Analysis of Arecoline in Rat Urine, and Identification of its Metabolites, by Liquid Chromatography–Tandem Mass Spectrometry

A simple and rapid high-performance liquid chromatographic–electrospray ionization (ESI) tandem mass spectrometric method has been developed for elucidation of the structures of the metabolites of arecoline in rat urine after administration of a single dose (20 mg kg^?1). The urine samples were purified on a C₁₈ solid-phase extraction cartridge and analysis was then performed on a reversed-phase C₁₈ column with 60:40 (v/v) methanol–0.01% triethylamine solution (2 mmol L^?1, adjusted to pH 3.5 with formic acid) as mobile phase and detection by on-line MS–MS. Identification of the metabolites and elucidation of their structures were performed by comparing molecular masses (ΔM), retention-times, and product ion spectra with those of the parent drug. The parent drug arecoline, four phase-I metabolites, and one phase-II metabolite were identified in rat urine.     

A Validated, Stability-Indicating, LC Method for Rabeprazole Sodium

We present a simple and reliable method for simultaneous determination of voriconazole and its main metabolite resulting from N-oxidation (UK-121,265), in human plasma. The work-up procedure used acetonitrile and potassium salts to precipitate plasma proteins. No internal standard was used. The chromatographic system used a LiChroCART^® 250-4 cartridge packed with LiChrospher^® 100 RP-8 (diameter particules, 5 μm). The UV monochromatic detector was set on 260 nm. The mobile phase consisted of a 60/40 (v/v) mixture of acetonitrile and water. The flow rate was 1 mL min^?1. The retention times for voriconazole and its metabolite were 8.98 and 4.02 min respectively, and total run time was 12 min. The linearity of the method was investigated from 0.31 to 10.0 mg L^?1; the lowest limit of quantification was 0.30 mg L^?1. Precision ranged from 2.41% to 6.32% for voriconazole and 0.80% to 11.6% for the N-oxide voriconazole metabolite. Accuracy was between 93.0% and 101% for voriconazole and 90.0% and 101% for the N-oxide voriconazole metabolite. This rapid and accurate method could be interesting to investigate metabolite/voriconazole ratio with respect to CYP2C19 genetic status and CYP3A4 activity changes.     

Determination of Piribedil in Human Serum, Urine and Pharmaceutical Dosage Form by LC-DAD

A rapid, selective and sensitive reversed-phase liquid chromatographic (LC) method was developed for the determination of piribedil in human serum, urine and pharmaceutical dosage form. LC analysis was carried out using reversed-phase isocratic elution with a C₁₈ column and a mobile phase of 0.01 M phosphate buffer-acetonitrile (50:50, v/v). The chromatograms showed good resolution and sensitivity with no interference of human serum and urine. Piribedil concentrations were determined using diode array detection at 240 nm. Sildenafil citrate was used as internal standard. The limit of quantification (LOQ) and limit of detection (LOD) concentrations were 107.2 and 321.6 pg mL^?1, 96.6 and 290.4 pg mL^?1, 161.7 and 53.9 pg mL^?1 for urine, serum and pharmaceutical dosage forms, respectively. The method was validated for its linearity, precision and accuracy and applied to the tablets, urine and human serum. In addition, the results were compared to those obtained from UV-spectrophotometry.     

Molecularly Imprinted Monolithic Column for Selective On-Line Extraction of Enrofloxacin and Ciprofloxacin from Urine

A simple on-line molecularly imprinted solid-phase extraction coupled with liquid chromatographic separation was developed for the simultaneous determination of enrofloxacin and its active metabolite ciprofloxacin from urine samples. The molecularly imprinted monolithic columns were synthesized in water-containing systems using norflorxacin as dummy template and methanol–water (5:1, v/v) as porogenic solvent, which reveal high affinity to enrofloxacin and ciprofloxacin in the aqueous environment. Due to the unique porous structure and flow-through channels existing in the network skeleton of the imprinted monolith, urine samples could be injected directly into the imprinted column, proteins and other biological matrix were quickly washed out and the analytes were selectively retained and enriched. Good linearity was obtained from 0.05 to 200 mg L^?1 with relative standard deviations less than 3.1%. The recoveries were higher than 87% at three different concentrations and the limit of detection of the method was 0.01 mg L^?1. Moreover, by increasing the injection volume, the sensitivity of the method could be further improved.     

Determination of Propylene Glycol in Low Volume Plasma and Urine Samples of Neonates by LC with Photodiode Array Detection

In order to enhance the sensitivity and to develop a method suitable for quantification of propylene glycol (PG) in low volume neonate plasma and urine samples, several steps in earlier described high performance liquid chromatographic methods were optimised. Chromatographic separation on a reversed-phase column and ultraviolet detection resulted in cleaner chromatograms without interfering compounds. Linearity of the standard curves was validated in the concentration range 0.25?C50 mg L^?1. The lower limit of quantification was 20 times lower than in earlier described methods. Presented method was suitable for quantification of PG concentrations in low volume neonate plasma (15?C46 mg L^?1) and urine samples (20?C175 mg L^?1) enabling us to document very low renal versus non-renal contribution of PG clearance in neonates.     

A Simple Method for Determination of Homologue Imidazolium Cations in Ionic Liquids Using IC with Direct Conductivity Detection

An investigation was carried out into the fast determination of five homologue imidazolium cations in ionic liquids by ion chromatography using a cation-exchange column and direct conductivity detection. Ethylenediamine, complex organic acid (citric acid, oxalic acid and tartaric acid) and organic modifiers (acetonitrile) were used as mobile phase. The influences of the eluent types, eluent concentration, eluent pH and column temperature on separation of the cations were discussed. Simultaneous separation and determination of the five homologue imidazolium cations in ionic liquids were achieved under an optimum condition. The optimized mobile phase was consisted of 0.25 mmol L^?1 ethylenediamine + 0.5 mmol L^?1 citric acid + 3% acetonitrile (v/v) (pH 4.1), set at a flow rate of 1.0 mL min^?1. The column temperature was 40 °C and detection limits were obtained in the range of 1.1–45.6 mg L^?1. The relative standard deviations of the chromatographic peak areas for the cations were <3.0% (n = 5). This method was successfully applied to separate imidazolium cations in ionic liquids produced by organic synthesis. The recoveries of spiked components were 92.5–101.9%.     

Determination of Racemic Ketorolac,Ketorolac Enantiomers and Their Metabolites in Human Plasma and Urine by LC–UV,Applied in Clinical Study During and After Pregnancy

In order to enhance the sensitivity and to develop a faster direct method for plasma and urine quantification of racemic ketorolac, its metabolites (p-hydroxy-ketorolac and ketorolac glucuronides) and ketorolac enantiomers, we developed an extraction procedure based on solid-phase extraction combined with specific and fast chromatographic separation. Extraction and chromatography resulted in cleaner chromatograms without interfering compounds. In both plasma and urine, linearity of the standard curves for racemic ketorolac and p-hydroxy-ketorolac was validated in the concentration range 0.025–10 mg L^?1, while for ketorolac enantiomers in the concentration range 0.025–5 mg L^?1. The lower limit of quantification was two times lower than in earlier described methods. The developed method was suitable for direct quantification of racemic ketorolac, p-hydroxy-ketorolac and ketorolac enantiomers in plasma and urine samples in women at delivery and in postpartum, enabling us to document significant intra-individual differences in pharmacokinetics between these physiological states.     

LC�CMS�CMS Method for Quantification of Hydralazine in BALB/C Mouse Plasma and Brain: Application to Pharmacokinetic Study

A rapid, sensitive and accurate high performance liquid chromatography method using tandem mass spectrometry detection for hydralazine in BALB/C mouse plasma and brain was developed and validated. The method involved a derivatization with 2,4-pentanedione at 50 °C for 1 h, and a step of solid phase extraction to purify and concentrate hydralazine derivative. Chromatographic separation was carried out on an Agilent ZORBAX SB-C18 column by elution with methanol?C0.01 mol L^?1 ammonium acetate (60:40, v/v). The multiple reaction monitoring transition used for quantification was m/z 225.2 ?? 129.5 in the electrospray positive ionization mode. Good linearity was obtained over the concentration range of 10?C200 ng mL^?1. The limits of detection were 0.49 and 1.05 ng mL^?1 for hydralazine in mouse plasma and brain, respectively. The limits of quantitation were 1.5 and 3.18 ng mL^?1 for hydralazine in mouse plasma and brain, respectively. Sample analysis time was 6 min including sample separation. The method was successfully applied to a pharmacokinetic study following intraperitoneal injection of hydralazine in BALB/C mice at the dose of 20 mg kg^?1.     

Simultaneous Determination of Ibuprofen and Diphenhydramine Citrate in Tablets by Validated LC

A stability-indicating LC method was developed for the simultaneous determination of ibuprofen and diphenhydramine citrate in pharmaceutical dosage forms. The chromatographic separation was achieved on an Inertsil ODS 3V, 150 × 4.6 mm, 5 μm, column. The mobile phase contained a mixture of 50 mM potassium dihydrogen phosphate buffer:acetonitrile:triethylamine:glacial acetic acid (55:45:0.2:0.2, v/v/v/v). This method allowed the determination of 2.85–9.14 mg mL^?1 of ibuprofen and 0.54–1.73 mg mL^?1 of diphenhydramine citrate, in a diluent consisting of pH 7.2, 50 mM potassium dihydrogen phosphate buffer:acetonitrile (40:60, v/v). The flow rate was 1.2 mL min^?1 and the detection wavelength was 260 nm. The limit of detection for ibuprofen and diphenhydramine citrate was 1.72 and 0.54 μg mL^?1 and the limit of quantification was 5.73 and 1.64 μg mL^?1, respectively. This method was validated for accuracy, precision and linearity. The method was also found to be stability indicating.     

Improving the Determination of Nitrovin in Feeds by Reversed-Phase LC with SPE

A simple reversed-phase liquid chromatographic method with ultraviolet detector (378 nm) for the determination of nitrovin in feeds was improved and validated. The mobile phase was a mixture of acetonitrile and 0.1% formic acid solution (v/v) in the ratio of 50:50 (v/v), and the flow rate was set at 1.2 mL min^?1. The extraction solution was a mixture of dimethyl formamide, acetonitrile and methanol (50:25:25, v/v), the sample was cleaned-up with reversed-phase solid phase extraction cartridge. The standard nitrovin was purified with crude nitrovin product by ethylene glycol monoethyl ether and identified by elemental analyzer. The limit of detection was 0.05 mg kg^?1 and the limit of quatification was 0.2 mg kg^?1 in feeds. The assay had satisfactory selectivity, recovery, linearity and precise repeatability and trueness.     

Matrix Solid-Phase Dispersion Extraction and UPLC Determination of Sudan Dyes in Chili Powder

A new method involving matrix solid-phase dispersion (MSPD) extraction and UPLC in conjunction with photodiode array detection was developed for the rapid and simple determination of Sudan dyes in chili powder. Separation of Sudan I, Sudan II, Sudan III, and Sudan IV was achieved within 2 min on the 1.7 μm Acquity UPLC BEH C18 column by using gradient elution with a mobile phase consisting of acetonitrile–water at a flow rate of 0.5 mL min^?1. Optimization of MSPD extraction parameters, such as type of solid sorbent and elution solvent were carried out. Optimal conditions selected for MSPD extraction were 0.25 g of sample, 0.5 g of silica gel as solid sorbent, and 7 mL of acetonitrile–methanol (9:1, v/v) as eluting solvent. Limits of detection ranged between 0.25 and 0.30 mg kg^?1 depending on the dye involved. All analytes provided average recoveries from spiked (at 1, 1.5, and 2 mg kg^?1) chili powder samples ranging from 81 to 106%. The method was applied to the analysis of chili powder samples obtained from different countries.     

Determination of Five Polyphenols by HPLC/DAD and Discrimination of Apple Varieties

The objective of this study was to set up a method to detect five compounds in fresh smashed apples by HPLC/DAD simultaneously. Different methods have been tested to control browning and ascorbic acid with ultrasonication was adopted. Methanol–water–acetic acid (30:69:1, v/v) containing 2.0 g of ascorbic acid L^?1 was chosen as the extract solvent. The method effectively simplified the sample treatment compared with the traditional ways. And primarily, the results were used to identify between different varieties. The chromatographic separation was performed on an Atlantis C18 (250 mm × 4.5 mm, particle size 5 μm) with a gradient elution program using a mixture of acetonitrile and 2% aqueous acetic acid (v/v) as mobile phase within 20 min at 270 nm wavelength. The variation of the content of five compounds was gallic acid (ND ~1.81 μg g^?1), protocatechuic acid (ND ~1.79 μg g^?1), chlorogenic acid (13.81–189.4 μg g^?1), caffeic acid (6.82–45.02 μg g^?1) and rutin (0.96–18.55 μg g^?1). The results could successfully be used to discriminate between different apple varieties (Gala, Fuji, Delicious, 8th Apple US, Golden Apple, Green Apple and Red Rose); chlorogenic acid and rutin being the polyphenols that contribute most to the differentiation.     

A Rapid Method for the Analysis of Ten Compounds in Psoralea corylifolia L. by UPLC

The objective of this study was to develop a simple, efficient and reliable method for routine quantitative analysis for Psoralea corylifolia L. An ultra performance liquid chromatography with DAD detector system was employed for simultaneous quantification of ten compounds. The chromatographic analysis was performed by UPLC with C₁₈ column and gradient elution of 0.05% formic acid aqueous solution and acetonitrile in 16 min. All calibration curves were linear (R ² ≥ 0.9990) over the tested ranges. The LOD and LOQ were lower to 13.07 and 39.22 ng mL^?1 with 2 μL of injection volume, respectively. The intra- and inter-day precisions as determined from sample solutions were below 4.1 and 4.2%. The average recoveries were ranged from 94.2 to 108.8% with RSDs ≤ 4.6%. This validated method was applied for the analysis of ten analytes in P. corylifolia L. from different origins. The variation of the content of ten compounds was remarkable among the tested samples: psoralenoside increased from 7.42 to 17.04 mg g^?1, isopsoralenoside from 6.05 to 14.34 mg g^?1, psoralen from 2.37 to 3.90 mg g^?1, isopsoralen from 2.53 to 3.65 mg g^?1, neobavaisoflavone from 1.59 to 2.96 mg g^?1, bavachin from 1.02 to 2.35 mg g^?1, psoralidin from 0.45 to 1.91 mg g^?1, isobavachalcone from 1.33 to 4.71 mg g^?1, corylifol A from 1.02 to 2.40 mg g^?1 and bakuchiol from 28.10 to 63.89 mg g^?1.     

Ion Chromatography in Combination with ESI Ion Trap MS for the Analysis of Diphacinone in Animal Livers

Analysis of diphacinone is often performed on C18 column by reserved phase liquid chromatographic methods coupled with ultraviolet detection or by ion-pair chromatography coupled with mass spectrometry. In this work, a sensitive and selective method using eluent generator reagent free ion chromatography coupled with electrospray ionization ion trap mass spectrometric detection for the quantification of diphacinone in animal livers has been developed. Samples were extracted with 10% (v/v) methanol in acetonitrile after spiking with warfarin (internal standard, IS) and cleaned by solid-phase extraction. The analytes were separated on an IonPac AS11 analytical column (250 mm × 4.0 mm) using 30.0 mmol L^?1 KOH (achieved by a KOH eluent generator) containing 10% (v/v) methanol as organic modifier at a flow rate of 1.0 mL min^?1. Detection was performed by negative electrospray ionization and ion trap tandem mass spectrometry in multiple reaction monitoring mode with a transition of m/z 339 → 167 for diphacinone, and 307 → 161 for IS, respectively. The calibration curve was linear (r ² = 0.993) in the concentration range of 1.0–200.0 ng mL^?1 with a lower limit of quantification of 0.3 ng g^?1 in animal livers. Intra- and inter-day relative standard deviations (RSDs) were less than 8.7 and 12.5%, respectively. Recoveries of diphacinone ranged from 81.2 to 89.5%. It was confirmed that this method could be used in clinical diagnosis or forensic toxicology.     

Determination of Veterinary Pharmaceuticals in Production Wastewater by HPTLC-Videodensitometry

An SPE-HPTLC method for simultaneous identification and quantification of seven pharmaceuticals in production wastewater was optimized and validated. The studied compounds were enrofloxacine, oxytetracycline, trimethoprim, sulfamethazine, sulfadiazine, sulfaguanidine and penicillin G/procaine. The method involves solid-phase extraction on hydrophilic-lipophilic balance cartridges with methanol and HPTLC analysis of extracts on CN modified chromatographic plates followed by videodensitometry at 254 and 366 nm. Optimization of chromatographic separation was performed by systematic variation of the mobile phase composition using genetic algorithm approach and the optimum mobile phase composition for TLC separation was 0.05 M H₂C₂O₄:methanol = 0.81:0.19 (v/v). Linearity of the method was demonstrated in the ranges from 1.5 to 15.0 μg L⁻¹ for enrofloxacine, 100–500 μg L⁻¹ for oxytetracycline, 150–600 μg L⁻¹ for trimethoprim, 300–1100 μg L⁻¹ for sulfaguanidine and 100–400 μg L⁻¹ for sulfamethazine, sulfadiazine and penicillin G/procaine with coefficients of determination higher than 0.991. Mean recoveries ranged from 74.6 to 117.1% and 55.1 to 108.0% for wellspring water and production wastewater, respectively. Only sulfaguanidine showed lower results. The described method has been applied to the determination of pharmaceuticals in wastewater samples from pharmaceutical industry.     

Simultaneous Determination of Acetaminophen, Caffeine, and Chlorphenamine Maleate in Paracetamol and Chlorphenamine Maleate Granules

A simple and rapid HPLC method using phenacetin (PHN) as internal standard has been developed for simultaneous determination of acetaminophen, caffeine, and chlorphenamine maleate in the product compound paracetamol and chlorphenamine maleate granules. Separation and quantitation were achieved on a 250 mm × 4.6 mm, 5 μm particle, C₁₈ column. The mobile phase was methanol 0.05 mol L^?1 aqueous KH₂PO₄ solution, 45:55 (v/v), containing 0.1% triethylamine and adjusted to pH 3.6 by addition of phosphoric acid; the flow rate was 1.0 mL min^?1. Detection of all compounds was by UV absorbance at 260 nm and elution of the analytes was achieved in less than 12 min. The linearity, accuracy, and precision of the method were acceptable to good over the concentration ranges 6.4–153.6 μg mL^?1 for acetaminophen, 5.0–120.0 μg mL^?1 for caffeine, and 9.6–230.4 μg mL^?1 for chlorphenamine maleate.     

LC–MS–MS Determination of Troxerutin in Plasma and Its Application to a Pharmacokinetic Study

A highly sensitive liquid chromatography–tandem mass spectrometry (LC–MS–MS) method for the determination of troxerutin in human plasma using tramadol as internal standard (IS) has been developed and validated. Sample preparation involved liquid–liquid extraction with ethyl acetate–isopropanol (95:5, v/v). The analyte and IS were separated by RP–LC with gradient elution using 10 mM ammonium acetate containing 0.1% formic acid and methanol at a flow rate of 0.9 mL min^?1. LC–MS–MS in the positive ion mode employed multiple reaction monitoring of the transitions at m/z 743.2→435.3 and m/z 264.1→58.0 for troxerutin and IS, respectively. The assay was linear in the concentration range 0.01–10 ng mL^?1 with precision and accuracy within assay variability limits as per FDA guidelines. The assay was successfully applied to a pharmacokinetic study involving oral administration of 300 mg troxerutin to eight healthy Chinese volunteers.     

Determination of Vitamins A and E Exploiting Cloud Point Extraction and Micellar Liquid Chromatography

Cloud point extraction and micellar chromatographic methods were developed for determination of vitamins A and E. The stationary phase was C18 and the mobile phase was 3.00% (w/v) SDS, 15.0% (v/v) butyl alcohol and 0.02 mol L^?1 phosphate buffer solution at pH 7.0. The retention times for vitamins A and E were 9.6 and 13.0, respectively. The extraction solution was 100 mmol L^?1 Triton X-100, 650 mg NaCl and 1.0% ascorbic acid at 70°C for 30 min. The method is precise (r.s.d. < 7%), the linear range was from 5.0 up to 360.0 mg L^?1 for both vitamins. Recovery test showed recuperation between 90.2 and 99.2%, and LOD and LOQ of 0.234 and 0.108 mg L^?1, 0.780 and 0.360 mg L^?1 to vitamins A and E were found.     

Fast Determination of Phenolic Compounds in Brazilian Wines from Vale do São Francisco Region by CE

Six phenolic compounds were separated and determined by capillary zone electrophoresis in red wine from Brazil’s region Vale do São Francisco with total analysis time of 12 min. The limit of detections varied from 1.59 to 2.24 mg L^?1. The relative standard deviations (for n = 6) varied from 0.28 to 3.50 %. The red wine samples analyzed were bought in the local market and the phenolic compound recoveries were in the range of 98–101 %. The concentrations of gallic acid in the samples of wines varied from 16.0 to 42.0 mg L^?1, caffeic acid (3.16–5.18 mg L^?1), syringic acid (5.73–13.0 mg L^?1), kaempferol (2.32–4.33 mg L^?1), quercetin (1.68–4.03 mg L^?1), myricetin (7.52–25.1 mg L^?1). The concentrations found agree with data reported in the literature.     

Quantification of Quetiapine in Human Plasma by LC–MS–MS

A rapid liquid chromatographic method with electrospray ionization tandem mass spectrometric detection has been developed and validated for quantification of quetiapine in heparinized human plasma. Plasma samples, without a drying and reconstitution step, were extracted by solid-phase extraction and eluted with acetonitrile. The analyte and zolpidem tartrate (internal standard, IS) were chromatographed on a C₁₈ column; the mobile phase was 85:15 (v/v) acetonitrile–5 mM ammonium formate, pH adjusted to 4.5 with formic acid, at a flow rate of 0.5 mL min^?1. The retention times of quetiapine and the IS were 1.25 and 1.05 min, respectively, and the run time was 1.8 min per sample. Selected reaction monitoring of MH⁺ at m/z 384.12 and 308.11 resulted in stable fragment ions with m/z 253.02 and 235.09 for quetiapine and the IS, respectively. Response was a linear function of concentration in the range 1.0–240.0 ng mL^?1, with r ≥ 0.9994. Recovery of quetiapine and the IS ranged from 74.82 to 85.57%. The assay has excellent characteristics and has been successfully used for analysis of quetiapine in healthy human subjects in a bioequivalence study.