; see also http://www.maik.ru.     

Determination of salicylic acid in human serum with capillary zone electrophoresis

The determination of salicylic acid (SA), a metabolite of aspirin, in human serum was developed using capillary zone electrophoresis (CZE) with diode array detection. The reproducibility of separation and quantification with CZE analysis of the extract of SA from human serum was appropriate for the intra- and inter-day assay coefficients. A high correlation was revealed between the serum SA levels in volunteers determined by CZE and those determined by a fluorescence polarization immunoassay (r=0.973, n=12), although the former values were slightly higher than the latter. There were no peaks interfering with the assay of SA by internal standard method. This CZE method could provide a simple and efficient method for monitoring SA in patients.     

东北虎幼体血清蛋白及胆汁酸的测定与分析

用HITACHI 7600-20全自动生化分析仪对13只东北虎幼体进行了总蛋白、白蛋白、球蛋白、前白蛋白及胆汁酸5项生化指标的测定,实验数据经SPSS软件统计分析.其结果为:总蛋白含量66.06±6.80 g/L,白蛋白含量34.19±2.51 g/L,球蛋白含量31.89±4.46 g/L,前白蛋白含量13.22±11.64 g/L,胆汁酸含量4.48±3.89 μmol/L.东北虎幼体雌雄个体间5项生化指标均无显著差异(P0.05).东北虎幼体与成体间,总蛋白、白蛋白、球蛋白含量均呈极显著差异(P<0.01).东北虎幼体与华南虎幼体间,总蛋白、球蛋白含量呈显著差异(P<0.05);白蛋白含量无显著差异(P0.05).     

Profound difference of metabolic pharmacokinetics between pure glycyrrhizin and glycyrrhizin in licorice decoction

To investigate the difference of metabolic pharmacokinetics between pure glycyrrhizin (GZ) and GZ in licorice decoction, six New Zealand White rabbits were orally given pure GZ and licorice decoction containing equivalent content of GZ in a randomized crossover design. HPLC methods were used for the quantitation of GZ and glycyrrhetic acid (GA) in serum. The results indicated that the areas under curves (AUCs) of GZ and GA after administration of licorice decoction were significantly higher than those after pure GZ. This result was contradictory with that obtained in rats. To explore the mechanism of the pharmacokinetic difference, feces of rabbits, rats, pigs and humans were used to investigate the presystemic metabolism of pure GZ and GZ in licorice decoction. The results indicated that pure GZ was hydrolyzed to GA more rapidly and to a greater extent than that in licorice decoction by various feces. In addition, when pure GZ was fermented, the metabolic profiles of GA and 3-dehydroGA in rabbit feces were quite different from other feces. In conclusion, the bioavailabilities of GZ and GA are significantly better from licorice than from pure GZ in rabbits but the presystemic metabolism of pure GZ in rabbit is rather different from that in rat, pig and human.     

Determination of serum oxalate using peroxyoxalate chemiluminescence of free oxalic acid

We describe a new sensitive and specific method for determination of oxalate in human serum. By using the chemiluminescence decay of monoperoxyoxalic acid very low concentrations of oxalate (200 nmol/L) can be determined. The mean serum oxalate level in apparently healthy controls was 14.5 ± 8.5 m?mol/L. Supplementation of ascorbic acid leads to an increase in serum oxalate level. While serum oxalate concentrations of calcium oxalate stone formers (x = 16.4 ± 9.8 m?mol/L) are not significantly different from the control group, an extreme increase of serum oxalate is evident in haemodialysis patients. The serum oxalate concentration decreased during dialysis treatment from 141.4 ± 32.1 m?mol/L to 36.4 ± 12.7 m?mol/L.     

Determination of uric acid in human urine and serum by capillary electrophoresis with chemiluminescence detection

A simple and sensitive method based on capillary electrophoresis (CE) with chemiluminescence (CL) detection has been developed for the determination of uric acid (UA). The sensitive detection was based on the enhancement effect of UA on the CL reaction between luminol and potassium ferricyanide (K₃[Fe(CN)₆]) in alkaline solution. A laboratory-built reaction flow cell and a photon counter were deployed for the CL detection. Experimental conditions for CL detection were studied in detail to achieve a maximum assay sensitivity. Optimal conditions were found to be 1.0 × 10⁻⁴ M luminol added to the CE running buffer and 1.0 × 10⁻⁴ M K₃[Fe(CN)₆] in 0.2 M NaOH solution introduced postcolumn. The proposed CE-CL assay showed good repeatability (relative standard deviation [RSD] = 3.5%, n = 11) and a detection limit of 3.5 × 10⁻⁷ M UA (signal/noise ratio [S/N] = 3). A linear calibration curve ranging from 6.0 × 10⁻⁷ to 3.0 × 10⁻⁵ M UA was obtained. The method was evaluated by quantifying UA in human urine and serum samples with satisfactory assay results.     

Determination of 3 beta-hydroxy-5-cholenoic acid in serum of hepatobiliary diseases--its glucuronidated and sulfated conjugates

3 beta-Hydroxy-5-cholenoic acid in the serum of control subjects and 62 patients with various hepatobiliary diseases was quantitated by mass fragmentography after separation into nonglucuronidated-nonsulfated, glucuronidated, and sulfated fractions. Deuterium-labeled deoxycholic acid and its glucuronide and sulfate were used as internal standards. Mean concentrations of total 3 beta-hydroxy-5-cholenoic acid in serum (mumole/liter) were as follows: Control subjects (14), 0.184; obstructive jaundice (15), 6.783; liver cirrhosis, compensated (12), 0.433, and decompensated (12), 1.636; chronic hepatitis (12), 0.241; and acute hepatitis (11), 2.364. Most of the 3 beta-hydroxy-5-cholenoic acid was glucuronidated or sulfated. Only in patients with obstructive jaundice did glucuronidation (60 +/- 14%) exceed sulfation (31 +/- 14%), sulfation exceeding glucuronidation in the others. The UDP-glucuronyltransferase might have different substrate specificities for 3 beta-hydroxy-5-cholenoic acid and other common bile acids, especially in the cholestatic state.     

Determination of lactic acid and pyruvic acid in serum and cerebrospinal fluid by ion-exclusion chromatography with a bulk acoustic wave detector

A chromatographic method based on a combination of ion-exclusion chromatography separation and bulk acoustic wave series piezoelectric quartz crystal detector quantification for the determination of pyruvic acid and lactic acid in serum and cerebrospinal fluid (CSF) was developed. The separation was carried out using a Shim-pak SCR-102H ion-exclusion column with phosphoric acid solution as eluent. The method shows an acceptable detection limit and anti-interference ability. Serum and CSF from healthy individuals and patients were analysed successfully.     

Licorice compounds glycyrrhizin and 18beta-glycyrrhetinic acid are potent modulators of bile acid-induced cytotoxicity in rat hepatocytes

The accumulation of hydrophobic bile acids results in cholestatic liver injury by increasing oxidative stress, mitochondrial dysfunction, and activation of cell signaling pathways. Licorice root and its constituents have been utilized as antihepatotoxic agents. The purpose of this study was to evaluate the potential modulation by a primary component of licorice root, glycyrrhizin (GL), and its metabolite, 18beta-glycyrrhetinic acid (GA), in a hepatocyte model of cholestatic liver injury. Preincubation of fresh rat hepatocyte suspensions with GL or GA reduced glycochenodeoxycholic acid (GCDC)-dependent reactive oxygen species generation, with GA more potent than GL. Interestingly, GL and GA had opposing effects toward GCDC-induced cytotoxicity; GA prevented both necrosis and apoptosis, whereas GL enhanced apoptosis. GCDC promoted activation of caspase 10, caspase 3, and PARP; all were inhibited by GA but not GL. Induction of apoptosis by GCDC was also associated with activation of JNK, which was prevented by GA. Activation of caspase 9 and dissipation of mitochondrial membrane potential were prevented by GA but not GL. In liver mitochondrial studies, GL and GA were both potent inhibitors of the mitochondrial permeability transition, reactive oxygen species generation, and cytochrome c release at submicromolar concentrations. Results from this study suggest that GL exhibits pro-apoptotic properties, whereas GA is a potent inhibitor of bile acid-induced apoptosis and necrosis in a manner consistent with its antioxidative effect.