RESUMEN
The objective of this paper was to develop a preparative method for the isolation and purification of liquiritigenin and glycyrrhetic acid from Glycyrrhiza uralensis Fisch using hydrolytic extraction combined with high-speed countercurrent chromatography (HSCCC). Liquiritigenin and glycyrrhetic acid were well hydrolyzed from liquiritin and glycyrrhizic acid by hydrochloric acid, respectively. The optimal extraction conditions were obtained by single-factor and orthogonal experiments, which were 100% ethanol, 1.5 mol/L hydrochloric acid, 1:25 ratio of solid to liquid, and extracted 2 h for one time. Using the two-phase solvent system of n-hexane-ethyl acetate-methanol-water (4:5:4:5, v/v), 2.1 mg liquiritigenin (the purity was 96.5% with a recovery of 87.6%) and 12.3 mg glycyrrhetic acid (the purity was 97.1% with a recovery of 74.4%) were obtained from 315-mg crude extraction by HSCCC. The retention ratio of stationary phase was 47.2%. Their structures were identified by HPLC, melting points, UV, Fourier-transform infrared, Electrospray ionization-MS, 1 H nuclear magnetic resonance (NMR), and 13 C NMR spectra. According to the antioxidant activity assays, liquiritigenin and glycyrrhetic acid had some scavenging abilities on 1,1-diphenyl-2-picrylhydrazyl free radicals; liquiritigenin had stronger scavenging ability on hydroxyl radicals.
Asunto(s)
Distribución en Contracorriente/métodos , Flavanonas/aislamiento & purificación , Ácido Glicirretínico/aislamiento & purificación , Glycyrrhiza uralensis/química , Extracción Líquido-Líquido/métodos , Antioxidantes/química , Antioxidantes/aislamiento & purificación , Flavanonas/química , Ácido Glicirretínico/química , Extractos Vegetales/químicaRESUMEN
Enhancing soluble receptor for advanced glycation endproducts (sRAGE) is considered as a potent strategy for diabetes therapy. sRAGE secretion is regulated by calcium and transient receptor potential canonical (TRPC) channels. However, the role of TRPC channels in diabetes remains unknown. 18ß-Glycyrrhetinic acid (18ß-GA), produced from liquorice, has shown antidiabetic properties. This study was aimed to investigate the effect of 18ß-GA on sRAGE secretion via TRPC channels in high glucose (HG)-induced THP-1 cells. HG treatment enhanced TRPC3 and TRPC6 expression and consequently caused reactive oxygen species (ROS) accumulation mediated through p47 nicotinamide-adenine dinucleotide phosphate oxidase and inducible nitric oxide synthase (iNOS) associated with uncoupling protein 2 (UCP2) decline and lower sRAGE secretion. Interestingly, 18ß-GA showed the dramatic effects similar to Pyr3 or 2-aminoethyl diphenyl borinate inhibitors and effectively reversed HG-elicited mechanisms including that blocking TRPC3 and TRPC6 protein expressions, suppressing intracellular [Ca2+] concentration, decreasing expressions of ROS, p47s, and iNOS, but increasing UCP2 level and promoting sRAGE secretion. Therefore, 18ß-GA provides a potential implication to diabetes mellitus and its complications.
Asunto(s)
Glucosa/antagonistas & inhibidores , Ácido Glicirretínico/análogos & derivados , Glycyrrhiza/química , Hipoglucemiantes/farmacología , Receptor para Productos Finales de Glicación Avanzada/genética , Canales Catiónicos TRPC/genética , Canal Catiónico TRPC6/genética , Compuestos de Boro/farmacología , Calcio/metabolismo , Regulación de la Expresión Génica , Glucosa/toxicidad , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirretínico/farmacología , Humanos , Hipoglucemiantes/aislamiento & purificación , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico Sintasa de Tipo II/metabolismo , Extractos Vegetales/química , Pirazoles/farmacología , Especies Reactivas de Oxígeno/antagonistas & inhibidores , Especies Reactivas de Oxígeno/metabolismo , Receptor para Productos Finales de Glicación Avanzada/antagonistas & inhibidores , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Transducción de Señal , Células THP-1 , Canales Catiónicos TRPC/antagonistas & inhibidores , Canales Catiónicos TRPC/metabolismo , Canal Catiónico TRPC6/antagonistas & inhibidores , Canal Catiónico TRPC6/metabolismo , Proteína Desacopladora 2/genética , Proteína Desacopladora 2/metabolismoRESUMEN
Hizikia fusiformis (Harvey) Okamura is a brown seaweed widely used in Korea and Japan, and it contains different therapeutically active constituents. In the present study, we investigated the activities of glycyrrhizin isolated from H. fusiformis, including its metabolites, 18α- and 18ß-glycyrrhetinic acid against Alzheimer's disease (AD) via acetyl and butyrylcholinesterase and ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibition. Among these three compounds, 18ß-glycyrrhetinic acid (IC50 = 8.93 ± 0.69 µM) demonstrated two fold potent activity against BACE1 compared to the positive control, quercetin (IC50 = 20.18 ± 0.79 µM). Additionally, glycyrrhizin with an IC50 value of 20.12 ± 1.87 µM showed similarity to quercetin, while 18α-glycyrrhetinic acid showed moderate activity (IC50 = 104.35 ± 2.84 µM). A kinetic study revealed that glycyrrhizin and 18ß-glycyrrhetinic acid were non-competitive and competitive inhibitiors of BACE1, demonstrated via K i values of 16.92 and 10.91 µM, respectively. Molecular docking simulation studies evidently revealed strong binding energy of these compounds for BACE1, indicating their high affinity and capacity for tighter binding to the active site of the enzyme. These data suggest that glycyrrhizin isolated from the edible seaweed, H. fusiformis and its metabolite, 18ß-glycyrrhetinic acid demonstrated selective inhibitory activity against BACE1 to alleviate AD.
Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/antagonistas & inhibidores , Antiinflamatorios/farmacología , Ácido Aspártico Endopeptidasas/antagonistas & inhibidores , Simulación por Computador , Ácido Glicirretínico/análogos & derivados , Ácido Glicirrínico/farmacología , Simulación del Acoplamiento Molecular/métodos , Algas Marinas , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Animales , Antiinflamatorios/aislamiento & purificación , Antiinflamatorios/metabolismo , Ácido Aspártico Endopeptidasas/metabolismo , Relación Dosis-Respuesta a Droga , Electrophorus , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirretínico/metabolismo , Ácido Glicirretínico/farmacología , Ácido Glicirrínico/aislamiento & purificación , Ácido Glicirrínico/metabolismo , Caballos , HumanosRESUMEN
OBJECTIVE: To develop a simple and sensitive high performance liquid chromatographic method for simultaneous determination of catechin hydrate, epicatechin, epigallocatechin, epicatechin gallate, epigallocatechin gallate, dihydromyricetin, glycyrrhizic acid and glycyrrhetinic acid in healthy food for anti-hangover and hepatoprotection, and compare with the capillary electrophoresis method established by our laboratory. METHODS: The samples were ultrasonically extracted by using methanol-water( 4ⶠ1, V/V) for 30 minutes and then centrifuged at 10 000 r/min for 10 minutes. The supernatant was filtered and injected into the HPLC system and then separated on a C_(18) column( 5 µm × 250 mm × 4. 6 mm) at 30â with gradient elution at a flow rate of 0. 8mL/min. Catechins and dihydromyricetin were detected at the wavelength of 210 nm, glycyrrhizic acid and glycyrrhetinic acid were detected at 250 nm. RESULTS: Under the optimal analytical conditions, the peak area of each analyte and its concentration had agood correlation within the linear range( r ≥ 0. 9996). The limits of detection and quantification of the method were 0. 07-1. 25 µg/g( S/N = 3) and 0. 22-4. 18 µg/g( S/N = 10), respectively. The intra-and inter-day relative standard deviations( RSDs)of the mixed standard solution were 0. 26%-1. 95% and 1. 17%-3. 89%, respectively. The spiked recoveries of the analytes were 86. 15%-98. 61%. CONCLUSION: The established method is sensitive and reliable, and could be used for quality control of the healthy food for anti-hangover and hepatoprotection.
Asunto(s)
Catequina/análisis , Cromatografía Líquida de Alta Presión/métodos , Flavonoles/análisis , Ácido Glicirretínico/análisis , Ácido Glicirrínico/análisis , Catequina/análogos & derivados , Catequina/aislamiento & purificación , Electroforesis Capilar , Flavonoles/aislamiento & purificación , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirrínico/aislamiento & purificación , Humanos , Sensibilidad y EspecificidadRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Glycyrrhiza uralensis Fisch. and processed radix Aconiti kusnezoffii are the main components in many Chinese traditional patent medicines with the ratio of 1:1, which are used for treatment of rheumatoid arthritis, heart failure and so on. Glycyrrhizic acid, glycyrrhetic acid, liquiritigenin and isoliquiritigenin are the essential bioactive triterpenes and flavones in the extract of G. uralensis, which were analysis by a simple but accurate method. MATERIALS AND METHODS: In the present study, a specific HPLC method was developed and validated for simultaneous determination of pharmacokinetic parameters of glycyrrhizic acid, glycyrrhetic acid, liquiritigenin and isoliquiritigenin in G. uralensis after oral administration of single herb extract and a combination of two herbs extracts respectively. RESULTS: The calibration curves of the four components had good linearity higher than 0.9991 in the measured range. The intra-day and inter-day precisions (RSD) at different levels were both within 9.73%, and the accuracies (RE) were in the range of -7.9-8.0%. Compared with pharmacokinetic parameters of G. uralensis administered orally, values of AUC and Cmax of liquiritigenin and isoliquiritigenin decreased significantly (p<0.05), plasma concentrations of glycyrrhizic acid rose slightly and bimodal phenomenon of concentration-time of isoliquiritigenin and glycyrrhetinic acid disappeared after combined administration. DISCUSSION AND CONCLUSIONS: Some components in the extract of processed radix A. kusnezoffii showed different effects on the pharmacokinetics of the four ingredients in G. uralensis.
Asunto(s)
Aconitum , Chalconas/farmacocinética , Flavanonas/farmacocinética , Ácido Glicirretínico/farmacocinética , Glycyrrhiza uralensis , Ácido Glicirrínico/farmacocinética , Administración Oral , Animales , Chalconas/administración & dosificación , Chalconas/aislamiento & purificación , Medicamentos Herbarios Chinos/administración & dosificación , Medicamentos Herbarios Chinos/aislamiento & purificación , Medicamentos Herbarios Chinos/farmacocinética , Flavanonas/administración & dosificación , Flavanonas/aislamiento & purificación , Flavonas/administración & dosificación , Flavonas/aislamiento & purificación , Flavonas/farmacocinética , Ácido Glicirretínico/administración & dosificación , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirrínico/administración & dosificación , Ácido Glicirrínico/aislamiento & purificación , Masculino , Raíces de Plantas , Ratas , Ratas Sprague-DawleyRESUMEN
The aim of this work was to study an application of a previously developed expedient acid-free technique for the preparation of glycyrrhetinic acid from ammonium glycyrrhizinate that requires no use of acids and toxic organic solvents. Subcritical water that serves as a reactant and a solvent was used in order to obtain glycyrrhetinic acid in good yields starting from ammonium glycyrrhizinate. It has been shown that variation of only one parameter of the process (temperature) allows alteration to thecomposition of the hydrolysis products. A new method was used for the synthesis of glycyrrhetinic acid (glycyrrhizic acid aglycone) and its monoglycoside. HPLC combined with mass spectrometry and NMR spectroscopy were used to determine the quantitative and qualitative compositions of the obtained products. The method developed for the production of glycyrrhetinic acid in subcritical water is environmentally friendly and faster than conventional hydrolysis methods that use acids and-expensive and toxic organic solvents. The proposed technique has a potential for the future development of inexpensive and environmentally friendly technologies for production of new pharmaceutical plant-based substances.
Asunto(s)
Compuestos de Amonio/química , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirrínico/química , Agua/química , Ácido Glicirretínico/química , Espectrometría de MasasRESUMEN
Septic shock and associated vascular hyporeactivity to vasoconstrictor agonists remain a major problem of critical care medicine. Here we report that glycyrrhetinic acid (GA), the active component of licorice, effectively restores vascular contractility in the model of lipopolysaccharide (LPS)-treated rat aorta. GA was as effective as the NO synthase inhibitor N(G)-nitroarginine methylester. GA did not affect the vascular NO levels (measured by EPR spin trapping) and relaxations to L-arginine in LPS-treated rings as well as relaxation to S-nitroso-N-acetylpenicillamine in control rings. Thus, GA may represent an interesting alternative to NO synthase inhibitors in sepsis-associated vascular dysfunction.
Asunto(s)
Aorta/efectos de los fármacos , Inhibidores Enzimáticos , Ácido Glicirretínico/farmacología , Ácido Glicirretínico/uso terapéutico , Lipopolisacáridos/efectos adversos , Norepinefrina/antagonistas & inhibidores , Norepinefrina/farmacología , Fitoterapia , Choque Séptico/tratamiento farmacológico , Choque Séptico/etiología , Vasoconstricción/efectos de los fármacos , Animales , Ácido Glicirretínico/aislamiento & purificación , Glycyrrhiza/química , Masculino , Óxido Nítrico Sintasa/antagonistas & inhibidores , Ratas WistarRESUMEN
Triptolide (TP) is a major active component in Tripterygium root, but its therapeutic window was very narrow due to its severe multi-organ toxicity. In this work, the effect of TP combined with glycyrrhetic acid (GA) on mRNA expression and activity of four cytochrome P450 (CYP) enzymes in rat liver was studied after intragastric administration of TP (0.05, 0.3 and 0.6 mg x kg(-1) x day(-1)) and TP (0.6 mg x kg(-1) x day(-1)) combined with GA (30 mg x kg(-1) x day(-1)) for 7 consecutive days. Compared with the control, the high dose of TP significantly up-regulated the mRNA expression levels of CYP2E1, 1A2, 3A1 and 2C11, the co-administration of TP and GA further up-regulated the mRNA expression levels of CYP3A1, 2C11 and 2E1 as compared with the high dose of TP. Meanwhile, TP at high dose and combined with GA significantly increased CYP3A-associated testosterone 6beta-hydroxylation activity (2.2-fold and 4.1-fold, respectively) as compared with the control. Because TP is mainly metabolized by CYP3A2 in male rats, the present work indicated that TP-induced increase of CYP3A activity might be an important reason for the rapidly metabolic clearance of TP in rat liver, and GA can reduce the hepatotoxicity of TP by promoting its hepatic metabolic clearance. Furthermore, the results also suggest that the drug interactions might be occurred when TP and GA were co-administered with other CYP3A substrate drug.
Asunto(s)
Sistema Enzimático del Citocromo P-450/metabolismo , Diterpenos/farmacología , Ácido Glicirretínico/farmacología , Hígado/enzimología , Fenantrenos/farmacología , Animales , Hidrocarburo de Aril Hidroxilasas/genética , Hidrocarburo de Aril Hidroxilasas/metabolismo , Citocromo P-450 CYP1A2/genética , Citocromo P-450 CYP1A2/metabolismo , Citocromo P-450 CYP2E1/genética , Citocromo P-450 CYP2E1/metabolismo , Citocromo P-450 CYP3A/genética , Citocromo P-450 CYP3A/metabolismo , Sistema Enzimático del Citocromo P-450/genética , Familia 2 del Citocromo P450 , Diterpenos/administración & dosificación , Diterpenos/aislamiento & purificación , Relación Dosis-Respuesta a Droga , Combinación de Medicamentos , Interacciones Farmacológicas , Activación Enzimática , Compuestos Epoxi/administración & dosificación , Compuestos Epoxi/aislamiento & purificación , Compuestos Epoxi/farmacología , Ácido Glicirretínico/aislamiento & purificación , Masculino , Fenantrenos/administración & dosificación , Fenantrenos/aislamiento & purificación , Raíces de Plantas/química , Plantas Medicinales/química , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Esteroide 16-alfa-Hidroxilasa/genética , Esteroide 16-alfa-Hidroxilasa/metabolismo , Tripterygium/químicaRESUMEN
Malaria is one of the most prevailing fatal diseases causing between 1.2 and 2.7 million deaths all over the world each year. Further, development of resistance against the frontline anti-malarial drugs has created an alarming situation, which requires intensive drug discovery to develop new, more effective, affordable and accessible anti-malarial agents possessing novel modes of action. Over the past few years triterpenoids from higher plants have shown a wide range of anti-malarial activities. As a part of our drug discovery program for anti-malarial agents from Indian medicinal plants, roots of Glycyrrhizaglabra were chemically investigated, which resulted in the isolation and characterization of 18ß-glycyrrhetinic acid (GA) as a major constituent. The in vitro studies against P. falciparum showed significant (IC50 1.69 µg/ml) anti-malarial potential for GA. Similarly, the molecular docking studies showed adequate docking (LibDock) score of 71.18 for GA and 131.15 for standard anti-malarial drug chloroquine. Further, in silico pharmacokinetic and drug-likeness studies showed that GA possesses drug-like properties. Finally, in vivo evaluation showed a dose dependent anti-malarial activity ranging from 68-100% at doses of 62.5-250 mg/kg on day 8. To the best of our knowledge this is the first ever report on the anti-malarial potential of GA. Further work on optimization of the anti-malarial lead is under progress.
Asunto(s)
Antimaláricos/farmacología , Simulación por Computador , Ácido Glicirretínico/análogos & derivados , Glycyrrhiza/química , Animales , Antimaláricos/uso terapéutico , Disponibilidad Biológica , Ácido Glicirretínico/química , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirretínico/farmacología , Ácido Glicirretínico/uso terapéutico , L-Lactato Deshidrogenasa/química , Malaria Falciparum/tratamiento farmacológico , Ratones , Modelos Moleculares , Raíces de Plantas/química , Plasmodium falciparum/efectos de los fármacos , Plasmodium falciparum/enzimologíaRESUMEN
A triterpene saponin, glucoglycyrrhizin, was isolated from a glycyrrhizin-deficient strain 83-555 of Glycyrrhiza uralensis (Leguminosae), and the structure was determined by chemical and spectral data to be 3-O-[ß-D-glucopyranosyl-(1â2)-ß-D-glucuronopyranosyl]-glycyrrhetinic acid. Since this saponin has a 2'-O-ß-D-glucopyranosyl moiety instead of the 2'-O-ß-D-glucuronopyranosyl moiety of glycyrrhizin, the glucuronidation of 3-O-ß-D-glucuronopyranosyl-glycyrrhetinic acid leading to glycyrrhizin is inhibited in this strain. All 4 offspring of the 83-555 strain produced glucoglycyrrhizin. Interestingly, 2 of the offspring produced both glycyrrhizin and glucoglycyrrhizin, and sequence analysis of the pkr gene suggested that these 2 offspring were hybrids of 83-555 strain and glycyrrhizin-producing strains.
Asunto(s)
Ácido Glicirretínico/análogos & derivados , Ácido Glicirretínico/aislamiento & purificación , Glycyrrhiza uralensis/química , Oxidorreductasas de Alcohol/genética , Proteínas Bacterianas/genética , ADN de Plantas/análisis , Genes de Plantas/genética , Ácido Glicirretínico/química , Glycyrrhiza uralensis/genética , Ácido Glicirrínico , Datos de Secuencia Molecular , Raíces de Plantas/química , Análisis de Secuencia de ADNRESUMEN
Although a number of chemicals have been isolated from Glycyrrhiza glabra, only a few have been evaluated for their biological significance. As part of our drug discovery program for antifilarial agents from Indian medicinal plants, the roots of G. glabra were chemically investigated, which resulted in the isolation and characterization of an antifilarial agent, glycyrrhetinic acid (GA, 1a) effective against microfilariae (mf) in vitro (LC100: 12.5 µM; IC50: 1.20 µM), but was inactive against adult worms. Further, GA (1a) was converted into six analogs (2a-7a) and their antifilarial potential was evaluated by studying in vitro motility and MTT reduction assays employing mf and adult worms of Brugia malayi. The results showed that out of six GA analogs, the benzyl amide analog (6a) killed adults and mf at 25 and 50 µM concentration, respectively, and inhibited 49% MTT reduction potential of the adult parasites. The IC50 values were found to be 8.8 and 2.2 µM for adults and mf, respectively. The SI of the compound was >60. On the other hand the octylamide analog (7a) required much higher concentration to adversely affect the parasites. Finally, both active amide analogs (6a and 7a) were in vivo evaluated using B. malayi-jird model, which showed that analog 6a possesses promising macrofilaricidal activity at 100mg/kg, s.c. ×5 days and around 40% of the treated animals showed calcified masses of worm fragments in peritoneal cavity of the animals. To the best of our knowledge this is the first ever report on the antifilarial potential of GA analogs. Further work on optimization of the antifilarial lead is under progress.
Asunto(s)
Filaricidas/química , Ácido Glicirretínico/análogos & derivados , Animales , Brugia Malayi/efectos de los fármacos , Femenino , Filaricidas/aislamiento & purificación , Filaricidas/farmacología , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirretínico/farmacología , Glycyrrhiza/química , Microfilarias/efectos de los fármacos , Raíces de Plantas/químicaRESUMEN
18ß-Glycyrrhetinic acid (ß-GA) is a natural triterpenoid compound derived from licorice root. ß-GA has been demonstrated to exert antiviral and antitumor effects. However, the effects of the maturation and immunostimulatory functions of dendritic cells (DCs) remain to be clearly elucidated. In this study, we attempted to determine whether ß-GA could influence DCs surface molecule expression, antigen uptake capacity, cytokine production and capacity to induce T-cell differentiation. The DCs used in this study were derived from murine bone marrow cells, and were used as immature or lipopolysaccharide (LPS)-stimulated mature DCs. The DCs were then assessed with regard to surface molecules expression, cytokine production, capacity to induce T-cell differentiation and proliferation. ß-GA was shown to significantly suppress the expression of surface molecules CD80, CD86, major histocompatibility complex (MHC) class I and MHC class II as well as the levels of interleukin-12 production in LPS-stimulated DCs. Moreover, ß-GA-treated DCs showed an impaired induction of the T helper type 1 immune response. These findings provide important understanding of the immunopharmacological functions of ß-GA and have ramifications for the development of therapeutic adjuvants for the treatment of DCs-related acute and chronic diseases.
Asunto(s)
Diferenciación Celular/inmunología , Citocinas/inmunología , Células Dendríticas/inmunología , Ácido Glicirretínico/análogos & derivados , Glycyrrhiza/química , Células TH1/inmunología , Animales , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Técnicas de Cocultivo , Citocinas/metabolismo , Células Dendríticas/citología , Células Dendríticas/efectos de los fármacos , Endocitosis/efectos de los fármacos , Endocitosis/inmunología , Citometría de Flujo , Ácido Glicirretínico/efectos adversos , Ácido Glicirretínico/aislamiento & purificación , Lipopolisacáridos/farmacología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Raíces de Plantas/química , Células TH1/citología , Células TH1/efectos de los fármacosRESUMEN
Two new phenolic glycosides were isolated from the ethanol extract of the roots of Inula cappa DC. Their structures were defined as 4-[(6-O-(E)-caffeoyl)-ß-D-glucopyranosyl]vanillic acid (1) and 3-O-[ß-D-apiofurarnosyl-(1-6)-ß-D-glucopyranoxy]-6-hydroxy-p-cymene (2) on the basis of spectral analysis.
Asunto(s)
Glicósidos/aislamiento & purificación , Ácido Glicirretínico/análogos & derivados , Inula/química , Fenoles/aislamiento & purificación , Ácido Vanílico/análogos & derivados , Glicósidos/química , Ácido Glicirretínico/química , Ácido Glicirretínico/aislamiento & purificación , Fenoles/química , Raíces de Plantas/química , Plantas Medicinales/química , Ácido Vanílico/química , Ácido Vanílico/aislamiento & purificaciónRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Shakuyakukanzoto (SKT) composed of Glycyrrhizae radix (G. radix) and Paeoniae radix (P. radix) has been traditionally used in Japan, Korea and China as an antispasmodic drug for the treatment of skeletal muscle cramps and intestinal cramps. AIM OF THIS STUDY: To evaluate the antispasmodic activity of SKT and its two components, as well as to identify the key constituents of the components which mediate this effect in skeletal muscles in vivo. MATERIALS AND METHODS: An experimental cramp model was constructed to evaluate the effects of peripherally-acting muscle relaxants on electrically-induced cramps under physiological conditions. This was accomplished by surgically isolating the motor supply to the gastrocnemius muscle in an anesthetized rat and delivering electrical stimuli to an isolated tibial nerve to induce tetanic contractions. We first tested dantrolene, a well-known peripherally-acting relaxant, to determine the sensitivity and reliability of our experimental model. We then evaluated the effects of SKT, P. radix, G. radix, and the eight active constituents of G. radix against tetanic contractions. RESULTS: We found that dantrolene (10 and 30 mg/kg, i.d.) rapidly and significantly inhibited tetanic contractions (P<0.01) irrespective of dose. SKT (0.5, 1.0, and 2.0 g/kg, i.d.) and G. radix (0.5 and 1.0 g/kg, i.d.) also significantly inhibited tetanic contractions (P<0.01) but in a dose-dependent manner owing to the actions of six of the eight active constituents in G. radix (liquiritin apioside, liquiritigenin, isoliquiritin apioside, isoliquiritigenin, glycycoumarin, and glycyrrhetinic acid, 20 µmol/kg, i.v.). These constituents, which include flavonoids, a triterpenoid, and a courmarin derivative, demonstrated temporal variations in their inhibitory activity. In contrast, P. radix (0.5 and 1.0 g/kg, i.d.) did not show a statistically significant antispasmodic effect in our study; however, we previously found that it had a significant antinociceptive effect. CONCLUSIONS: Our findings show that SKT inhibits tetanic contractions in vivo and that G. radix is the main antispasmodic component due to the actions of its active constituents, thus supporting the traditional use of SKT. We further propose that SKT containing the antispasmodic G. radix and antinociceptive P. radix is a pharmaceutically elegant option for muscle cramps as treatment requires a two-pronged approach, i.e., inhibition of hyperexcitable skeletal tissues and modulation of the pain accompanying cramps.
Asunto(s)
Medicamentos Herbarios Chinos/uso terapéutico , Calambre Muscular/tratamiento farmacológico , Fitoterapia/métodos , Animales , Animales no Consanguíneos , Chalcona/análogos & derivados , Chalcona/farmacología , Chalconas/aislamiento & purificación , Chalconas/farmacología , Cumarinas/aislamiento & purificación , Cumarinas/farmacología , Dantroleno/farmacología , Dantroleno/uso terapéutico , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Combinación de Medicamentos , Medicamentos Herbarios Chinos/química , Medicamentos Herbarios Chinos/farmacología , Estimulación Eléctrica/métodos , Flavanonas/aislamiento & purificación , Flavanonas/farmacología , Glucósidos/aislamiento & purificación , Glucósidos/farmacología , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirretínico/farmacología , Glycyrrhiza/química , Masculino , Ratones , Contracción Muscular/efectos de los fármacos , Contracción Muscular/fisiología , Calambre Muscular/fisiopatología , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/fisiología , Paeonia/química , Parasimpatolíticos/química , Parasimpatolíticos/farmacología , Raíces de Plantas/química , Ratas , Ratas Wistar , Prueba de Desempeño de Rotación con Aceleración Constante/métodos , Nervio Tibial/efectos de los fármacos , Nervio Tibial/fisiologíaRESUMEN
The sensitivity, specificity and selectivity of liquid chromatography/mass spectrometry tandem mass spectrometry (LC-MS/MS) make it an essential tool for the characterization and identification of low molecular compounds such as fatty acids, sterols, cholastane derivatives, nucleosides etc. In the current work, the marine sponge Spongosorites halichondriodes (order Halichondrida, Family Halichondriidae); a particularly rich source of cytotoxic compounds is studied for the initial characterization of bioactive compounds. The composition of ethyl acetate and butanol extracts were subjected to LC-MS and LC-MS/MS. Many novel sterol derivatives compounds which were not reported in any marine sponge mainly belonged to the group of C25-C28 saturated and unsaturated esters like 3ß, 4ß, 7α, 12α-tetrahydroxy-5ß-cholan-24-oic acid methyl ester, 7 α, 12 ß-dihydroxy-5 ß-cholan24-oic acid methyl ester, novel isocoumarin citrinolactone A, a triterpenoid glycyrrhetinic acid as well as other unknown compounds in this species such as nucleoside inosine was identified. Other compound investigated was 3ß, 6ß, 7α-trihydroxy-5ß-cholan-24-oic acid methyl ester. All the sterol ester derivatives are reported here for the first time in marine sponge belonging to family Halichondriidae. However, the literature report supports the occurrence of 3ß-hydroxy sterols which is considered as a biomarker for this family.
Asunto(s)
Cromatografía Liquida/métodos , Poríferos/química , Espectrometría de Masas en Tándem/métodos , Animales , Benzopiranos/análisis , Benzopiranos/aislamiento & purificación , Ésteres/análisis , Ésteres/aislamiento & purificación , Ácido Glicirretínico/análisis , Ácido Glicirretínico/aislamiento & purificación , Inosina/análisis , Inosina/aislamiento & purificación , Isocumarinas/análisis , Isocumarinas/aislamiento & purificación , Nucleósidos/análisis , Nucleósidos/aislamiento & purificación , Esteroles/análisis , Esteroles/aislamiento & purificación , Triterpenos/análisis , Triterpenos/aislamiento & purificaciónRESUMEN
18ß-Glycyrrhetinic acid (GA) is a major metabolite of glycyrrhizin (GL), which is one of the components of glycyrrhiza root, a constituent herb of the traditional Japanese medicine yokukansan. It is well known that most GL is metabolized to GA in the intestine by bacteria. A previous in vitro study using cultured rat cortical astrocytes suggested that GA activates glutamate transport, which is a putative mechanism of the psychotropic effect of yokukansan. To activate the glutamate transport in the brain, GA must be absorbed into the blood after oral administration of yokukansan and then cross the blood-brain barrier (BBB) to reach the brain. However, there is no data on the BBB permeability of GA derived from yokukansan. In the present study, the BBB permeability of GA was investigated in both in vivo and in vitro studies. In the in vivo study, GA was detected in the plasma, brain, and cerebrospinal fluid of rats orally administered yokukansan. In the in vitro study using a BBB model composed of co-culture of endothelial cells, pericytes, and astrocytes, the permeability rate and apparent permeability coefficient of GA were found to be 13.3 ± 0.5 % and 16.5 ± 0.7 × 10(-6) cm/s. These in vivo and in vitro results suggest that GL in orally administered yokukansan is absorbed into the blood as GA, and then reaches the brain through the BBB. This evidence further supports the possibility that GA is an active component in the psychotropic effect of yokukansan.
Asunto(s)
Barrera Hematoencefálica/metabolismo , Permeabilidad Capilar/fisiología , Medicamentos Herbarios Chinos/metabolismo , Ácido Glicirretínico/análogos & derivados , Glycyrrhiza , Medicina Tradicional de Asia Oriental , Raíces de Plantas , Animales , Barrera Hematoencefálica/efectos de los fármacos , Permeabilidad Capilar/efectos de los fármacos , Medicamentos Herbarios Chinos/farmacología , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirretínico/metabolismo , Ácido Glicirretínico/farmacología , Ácido Glicirrínico/aislamiento & purificación , Ácido Glicirrínico/metabolismo , Ácido Glicirrínico/farmacología , Japón , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
Glycyrrhetinic acid (GA) is an active component of licorice root that has long been used as a herbal medicine for the treatment of peptic ulcer, hepatitis, and pulmonary and skin diseases in Asia and Europe. In this study, we analyzed the effect of GA extracted from Glycyrrhiza uralensis Fisch. on the expression of Toll-like receptors (TLRs) that play key roles in regulating the innate immune response against invading pathogens. Stimulation of Ana-1 murine macrophages with GA induced a significant dose-dependent expression of TLR-4, and its mRNA expression that increased from 3-h post-treatment was approximately fivefold over the level in the mock-treated cells. No endotoxin contamination contributed to the GA-induced TLR-4 expression, because polymyxin B treatment did not alter the upregulated expression of TLR-4 in GA-treated cells. Several molecules, such as myeloid differentiation factor 88, interferon-ß, and interleukin-6, which are involved in the TLR-4 downstream signaling pathway, were upregulated significantly in response to GA stimulation. Our findings demonstrate that GA is able to induce the expression of TLR-4 and activate its downstream signaling pathway.
Asunto(s)
Ácido Glicirretínico/aislamiento & purificación , Glycyrrhiza uralensis/química , Macrófagos/efectos de los fármacos , Receptor Toll-Like 4/efectos de los fármacos , Animales , Ácido Glicirretínico/química , Ácido Glicirretínico/inmunología , Humanos , Ratones , Estructura Molecular , Transducción de Señal/efectos de los fármacos , Receptor Toll-Like 4/genéticaRESUMEN
Licorice root has been frequently used as antitode in traditional Chinese medicine. As the main active component of Licorice root, glycyrrhetic acid (GA) is mainly metabolized in liver. This study was designed to investigate the in vitro metabolism of GA by human liver microsomes (HLM) and human recombinant cytochrome P450 (CYP) isoforms. The results indicated that GA was metabolized mainly by CYP3A4. The K(m), V(max) and CL(int) of GA in HLM were 18.6 micromol x L(-1), 4.4 nmol x mg(-1) (protein) x min(-1) and 0.237 mL x mg(-1) (protein) x min(-1), respectively. At concentration up to 50 micromol x L(-1), GA inhibited CYP2C19, CYP2C9 and CYP3A4 enzyme activities with the inhibitory potencies up to 50%.
Asunto(s)
Citocromo P-450 CYP3A/metabolismo , Ácido Glicirretínico/farmacocinética , Microsomas Hepáticos , Hidrocarburo de Aril Hidroxilasas/antagonistas & inhibidores , Hidrocarburo de Aril Hidroxilasas/metabolismo , Citocromo P-450 CYP2C19 , Citocromo P-450 CYP2C9 , Inhibidores del Citocromo P-450 CYP3A , Inhibidores Enzimáticos/farmacología , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirretínico/farmacología , Glycyrrhiza/química , Humanos , Isoenzimas/metabolismo , Microsomas Hepáticos/enzimología , Microsomas Hepáticos/metabolismo , Raíces de Plantas/química , Plantas Medicinales/químicaRESUMEN
In this study, an automated system for phytochemical analysis was successfully fabricated for the first time in our laboratory. The system included on-line decocting, filtering, cooling, sample introducing, separation, and detection, which greatly simplified the sample preparation and shortened the analysis time. Samples from the decoction extract were drawn every 5 min through an on-line filter and a condenser pipe to the sample loop from which 20-µL samples were injected into the running buffer and transported into a split-flow interface coupling the flow injection and capillary electrophoresis systems. The separation of glycyrrhetinic acid (GTA) and glycyrrhizic acid (GA) took less than 5 min by using a 10 mM borate buffer (adjusted pH to 8.8) and +10 kV voltage. Calibration curves showed good linearity with correlation coefficients (R) more than 0.9991. The intra-day repeatabilities (n = 5, expressed as relative standard deviation) of the proposed system, obtained using GTA and GA standards, were 1.1% and 0.8% for migration time and 0.7% and 0.9% for peak area, respectively. The mean recoveries of GTA and GA in the off-line extract of Glycyrrhiza uralensis Fisch root were better than 99.0%. The limits of detection (signal-to-noise ratio = 3) of the proposed method were 6.2 µg/mL and 6.9 µg/mL for GTA and GA, respectively. The dynamic changes of GTA and GA on the decoction time were obtained during the on-line decoction process of Glycyrrhiza uralensis Fisch root.
Asunto(s)
Electroforesis Capilar/métodos , Análisis de Inyección de Flujo/métodos , Glycyrrhiza/química , Extractos Vegetales/aislamiento & purificación , Automatización , Electroforesis Capilar/instrumentación , Ácido Glicirretínico/análisis , Ácido Glicirretínico/aislamiento & purificación , Ácido Glicirrínico/análisis , Ácido Glicirrínico/aislamiento & purificación , Extractos Vegetales/análisisRESUMEN
Five hydroxylated derivatives of glycyrrhetinic acid by Mucor polymorphosporus were isolated. Among them, 6beta, 7beta-dihydroxyglycyrrhentic acid (2) and 27-hydroxyglycyrrhentic acid (3) are new compounds. Their chemical structures were identified by spectral methods including 2D-NMR.