Bharathi Avula

Ginkgo biloba does not alter clearance of flurbiprofen, a cytochrome P450-2C9 substrate.

The effect of Ginkgo biloba on the activity of CYP2C9, the isoform responsible for S‐warfarin clearance, was assessed in 11 healthy volunteers who received single 100‐mg doses of flurbiprofen, a probe substrate for CYP2C9. Subjects also received either a standardized G biloba leaf preparation (Ginkgold, 3 doses of 120 mg) or matching placebo in a randomized, double‐blind, 2‐way crossover study. Mean kinetic variables for flurbiprofen with either placebo or G biloba were elimination half‐life, 3.9 versus 3.5 hours; total AUC, 57 versus 55 μg/mL•h; and oral clearance, 32.9 versus 31.6 mL/min. None of these differences was significant. Based on high‐performance liquid chromatography analysis, each 60‐mg Ginkgold tablet contained 6.6 μg of amentoflavone and 61.2 μg of quercetin, both previously identified as CYP2C9 inhibitors. These amounts were apparently too low to inhibit CYP2C9 function in vivo. The results confirm previous controlled clinical studies showing no effect of ginkgo on the kinetics or dynamics of warfarin.

New oxypregnane glycosides from appetite suppressant herbal supplement Hoodia gordonii

Hoodigosides A-K (1-11), eleven new oxypregnane glycosides and a previously reported oxypregnane glycoside P57AS3 were isolated from the aerial parts of Hoodia gordonii. The structures of these 12-O-beta-tigloyl isoramanone glycosides were determined on the basis of chemical evidence and extensive spectroscopic methods that include one-dimensional and two-dimensional NMR. Cytotoxicity and antioxidant activities of these compounds were tested in cell based assays where they were found to be inactive.

A rapid method for chemical fingerprint analysis of Hoodia species, related genera, and dietary supplements using UPLC-UV-MS.

Recently, ultra-performance liquid chromatography (UPLC) has proven to be one of the most promising developments in the area of high-speed chromatographic separations with increased sensitivity and resolution. In this work, a reverse phase chromatographic method was developed using UPLC for the chemical fingerprint analysis of 12 hoodigosides, related genera and dietary supplements. The method is also used for the quantification of P57 in Hoodia species and dietary supplements that claim to contain Hoodia. The analysis was performed on a Waters Acquity UPLC system with an Acquity UPLC BEH C18 column (100 mm x 2.1mm I.D., 1.7 microm) and a gradient elution of water and acetonitrile, both containing 0.05% formic acid with a run time of 15 min. The calibration curve of P57 showed good linearity (r(2)>0.999) within the established range (1-100 microg/mL). The limit of detection (LOD) and limit of quantification (LOQ) were found to be 0.3 and 0.9 microg/mL, respectively. The RSD for intra- and inter-day were less than 3.0%, and the recovery efficiency as 97-103%. LC-mass spectrometry coupled with electrospray ionization (ESI) interface method is described for the identification of P57. The developed method was successfully applied to the identification of 12 oxypregnane glycosides in four different species of Hoodia, 23 related genera and 35 dietary supplements that claim to contain H. gordonii. The UPLC profiles of various plant samples were compared for the presence of oxypregnane glycosides. Different sample matrices were successfully analyzed, providing the wide range of applicability of this method, including gels, capsules, tablets, sprays, tea bags, snack bars, powders and juices.

Flavonol glycosides from the south African medicinal plant Sutherlandia frutescens.

Phytochemical investigation of the leaves of Sutherlandia frutescens led to the isolation of four new 3-hydroxy-3-methylglutaroyl-containing flavonol glycosides, sutherlandins A-D ( 1- 4). Their structures were elucidated by chemical and spectroscopic methods as quercetin 3- O- beta- D-xylopyranosyl(1 --> 2)-[6- O-(3-hydroxy-3-methylglutaroyl)]- beta- D-glucopyranoside ( 1), quercetin 3- O- beta- D-apiofuranosyl(1 --> 2)-[6- O-(3-hydroxy-3-methylglutaroyl)]- beta- D-glucopyranoside ( 2), kaempferol 3- O- beta- D-xylopyranosyl(1 --> 2)-[6- O-(3-hydroxy-3-methylglutaroyl)]- beta- D-glucopyranoside ( 3), and kaempferol 3- O- beta- D-apiofuranosyl(1 --> 2)-[6- O-(3-hydroxy-3-methylglutaroyl)]- beta- D-glucopyranoside ( 4).

Quantitative determination of flavonoids and cycloartanol glycosides from aerial parts of Sutherlandia frutescens (L.) R. BR. by using LC-UV/ELSD methods and confirmation by using LC-MS method.

This paper describes the first analytical method for the determination of four flavonoids (sutherlandins A-D) and four cycloartanol glycosides (sutherlandiosides A-D) from the aerial parts of Sutherlandia frutescens (L.) R. Br. A separation by HPLC was achieved by using a reversed phase (RP-18) column, PDA with ELS detection, and a water/acetonitrile gradient as the mobile phase. The wavelength used for quantification of four flavonoids with the diode array detector was 260 nm. Owing to their low UV absorption, the cycloartanol glycosides were detected by evaporative light scattering. The method was validated for linearity, repeatability, limits of detection (LOD) and limits of quantification (LOQ). The limits of detection and limits of quantification of eight compounds were found to be in the range from 0.1 to 7.5 microg/mL and 0.5 to 25 microg/mL, respectively. The analysis of products showed considerable variation of 1.099-5.224 mg/average weight for the major compound, sutherlandioside B. The eight compounds in plant sample and products of S. frutescens were further confirmed by LC-ESI-TOF. This method involved the use of the [M+H](+) and [M+Na](+) ions in the positive ion mode with extractive ion monitoring (EIM).

Isobolographic analysis of the sedative interaction between six central nervous system depressant drugs and Valeriana edulis hydroalcoholic extract in mice

It has been declared frequently that valerian may potentiate the effect of other central nervous system (CNS) depressant drugs, however there has been a lack of experimental data. We have evaluated the profile of the interactions between the ethanol extract of Valeriana edulis spp procera and six CNS depressant drugs using an exploratory model to test the sedative effect in mice. All the compounds tested showed a dose‐dependent sedative effect with the following ED50 values: valerian 181.62, diazepam 1.21, ethanol 1938, pentobarbital 11.86, buspirone 1.04, haloperidol 0.41 and diphenhydramine 17.06 mg kg−1. An isobolographic analysis was used to evaluate the sedative interaction of the intraperitoneal co‐administration of 1:1 fixed‐ratio combination of equi‐effective doses of valerian extract with each CNS depressant drug. The ED50 theoretical (Zadd) and experimental (Zexp) for each combination were: valerian + diazepam, Zadd = 91.41 mg kg−1, Zexp = 81.64 mg kg−1; valerian + ethanol, Zadd = 1060.22 mg kg−1, Zexp = 687.89 mg kg−1; valerian + pentobarbital, Zadd = 96.74 mg kg−1, Zexp = 151.83 mg kg−1; valerian + buspirone, Zadd = 91.33 mg kg−1, Zexp = 112.73 mg kg−1; valerian + haloperidol, Zadd = 91.01 mg kg−1, Zexp = 91.52 mg kg−1; valerian + diphenhydramine, Zadd = 99.34 mg kg−1, Zexp = 123.52 mg kg−1. Neither synergistic nor attenuate effects were found in any of the combinations evaluated. We concluded that the valerian extract did not potentiate the sedative effect of commonly prescribed CNS depressant drugs as was expected. The additive effect found through the isobolographic analysis suggested that the sedative effect of V. edulis resulted from the activation of common mechanisms of haloperidol, diazepam, buspirone, pentobarbital, diphenhydramine and ethanol.

Simultaneous determination of sesquiterpenes and pyrrolizidine alkaloids from the rhizomes of Petasites hybridus (L.) G.M. et Sch. and dietary supplements using UPLC-UV and HPLC-TOF-MS methods.

UPLC-UV and HPLC-TOF-MS methods have been developed for the analysis of major sesquiterpenes and pyrrolizidine alkaloids from rhizomes of Petasites hybridus (L.) G.M. et Sch. (Family, Asteracea) and dietary supplements claiming to contain P. hybridus. The best results were obtained with Acquity UPLC™ HSS T3 (100 mm × 2.1 mm, I.D., 1.8 μm) column system using a gradient elution with a mobile phase consisting of ammonium formate (50mM) and acetonitrile (0.05% formic acid) at a constant flow rate of 0.25 mL/min via UPLC-UV. The newly developed method was validated according to the ICH guidelines with respect to specificity, linearity, accuracy and precision. The limits of detection were found to be 5 μg/mL and 0.1 μg/mL for pyrrolizidine alkaloids and sesquiterpenes, respectively by UPLC-UV and 0.001 and 0.01 μg/mL, respectively using HPLC-TOF-MS. The methods were successfully used to analyze different P. hybridus market products, as well as to distinguish between two other Petasites species. The total content of petasins was found to be in the range of 0.02-11.6 mg/dosage form for 15 dietary supplements and no petasins were detected in an additional six dietary supplements. Additionally, pyrrolizidine alkaloids, which are considered to be toxic for the liver, were detected in seven dietary supplements. The amount of petasin in seven dietary supplements was found to be within limits of label claim and no pyrrolizidine alkaloids were detected. HPLC-mass spectrometry coupled with electrospray ionization (ESI) interface method is described for the identification and confirmation of sesquiterpenes and pyrrolizidine alkaloids from plant extracts and dietary supplements that claim to contain P. hybridus as well as different species of Petasites.

Quantitative Determination of Triterpenoids and Formononetin in Rhizomes of Black Cohosh (Actaea racemosa) and Dietary Supplements by Using UPLC-UV/ELS Detection and Identification by UPLC-MS

A UPLC-UV/ELSD method has been developed for analysis of major triterpenoids and formononetin in ACTAEA RACEMOSA L. (family Ranunculaceae) samples. The best results were obtained with an Acquity UPLC BEH C18 (100 mmx2.1 mm, i. d., 1 microm) column system using gradient elution with a mobile phase consisting of water and acetonitrile:methanol (7:3) at a constant flow rate of 0.3 mL/min. Owing to their low UV absorption, the triterpene saponins were detected by evaporative light scattering. Within 5.5 minutes, three main triterpenoid glycosides [cimiracemoside A, 23- EPI-26-deoxyactein, and actein] and an isoflavonoid, formononetin, could be separated, with detection limits of 5, 5, 10, and 0.01 microg/mL, respectively. The method was successfully used to analyze different Actaea racemosa market products as well as to distinguish between two other ACTAEA species. There was a significant variability in the amounts of the selected triterpene glycosides for the products containing black cohosh and rhizomes of black cohosh. The isoflavone formononetin was not detected in the samples analyzed. LC-MS coupled with the electrospray ionization (ESI) interface method is described for the identification of formononetin and triterpenoid glycosides in plant samples and dietary supplements that claim to contain black cohosh and different species of Actaea.

Transport of Schisandra chinensis extract and its biologically-active constituents across Caco-2 cell monolayers — an in-vitro model of intestinal transport

We have determined the intestinal transport of Schisandra chinensis extract and its lignans (gomisin A, gomisin N and schisandrin C) in the Caco‐2 cell monolayer model. The transport across monolayers was examined for 2 h in absorptive and secretory directions. Quantitation of lignans was performed by HPLC. Out of the three lignans, gomisin A exhibited bi‐directional transport, with Papp values in the range of 25–29 × 10−6 cm s−1, indicating a passive diffusion. Gomisin N, mixture and Schisandra extract displayed a higher transport in the secretory direction with efflux ratios in the range of 2.2–5.2. The efflux was decreased in the presence of inhibitors of multidrug resistance protein (MRP) transporter (MK‐571) and P‐glycoprotein (verapamil) indicating a possible involvement of an efflux pump and MRP in the transport of Schisandra lignans. Poor transport of schisandrin C was observed which could not be quantitated. The permeability of gomisin A in the isolated form was significantly different compared with the mixture or extract.

Evaluation of In Vitro Absorption, Distribution, Metabolism, and Excretion (ADME) Properties of Mitragynine, 7-Hydroxymitragynine, and Mitraphylline

Mitragyna speciosa (kratom) is a popular herb in Southeast Asia, which is traditionally used to treat withdrawal symptoms associated with opiate addiction. Mitragynine, 7-hydroxymitragynine, and mitraphylline are reported to be the central nervous system active alkaloids which bind to the opiate receptors. Mitraphylline is also present in the bark of Uncaria tomentosa (cats claw). Several therapeutic properties have been reported for these compounds but limited information is available on the absorption and distribution properties. This study focuses on evaluating the absorption, distribution, metabolism, and excretion (ADME) properties of these compounds and their effect on major efflux transporter P-glycoprotein, using in vitro methods. Quantitative analysis was performed by the Q-TOF LC-MS system. Mitragynine was unstable in simulated gastric fluid with 26 % degradation but stable in simulated intestinal fluid. 7-Hydroxymitragynine degraded up to 27 % in simulated gastric fluid, which could account for its conversion to mitragynine (23 %), while only 6 % degradation was seen in simulated intestinal fluid. Mitraphylline was stable in simulated gastric fluid but unstable in simulated intestinal fluid (13.6 % degradation). Mitragynine and 7-hydroxymitragynine showed moderate permeability across Caco-2 and MDR-MDCK monolayers with no significant efflux. However, mitraphylline was subjected to efflux mediated by P-glycoprotein in both Caco-2 and MDR-MDCK monolayers. Mitragynine was found to be metabolically stable in both human liver microsomes and S9 fractions. In contrast, both 7-hydroxymitragynine and mitraphylline were metabolized by human liver microsomes with half-lives of 24 and 50 min, respectively. All three compounds exhibited high plasma protein binding (> 90 %) determined by equilibrium dialysis. Mitragynine and 7-hydroxymitragynine inhibited P-glycoprotein with EC50 values of 18.2 ± 3.6 µM and 32.4 ± 1.9 µM, respectively, determined by the calcein-AM fluorescent assay, while no inhibition was seen with mitraphylline. These data indicate the possibility of a drug interaction if mitragynine and 7-hydroxymitragynine are coadministered with drugs that are P-glycoprotein substrates.