Column-switching high-performance liquid chromatographic assay for the determination of quercetin in human urine with ultraviolet absorbance detection

There were analysed changes of circadian, circannual and decasecond biorhythms of the test tasks' performance of the control room operators in power stations for the age diapason 22-53 years. The decasecond rhythms' significance of the mental activity rate gradually increases with ageing. The circadian and circannual rhythms' significance for indices of attention and activity rate is minimum in the middle age group and it is maximum in the senior one. The circadian and circannual rhythms' significance for the logic-combinatorial task performance quality is maximum in the middle age group. The revealed differences have been connected with effect of the professional activity specificity on age changes of middle levels of different indices of the operators' mental activity. The complex analysis' results of the biorhythms' significance for the different indices of mental activity are proposed as a (prognostic) criterion of the operators' adjustment to the effective performance of the industrial activity at different times of day and year.     

Column-switching high-performance liquid chromatographic assay for determination of apigenin and acacetin in human urine with ultraviolet absorbance detection

Postweaning social isolation can influence the sensitivity of rats to several effects of drugs of abuse. The present study investigated the influence of postweaning housing conditions on the sensitivity of rats to the aversive effects of a number of psychoactive agents using a conditioned taste aversion (CTA) test procedure. Development of a CTA was assessed by pairing administration of the drug with the consumption of a 0.05% (weight/volume) saccharin solution in water-deprived (18 h) rats in a 20 min drinking period. Saccharin consumption was then measured in 20 min test sessions over the next 4 consecutive days. Consumption of saccharin solution was significantly reduced in both isolated and enriched rats following administration of d-amphetamine (2 mg/kg), cocaine (30 mg/kg), morphine (10 mg/kg), nicotine (1.0 mg/kg), caffeine (20 mg/kg), alcohol (1.5 g/kg), and LiCl (0.15 M, 4 ml/kg). There was no significant effect of housing conditions on the CTA induced by cocaine, nicotine, alcohol, or LiCl; however, isolation-reared rats were found to be less sensitive to the aversive effects of d-amphetamine, morphine, and caffeine in this paradigm. These results suggest that rearing rats in social isolation induces an attenuation in sensitivity to the aversive effects of some psychoactive agents.     

Automated high-performance liquid chromatographic determination of hydroxylysylpyridinoline and lysylpyridinoline in urine using a column-switching method

17 right-handed males volunteered for an experiment that compared task-related patterns of electromyographic (EMG) activation with data from muscle biopsy on proportion of slow-twitch ((ST) aerobic) to fast-twitch ((FT) anaerobic) muscle fibers. The biopsy was taken from the right-leg gastrocnemius muscle after EMG measurement from that area of the leg muscle. EMG was also recorded from the left forearm flexor carpi radialis area. Recordings were obtained from pre- and post-task resting periods and during 150 s of video-task performance when the right hand operated a joy-stick. The results showed a highly significant tonic EMG activation in the leg muscle of subjects with predominance of ST fibers, and this relationship generalized to the EMG from the 'passive' forearm. The proportion of ST to FT fibers is genetically defined and not altered by exercise. Therefore, our results lend support to a genetic differentiation between individuals with high vs. low probability of unintended build-up of muscle tension during perceptual-motor task performance.     

Simple high-performance liquid chromatographic method for determination of pentoxifylline in human plasma

A simple high-performance liquid chromatographic method using ultraviolet detection was developed for the determination of pentoxifylline in human plasma. Prior to analysis, pentoxifylline and the internal standard (chloramphenicol) were extracted from plasma sample using dichloromethane. The mobile phase comprised 0.02 M phosphoric acid adjusted to pH 4, methanol and tetrahydrofuran (55:45:1, v/v). Analysis was run at a flow-rate of 1.4 ml/min with the detector operated at a wavelength of 273 nm. The method was specific and sensitive with a detection limit of approximately 3.0 ng/ml at a signal to noise ratio of 3:1, while the limit of quantification was 12.5 ng/ml. Mean recovery value of the extraction procedure was about 99.9%, while the within-day and between-day coefficient of variation and percent error values of the assay method were all less than 10.0%. The calibration curve was linear over a concentration range of 12.5-400.0 ng/ml.     

Determination of 5-hydroxytryptophol in urine by high-performance liquid chromatography: application of a new post-column derivatization method with fluorometric detection

The aim of the present study was to develop a high-performance liquid chromatographic (HPLC) method for determination of the serotonin metabolite 5-hydroxytryptophol (5HTOL) in human urine. 5HTOL was liberated from its conjugated form by enzymatic hydrolysis and isolated by a sample clean-up procedure on a small Sephadex G-10 column. The eluate was injected onto an isocratically eluted C18 reversed-phase column and 5HTOL was converted into a fluorescent oxazole derivative by on-line post-column reaction with benzylamine in the presence of potassium hexacyanoferrate(III). The limit of detection was about 10 nM and the intra-assay coefficients of variation were below 4% with urine samples and standard solutions. The results indicate that the method can be used as a screening method to discriminate between normal and elevated levels of total (free + conjugated) 5HTOL in urine.     

Simple high-performance liquid chromatographic method for determination of losartan and E-3174 metabolite in human plasma, urine and dialysate

A simple high-performance liquid chromatographic (HPLC) method was developed for the determination of losartan and its E-3174 metabolite in human plasma, urine and dialysate. For plasma, a gradient mobile phase consisting of 25 mM potassium phosphate and acetonitrile pH 2.2 was used with a phenyl analytical column and fluorescence detection. For urine and dialysate, an isocratic mobile phase consisting of 25 mM potassium phosphate and acetonitrile (60:40, v/v) pH 2.2 was used. The method demonstrated linearity from 10 to 1000 ng/ml with a detection limit of 1 ng/ml for losartan and E-3174 using 10 microl of prepared plasma, urine or dialysate. The method was utilized in a study evaluating the pharmacokinetic and pharmacodynamic effects of losartan in patients with kidney failure undergoing continuous ambulatory peritoneal dialysis (CAPD).     

Sensitive and selective determination of methylenedioxylated amphetamines by high-performance liquid chromatography with fluorimetric detection

A rapid, sensitive and selective liquid chromatographic method with fluorimetric detection was developed for the separation and quantification of four methylenedioxylated amphetamines without interference of other drugs of abuse and common substances found in illicit tablets. The method was validated by examining linearity, precision and accuracy as well as detection and quantification limits. Methylenedioxylated amphetamines were quantified in eight tablets from illicit drug seizures and results were quantitatively compared to HPLC-UV analyses. To demonstrate the better sensitivity of the fluorimetric detection, methylenedioxylated amphetamines were analyzed in serum after a liquid-liquid extraction procedure and results were also compared to HPLC-UV analyses.     

Sensitive high-performance liquid chromatographic determination of cyclizine and its demethylated metabolite, norcyclizine, in biological fluids using coulometric detection

An accurate, sensitive, selective and reproducible high-performance liquid chromatographic method with coulometric detection for the determination of cyclizine and its inactive demethylated metabolite, norcyclizine, in biological fluids has been developed. The drugs were separated using a custom packed reversed-phase C18 analytical column and phosphate buffer (0.05 M, pH 3)-acetonitrile (7:3) as mobile phase. The dual electrode coulometric detector was operated in the "oxidative-screen" mode with the upstream electrode (detector 1) set at 0.55 V and the downstream electrode (detector 2) set at 0.90 V. Serum and urine samples were prepared for analysis by solid-phase extraction, followed by a simple phase-separation step. The limit of quantitation was 1 ng/ml for both cyclizine and norcyclizine in serum and urine.     

Sensitive chiral high-performance liquid chromatographic assay for labetalol in biological fluids

The four stereoisomers of the combined alpha- and beta-adrenoceptor antagonist labetalol were separated and quantified at therapeutic concentrations by normal-phase high-pressure liquid chromatography using a chiral stationary phase and fluorescence detection. Drug in plasma or urine was recovered by solid-phase extraction with 83+/-5% efficiency. Limits of detection from biological samples (3 ml) were between 1.5-1.8 ng ml(-1). Intra-day and inter-day variation at 25 ng ml(-1) were < or = 2.7% and < or = 5.80% respectively for all stereoisomers. The assay was applied to an examination of the disposition of labetalol stereoisomers after a single oral dose of racemate to a human volunteer. Labetalol appears to undergo enantioselective metabolism leading to relatively low plasma concentrations of the pharmacologically active enantiomers.     

Column-switching liquid chromatographic method for the simultaneous determination of iothalamic acid and creatinine in biological fluids

A column-switching liquid chromatographic method for simultaneous determination of iothalamate and creatinine in human serum and urine was developed. Iothalamate and creatinine were separated on a weakly acidic ion-exchange column (C1) by ion-exclusion chromatography and iothalamate excluded from the column was purified by gel chromatography on a hydrophilic gel column (C2) and then by ion-exchange chromatography on a weakly basic ion-exchange column (C3). Creatinine that was eluted from C1 after iothalamate was transferred to a hydrophilic gel column (C4) and then to a strongly acidic ion-exchange column (C5). The mobile phase for C1-C4 was a pH 3.8 propionate buffer (propionic acid-NaOH = 0.35 + 0.035 mol/kg in water) and a pH 5.6 propionate buffer (propionic acid-NaOH = 0.04 + 0.035 mol/kg in water) was used for C5. Diluted serum and urine samples could be injected directly on to C1, as the matrix of C1 is hydrophilic and C1 is backflushed after the transfer of the creatinine fraction from C1 to C4. Iothalamate and creatinine in the eluates were determined by measuring their ultraviolet absorption at 245 and 234 nm, respectively. The precision (R.S.D.) of the chromatographic method was 1.6% (n = 7) and 0.36% (n = 6) for diluted serum and urine with iothalamate concentrations of 1.0 and 10.0 mumol/l, respectively, and 0.85% (n = 7) and 0.55% (n = 7) for diluted serum and urine with creatinine concentrations of 5.77 and 272 mumol/l, respectively.     

Enzymatic method for the simultaneous determination of hyaluronan and chondroitin sulfates using high-performance liquid chromatography

Biological samples have a high dielectric constant that can shorten RF wavelengths by a factor of 8 relative to the vacuum. At high field strengths, finite wavelength effects within larger samples are the dominant cause of RF field nonuniformity. A coil design is presented that can reduce and even eliminate this inhomogeneity; 4-T images in phantoms and in the head of a normal volunteer are presented, which demonstrate improved homogeneity relative to a standard coil. This coil design should aid in realizing the potential advantages of imaging large samples at high field strengths.     

Simple high-performance liquid chromatographic method for the detection of phenylpropionylglycine in urine as a diagnostic tool in inherited medium-chain acyl-coenzyme A dehydrogenase deficiency

Deficiency of medium-chain acyl-CoA dehydrogenase is a frequent and treatable metabolic defect, which can be diagnosed by detection of phenylpropionylglycine in urine after an oral load of phenylpropionic acid. We studied the determination of phenylpropionylglycine in urine by isocratic ion-exclusion chromatography on a cation-exchange column using water-sulphuric acid (pH values between 2 and 4) as mobile phase. Phenylpropionylglycine, phenylpropionic acid and hippuric acid exhibited high retention factors with only a slight decline at increasing solvent pH. This resulted in a good separation from interfering substances after direct injection of urine. We hypothesize that pi-pi interactions between the aromatic carbonic acids and the ion-exchange resin are responsible for the strong retention on the stationary phase. We conclude that, even in asymptomatic patients, determination of phenylpropionylglycine in urine after a phenylpropionic acid load by ion-exclusion chromatography is a rapid and reliable diagnostic tool for the detection of medium-chain acyl-CoA dehydrogenase deficiency.     

Total homocysteine levels in plasma: high-performance liquid chromatographic determination with electrochemical detection and glassy carbon electrode

The suitability of the vacancy affinity capillary electrophoretic method (VACE) for study of displacement of a target drug from a protein by simultaneously administered drugs was investigated. As test system, the displacement of warfarin from bovine serum albumin (BSA) by furosemide and phenylbutazone was selected. It appears that the displacement can be observed well from the shift of the actual mobility of warfarin when a displacer drug is added. Also, the competitive action of the displacer drugs (affinity for BSA) is clearly visible. The VACE method seems to be attractive for rapid assessment of information about the competitive properties of coadministered compounds.     

High-performance liquid chromatographic determination of phenylephrine in human serum using column switching with fluorescence detection

A method for the determination of total phenylephrine (free plus conjugated) in human serum was developed using column-switching high-performance liquid chromatography (HPLC) with fluorescence detection. After serum was deproteinized with acetonitrile, the conjugated phenylephrine was hydrolyzed with diluted hydrochloric acid. The solution was evaporated to dryness. The reconstituted residue was analyzed with HPLC using a heart-cut technique. Good recoveries of the analytes from spiked human serum samples were obtained with small coefficients of variation. A good linear response was obtained for the concentration range 5-500 ng/ml. The lower limit of quantitation for phenylephrine in human serum was 5 ng/ml. The method was applied to the determination of phenylephrine in human serum after oral administration of phenylephrine hydrochloride.     

Comparison of high-performance liquid chromatographic serum profiles of humans and dogs

The sera of 30 healthy male beagles were analyzed by reversed-phase high-performance liquid chromatography. The profiles were compared with those obtained from the sera of 30 healthy human donors. The chromatograms of each group were very reproducible; however, there were characteristic differences between the two groups. The compounds observed in both the human and canine profiles were identified as creatinine, uric acid, tyrosine, hypoxanthine, xanthine, kynurenine, inosine and tryptophan. Compounds present only in the canine profiles were identified as cytindine, riboflavin and 5-methyl-cytidine. Compounds present only in the human profiles include uridine, guanosine, hippuric acid and the dietary dependent compounds theobromine and caffeine. The compounds present in both human and canine sera were quantitated and compared statistically. The amounts of these compounds were very similar, except for uric acid.     

Sensitive determination of a new antiarrhythmic agent, trecetilide, in plasma by high-performance liquid chromatography with fluorescence detection

Two high-performance liquid chromatography (HPLC) methods were developed for the determination of trecetilide in plasma samples. Differing only in the addition of a derivatization step and different detection wavelengths, the two methods encompassed a wide concentration range. In both methods, plasma samples (0.1 ml) with added internal standard were applied to solid-phase extraction discs containing a non-polar/strong cation mixed-phase, washed and eluted with an acetone-acetonitrile triethylamine mixture. The eluate was evaporated to dryness, and either reconstituted and directly injected onto an HPLC column or first derivatized with 1-naphthyl isocyanate before HPLC analysis. In both methods, the separation was performed isocratically on a cyano analytical column utilizing a mobile phase composed of acetonitrile-pH 7.9 phosphate buffer (70:30, v/v). The column effluent was monitored by fluorescence detection at 290/345 nm (with derivatization) or 235/320 nm (without derivatization). The limits of detection and quantitation of the assay were 0.57 and 1.9 ng/ml, respectively, when derivatization was used, or 4.3 and 14 ng/ml, respectively, without derivatization.     

High-performance liquid chromatographic determination of human serum albumin in plasma and urine by post-column fluorescence enhancement detection using 8-anilino-1-naphthalenesulfonic acid

A method for the quantitative determination of human serum albumin in plasma and urine by post-column high-performance liquid chromatography with fluorescence detection is presented. Human serum albumin in plasma and urine is separated by gel-permeation column chromatography, which is followed by fluorescence detection. The detection uses the fluorescence enhancement observed when 8-anilino-1-naphthalenesulfonic acid binds to human serum albumin. The method is rapid to perform and is highly sensitive (detection limit is 2.5 ng/injection volume at a signal-to-noise ratio of three.