Reversed-phase ODS Column Research Articles

Environmental benign AQbD based estimation of ketoconazole and beclomethasone by RP-HPLC and multi-analytical UV spectrophotometric method

Hazardous chemicalsare one of the most significant adverse effects of drug analysis. Chemicalwaste generatedat every stage of the analysisfound to bein significant concentrations during quality testing. Green analytical chemistry provides a safe environment for drug analysis. Three simple and environmentally sustainable UV spectrophotometric andliquid chromatographic techniques were established to simultaneously measure Ketoconazole and Beclomethasone dipropionate in their bulk and cream formulation. Firstly, the spectrophotometric estimation deals with three green methods: absorption ratio, the first-order absorption ratio, and the area under curve methods. The second approach was green rotatable centralcomposite analytical quality by design-based RP-HPLC method with an optimized mobile phase of ethanol: 0.1 M potassium dihydrogen phosphate buffer (pH 2.5) 33: 67 v/v. The separation was accomplished using an ODS reversed-phase column (250 × 4.6 mm, 5 µm) operating at a 1.0 mL/min flow rate. The determination was performed using the concentrations ranging for Ketoconazole 140–260 µg/mL and Beclomethasone 1.75–3.75 µg/mL (R2 = 0.9991–0.9999). The results showed that the Forced degradation experiments indicate that the drugs degrade under their intended circumstances. Green solvents were utilized in the developed techniques since they significantly impact environmentally benign analytical methods. The proposed techniques have been validated in linearity, accuracy, and precision based on International Council on Harmonization guidelines and green assessment tools, with acceptable results. These techniques shall be employed in place of methods that utilized toxic chemicals for determination and are easily adaptable for routine analysis.

Development of HPLC method for estimation of glyoxylic acid after pre-column fluorescence derivatization approach based on thiazine derivative formation: A new application in healthy and cardiovascular patients’ sera

Glyoxylic acid (GA) is the intermediate metabolite in various mammalian metabolic pathways. GA showed high reactivity towards formation of advanced glycation end-products (AGEs); the main cause of pathogenesis and complications of many diseases. The presented study aimed to detect GA in healthy and cardiovascular patients’ (CV) sera; however analysis of GA in biological fluid is a challenge and requires chemical derivatization. Hence, a new, highly sensitive, time saving and reproducible pre-column fluorescence derivatization procedure coupled with high performance liquid chromatography (HPLC) method was developed. The derivatization method was based on reaction of 2-aminobenzenthiol (2-ABT), a fluorogenic reagent, with GA in acidic medium to form highly fluorescent thiazine derivative (290 and 390 nm for excitation and emission wavelengths respectively). The fluorescent derivative was separated within 6 min on a reversed-phase ODS column using an isocratic elution with a mixture of methanol–water (70:30, v/v%). The proposed method parameters were optimized and the method was validated. A good linearity in the concentration range (0.05–5.0 µM) was obtained with detection limit (LOD) of 10 nM (200 fmol/injection), which is more sensitive than several previous methods. Moreover, the recovery results were within the range of 85.0–95.5 % and the intra- and inter-day precision results were ≤3.5%. It should be emphasized that this method is the first one for monitoring of GA in CV patients; to investigate its role for diagnosis and monitoring the severity and complications of this disease in clinical laboratory.

Frying influence on virgin olive oils by monitoring change of aliphatic monoaldehydes

When virgin olive oil (VOO) have been fried long time at high temperature, some aliphatic monoaldehydes including hexanal, heptanal, octaldehyde, nonanal and decanal were found in fried VOO and food samples. The HPLC method has been developed for determination of aliphatic monoaldehydes in VOO and food samples after different fried conditions. These five aliphatic monoaldehydes could be separated and analyzed in 60 min on reversed phase ODS column with methanol/0.1% acetic acid water solution mobile phase at 1.0 mL min-1 flow rate, after being derivatized with 2,4-dinitrophenylhydrazine (DNPH). The method was validated with recovery results ranging from 86% to 114%. Calibration plots of aliphatic aldehydes were linear (r≥0.9991) in the concentration range from 1.0×10-6 to 1.5×10-4 mol L-1. The LODs were between 6.5×10-8 and 3.98×10-7 mol L-1. The proposed method provides a reliable and sensitive quantitative evaluation for aliphatic monoaldehydes in fried oils and fried potatoes samples. The experiments results verify that the VOOs are not suitable to be fried long time at high temperature. The frying time should be less than 3.0 min, and the frying temperature should be below 150 °C for the healthy diet.

Rapid analysis of neomycin in cochlear perilymph of guinea pigs using disposable SPE cartridges and high performance liquid chromatography-tandem mass spectrometry

Irreversible hearing loss induced by aminoglycoside in human through local or systemic administration route negatively impacts quality of life. The aim of this work was to develop and validate an analytical method suitable for the detection and quantification of neomycin in cochlear perilymph of guinea pig after local application. The SupelMIP SPE column was used for the pre-treatment of matrix. Chromatographic separation was conducted by a reversed phase ODS column (100 × 2.1 mm, 3 μm) at 40 °C in gradient mode with 0.2‰ (v/v) HFBA in water and 0.2‰ (v/v) HFBA in acetonitrile as mobile phase, at a flow rate of 0.30 mL/min, with retention time of 3.50 and 3.62 min for internal standard tobramycin and analyte neomycin, respectively. The MS was performed with positive ionization mode, with data acquisition in Multi Reaction Monitor (MRM) mode. This method was proved to be specific, accurate (97.1–115% of nominal values) and precise (CV% < 15%). Calibration curves for matrix matched standard of neomycin ranged from 1.25 to 200 μg/mL, with LOD and LLOQ of 0.625 and 1.25 μg/mL in blank matrix. The matrix effect was corrected to (−0.1) - 1.33 by adding internal standard. The relative SPE recovery values were ≥98.9% in low, medium and high QC samples. Neomycin in matrix proved to be stable under room temperature - and −20 °C, or under three freeze-thawing cycles, or under processing as well. Finally, the proposed method was successfully applied to a toxicokinetics study of neomycin in perilymph after round window membrane (RWM) administration, which was in accordance with threshold shift of auditory brainstem response (ABR) test related to hearing loss.

Improved Method for Determination of Raspberry Ketone in Fragrance Mist by HPLC-Fluorescence Analysis after Pre-Column Derivatization with 4-(&amp;lt;i&amp;gt;N,N&amp;lt;/i&amp;gt;-Dimethylaminosulfonyl)-7-(&amp;lt;i&amp;gt;N&amp;lt;/i&amp;gt;-chloroformylmethyl-&amp;lt;i&amp;gt;N&amp;lt;/i&amp;gt;-methylamino)-2,1,3-benzoxadiazole

Raspberry ketone {RK, 4-(4-hydroxyphenyl)butan-2-one} is structurally resembles 4-(4-hydroxyphenyl)-2-butanol, which causes leukoderma on consumers’ skin. Therefore, it is important to measure in cosmetics for quality assessment. Very recently, an HPLC-fluorescence method for determination of RK in a fragrance mist by pre-column derivatization with 4-hydrazino-7-nitro-2,1,3-benzoxadiazole hydrazine was established. However, the derivatization conditions (80°C, 20 min) were severe. In this study, an improved pre-column derivatization with 4-(N,N-dimethylaminosulfonyl)-7-(N-chloro-formylmethyl-N-methylamino)-2,1,3-benzoxadiazole (DBD-COCl) is presented by HPLC-fluorescence method for determination of RK. The DBD-CO-RK derivative was eluted from a reversed-phase ODS column, and detected with excitation at 440 nm and emission at 543 nm. Derivatization was performed at room temperature for 3 min. The retention time of DBD-CO-RK derivative was 16.8 min. The standard curve was linear in the range of 0.05 to 2.5 μg/mL, with a correlation coefficient (r2) value of 0.9988. The lower limit of detection was 0.01 μg/mL (absolute amount of 0.3 pmol). The coefficients of variation were less than 10.0%. The content of RK in fragrance mist (1.00 mL) was 1.20 ± 0.08 mg (range, 1.10 to 1.31 mg, n = 5). Recovery tests were satisfactory (91.8 ± 5.4%; range, 84.2 to 98.2%, n = 5).

Protein cross linker enzymes and food applications

Irreversible hearing loss induced by aminoglycoside in human through local or systemic administration route negatively impacts quality of life. The aim of this work was to develop and validate an analytical method suitable for the detection and quantification of neomycin in cochlear perilymph of guinea pig after local application. The SupelMIP SPE column was used for the pre-treatment of matrix. Chromatographic separation was conducted by a reversed phase ODS column (100 × 2.1 mm, 3 μm) at 40 °C in gradient mode with 0.2‰ (v/v) HFBA in water and 0.2‰ (v/v) HFBA in acetonitrile as mobile phase, at a flow rate of 0.30 mL/min, with retention time of 3.50 and 3.62 min for internal standard tobramycin and analyte neomycin, respectively. The MS was performed with positive ionization mode, with data acquisition in Multi Reaction Monitor (MRM) mode. This method was proved to be specific, accurate (97.1–115% of nominal values) and precise (CV% < 15%). Calibration curves for matrix matched standard of neomycin ranged from 1.25 to 200 μg/mL, with LOD and LLOQ of 0.625 and 1.25 μg/mL in blank matrix. The matrix effect was corrected to (−0.1) - 1.33 by adding internal standard. The relative SPE recovery values were ≥98.9% in low, medium and high QC samples. Neomycin in matrix proved to be stable under room temperature - and −20 °C, or under three freeze-thawing cycles, or under processing as well. Finally, the proposed method was successfully applied to a toxicokinetics study of neomycin in perilymph after round window membrane (RWM) administration, which was in accordance with threshold shift of auditory brainstem response (ABR) test related to hearing loss.

Simple HPLC-Fluorescence Determination of Raspberry Ketone in Fragrance Mist after Pre-Column Derivatization with 4-Hydrazino-7-nitro-2,1,3-benzoxadiazole

Raspberry ketone {RK, 4-(4-hydroxyphenyl)butan-2-one} is a natural compound contained in raspberry, and is added to cosmetics for skin whitening. It is very important to measure the RK level in cosmetics for quality assessment, since RK structurally resembles 4-(4-hydroxyphenyl)-2-butanol, which causes leukoderma on consumers’ skin. Here, we present a simple HPLC-fluorescence method for determination of RK in a fragrance mist by pre-column derivatization with 4-hydrazino-7-nitro-2,1,3-benzoxadiazole hydrazine (NBD-H), which reacts with the carbonyl group of RK. The NBD-RK derivative was eluted from a reversed-phase ODS column, and detected with excitation at 470 nm and emission at 550 nm. The retention time of NBD-RK derivative obtained by reaction with NBD-H at 80°C for 20 min was 10.3 min. The standard curve was linear in the range of 0.2 to 10 μg/mL, with a correlation coefficient (r2) value of 0.9980. The lower limit of detection was 0.018 μg/mL (absolute amount of 1.8 pmol). The coefficients of variation were less than 8.1%. The content of RK in fragrance mist (1.00 mL) was 1.18 ± 0.07 mg (range: 1.12 to 1.28 mg, n = 5). Recovery tests were satisfactory (83.9% ± 3.9%; range: 79.6 to 88.8%, n = 5).

Reversed Phase Column HPLC-ICP-MS Conditions for Arsenic Speciation Analysis of Rice Flour

New measurement conditions for arsenic speciation analysis of rice flour were developed using HPLC-ICP-MS equipped with a reversed phase ODS column. Eight arsenic species, namely, arsenite [As(III)], arsenate [As(V)], monomethylarsonic acid (MMAA), dimethylarsinic acid (DMAA), trimethylarsine oxide (TMAO), tetramethylarsonium (TeMA), arsenobetaine (AsB) and arsenocholine (AsC), were separated and determined under the proposed conditions. In particular, As(III) and MMAA and DMAA and AsB were completely separated using a newly proposed eluent containing ammonium dihydrogen phosphate. Importantly, the sensitivity changes, in particular those of As(V) and As(III) caused by coexisting elements and by complex matrix composition, which had been problematical in previously reported methods, were eliminated. The new eluent can be applied to C8, C18 and C30 ODS columns with the same effectiveness and with excellent repeatability. The proposed analytical method was successfully applied to extracts of rice flour certified reference materials.

Chromatographic determination of aliphatic aldehydes in human serum after pre-column derivatization using 2,2′-furil, a novel fluorogenic reagent

A novel, highly sensitive and selective fluorimetric liquid chromatographic method for simultaneous determination of medium chain aliphatic aldehydes was developed. The method was based on the derivatization of aliphatic aldehydes with 1,2-di(2-furyl)-1,2-ethanedione (2,2′-furil), a novel fluorogenic reagent, to form highly fluorescent difurylimidazole derivatives. The fluorescence derivatives were separated in less than 20min on a reversed-phase ODS column using an isocratic elution with a mixture of methanol–water (80:20, v/v%). The detection limits were from 0.19 to 0.50nM (1–10fmol/injection) at a signal-to-noise ratio (S/N) of 3. This method was successfully applied for monitoring of aliphatic aldehydes in healthy human sera by a simple pretreatment procedure without interferences from serum constituents.

Analysis of naltrexone and its metabolite 6-beta-naltrexol in serum with high-performance liquid chromatography.

BackgroundNaltrexone has been proven to be an effective treatment option for the treatment of alcohol dependency. In this article we introduce a reliable and simple method developed for the simultaneous determination of naltrexone and 6-β-naltrexol in human serum by using high-performance liquid chromatography (HPLC).FindingsLiquid-liquid extraction with butyl acetate from basic solutions (pH 9) was chosen for extraction with nalorphine as an internal standard (IS). Analytes were back-extracted from organic solvent into perchloric acid. The acid extract was chromatographed by HPLC with a reverse-phase ODS-column and electrochemical detector. The mobile phase was a NaH2PO4-solution with acetonitrile as an organic modifier and octanesulphonic acid and tetraethylammonium hydrogen sulphate as ion-pair reagents. The recovery of the extraction method was 48% for naltrexone and 75% for 6-β-naltrexol. The limit of quantification was 5.0 ng/ml for naltrexone and 1.0 ng/ml for 6-β-naltrexol. The analysed concentrations of naltrexone differed from the theoretic concentrations by 0.7 to 2.3% and those of 6-β-naltrexol by 2.6%. The relative standard deviation of within-day assay was from 0.9 to 5.7% for naltrexone and from 0.8 to 4.2% for 6-β-naltrexol; for the between-day assay it was 5.7% and 4.2%, respectively.ConclusionsOur results indicate that the developed method is suitable for determination of naltrexone and 6-β-naltrexol in human serum.

Determination of polycyclic aromatic hydrocarbons in edible oils and barbecued food by HPLC/UV–Vis detection

Determination of nine polycyclic aromatic hydrocarbons in corn, sunflower, olive oils and barbecued meat and fish by HPLC/UV–Vis method is described. The extraction procedure included a saponification, liquid–liquid extraction and finally purification of PAHs through a house-made silica–alumina column. Chromatographic determination was based on separation of PAHs on ODS column and measurement at 254nm. All polycyclic aromatic hydrocarbons were separated and analyzed in 12min on reversed phase ODS column with acetonitrile/water mobile phase at 1.5mLmin−1 flow rate. The detection limits of nine polycyclic aromatic hydrocarbons ranged from 0.26 to 1.15μgL−1 at a signal/noise ratio of 3. The linearity of the method was between 0.9951 and 0.9996. Oil samples contain different PAHs ranging from 0.44 to 98.92μgL−1. Barbecuing process increased the concentration (in the range of 2- to 8-fold) and caused the formation of PAHs in food samples.

Bioactive metabolites from Guignardia sp., an endophytic fungus residing in Undaria pinnatifida

Aim To isolate new and/or bioactive constituents from EtOAc extract of liquid culture of endophyte Guignardia sp. from the leaves of Undaria pinnatifida (Harv.) Sur. Methods Isolation and purification were performed through silica gel column chromatograph, Sephadex LH-20 and reversed-phase ODS column and the structures of the compounds obtained were identified through a combination of spectral and chemical methods (IR, MS, 1H and 13C NMR). In vitro bioactive assays including antifungal activity against three human pathogenic fungi Microsporum canis, Tricophyton rubrum and Epidermophyton floccosom and cytotoxic activity against the human nasopharyngeal epidermoid tumor KB cell line were evaluated. Results Seven compounds have been obtained from the liquid culture of the title endophyte: ergosterol peroxide (6, 22-diene-5, 8-epidioxyergosta-3-ol) ( 1), ergosterol ( 2), cyclo-(Tyr-Leu) ( 3), cyclo-(Phe-Phe) ( 4), cyclo-(Val-Leu) ( 5), cyclo-(Phe-Pro) ( 6) and cyclo-(Leu-Ile) ( 7). Compounds 1–3 and 6 inhibited the growth of M. canis with MICs of 10.0, 20.0, 50.0 and 5.0 μg·mL −1, respectively and compounds 1, 2 and 6 against T. rubrum with MICs of 15.0, 20.0 and 10.0 μg·mL −1, respectively and 1 and 6 against E. floccosom with MICs of 20.0 and 50.0 μg·mL −1, respectively. In addition, compounds 1, 3 and 6 exhibited cytotoxic activity against KB cell line with IC 50 of 20.0, 10.0, 10.0 μg·mL −1, respectively. Conclusion Compounds 1–7 were obtained from Guignardia sp. of U. pinnatifida for the first time, and compounds 1 and 6 had potent cytotoxic and antifungal activity.

Determination of tryptophan and kynurenine in human plasma by liquid chromatography–electrochemical detection with multi‐wall carbon nanotube‐modified glassy carbon electrode

A novel method was developed for the simultaneous determination of kynurenine and tryptophan by high-performance liquid chromatography with electrochemical detection at multi-wall carbon nanotube (MWCNT)-modified glassy carbon electrode. The separation and detection conditions were optimized. The typical HPLC experiments were conducted by using a reversed-phase ODS column with a mobile phase consisting of stock acetate buffer (pH 5)-methanol (4:1, v/v) using an isocratic elution at the flow rate of 1.0 mL/min. The obtained LODs for kynurenine and tryptophane were 0.5 and 0.4 µmol/L, respectively. The analytical method for human plasma samples was validated and confirmed by LC-UV and LC-MS. The recoveries were in the range of 84.8-110%, and the precision was lower than 5.9%.

Pharmacokinetics of Laetispicine and Its Brain Distribution in Rats

Laetispicine, which is a novel amide alkaloid isolated from the stem of Piper laetispicum, has been proven to possess antidepressant and antinociceptive effects. This study examined the pharmacokinetic characteristics of laetispicine in plasma and brain distribution in rats by a simple sensitive HPLC-UV method. The separation was performed on a reverse-phase ODS column (250 mm x 4.6 mm, i.d., 5 microm) with a mobile phase composed of acetonitrile-water (75:25, v/v) as the mobile phase at a flow rate of 1.0 ml/min with UV detection at 259 nm. The calibration curve of laetispicine in rat plasma showed excellent linear behavior between 0.005-5.0 microg/ml (r2 = 0.9992), and between 0.02-0.5 microg/ml (r2 = 0.9952) in rat brain samples. The lower limit of quantification (LLOQ) was found to be 0.005 microg/ml in rat plasma and 0.02 microg/ml in rat brain samples. This HPLC assay was a precise and reliable method for the analysis of laetispicine in pharmacokinetic studies. Laetispicine was rapidly and extensively transported across the blood-brain barrier (BBB) and distributed into different brain regions.

Simultaneous HPLC–UV determination of rhein and aceclofenac in human plasma

Diacerein and aceclofenac are prescribed for reducing the symptoms associated with osteoarthritis. We present a simple HPLC method with UV detection for simultaneous determination of rhein (the immediate metabolite of diacerein) and aceclofenac from human plasma samples. Sample preparation was accomplished through liquid–liquid extraction with ethyl acetate and chromatographic separation was performed on a reversed-phase ODS column. Mobile phase consisted of a mixture of acetate buffer and acetonitrile run under gradient at flow rate of 1.0 ml/min. Wavelength was set at 258 nm. The method was validated for linearity, accuracy, precision and stability. The calibration was linear over the range of 0.1–7.0 μg/ml for rhein and 0.5–20 μg/ml for aceclofenac using 500 μl plasma samples. Extraction recoveries were 85% for rhein and 70% for aceclofenac. The method can easily be adopted for high-throughput clinical and pharmacokinetic studies of above two-drug fixed dose combination formulations.

Validated LC Method, with a Chiral Mobile Phase, for Separation of the Isomers of Fexofenadine Hydrochloride

A high-performance liquid chromatographic method has been developed for resolution of the structural isomers of fexofenadine hydrochloride in the bulk drug. The isomers were resolved to baseline on a reversed-phase ODS column with pH 3 aqueous buffer–acetonitrile 60:40 containing 5 g L−1 β-cyclodextrin as mobile phase additive. The aqueous buffer was prepared by dissolving 6.8 g KH2PO4 in 1,000 mL water and adjusting to the pH 3.0 with orthophosphoric acid. Resolution for the structural isomers was not less than 3.0. The method was extensively validated and proved to be accurate and precise. The calibration plot was indicative of an excellent linear relationship between response and concentration over the range 0.025–3.750 ppm for the meta isomer. The limits of detection and quantification for the meta isomer were 0.003 and 0.009 ppm, respectively, for an injection volume of 20 μL. Recovery of the meta isomer from bulk drug samples of fexofenadine HCl was 98.43, 96.52, and 97.72% for addition of 0.5, 0.6, and 0.7%, respectively. The analytical solution was stable for 48 h. The method was found to be accurate, and suitable for quantitative determination of the meta isomer in the bulk drug; it can also be used as a stability-indicating method for assay of fexofenadine.

Determination and pharmacokinetic study of indirubin in rat plasma by high-performance liquid chromatography

A specific, simple and sensitive HPLC method with UV detection was developed and validated for the pharmacokinetic studies of indirubin in rat plasma for the first time. Indirubin, with osthole as the internal standard, was extracted from plasma samples by liquid-liquid extraction. Chromatographic separation was conducted on a reverse-phase ODS column (200 mm x 4.6 mm, i.d., 5 microm), using a mixture of methanol-water (75:25, v/v) as the mobile phase at a flow rate of 1.0 ml/min with UV detection at 289 nm. The calibration curve of indirubin was linear over the range of 6.5-1950 ng/ml in rat plasma. The lower limit of quantification (LLOQ) was found to be 6.5 ng/ml. The present method was successfully applied for estimating the pharmacokinetic parameters of indirubin following intravenous and intraperitoneal administration of indirubin to rats.

Validated Enantioselective LC Method, with Precolumn Derivatization with Marfey’s Reagent, for Analysis of the Antiepileptic Drug Pregabalin in Bulk Drug Samples

An enantioselective high-performance liquid chromatographic method, with precolumn derivatization with Marfey’s chiral reagent, sodium 2,4-dinitro-5-fluorophenyl-l-alanine amide, has been developed for resolution of the enantiomers of a new antiepileptic drug, pregabalin, in the bulk drug. The diastereomers of the pregabalin enantiomers were resolved to baseline on a reversed-phase ODS column with a 60:40 (v/v) mixture of aqueous 0.2% triethylamine (pH adjusted to 3.5 with dilute orthophosphoric acid) and acetonitrile as mobile phase. Resolution between the diastereomers was not less than five. The method was extensively validated and proved to be robust. The calibration plot was indicative of an excellent linear relationship between response and concentration over the range 750 (LOQ) to 7,500 ng L−1 for the R enantiomer. The limits of detection and quantification of the R enantiomer were 250 and 750 ng L−1, respectively, for an injection volume of 10 μL. Recovery of the R enantiomer from bulk drug samples of pregabalin ranged from 97.5 to 101.76%. Solutions of pregabalin in water and in the mobile phase were found to be stable for at least 48 h. The method was found to be suitable and accurate for quantitative determination of the R enantiomer in the bulk drug. It can be also used to test the stability of samples of pregabalin.

A rapid high performance liquid chromatographic determination of methyldopa in human serum with fluorescence detection and alumina extraction: Application to a bioequivalence study

A simple and ultra rapid high performance liquid chromatographic (HPLC) method coupled with alumina extraction and fluorescence detection was described for determination of methyldopa in human serum. The drug and an internal standard were adsorbed onto alumina and eluted using acidic methanol. The eluate was directly injected onto ODS reverse phase column using a mixture of phosphate buffer (0.05 M) containing triethylamine (100 microl/l, v/v; pH 2.3) and methanol (92:8, v/v) at a flow rate of 2.1 ml/min as the mobile phase. The fluorescence detector excitation and emission wavelengths were set at 270 and 320 nm, respectively. No interference in the assay from any endogenous substances or other concurrently used drugs was observed and the retention times of I.S. and the drug were 1.7 and 2.4 min, respectively with total run time (injection to injection) of less than 3.5 min. The limit of quantification was evaluated to be 2 ng/ml. Validity of the method was studied and the method was precise and accurate with a linearity range from 20 ng/ml to 5000 ng/ml. This method has been used in a randomized crossover bioequivalence study of two different methyldopa preparations in 24 healthy volunteers.

Rapid determination of the antiparasitic drugs flubendazole and febantel in feeds by HPLC with ultraviolet detection

A rapid and very effective analytical procedure for the simultaneous HPLC determination of two anthelmintics, Flubendazole (FLUB) and Febantel (FEBA), in swine and poultry feeds was developed and tested. The ground feed samples were extracted using dimethylformamide. The extracts were directly analyzed, without any purification, on a reversed-phase ODS column (150 mm × 4.6 mm, 5 μm) with acetonitrile–phosphate buffer (pH 3; 0.017 M) as mobile phase. Ultraviolet detection of FLUB and FEBA was carried out at 300 nm. The method was validated for specificity, linearity, accuracy, repeatability, limit of detection and limit of quantification. The limits of detection (LOD) of FLUB and FEBA in feeds, based on a detector signal-to-noise ratio of 3, were 3 mg kg −1 and the lowest levels tested in feeds by this procedure (limit of quantification, LOQ) were 10 mg kg −1. The mean recovery of FLUB and FEBA from spiked samples, in a concentration range of 10–200 mg kg −1, was 98% with a CV% of 4% and 99% with a CV% of 2%, respectively.