Validation of HPLC Analysis of Aspartate and Glutamate Neurotransmitters Following <b> <i>o</i> </b>‐Phthaldialdehyde‐Mercaptoethanol Derivatization

Rabouan, Sylvie; Olivier, Jean‐Christophe; Guillemin, Hélène; Barthes, D.

doi:10.1081/jlc-120021286

Cited by 7 publications

(3 citation statements)

References 16 publications

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“…[11][12][13] As a result, a highly sensitive analytical technique is required. To date, a variety of techniques have been utilized to analyze AA neurotransmitters in the microdialysate such as biosensors, [14] microchip electrophoresis, [15] CE-LIF, [16] LC-MS, [17] HPLC, [18,19] etc… Among these, HPLC is most common due to its advantages in operation, cost, and repeatability. Fluorescence detector (FLD) is most widely used in the HPLC detection of AA neurotransmitters, [20][21][22] but the electrochemical detector (ECD) is a better choice partly because of its sensitivity.…”

Section: Introductionmentioning

confidence: 99%

Assessment of Amino Acid Neurotransmitters in Rat Brain Microdialysis Samples by High-Performance Liquid Chromatography with Coulometric Detection

Chen

Deng

Yao

et al. 2015

Journal of Liquid Chromatography & Related Technologies

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A simple method was developed for simultaneous determination of five amino acid neurotransmitters (taurine, glutamate, aspartate, glycine, and γ-aminobutyric acid) in brain microdialysates by high performance liquid chromatography with coulometric detection.The microdialysates were derivatized with o-phthalaldehyde in 5 mM sodium sulfite and separated within 17 min using isocratic elution. Detection parameters for these transmitters were linearity (r 2 ) > 0.999 over the concentration range from 0.1 to 15.00 μM (0.01 to 1.00 μM for GABA); reproducibility 2.88 -11.67% (relative standard deviation); the limits of detection 4.4 -17.2 fmol and recovery 86.66% -103.42%. This method was successfully applied to analyze these neurotransmitters in the brain. The changes of neurotransmitters were also investigated via local administration of potassium chloride and bicuculline, which was a γ-aminobutyric acid receptor antagonist. It was found that the developed method is more efficient as compared to previously reported methods in simultaneously Downloaded by [New York University] at 06:06 26 May 2015 2 determining five major neurotransmitters because it shows quick separation, sensitive detection, and is easy to operate. This method can be a practical choice for continuous monitoring of the amino acid neurotransmitters in vivo.

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Section: Introductionmentioning

confidence: 99%

Assessment of Amino Acid Neurotransmitters in Rat Brain Microdialysis Samples by High-Performance Liquid Chromatography with Coulometric Detection

Chen

Deng

Yao

et al. 2015

Journal of Liquid Chromatography & Related Technologies

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“…Blood samples were drawn for norfloxacin high-performance liquid chromatography determinations in plasma (2). Dialysates were collected every 15 min for glutamate determinations (8). PK-PD modeling was conducted as previously described (2).…”

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confidence: 99%

Norfloxacin-Induced Electroencephalogram Alteration and Seizures in Rats Are Not Triggered by Enhanced Levels of Intracerebral Glutamate

Chenel¹,

Limosin²,

Marchand³

et al. 2003

Antimicrob Agents Chemother

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Pharmacokinetic-pharmacodynamic modeling of the electroencephalogram (EEG) effect was combined with intracerebral glutamate determinations using microdialysis for rats receiving norfloxacin intravenously (150 mg/kg of body weight). The EEG effect (accompanied by tremors and seizures) was consistently observed without glutamate level modifications. Therefore, norfloxacin-inducted seizures are not triggered by intracerebral glutamate level enhancement.Fluoroquinolone antibiotics may induce central nervous system excitatory side effects, including anxiety, nervousness, hallucination, and even seizures on rare occasions (3, 13). It is usually asserted that these excitatory side effects result from inhibition of GABA (␥-aminobutyric acid) binding to its receptors (1, 11). Using intracerebral microdialysis, however, Smolders et al. have previously observed in conscious rats norfloxacin-induced convulsions accompanied not by any apparent effect on the extracellular hippocampal GABA levels but rather by enhanced intracerebral glutamate concentrations (10). Studies of the relationship between glutamate concentrations and epileptic seizures have yielded conflicting observations, as recently reviewed (12). Smolders et al. have hypothesized that enhanced glutamate levels might be essential for the induction of norfloxacin seizures (10), but it could also be argued that the generation, spreading, and maintenance of seizures eventually leads to glutamate release and elevation (9). Quantitative electroencephalogram (EEG) recording combined with intracerebral microdialysis offers a unique opportunity to address this question. In rats treated with imipenem (5) and norfloxacin (2), increases in the total power of the EEG signal are related to behavioral modifications and the occurrence of tremor and partial seizures and can therefore be considered an appropriate surrogate pharmacodynamic (PD) end point for the investigation of the convulsant activity of these antibiotics in rats. Furthermore, the kinetics of this effect can be very precisely evaluated using pharmacokinetic (PK)-PD modeling techniques (2, 5).The objective of this study was therefore to combine quantitative EEG recording (for the characterization of seizure induction and maintenance) with intracerebral microdialysis using the hippocampus (for the measurement of glutamate levels) of rats (after the administration of norfloxacin) to look for a causality relationship between the two factors. The experiment was conducted in accordance with the Principles of Laboratory Animal Care (National Institutes of Health), and ethical approval was obtained from the Animal Ethics Committee of the Faculty of Pharmacy (BHE 2001/12/ AE). Norfloxacin was obtained from Sigma (Saint-Quentin Fallavier, France), and a salt was prepared as previously described (4). L-glutamate standards were supplied by Sigma. All chemicals used were of analytical grade, and the analytical solvents used were of high-performance liquid chromatography grade. Six male Sprague-Dawley rats (weighing 299 Ϯ 15 g)...

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“…A detectabilidade do detector acoplado no cromatógrafo líquido de alta eficiência foi um importante parâmetro avaliado neste estudo, já que os aminoácidos A desvantagem deste reagente é a instabilidade dos derivativos isoindoles formados em temperatura ambiente sendo necessário limitar seu tempo de reação (RABOUAN et al, 2003). Neste contexto, para se conseguir uma reprodutibilidade alta o derivativo formado permaneceu exatamente por 3 minutos em temperatura ambiente.…”

Section: Otimização Da Detecção Dos Analitosunclassified

Desenvolvimento e validação de metodologia para análise simultânea de aminoácidos como possíveis marcadores neurobiológicos da esquizofrenia utilizando a cromatografia líquida de alta eficiência com detecção por fluorescência

Alcantara¹

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Desenvolvimento e validação de metodologia para análise simultânea de aminoácidos como possíveis marcadores neurobiológicos da esquizofrenia utilizando a cromatografia líquida de alta eficiência com detecção por fluorescência As teorias neurobiológicas defendem que a esquizofrenia é essencialmente causada por alterações bioquímicas e estruturais do cérebro. A importância que os aminoácidos tem com a fisiopatologia da esquizofrenia e, por este envolvimento encontrar-se pouco esclarecido é que esta pesquisa teve como propósito desenvolver e validar uma metodologia analítica para a análise simultânea dos aminoácidos: glutamato, aspartato, serina, glicina, arginina e lisina em plasma de pacientes com diagnóstico de esquizofrenia, utilizando para isto a cromatografia líquida de alta eficiência com detecção por fluorescência. Os analitos foram inicialmente extraídos através do processo de precipitação protéica, com o solvente orgânico acetonitrila. Para que pudessem ser detectados por fluorescência os aminoácidos foram derivatizados empregando orto-ftalaldeído na presença de Bmercaptoetanol. O tempo total de separação foi de 18 minutos em coluna analítica LiChroCART ® RP-18 (Merck, 125mm, 4mm d.i., 5µm) no modo de eluição gradiente com tampão acetato de sódio 0,1 mol/L (pH 7,0, A) e acetonitrila e água (B), na proporção 70:30, respectivamente, para posterior detecção por fluorescência (excitação 340 nm, emissão 450 nm). O método foi validado segundo os critérios estabelecidos pela Agência Nacional de Vigilância Sanitária (ANVISA). A resposta do detector encontrou-se linear na faixa de 9,6 a 192 pmol/mL para todos os aminoácidos. O limite de detecção estabelecido foi de 7,68 pmol/mL e o limite de quantificação foi de 9,6 pmol/mL. A recuperação foi superior a 70%. A precisão e exatidão intra-ensaio variou de 3,6% a 5,3% e 3,1% a 8,3%, respectivamente. A precisão e exatidão interensaio variou de 3,6% a 7,1% e 3,1% a 11,4%, respectivamente. A especificidade foi determinada para os seguintes interferentes: lorazepam, diazepam, clonazepam, alprazolam, haloperidol, levomepromazina, propranolol, ranitidina, fluoxetina, olanzapina, carbamazepina, diclofenaco, amiodarona, sulfametoxazol e, ainda, plasma hemolisado, normal e lipêmico. O método desenvolvido e validado mostrou ser eficiente na determinação simultânea de aminoácidos podendo ser aplicado em amostras de pacientes esquizofrênicos a fim de investigar seu envolvimento com esta desordem psiquiátrica tão intrigante.Palavras-chave: aminoácidos; plasma; cromatografia líquida de alta eficiência; esquizofrenia. Schizophrenia is primarily caused by structural and biochemical changes in the brain according to the main neurobiological theory, including the amino acids concentrations in the human plasma. The relation between these amino acids concentrations and the schizophrenia needs more clarification. Therefore, an analytical tool is necessary to provide which of these amino acids are related with this mental disease. The aim of this research was to develop and to validate an ...

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Validation of HPLC Analysis of Aspartate and Glutamate Neurotransmitters Following o ‐Phthaldialdehyde‐Mercaptoethanol Derivatization

Cited by 7 publications

References 16 publications

Assessment of Amino Acid Neurotransmitters in Rat Brain Microdialysis Samples by High-Performance Liquid Chromatography with Coulometric Detection

Assessment of Amino Acid Neurotransmitters in Rat Brain Microdialysis Samples by High-Performance Liquid Chromatography with Coulometric Detection

Norfloxacin-Induced Electroencephalogram Alteration and Seizures in Rats Are Not Triggered by Enhanced Levels of Intracerebral Glutamate

Desenvolvimento e validação de metodologia para análise simultânea de aminoácidos como possíveis marcadores neurobiológicos da esquizofrenia utilizando a cromatografia líquida de alta eficiência com detecção por fluorescência

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