The simultaneous separation and determination of six flavonoids and troxerutin in rat urine and chicken plasma by reversed-phase high-performance liquid chromatography with ultraviolet-visible detection

The method of high-performance liquid chromatography (HPLC) with UV-vis detection was used and validated for the simultaneous determination of six flavonoids (puerarin, rutin, morin, luteolin, quercetin, kaempferol) and troxerutin in rat urine and chicken plasma. Chromatographic separation was performed using a VP-ODS column (150 mm x 4.6 mm, 5.0 microm) maintained at 35.0 degrees C. The mobile phase was a mixture of water, methanol and acetic acid (57:43:1, v/v/v, pH 3.0) at the flow rate of 0.8 mL/min. Six flavonoids and troxerutin were analyzed simultaneously with good separation. On optimum conditions, calibration curves were found to be linear with the ranges of 0.10-70.00 microg/mL (puerarin, rutin, morin, luteolin, quercetin, kaempferol) and 0.50-350.00 microg/mL (troxerutin). The detection limits were 0.010-0.050 microg/mL. The method was validated for accuracy and precision, and it was successfully applied to determine drug concentrations in rat urine and chicken plasma samples from rat and chicken that had been orally administered with six flavonoids and troxerutin.     

High-performance liquid chromatography spectrometric analysis of tripterin in rat plasma

A quick, precise and reliable HPLC method has been developed to determine tripterin in rat plasma. After liquid-liquid extraction, the analytes was analyzed on a Discovery ODS C(18) column (5microm, 4.6mmx250mm) with an isocratic elution consisting of methanol-water-phosphoric acid (87:13:0.2, v/v/v). Ultraviolet detection was at 425nm. Using trioxymethylanthraquinone as an internal standard, the assay was linear over the concentration range of 0.025-1.60microg/mL (r(2)=0.9988). The extraction recovery of tripterin in rat plasma was more than 62%. The intra- and inter-day precision was less than 13% (CV). This validated method was successfully applied to the pharmacokinetics of tripterin in rats.     

An HPLC method for determination of oridonin in rabbits using isopsoralen as an internal standard and its application to pharmacokinetic studies for oridonin-loaded nanoparticles

A simple and sensitive HPLC method has been developed and validated for the determination of oridonin (ORI) in rabbit plasma. A simple liquid-liquid extraction (LLE) method was applied to extract ORI and the internal standard (IS), isopsoralen, from rabbit plasma. Chromatographic separation of ORI and the IS was achieved with a Kromasil C18 5-mum column (250 mm x 4.6 mm) using methanol-water (50:50, v/v) as mobile phase at a flow rate of 1 mL/min. The ultraviolet (UV) detection wavelength was set at 241 nm. The lower limit of quantification (LLOQ) was 0.02 microg/mL. The calibration curves were linear over a concentration range of 0.02-10 microg/mL. The assay accuracy and precision were within the range of 95.1-113.5% and 5.4-8.6%, respectively. This HPLC method was applied successfully to the pharmacokinetic study of ORI-loaded poly(caprolactone)-poly(ethylene oxide)-poly(caprolactone) copolymer nanoparticles (ORI-PCL-PEO-PCL-NP) in rabbits, given as a single intravenous injection at the dose equivalent to 2mg of ORI/kg, and the pharmacokinetic parameters for ORI were compared with a single intravenous injection of a ORI solution at the same dose.     

Quantitation of niflumic acid in human plasma by high-performance liquid chromatography with ultraviolet absorbance detection and its application to a bioequivalence study of talniflumate tablets

A rapid and simple HPLC method with UV detection (288 nm) was developed and validated for quantitation of niflumic acid in human plasma, the active metabolite of talniflumate. After precipitation with 100% methanol containing the internal standard, indomethacin, the analysis of the niflumic acid level in the plasma samples was carried out using a reverse phase C18 CAPCELL PAK (5 microm, 4.6 mm x 250 mm) column. The chromatographic separation was accomplished with an isocratic mobile phase consisting of a mixture of 0.1M sodium acetate in water and acetonitrile (37:63, v/v), adjusted to pH 6.4. This HPLC method was validated by examining its precision and accuracy for inter- and intra-day runs in a linear concentration range of 0.02-5.00 microg/mL. Stability of niflumic acid in plasma was excellent, with no evidence of degradation during sample processing (autosampler) and 30 days storage in a freezer. This validated method was successfully applied to the bioequivalence study of talniflunate in healthy volunteers.     

Determination of epimedin C in rat plasma by reversed-phase high-performance chromatography after oral administration of Herba Epimedii extract

A simple high-performance liquid chromatographic (HPLC) method has been developed for the determination of epimedin C in rat plasma and applied to a pharmacokinetic study in rats after administration of Herba Epimedii extract. After addition of carbamazepine as an internal standard plasma samples were extracted with ethyl acetate. HPLC analysis of the extracts was performed on a Hypersil ODS2 analytical column using acetonitrile -0.4% acetic acid (25:75, v/v) as the mobile phase. The UV detector was set at 260 nm. The standard curve was linear over the range 0.05-4.0 microg/mL. The lower limit of quantification was 0.05 microg/mL. The HPLC method developed could be easily applied to the determination and pharmacokinetic study of epimedin C in rat plasma after giving the animals Herba Epimedii extract.     

High-performance liquid chromatography spectrometric analysis of trans-resveratrol in rat plasma

A HPLC method for determination of trans-resveratrol concentrations in rat plasma was developed. Plasma samples were treated with acetonitrile to deposit proteins. The analysis used a Hypersil ODS(2) C(18) column (5 microm, 4.6 mm x 250 mm) and methanol/distilled water as the mobile phase (flow-rate=1 mL/min). The UV detection wavelength was 303 nm, and chlorzoxazone was used as the internal standard. The calibration curve was linear over the range of 0.02-40 microg/mL with a correlation coefficient of 0.9997. This concentration range corresponds well with the plasma concentrations of resveratrol in pharmacokinetic studies. There was 98.7%, 91.3% and 84.4% recovery from 0.02, 0.4 and 40 microg/mL plasma samples respectively. The R.S.D. of intra- and inter-day assay variations were all less than 12%. This HPLC assay is a quick, precise and reliable method for the analysis of resveratrol in pharmacokinetic studies.     

Measurement of amoxicillin in plasma and gastric samples using high-performance liquid chromatography with fluorimetric detection

A rapid, selective and sensitive HPLC assay has been developed for the routine analysis of amoxicillin in rat plasma, gastric juice aspirate and gastric tissue which is applicable to low concentrations of amoxicillin (<1 microg mL(-1)) or small sample volumes. Amoxicillin was converted, via an internal rearrangement, to form a fluorescent product which was subsequently recovered using liquid-liquid extraction. A Kromasil ODS 3 microm (150 x 3.2 mm I.D.) column was maintained at 40 degrees C and used with a mobile phase consisting of methanol-water (55:45, v/v). Fluorimetric detection was at an lambda(ex) of 365 nm and an lambda(em) of 445 nm. The limits of quantitation for amoxicillin were 0.1 microg mL(-1) for gastric juice aspirate (500 microL), 0.5 microg mL(-1) for plasma (50 microL) and 0.075 microg g(-1) for gastric tissue (250 mg). The method was linear up to at least 15 microg mL(-1) in gastric juice aspirate, up to 200 microg mL(-1) in plasma and up to 100 microg g(-1) in gastric tissue, with inter- and intra-day RSDs being less than 19%. The assay has been applied to the measurement of amoxicillin in rat plasma, gastric juice aspirate and gastric tissue for pharmacokinetic studies in individual rats.     

Determination and pharmacokinetics of orientin in rabbit plasma by liquid chromatography after intravenous administration of orientin and Trollius chinensis Bunge extract

A high-performance liquid chromatography (HPLC) method was developed and validated for the determination of orientin in rabbit plasma using ultraviolet (UV) absorbance detection. Orientin is the active constituent of purified herbal extract (TRO PE) from the flower of Trollius chinensis Bunge. Protein precipitation was used as the sample preparation technique. A Diamonsil C18 column (150 mmx4.6 mm, 5 microm) was equilibrated with a mobile phase composed of 0.1% acetic acid/methanol/acetonitrile (80/5/15, v/v/v). The calibration curve of orientin in rabbit plasma was linear in the concentration range of 0.530-53.0 microg/mL. This validated method was successfully applied to a pharmacokinetic study in rabbits after the intravenous administrations of orientin and TRO PE at three different doses.     

Evaluation of human neutrophil elastase inhibitory effect of iridoid glycosides from Hedyotis diffusa

Five iridoid glycosides were isolated from the MeOH extract of Hedyotis diffusa, and their structures were elucidated as E-6-O-p-methoxycinnamoyl scandoside methyl ester (1), Z-6-O-p-methoxycinnamoyl scandoside methyl ester (2), E-6-O-p-feruloyl scandoside methyl ester (3), E-6-O-p-coumaroyl scandoside methyl ester (4), and Z-6-O-p-coumaroyl scandoside methyl ester (5) by interpretation of their spectroscopic data. All the isolated compounds were evaluated for human neutrophil elastase inhibitory effect, and compound 1 showed potent activity with an IC₅₀ value of 18.0 μM. The molecular docking simulation suggested a structural model for the inhibition of human neutrophil elastase by compound 1.     

Selective and rapid liquid chromatography/negative-ion electrospray ionization mass spectrometry method for the quantification of valacyclovir and its metabolite in human plasma

A rapid, sensitive and specific method was developed for the quantification of valacyclovir and acyclovir in human plasma. Sample preparation was performed by protein precipitation with acetonitrile followed by filtration. Valacyclovir, acyclovir and ganciclovir (internal standard) were separated isocratically on a reversed-phase porous graphitized carbon analytical column (2.1 mm x 125.0 mm i.d., particle size 5 microm), using a mobile phase of acetonitrile/water with 0.05% (v/v) diethylamine (50:50, v/v) at a flow rate of 0.15 mL min(-1) in 4.0 min. Detection was performed by negative electrospray ionization using the selected ion monitoring mode of the deprotonated molecular ions at m/z 323.0 for valacyclovir, 224.0 for acyclovir and 254.0 for ganciclovir. The assay had linear calibration curves over the range 0.020-0.800 microg mL(-1) for valacyclovir and 0.100-20.00 microg mL(-1) for acyclovir. Accuracy and precision were within the acceptance limit of 15%. The method was successfully applied to the analysis of plasma samples obtained from patients after oral administration of valacyclovir.     

Determination of inositol hexanicotinate in rat plasma by high performance liquid chromatography with UV detection

A HPLC method with UV detection at 262nm was developed to analyze inositol hexanicotinate in rat plasma. Plasma samples were extracted with an equal volume of acetonitrile, followed by dilution with mobile phase buffer (5mM phosphate buffer, pH 6.0) to eliminate any solvent effects. Inositol hexanicotinate and the internal standard (mebendazole) were separated isocratically using a mobile phase of acetonitrile/phosphate buffer (35:65, v/v, pH 6.0) at a flow rate of 1.0mL/min and a reverse-phase XTerra MS C(18) column (4.6mmx150mm, 3.5microm). The standard curve was linear over a concentration range of 1.5-100.0microg/mL of inositol hexanicotinate in rat plasma. The HPLC method was validated with intra- and inter-day precisions of 1.55-4.30% and 2.69-21.5%, respectively. The intra- and inter-day biases were -0.75 to 19.8% and 2.58-22.0%, respectively. At plasma concentrations of 1.5-100microg/mL, the mean recovery of inositol hexanicotinate was 99.6%. The results of a stability study indicated that inositol hexanicotinate was unstable in rat plasma samples, but was stable in acetonitrile extracts of rat plasma for up to 24h at 4 degrees C. The assay is simple, rapid, specific, sensitive, and reproducible and has been used successfully to analyze inositol hexanicotinate plasma concentrations in a pharmacokinetic study using the rat as an animal model.     

Determination of salirasib (S-trans,trans-farnesylthiosalicylic acid) in human plasma using liquid chromatography-tandem mass spectrometry

A liquid chromatography/tandem mass spectrometric (LC/MS/MS) assay was developed for the quantitative determination of salirasib (S-trans,trans-farnesylthiosalicylic acid, FTS) in human plasma. Sample pretreatment involved liquid-liquid extraction with methyl t-butyl ether of 0.5-mL aliquots of lithium heparin plasma spiked with the internal standard, S-trans,trans-5-fluoro-farnesylthiosalicylic acid (5-F-FTS). Separation was achieved on Waters X-Terra C(18) (50 mm x 2.1 mm i.d., 3.5 microm) at room temperature using isocratic elution with acetonitrile/10 mM ammonium acetate buffer mobile phase (80:20, v/v) containing 0.1% formic acid at a flow rate of 0.20 mL/min. Detection was performed using electrospray MS/MS by monitoring the ion transitions from m/z 357.2-->153.0 (salirasib) and m/z 375.1-->138.8 (5-F-FTS). Calibration curves were linear in the concentration range of 1-1000 ng/mL. A 5000 ng/mL sample that was diluted 1:10 (v/v) with plasma was accurately quantitated. The values for both within day and between day precision and accuracy were well within the generally accepted criteria for analytical method (<8.0%). This assay was subsequently used for the determination of salirasib concentrations in plasma of cancer patients after oral administration of salirasib at a dose of 400 mg.     

Development and validation of an HPLC method for the determination of dibenzoylmethane in rat plasma and its application to the pharmacokinetic study

A highly sensitive and simple high-performance liquid chromatographic (HPLC) assay has been developed and validated for the quantification of dibenzoylmethane (DBM) in rat plasma. DBM and internal standard (I.S.) 1-(5-chloro-2-hydroxy-4-methylphenyl)-3-phenyl-1,3-propanedione (CHMPP) were extracted from rat plasma by ethyl acetate/methanol (95:5, v/v) and analyzed using reverse-phase gradient elution with a Phenomenex Gemini C18 5-mum column. A gradient of mobile phase (mobile phase A: water/methanol (80:20, v/v) with 0.1% TFA and mobile phase B: acetonitrile with 0.1% TFA) at a flow rate of 0.2 mL/min, and ultraviolet (UV) detection at 335 nm were utilized. The lower limit of quantification (LLOQ) using 50 microL rat plasma was 0.05 microg/mL. The calibration curve was linear over a concentration range of 0.05-20 microg/mL. The mean recoveries were 80.6+/-5.7, 83.4+/-1.6 and 77.1+/-3.4% with quality control (QC) level of 0.05, 1 and 20 microg/mL, respectively. Intra- and inter-day assay accuracy and precision fulfilled US FDA guidance for industry bioanalytical method validation. Stability studies showed that DBM was stable in rat plasma after 4h incubation at room temperature, one month storage at -80 degrees C and three freeze/thaw cycles, as well as in reconstitute buffer for 48 h at 4 degrees C. The utility of the assay was confirmed by the successful analysis of plasma samples from DBM pharmacokinetics studies in the rats after oral and intravenous administrations.     

A sensitive assay for simultaneous determination of plasma concentrations of valganciclovir and its active metabolite ganciclovir by LC/MS/MS

A protein precipitation, liquid chromatography/tandem mass spectrometry (LC/MS/MS) method has been developed and validated for the simultaneous determination of valganciclovir and its active metabolite ganciclovir in human plasma. The solvent system also served as a protein precipitation reagent. The chromatographic separation was achieved on an Aquasil C18 column (50 mm x 2.1mm, 5 microm). A linear gradient mobile phase between 0.02% formic acid and methanol was used. Detection was by positive ion electrospray tandem mass spectrometry on a Sciex API3000. The standard curves, which ranged from 4 to 512 ng/mL for valganciclovir and from 0.1 to 12.8 microg/mL for ganciclovir, were fitted to a 1/x weighted quadratic regression model. The method was proved to be accurate, specific and sensitive enough and was successfully applied to a pharmacokinetic study.     

Determination of mitiglinide in rat plasma by high-performance liquid chromatography with UV detection

A selective and sensitive high-performance liquid chromatography method has been developed and validated for determination of mitiglinide (MGN) in rat plasma using 2-(4-biphenylyl) propionic acid (BPA) as internal standard. Liquid-liquid extraction was used for sample preparation. Chromatographic separation was achieved on a C(18) column using acetonitrile and 0.02 mol/l KH(2)PO(4) buffer (pH 4.0) (45:55, v/v) as mobile phase delivered at 1.0 ml/min. The UV detector was set at 210 nm. The assay was linear over the range 0.1-20 microg/ml for MGN. The average extraction recoveries of MGN and BPA from rat plasma were 98.6 and 97.4%, respectively. The developed method has been applied to the pharmacokinetic study of MGN in rats.     

Pharmacokinetics and tissue distribution of docetaxel by liquid chromatography-mass spectrometry: evaluation of folate receptor-targeting amphiphilic copolymer modified nanostructured lipid carrier

A novel amphiphilic copolymer, folate-poly(PEG-cyanoacrylate-co-cholesteryl cyanoacrylate) (FA-PEG-PCHL) was synthesized to modify docetaxel-loaded nanostructured lipid carrier to lead to a long blood circulating effect and targeting ability for the delivery of antitumor drug in cancer. To investigate the characteristics of modified docetaxel-loaded nanostructured lipid carrier in vivo, a liquid chromatography-mass spectrometry method was developed and validated for the determination of docetaxel in rat plasma and tumor-bearing mouse tissue samples. The biosamples were extracted by liquid-liquid extraction method with ether and separated on a C(18) column (150 mm×4.6 mm, 5 μm) using a mobile phase consisting of methanol-0.01% formic acid water (82:18, v/v). The standard curves were linear over the ranges of 0.01-4.0 μg/mL for plasma and 0.02-8.0 μg/g for tissue samples, respectively. The validated method was successfully applied to the pharmacokinetic study in rat plasma and tissue distribution study in mouse tissues of docetaxel after an intravenous administration of docetaxel injection (DTX injection), docetaxel-loaded nanostructured lipid carrier (DTX-NLC) and FA-PEG-PCHL-modified docetaxel-loaded nanostructured lipid carrier (FA-DTX-NLC), respectively. The results indicated that the FA-DTX-NLC led to significant differences in pharmacokinetic profile and tissue distribution. Nanostructured lipid carrier modified by FA-PEG-PCHL could be one of the promising suspensions for the delivery of docetaxel in cancer.     

顺式藁苯内酯口服给药在大鼠血浆和脑内的药动学特性(英文)

建立大鼠血浆和脑中Z-槀苯内酯(LIG)浓度测定的高效液相色谱法。采用Agilent Hypersil ODS C18色谱柱(150mm×4.6mm,5μm),流动相为甲醇-5%异丙醇水溶液(60:40,v/v),流速为1.0mL/min,检测波长为280nm。血浆与脑中槀苯内酯浓度线性检测范围分别为93.75~3750ng/m(r=0.9999)和93.75~3750ng/g(r=0.9997),日内及日间精密度RSD10%。本法适用于大鼠口服LIG后血浆及脑中药物浓度的研究。     

Development of a liquid chromatography/tandem mass spectrometry assay for the determination of bestatin in rat plasma and its application to a pharmacokinetic study

Bestatin is a low molecular weight aminopeptidase inhibitor originally isolated from culture filtrates of Streptomyces olivoreticuli. We have developed a sensitive, specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the quantitative determination of bestatin in rat plasma using granisetron as the internal standard. The analyte and internal standard were isolated from 50 microL plasma samples by solid phase extraction (SPE). Reverse-phase HPLC separation was accomplished on a Lichrospher C18 column (4.6 mm x 50 mm, 5 microm) with a mobile phase composed of methanol-water-formic acid (70:30:0.5, v/v/v) at a flow rate of 0.8 mL/min. The method had a chromatographic total run time of 3 min. A Varian 1200L electrospray tandem mass spectrometer equipped with an electrospray ionization source was operated in selected reaction monitoring (SRM) mode with the precursor-to-product ion transitions m/z 309.2-->120.0 (bestatin) and 313.4-->138.0 (granisetron) used for quantitation. The method was sensitive with a lower limit of quantitation (LLOQ) of 5 ng/mL, with good linearity (r2 >or= 0.999) over the linear range of 5-2000 ng/mL. All the validation data, such as accuracy, precision, and inter-day repeatability, were within the required limits. The method was successfully applied to pharmacokinetic study of bestatin in rats.     

Determination of lamivudine in plasma, amniotic fluid, and rat tissues by liquid chromatography

An HPLC method for the quantification of lamivudine (3TC) in rat plasma, amniotic fluid, placental and fetal tissues has been developed, validated and applied to the study of the placental transport of this drug in the pregnant rat. Placental and fetal tissues were processed using liquid-liquid extraction enhanced by salting out the sample using a saturated solution of ammonium sulfate. Plasma and amniotic fluid samples were processed by protein precipitation using 2 M perchloric acid. Reverse phase chromatography was performed using a phenyl column (5 microm, 150 mm x 2 mm i.d.) under a flow rate of 0.2 ml/min. The mobile phase consisted of 5% methanol in 20 mM dibasic phosphate buffer (pH 6). The method was validated over the range from 0.1 to 50 microg/ml for plasma and amniotic fluid and 0.2-50 microg/ml for the placental and fetal tissues.