Determination of two metabolites of calycosin-7-O-beta-D-glucopyranoside in rat urine by HPLC

A simple and specific analytical method for the simultaneous determination of the two metabolites of calycosin-7-O-beta-D-glucopyranoside, calycosin-7-O-beta-D-glucuronic acid methyl ester (M-1) and calycosin (M-2), in rat urine was developed using high-performance liquid chromatography. Quercetin was employed as an internal standard. The correlation coefficients of the calibration curves were higher than 0.999; both intra- and inter-day precisions of two metabolites were determined and their RSD did not exceed 10%. The accuracy and linear range were investigated in detail. The cumulative urinary excretions of two metabolites were measured.     

A simple and sensitive HPLC‐UV determination of 7‐O‐succinyl macrolactin A in rat plasma and urine and its application to a pharmacokinetic study

A simple, sensitive and reproducible isocratic reversed‐phase (C₁₈) high‐performance liquid chromatography (HPLC) method was developed to determine 7‐O‐succinyl macrolactin A (SMA) in rat plasma and urine samples using UV detector set at 230 nm. Lamotrigine was used as internal standards (IS) to ensure the precision and accuracy of the method. The retention times of SMA and IS for the plasma sample were 9.2 and 4.4 min, respectively, and those for the urine samples were 7.9 and 4.3 min, respectively. The intra‐ and inter‐day variations of the analytical responses, expressed in terms of relative standard deviation, were less than 14.9%. The accuracy, in terms of average analytical recovery, ranged from 90.4 to 119%. The lower limits of quantification of SMA in rat plasma and urine samples were 0.02 and 0.1 µg/mL, respectively. This method is applicable for the pharmacokinetic studies of SMA in rats. Copyright © 2012 John Wiley & Sons, Ltd.     

Solvent effect in 1H NMR spectra of 3'-hydroxy-4'-methoxy isoflavonoids from Astragalus membranaceus var. mongholicus

Four 3'-hydroxy-4'-methoxy-isoflavonoids: calycosin-7-O-beta-D-glucopyranoside (1), calycosin (2), pratensein-7-O-beta-D-glucopyranoside (3), and pratensein (4) were isolated from Astragalus membranaceus var. mongholicus, among which compound 4 was obtained from this plant for the first time. The solvent effect obscuring (1)H signal patterns of B ring of compounds 1-4 was reported to avoid mis-assignments. Previously reported NMR data of compounds 1 and 2 were corrected based on 1D and 2D NMR experiments.     

Determination of caudatin-2,6-dideoxy-3-O-methy-beta-d-cymaropyranoside in rat plasma using liquid chromatography-tandem mass spectrometry

A selective, sensitive and rapid liquid chromatography-tandem mass spectrometry method has been developed for the determination of caudatin-2,6-dideoxy-3-O-methy-beta-d-cymaropyranoside (CDMC) in rat plasma. This method involves a plasma clean-up step using liquid-liquid extraction, followed by LC separation and positive electrospray ionization mass spectrometry detection (LC/ESI-MS/MS). Chromatographic separation of the analytes was achieved using a C(18) column with a mobile phase of acetonitrile and water (70:30, v/v) at a flow rate of 1.0 mL/min. Low energy collision tandem mass spectrometric analysis (CID-MS/MS) using the multiple reaction monitoring (MRM) mode was used for analyte quantification. For the MRM analysis of CDMC, the following transition at m/z 658.4 --> 529.6 derived from the protonated molecule [M + Na](+). A calibration curve was linear in the 5-500 ng/mL range for CDMC, and the limit of detection was 5 ng/mL. The inter- and intra-day precisions (RSD) were 相似文献   

Determination of fluoxetine and norfluoxetine in rat plasma by HPLC with pre-column derivatization and fluorescence detection

Both fluoxetine (FLX) and its N-demethylated metabolite, norfluoxetine (NFLX), have been reported to be potent serotonin-reuptake inhibitors. A sensitive and reliable method that allows simultaneous quantification of their plasma levels would be valuable and was developed in this work. The procedure included extraction of FLX and NFLX from plasma, fluorescence derivatization with 4-(N-chloroformylmethyl-N-methyl) amino-7-nitro-2,1,3-benzoxadiazole (NBD-COCl), separation of the derivatives on an octadecylsilica column with acetonitrile-water (55:45,v/v) as mobile phase and fluorescence detection with emission at 537 nm and excitation at 478 nm. The calibration curves were linear for FLX and NFLX concentration over the range of 10-1000 nM (r = 0.9992 and r = 0.9997) and the limits of quantitation were 10 nM in 100 micro L of plasma. Precision of intra- and inter-day RSD of less than 12% and accuracy of intra- and inter-day RE within -6.0-13% were achieved. The method described was applied to analysis of the plasma samples from rats treated with FLX hydrochloride and to the pharmacokinetic study.     

Determination of the uronic acid composition of seaweed dietary fibre by HPLC

A high-performance liquid chromatographic (HPLC) method is described for determination of the ratio of beta-d-mannuronic acid to alpha-l-guluronic acid (M/G ratio) in dietary fibre of edible seaweeds. Total dietary fibre (TDF) content was determined gravimetrically. The TDF fraction was hydrolysed with 12 m and 1 m H(2)SO(4), then neutralized with AG 4 x 4 resin. The uronic acids were separated in a Tracer Extrasil SAX 5 micro m column (25 cm x 4 mm) at 35 degrees C, with 2 mm KH(2)PO(4) containing 5% methanol as mobile phase at a fl ow rate of 1.5 mL/min. The detection wavelength was UV 210 nm. The chromatographic identifications of beta-d-mannuronic acid and alpha-l-guluronic acid were confirmed by liquid chromatography-mass spectrometry (LC-MS). The method precision was 1.4% for beta-d-mannuronic acid and 3.5% for alpha-l-guluronic acid. The method was used to determine M/G ratio in canned seaweeds (Saccorhiza polyschides and Himanthalia elongata) and in dried seaweeds (H. elongata, Laminaria ochroleuca, Undaria pinnatifida, Palmaria sp. and Porphyra sp.).     

血、尿中氯硝西泮及其代谢物7-氨基氯硝西泮的GC-ECD法检测

报道了血、尿中氯硝西泮及其代谢物7－氨基氯硝西泮的GC－ECD检测方法。苯－异戊醇碱性条件下（pH10.8）液－液萃取，灵敏度较高，氯硝西泮和7－氨基氯硝西泮的检测限（LOD）分别为3．2ng/mL及1．7ng/mL。线性范围5－300ng/mL，RSD5．3％。     

High-performance liquid chromatographic method for the determination of mangiferin in rat plasma and urine

A reversed-phase high-performance liquid chromatography assay for mangiferin in rat plasma and urine was developed. Rutin was employed as an internal standard. The mobile phase consisted of acetonitrile-water (16:84, v/v) containing 3% acetic acid at a flow rate of 1 mL/min. Detection was at 257 and 365 nm for mangiferin in plasma and urine, respectively. The limit of quantitation (LOQ) of mangiferin was 0.6 microg/mL in plasma, and 0.48 microg/mL in urine. The standard curve was linear from 0.6 to 24 microg/mL in plasma, and 0.48 to 24 microg/mL in urine, both intra- and inter-day precision of the mangiferin were determined and their RSD did not exceed 10%. The method provides a technique for rapid analysis of mangiferin in rat plasma and urine, which can be used in pharmacokinetic studies.     

Simultaneous HPLC determination of 14 tricyclic antidepressants and metabolites in human plasma

A HPLC method has been developed for the simultaneous determination of seven tricyclic antidepressants (TCAs) and seven metabolites in human plasma. The analyte separation was obtained using a C8 reversed phase column and a mobile phase composed of 68% aqueous phosphate buffer at pH 3.0 and 32% ACN. The UV detector was set at 220 nm and loxapine was used as the internal standard. A careful pre‐treatment procedure for plasma samples was developed, using SPE on C2 cartridges, which gives satisfactory extraction yields (>80%) and good sample purification. The LOQs were always lower than 9.1 ng/mL and the LODs always lower than 3.1 ng/mL for all analytes. The method was successfully applied to plasma samples from depressed patients undergoing therapy with one or more TCA drugs. Precision data (RSD <8.1%), as well as accuracy results (recovery >80%), were satisfactory and no interference from other drugs was found. Hence the method seems to be suitable for the therapeutic drug monitoring of patients treated with TCAs under monotherapy or polypharmacy regimens.     

5-雄烯-3β, 7α, 17β-三醇和5-雄烯-3β, 7β, 17β-三醇的合成

以5-雄烯二醇为原料,用微生物转化的方法合成了两个重要的神经甾体5-雄烯-3β, 7α, 17β-三醇和5-雄烯-3β, 7β, 17β-三醇。所用菌种总枝毛霉为我们自己筛选,并首次应用于5-雄烯-3β, 7α, 17β-三醇和5-雄烯-3β, 7β, 17β-三醇的合成中。     

Determination of erythromycin A in rat plasma and urine by high-performance liquid chromatography with chemiluminescence detection using Tris(2,2'-bipyridine)ruthenium(II)

A simple and sensitive method was developed for the determination of erythromycin A (EA), decladinosyl erythromycin A (dClEA) and erythromycin B (EB) in rat plasma and urine by high-performance liquid chromatography with electrogenerated chemiluminescence detection using Tris(2,2'-bipyridine)ruthenium(II). The recovery rates of EA, dClEA and EB were 97, 94 and 85% from rat plasma and 89, 83 and 93% from rat urine, respectively. The calibration curves were linear over the concentration ranges 0.05-5 microg/mL for plasma and 0.5-50 microg/mL for urine. The precision and accuracy for all analytes in rat plasma were < or =9.0 and -6.3-7.2%, and those in urine were < or =9.4% and -6.1-7.6%, respectively. This method proved to be a powerful tool for determination of EA, dClEA and EB concentrations in samples from rats.     

Simple and rapid high-performance liquid chromatographic method for endogenous alpha-tocopherol determination in human plasma

A simple and rapid reversed-phase high-performance liquid chromatographic method was developed and validated for the determination of endogenous alpha-tocopherol in human plasma. Following addition of alpha-tocopheryl acetate as the internal standard, the plasma was deproteinized using acetonitrile and isopropanol mixture prior to HPLC analysis. Methanol was used as the mobile phase and the effluent was quantitated at 292 nm. By this developed method, the concentrations of alpha-tocopherol were linearly related to their responses in the range of 0.8-30 microg/mL. The relative standard deviations intra-day and inter-day for alpha-tocopherol in plasma were less than 10%. The percentage of bias was within +/-4%, which confirmed the accuracy of the method. The method has been successfully applied for determining endogenous alpha-tocopherol in healthy Thai male volunteers.     

A rapid and sensitive LC–MS/MS method for quantification of quercetin‐3‐O‐β‐d‐glucopyranosyl‐7‐O‐β‐d‐gentiobioside in plasma and its application to a pharmacokinetic study

A rapid and sensitive LC–MS/MS method with good accuracy and precision was developed and validated for the pharmacokinetic study of quercetin‐3‐O‐β‐d ‐glucopyranosyl‐7‐O‐β‐d ‐gentiobioside (QGG) in Sprague–Dawley rats. Plasma samples were simply precipitated by methanol and then analyzed by LC–MS/MS. A Venusil® ASB C₁₈ column (2.1 × 50 mm, i.d. 5 μm) was used for separation, with methanol–water (50:50, v/v) as the mobile phase at a flow rate of 300 μL/min. The optimized mass transition ion‐pairs (m/z) for quantitation were 787.3/301.3 for QGG, and 725.3/293.3 for internal standard. The linear range was 7.32–1830 ng/mL with an average correlation coefficient of 0.9992, and the limit of quantification was 7.32 ng/mL. The intra‐ and inter‐day precision and accuracy were less than ±15%. At low, medium and high quality control concentrations, the recovery and matrix effect of the analyte and IS were in the range of 89.06–92.43 and 88.58–97.62%, respectively. The method was applied for the pharmacokinetic study of QGG in Sprague–Dawley rats. Copyright © 2016 John Wiley & Sons, Ltd.     

Selective extraction of clonazepam from human plasma and urine samples by molecularly imprinted polymeric beads

A molecularly imprinted polymer (MIP) based on free‐radical polymerization was prepared with 1‐(N,N‐biscarboxymethyl)amino‐3‐allylglycerol and N,N‐dimethylacrylamide as functional monomers, N,N‐methylene diacrylamide as the cross‐linker, copper ion‐clonazepam as the template and 2,2‐azobis(2‐methylbutyronitrile) as the initiator. The imprinted polymer was characterized by Fourier transform infrared spectroscopy, elemental analysis, thermogravimetric analysis, and SEM. The MIP of agglomerated microparticles with multipores was used for SPE. The imprinted polymer sorbent was selective for clonazepam. The optimum pH and sorption capacity were 5 and 0.18 mg/g at 20°C, respectively. The profile of the drug uptake by the sorbent reflects good accessibility of the active sites in the imprinted polymer sorbent. The MIP‐SPE was the most feasible technique for the extraction of clonazepam with a high recovery from human plasma and urine samples.     

Molecularly imprinted solid-phase extraction for the selective HPLC determination of ractopamine in pig urine

A method was developed for the determination of ractopamine in pig urine using molecularly imprinted solid-phase extraction (MISPE) as the sample clean-up technique combined with high-performance liquid chromatography. The molecularly imprinted polymer (MIP) was synthesized in acetonitrile-triethylamine system using ractopamine (RAC) as the template and acrylamide as the monomer. The binding capacity of the polymer toward RAC was found to be about 2.57 mg of ractopamine/g of polymer. The optimal procedures for MISPE consisted of conditioning with 3 mL methanol, equilibrating with 3 mL of water, loading volume of <10 mL of aqueous sample (pH 7), washing with 3 mL water and 3 mL methanol, and eluting with 5 mL of 5% ammonia in methanol. In the four spiked samples with the levels of 0.01, 0.1, 1.0 and 5.0 μg/mL, the mean recoveries of analyte on the MIP were higher than 90% with relative standard deviation <10%, and significant differences between imprinted and non-imprinted materials were observed. The MIP selectivity was evaluated by checking 11 drugs with similar and different molecular structures to that of RAC. The characteristics of three-dimensional cavities and hydrogen bond interaction were regarded as the main factors that dominated the retention of RAC on the MISPE cartridge.     

Separation and quantitation of several angiotensin II receptor antagonist drugs in human urine by a SPE-HPLC-DAD method

In this work, an SPE-HPLC method coupled to photodiode array detection was validated in human urine matrix, in order to monitor four antihypertensive angiotensin II receptor antagonist drugs in patients under cardiovascular treatment. For that purpose, experimental design was used. Quantitation was accomplished by the internal standard method. The obtained LOQs were 95, 113, 125, and 85 ng/mL for eprosartan, telmisartan, irbesartan, and valsartan, respectively. The intraday and interday precision and accuracy at four concentration levels in the working range (LOQ-15 microg/mL) were always lower than 11% RSD and 8% relative error. The urine samples proved to be stable during 4 h at room temperature, after three thaw-freeze cycles, and for 2 months at -20 degrees C. No interferences from other endogenous compounds or co-administered drugs were found. The method has been successfully applied to monitor the renal elimination of eprosartan and valsartan during 24 h.     

Identification and determination of major flavonoids in rat urine by HPLC-UV and HPLC-MS methods following oral administration of Dalbergia odorifera extract

Flavonoids are the main active constituents of Dalbergia odorifera. The excretion of the major flavonoids in rat urine after oral administration of D. odorifera extract was investigated by HPLC-UV and HPLC-MS methods. Utilizing the HPLC-MS technique, 18 flavonoids, including five isoflavones, four isoflavanones, four neoflavones, two flavanones, two chalcones and one isoflavanonol were identified in free form in a urine sample based on the direct comparison of the corresponding tR, UV maximum absorbance (lambda(max)) values and MS data with the authentic standards. The amounts of the prominent flavonoids, (3R)-4'-methoxy-2',3,7-trihydroxyisoflavanone and vestitone, were determined by HPLC-UV with the internal standard method, and the validation procedure confirmed that it afforded reliable analysis of these two analytes in urine after oral administration of D. odorifera extract.     

Simultaneous determination of ampicillin, cefoperazone, and sulbactam in pharmaceutical formulations by HPLC with beta-cyclodextrin stationary phase

An accurate and reproducible method for the simultaneous determination of ampicillin (AMP), sulbactam (SUL), and cefoperazone (CFP) in pharmaceutical formulations by using HPLC with beta-CD stationary phase was developed. It involved the use of the added tetraethylammonium acetate (TEAA) reagent, pH, and methanol as the significant parameters to find the optimum separation condition. A high resolution and selectivity of analytes was obtained by running the mobile phase in methanol-5 mM TEAA buffer = 35:65 (v/v, pH 4.5) at 280 nm. The mean recoveries ranged from 96.6 to 103.3% for AMP in the synthetic mixture, 97.6 to 103.0% for SUL, and 97.0 to 104.0% for CFP. The low LOD (<1.8 microg/mL) and low CV (<0.9%) assured that this method was sensitive and reproducible. The assay of analytes in commercial products exhibited that it was convenient and reproducible for routine analyses of these components in sterilized H(2)O, saline, or 5% dextrose injection solutions.     

Simultaneous analysis of sarin, pyridostigmine bromide and their metabolites in rat plasma and urine using HPLC

Summary A method was developed for the separation and quantification of the warfare nerve agent sarin (O-isopropylmethylphosphonoflouridate), its metabolite methylphosphonic acid, the anti nerve agent drug pyridostigmine bromide (PB;3-dimethylaminocarbonyloxy-N-methyl pyridinium bromide) and its metaboliteN-methyl-3-hydroxypyridinium bromide in rat plasma and urine. The method involved using solid phase extraction and high performance liquid chromatography (HPLC) with reversed phase C₁₈ column, and UV detection at 280 nm. The compounds were separated using gradient of 1% to 55% acetonitrile in 0.1% triflouroacetic acid water solution (pH 3.20) at flow rate of 0.9 ml/min in a period of 15 min. The retention times ranged from 4.4–12.1 min. The limits of detection were 50 ng mL⁻¹ for PB andN-methyl-3-hydroxypyridinium bromide, and 10 μg mL⁻¹ for sarin and methylphosphonic acid, while limits of quantitation were between 100 ng mL⁻¹–12 μg mL⁻¹. Average percentage recovery of five spiked samples from plasma were 84.6±8.4, 86.5±9.0, 76.4±8.5, 81.3±8.2, and from urine 78.5±7.9, 76.4±7.8, 74.4±8.4, 80.6±6.8 for sarin, methylphosphonic acid, pyridostigmine bromide andN-methyl-3-hydroxypyridinium bromide, respectively. This method was applied to analyze the above chemicals and metabolites following combined administration in rats.