gamma-Aminobutyric acid, catecholamine and indoleamine determinations from the same brain region by high-performance liquid chromatography with electrochemical detection.

A new procedure for the measurement of gamma-aminobutyric acid, norepinephrine, dopamine, serotonin and 5-hydroxyindoleacetic acid from the same brain region was developed. In general, two separate high-performance liquid chromatographic runs were performed, one for the gamma-aminobutyric acid determination and one for the determination of the monoamines. The electrochemical detection of gamma-aminobutyric acid was determined by a new procedure that utilized a small aliquot of the brain sample prepared for monoamine measurement. This assay was linear and parallel between 6 and 200 ng per 20-microliters injection with 5-aminovaleric acid utilized as an internal standard. Inter-assay variability averaged 5% throughout the assay with gamma-aminobutyric acid values in the gerbil hypothalamus of 344 micrograms/g. The catecholamine assay has been characterized previously and utilizes 3,4-dihydroxybenzylamine as an internal standard with less than 5% variability. Norepinephrine, dopamine, serotonin and 5-hydroxyindoleacetic acid levels in the gerbil hypothalamus averaged 2922, 729, 797 and 272 ng/g, respectively. This new protocol allows a wide range of neurochemicals to be determined and evaluated from the same brain region.     

Derivatization chemistries for determination of serotonin, norepinephrine and dopamine in brain microdialysis samples by liquid chromatography with fluorescence detection

The present paper provides an overview on currently developed derivatization chemistries and techniques for determination of monoamine neurotransmitters serotonin (5-HT), norepinephrine (NE) and dopamine (DA) in microdialysis samples by microbore liquid chromatography with fluorescence detection. In mild alkaline conditions, 5-hydroxyindoles and catecholamines react with benzylamine (BA), forming highly fluorescent 2-phenyl-4,5-pyrrolobenzoxazoles and 2-phenyl(4,5-dihydropyrrolo) [2,3-f]benzoxazoles, respectively. However, for derivatization of DA a higher fluorescence intensity was achieved for reaction with 1,2-diphenylethylenediamine (DPE) rather than with BA, therefore for simultaneous determination of 5-HT, NE and DA in brain microdialysates, a two-step derivatization with BA followed by DPE was developed. The detection limits for 5-HT, NE and DA were 0.2, 0.08 and 0.13 fmol, respectively, in an injection volume of 20 microL, which corresponds to concentrations of 30, 12 and 19.5 pm, respectively in standard solution prior to derivatization. The experimental data presented demonstrate the ability of the technique to simultaneously monitor neuronally releasable pools of monoamine neurotransmitters in the rat and mouse brains at basal conditions and following pharmacological treatments or physiological stimuli. These techniques play an important role in drug discovery and clinical investigation of psychiatric and neurological diseases such as depression, schizophrenia and Parkinson's disease.     

Electrospray ionization tandem mass spectrometry for the simultaneous determination of opiates and cocaine in human hair

A fast and highly sensitive electrospray ionization tandem mass spectrometry (ESI-MS/MS) method has been developed for the simultaneous determination of morphine, 6-methylacetylmorphine (6-MAM), codeine, cocaine and benzoylecgonine (BZE) in hair from drug abusers. Pulverized hair samples were subjected to an optimized matrix solid phase dispersion (MSPD) procedure with alumina, followed by diluted hydrochloric acid elution on column solid-phase extraction (SPE) clean-up/pre-concentration. Alternatively, samples were also subjected to an optimized ultrasound assisted enzymatic hydrolysis (USEH) with Pronase E, followed by an off-line SPE clean up/pre-concentration procedure. Positive electrospray ionization and multiple reaction monitoring (MRM) with one precursor ion/product ion transition were used for the identification and quantification (deuterated analogues of each target as internal standards) of each analyte. The chromatographic pump and the autosampler were used for injecting the standards and the hair extracts (20 μL) as a flow injection analysis mode. The highest sensitivity was achieved when delivering the targets with an acetonitrile/water/formic acid (80/19.875/0.125) mixture. The limits of detection of the method were 39.2, 4.4, 6.8, 7.0 and 7.4 ng g(-1) for morphine, 6-MAM, codeine, cocaine and BZE, respectively. Relative standard deviations of intra- and inter-day precision were lower than 9 and 12%, respectively; whereas, analytical recoveries ranged from 96±5 to 106±4%. The developed method (MSPD-ESI-MS/MS) was applied to different hair samples from polydrug abusers, and results were statistically compared to those obtained after a conventional gas chromatography-mass spectrometry (GC-MS) analysis and also after USEH and ESI-MS/MS or GC-MS determinations.     

Determination of trace amounts of morphine in human plasma by anodic adsorptive stripping differential pulse voltammetry

New adsorptive anodic differential pulse stripping voltammetry method for the direct determination of morphine at trace levels in human plasma of addicts is proposed.The procedure involves an adsorptive accumulation of morphine on a HMDE,followed by oxidation of adsorbed morphine by voltammetry scan using differential pulse modulation.The optimum conditions for the analysis of morphine are pH 10.5,Eacc of -100 mV（vs.Ag/AgCl）,and tacc of 120 s.The peak current is proportional to the concentration of morphine,and a Linear calibration graph is obtained at 0.01-3.10μg mL^-1.A relative standard deviation of 1.06%（n=5）was obtained,and the limit of detection was 3 ng mL^-1.The capabiLity of the method for the analysis of real samples was evaluated by the determination of morphine in spiked human plasma and addicts human plasma with satisfactory results.     

Sensitive assay for catecholamines in pharmaceutical samples and blood plasma using flow injection chemiluminescence analysis.

A rapid and sensitive chemiluminescence (CL) method using flow injection analysis was described for the determination of three catecholamines: dopamine, adrenaline and dobutamine, based on their greatly enhancing effects on the CL reaction of luminol-potassium periodate in basic solutions. Under the optimized conditions, the calibration graphs relating the increase of CL intensity to the concentration of the analytes were linear. The present method allows for the determination of dopamine, adrenaline, and dobutamine over the range of 1.0 x 10(-10) - 1.0 x 10(-7) g/ml. The relative standard deviations for measurements (n=11) of dopamine, adrenaline and dobutamine were 2.9, 2.3 and 1.8% when the concentrations of three catecholamines were at 1.0 x 10(-9) g/ml, respectively. The detection limits of the method were 2.0 x 10(-11) g/ml dopamine, 1.0 x 10(-11) g/ml adrenaline and 4.0 x 10(-11) g/ml dobutamine. The method was successfully applied to the determination of three catecholamines in pharmaceutical samples and blood plasma.     

Selective trace determination of dithiocarbamate fungicides in fruits and vegetables by reversed-phase ion-pair liquid chromatography with ultraviolet and electrochemical detection

The predominant methods for determination of dithiocarbamate fungicides (DTC) have been based on quantitation of carbon disulfide released by hot acid digestion. Because the subgroups of the DTC family differ in their chemical properties and toxicological behavior, selective determination is required. To meet the demand for a fast, simple, and sensitive procedure, a new reversed-phase ion-pair chromatographic method was developed, consisting of surface extraction followed by direct injection into a liquid chromatographic system equipped with ultraviolet and electrochemical detectors, connected in series. The procedure is applicable to residues of N-methyldithiocarbamates (metam-sodium), N,N-dimethyldithiocarbamates (e.g., ziram and ferbam), ethylenebisdithiocarbamates (e.g., nabam, maneb, zineb, and mancozeb), and propylenebisdithiocarbamates (e.g., propineb) in fruits and vegetables. Limits of quantitation, calculated according to the procedure of the Deutsche Forschungsgemeinschaft, are 9, 12, 8, and 12 microg CS2/L for N-methyl-DTC, N,N-dimethyl-DTC, ethylenebis-DTC, and propylenebis-DTC, respectively, when electrochemical detection is used.     

Determination of 6-acetylmorphine in urine as a specific marker for heroin abuse by high-performance liquid chromatography with fluorescence detection

A sensitive method is described for the determination of 6-acetylmorphine in urine, to detect the use of heroin by drug addicts participating in a morphine-dispensing experiment. The method is based on an extraction procedure, reaction of the isolated 6-acetylmorphine with excess of morphine to give a highly fluorescent mixed dimer, and determination by normal-phase high-performance liquid chromatography (h.p.l.c.) with fluorescence detection. The calibration graph was linear at low 6-acetylmorphine concentrations and the absolute detection limit was 0.4 ng. In urine samples, 6 μg l^?1 of 6-acetylmorphine could be detected. A reversed-phase system is also outlined.     

Determination of 5,5-diphenylhydantoin and its major metabolites in biological specimens by gas chromatography and selected ion-monitoring

Summary A specific and sensitive procedure for simultaneous determination of 5,5-diphenylhydantoin, 5-(4-hydroxyphenyl)-5-phenylhydantoin, 5-(3,4-dihydroxyphenyl)-5-phenylhydantoin and 5-(3-methoxy-4-hydroxyphenyl)-5-phenylhydantoin in biological specimens is described. The method involves formation of butylated or trideuteromethylated derivatives of the drug and its metabolites and analysis by gas chromatography. The lower detectable concentration of these compounds per ml of body fluids or g of wet tissue is either 1 g or 20 ng, depending on the type of detector used, flame ionization or selected-ion monitoring. Data on the urinary elimination of 5,5-diphenylhydantoin and hydroxylated metabolites in male rats following a single intraperitoneal injection of the drug (25 mg/kg) are also reported.     

Microassay for N-acetyltransferase activity using high-performance liquid chromatography with electrochemical detection

A rapid, sensitive procedure has been developed for determination of N-acetyltransferase activity against octopamine, dopamine and 5-hydroxytryptamine. The assay, which is performed in a volume of 10 microliters, is based upon the separation and detection of monoamine substrates and their N-acetylated derivatives using high-performance liquid chromatography with electrochemical detection. The method has been used to measure N-acetyltransferase activity against octopamine, dopamine and 5-hydroxytryptamine in the cerebral ganglion of the American cockroach, Periplaneta americana and to study bi-substrate kinetics of the enzyme.     

流动注射双安培法测定多巴胺

通过偶合多巴胺在铂电极上的氧化和高锰酸钾在铂电极上的还原,建立了一个不施加电压的条件下的流动注射双安培法直接测定多巴胺的新方法。以0.05 mol/L硫酸为载液,多巴胺的氧化峰电流与其浓度在0.8~160 mg/L范围内呈线性关系,线性回归方程为i(nA)=652.9C-239.2(r=0.9998,n=10),检出限为0.2 mg/L;RSD为2.86%(N=80 mg/L,n=14);进样频率为80次/h。本方法具有很高的选择性和灵敏度,样品处理方法简单快速,适于连续自动测定。用于实际样品的测定,结果满意。     

Direct determination of celiprolol in human urine using on-line coupled ITP-CZE method with fiber-based DAD

The present work illustrated possibilities of column-coupling electrophoresis combined with DAD for the direct quantitative determination of trace drug (celiprolol, CEL) in clinical human urine samples. ITP, on-line coupled with CZE, served as an ideal injection technique (high sample load capacity, narrow and sharp drug zone). Moreover, the ITP provided an effective on-line sample pretreatment (preseparation, purification and preconcentration of the drug) producing analyte zone suitable for its direct detection and quantitation in CZE stage. Spectral DAD in comparison with single wavelength ultraviolet detection enhanced value of analytical information (i) verifying purity (i.e., spectral homogeneity) of drug zone (according to differences in spectrum profiles when compared tested and reference drug spectra) and (ii) indicating zones/peaks with spectra similar to the drug spectrum (potential structurally related metabolites). The characterization of trace analyte signals superposed on the baseline noise was more definite thanks to the application of background correction and smoothing procedure to the raw DAD spectra (producing relevant spectral response). The proposed ITP-CZE-DAD method was characterized by favorable performance parameters for CEL in urine matrices {e.g., the lower limit of quantification was 9.7 ng/mL, RSD and relative error of the determinations were lower than 3% (precision) and 1% (accuracy), respectively, analyte peak exhibited spectral homogeneity (reflecting separation selectivity), separation efficiency was 84,500 theoretical plates} and successfully applied in a trial pharmacokinetic study of CEL.     

Flow injection with inhibited chemiluminescence method for the determination of dopamine hydrochloride.

A simple rapid and accurate flow injection inhibitory chemiluminescence method has been developed for the determination of dopamine hydrochloride based on its inhibition of the chemiluminescence from the luminol-potassium hexacyanoferrate(III) system. The linear range of determination is 4.0 x 10(-9) - 4.0 x 10(-7) g ml(-1) for dopamine hydrochloride and the detection limit is 1.14 x 10(-9) g ml(-1). The method has been applied to determine the content of dopamine in pharmaceutical preparation with satisfactory results.     

A simple and sensitive high‐performance liquid chromatography–electrochemical detection assay for the quantitative determination of monoamines and respective metabolites in six discrete brain regions of mice

A rapid, sensitive, and reproducible assay is described for the quantitative determination of the monoamine neurotransmitters dopamine, norepinephrine and serotonin, their metabolites, and the internal standard 3,4‐dihydroxybenzlyamine hydro‐bromide in mouse brain homogenate using high‐performance liquid chromatography with electrochemical detection. The method was validated in the following brain areas: frontal cortex, striatum, nucleus accumbens, hippocampus, substantia nigra pars compacta and ventral tegmental area. Biogenic amines and relevant metabolites were extracted from discrete brain regions using a simple protein precipitation procedure, and the chromatography was achieved using a C₁₈ column. The method was accurate over the linear range of 0.300–30 ng/mL (r = 0.999) for dopamine and 0.300–15 ng/mL (r = 0.999) for norepinephrine, 3,4‐dihydroxybenzlyamine hydro‐bromide, homovanillic acid and 5‐hydroxyindolacetic acid, with detection limits of ~0.125 ng/mL (5 pg on column) for each of these analytes. Accuracy and linearity for serotonin were observed throughout the concentration range of 0.625–30 ng/mL (r = 0.998) with an analytical detection limit of ~0.300 ng/mL (12 pg on column). Relative recoveries for all analytes were approximately ≥90% and the analytical run time was <10 min. The described method utilized minimal sample preparation procedures and was optimized to provide the sensitivity limits required for simultaneous monoamine and metabolite analysis in small, discrete brain tissue samples.     

Simultaneous determination of monoamine transmitters, precursors and metabolites in a single mouse brain

A simple and sensitive procedure for simultaneous determination of monoamine transmitters and related substances including precursors and metabolites has been developed for a single mouse brain. The proposed procedure involves (1) primary butanol extraction, (2) separation of the substances into either acid or alkaline aqueous layers according to their physicochemical properties, and (3) determination by means of high-performance liquid chromatography with electrochemical detection. Three transmitters (noradrenaline, dopamine and 5-hydroxytryptamine) and their precursors (tyrosine, 3,4-dihydroxyphenylalanine and tryptophan) and major metabolites (normethanephrine, 3-methoxy-4-hydroxyphenylethylene glycol, 3-methoxytyramine, 3,4-dihydroxyphenylacetic acid, 3-methoxy-4-hydroxyphenylacetic acid and 5-hydroxyindoleacetic acid) were selectively separated and sensitively detected in mouse whole brain sample. Although 3-methoxy-4-hydroxymandelic acid was also separated from other substances by authentic chromatography, the substance was not detected in mouse brain. Changes in levels of these substances were examined for drugs whose effects had been previously confirmed. These changes reflected putative effects of the drugs and confirmed that the procedure is effective for neurochemical research into the transmitter system.     

Sensitive determination of norepinephrine,epinephrine, dopamine and 5‐hydroxytryptamine by coupling HPLC with [Ag(HIO6)2]5−–luminol chemiluminescence detection

Based on the enhancing effects of norepinephrine (NE), epinephrine (EP), dopamine (DA) and 5‐hydroxytryptamine (5‐HT) on the chemiluminescence (CL) reaction between [Ag(HIO₆)₂]^5? and luminol in alkaline solution, a high‐performance liquid chromatography (HPLC) method with CL detection was explored for the sensitive determination of monoamine neurotransmitters for the first time. The UV–visible absorption spectra were recorded to study the enhancement mechanism of monoamine neurotransmitters on the CL of [Ag(HIO₆)₂]^5? and luminol reaction. The HPLC separation of NE, EP, DA and 5‐HT was achieved with isocratic elution using a mixture of aqueous 0.2% phosphoric acid and methanol (5:95, v/v) within 11.0 min. Under the optimized conditions, the detection limits of NE, EP, DA, and 5‐HT were 4.8, 0.9, 1.9 and 2.3 ng/mL, respectively, corresponding to 17.6–96.0 pg for 20 μL sample injection. The recoveries of monoamine neurotransmitters in rat brain were >95.6% with the precisions expressed by RSD <5.0%. The validated HPLC‐CL method was successfully applied for the quantification of NE, EP, DA and 5‐HT in rat brain. This method has promising potential for some biological and clinical investigations focusing on the levels of monoamine neurotransmitters. Copyright © 2016 John Wiley & Sons, Ltd.     

Determination of aluminum in large volume parenteral drug products used in total parenteral nutrition therapy by ICP-MS with a dynamic reaction cell

The proposed method was developed for the determination of aluminum (Al) in large volume parenteral (LVP) drug products used in total parenteral nutrition (TPN) therapy. The determination of Al in LVP drug products was performed by an inductively coupled plasma mass spectrometer equipped with a dynamic reaction cell (DRC-ICP-MS). DRC-ICP-MS conditions for the analysis of Al were studied to obtain the best signal to background (S/N) ratios. The interfering polyatomic ions at mass 27 (Al) were reduced by using NH(3) as a reaction gas. The detection limit of Al in a 1% (v/v) HNO(3) aqueous solution was 2 ng/l. The Al contents in LVP drug products obtained by this method were in the range of 1.16-4.33 microg/l and were less than 25 microg/l, that is, the regulation value of Food and Drug Administration (FDA). In order to trace the origin of Al in LVP drug products, each part of the LVP drug product, which is composed of three chambers, was investigated. However, a clear difference of the Al contents in each chamber was not observed. Furthermore, the Al contents in injection bags were quantified. Although the Al contents in injection bags were relatively high (in the range of 27.5-33.6 microg/g), dissolution of Al from the injection bags was not observed in the stability testing. From all of these results, it was concluded that the Al contents in the LVP drug products investigated originated in the amount of the Al in each raw material.     

Kinetic Spectrophotometric Determination of Ranitidine

Two spectrophotometric methods were developed for the determination of ranitidine. The first method was a kinetic spectrophotometric method based on the catalytic effect of ranitidine on the reaction between sodium azide and iodine in an aqueous solution. The calibration graph was linear from 4–24 μg/mL. The drug was determined by measuring the decrease in the absorbance of iodine at 348 nm using a fixed time method. The decrease in the absorbance after 1 minute from the initiation of the reaction was related to the concentration of drug. The detection limit of the procedure was 0.76 μg/mL. The proposed procedure was successfully utilized in the determination of the drug in pharmaceutical preparations with mean recovery in the range of 99.83 ? 101.16%. The second method is a colorimetric method, which depends on the measurement of absorbances of tris (o‐phenanthroline) iron(II) [method 2A] and tris (bipyridyl) iron(II) [method 2B] complexes at 512 nm. The complexes obeyed Beer's law over the concentration range of 2–16 μg/mL and 4–40 μg/mL for methods 2A and 2B, respectively. The developed method has been successfully applied for the determination of ranitidine in bulk drugs and pharmaceutical formulations. The common excipients and additives did not interfere in its determination.     

中枢多巴胺系统正电子发射计算机断层扫描显像剂的研究进展

中枢多巴胺系统与多种神经行为障碍的病理生理学有关。一直以来,多巴胺系统正电子发射计算机断层扫描(PET)成像在研究活体大脑中多巴胺生物化学过程上有着重要价值。PET成像的基础是¹¹C、¹⁸F等发射正电子的放射性核素标记的显像剂,这些显像剂通过与多巴胺神经系统不同的靶点特异性结合从而反映多巴胺合成、囊泡储存、突触释放和受体结合以及再摄取过程,推动神经病学、精神病学、药物滥用和成瘾以及药物开发的研究进展。本文综述了以氨基酸脱羧酶、多巴胺转运体、多巴胺受体以及囊泡单胺转运体为靶点的¹¹C、¹⁸F标记的PET显像剂的研究进展。     

中枢多巴胺系统正电子发射计算机断层扫描显像剂的研究进展

中枢多巴胺系统与多种神经行为障碍的病理生理学有关。一直以来,多巴胺系统正电子发射计算机断层扫描(PET)成像在研究活体大脑中多巴胺生物化学过程上有着重要价值。PET成像的基础是¹¹C、¹⁸F等发射正电子的放射性核素标记的显像剂,这些显像剂通过与多巴胺神经系统不同的靶点特异性结合从而反映多巴胺合成、囊泡储存、突触释放和受体结合以及再摄取过程,推动神经病学、精神病学、药物滥用和成瘾以及药物开发的研究进展。本文综述了以氨基酸脱羧酶、多巴胺转运体、多巴胺受体以及囊泡单胺转运体为靶点的¹¹C、¹⁸F标记的PET显像剂的研究进展。     

High-throughput liquid-chromatography method with fluorescence detection for reciprocal determination of furosemide or norfloxacin in human plasma

A simple, high-throughput, highly selective and sensitive HPLC-FLD method for isolation and determination of furosemide and/or norfloxacin in human plasma samples following a simple organic solvent deproteinization step with acetonitrile as sample 'clean-up' procedure is reported. One of the two drug substances plays the internal standard role for the determination of the other. Separation of analyte and internal standard was achieved in less than 5.3 min (injection to injection) on a Chromolith Performance RP-18e column, using an aqueous component containing 0.015 mol/L sodium heptane-sulfonate and 0.2% triethylamine brought to pH = 2.5 with H(3)PO(4). The composition of the mobile phase was: acetonitrile-methanol-aqueous component = 70:15:15 (v/v/v) and the flow-rate was set up to 3 mL/min. The chromatographic method applied to the determination of furosemide relies on fluorescent detection parameters of 235 nm for the excitation wavelength, and 402 nm for the emission wavelength. In case of norfloxacin, the excitation wavelength is set up to 268 nm and the emission wavelength is set up to 445 nm. The overall method leads to quantitation limits of about 27 ng/mL for furosemide, and 19.5 ng/mL for norfloxacin, using an injection volume of 250 microL. The method was applied to the bioequivalence study of two furosemide-containing formulations.