Stacking and determination of phenazine-1-carboxylic acid with low p<Emphasis Type="Italic">K</Emphasis><Subscript>a</Subscript> in soil via moving reaction boundaryformed by alkaline and double acidic buffers in capillary electrophoresis

As shown herein, a normal moving reaction boundary (MRB) formed by an alkaline buffer and a single acidic buffer had poor stacking to the new important plant growth promoter of phenazine-1-carboxylic acid (PCA) in soil due to the leak induced by its low pK _a. To stack the PCA with low pK _a efficiently, a novel stacking system of MRB was developed, which was formed by an alkaline buffer and double acidic buffers (viz., acidic sample and blank buffers). With the novel system, the PCA leaking into the blank buffer from the sample buffer could be well stacked by the prolonged MRB formed between the alkaline buffer and blank buffer. The relevant mechanism of stacking was discussed briefly. The stacking system, coupled with sample pretreatment, could achieve a 214-fold increase of PCA sensitivity under the optimal conditions (15 mM (pH 11.5) Gly-NaOH as the alkaline buffer, 15 mM (pH 3.0) Gly-HCl-acetonitrile (20%, v/v) as the acidic sample buffer, 15 mM (pH 3.0) Gly-HCl as the blank buffer, 3 min 13 mbar injection of double acidic buffers, benzoic acid as the internal standard, 75 μm i.d. × 53 cm (44 cm effective length) capillary, 25 kV and 248 nm). The limit of detection of PCA in soil was decreased to 17 ng/g, the intra-day and inter-day precision values (expressed as relative standard deviations) were 3.17–4.24% and 4.17–4.87%, respectively, and the recoveries of PCA at three concentration levels changed from 52.20% to 102.61%. The developed method could be used for the detection of PCA in soil at trace level.     

A Novel Enhancing Flow-Injection Chemiluminescence Method for the Determination of Glutathione Using the Reaction of Luminol with Hydrogen Peroxide

 A rapid flow-injection method with chemiluminescence (CL) detection is described for the determination of glutathione (GSH). The method is based on the CL reaction of luminol and hydrogen peroxide. GSH can greatly enhance the chemiluminescence intensity in 0.1 mol/L borax–sodium hydroxide buffer solution (pH = 9.7). The maximum CL intensity was directly proportional to the concentration of GSH in the range 3.0 × 10⁻⁷–2.0 × 10⁻⁵ mol/L, and the detection limit was 6.8 × 10⁻⁸ mol/L. The relative standard deviation was 3.4% for 5.0 × 10⁻⁶ mol/L of GSH (n = 11). Received October 23, 2001; accepted June 18, 2002     

Poly(vinylferrocenium) perchlorate–polyaniline composite film-coated electrode for amperometric determination of hydroquinone

Poly(vinylferrocenium) perchlorate–polyaniline (PVF⁺–PANI) composite film was synthesized electrochemically on Pt electrode in a methylene chloride solution containing a mixture of poly(vinylferrocene) (PVF) polymer and aniline monomer. PVF⁺ polymer in the composite film was used as an electron transfer mediator. The composite coating showed significant electrochemical activity towards hydroquinone (HQ) at pH 4, with high sensitivity and a wide linearity range. The interaction of HQ with PVF⁺ and PANI homopolymer films was investigated electrochemically and spectroscopically. HQ molecules are accumulated on the electrode surface due to trapping by both polymers in the composite film and then oxidized catalytically by PANI. The most significant contribution of PVF⁺ polymer is that it facilitates electron transfer in the composite film. The linear response range was found to be between 1.60 × 10⁻⁴ mM and 115 mM (R ² = 0.999) at 0.45 V vs saturated calomel electrode. The limit of detection (LOD) was 4.94 × 10⁻⁵ mM.     

Chemiluminescence of Peracetic Acid in Alkaline Medium and its Application to Dihydralazine Sulfate Determination

The chemiluminescence (CL) of peracetic acid (PAA) in alkaline medium is very weak but is strongly enhanced after the addition of dihydralazine sulfate (DHZS). Based on this phenomenon, a simple, rapid and highly sensitive flow-injection CL method for the determination of DHZS was developed. The CL emission was linearly related to the DHZS concentration in the range of 20–4000 ng mL⁻¹ with a detection limit (3σ) of 1.2 ng mL⁻¹. As a preliminary application, the proposed method was successfully applied to the determination of DHZS in pharmaceutical preparations; the recovery of DHZS in human urine was between 96.5% and 102.2%. A detailed CL mechanism was proposed and singlet molecular oxygen (¹O₂) was suggested to be produced in the CL reaction process.     

Voltammetric sensor for glutathione determination based on ferrocene-modified carbon paste electrode

The electrocatalytic oxidation of glutathione (GSH) has been studied at the surface of ferrocene-modified carbon paste electrode (FMCPE). Cyclic voltammetry (CV), double potential step chronoamperometry, and differential pulse voltammetry (DPV) techniques were used to investigate the suitability of incorporation of ferrocene into FMCPE as a mediator for the electrocatalytic oxidation of GSH in buffered aqueous solution. Results showed that pH 7.00 is the most suitable for this purpose. In the optimum condition (pH 7.00), the electrocatalytic ability of about 480 mV can be found and the heterogeneous rate constant of catalytic reaction was calculated as . Also, the diffusion coefficient of glutathione, D, was found to be 3.61 × 10^–5 cm² s⁻¹. The electrocatalytic oxidation peak current of glutathione at the surface of this modified electrode was linearly dependent on the GSH concentration and the linear analytical curves were obtained in the ranges of 3.2 × 10^–5 M–1.6 × 10^–3 M and 2.2 × 10^–6 M–3.5 × 10^–3 M with cyclic voltammetry and differential pulse voltammetry methods, respectively. The detection limits (3σ) were determined as 1.8 × 10^–5 M and 2.1 × 10^–6 M using CV and DPV, respectively. Finally, the electrocatalytic oxidation of GSH at the surface of this modified electrode can be employed as a new method for the voltammetric determination of glutathione in real samples such as human plasma.     

Use of polyclonal antibodies to ochratoxin A with a quartz–crystal microbalance for developing real-time mycotoxin piezoelectric immunosensors

A piezoelectric immunosensor was tested for ochratoxin A (OTA) mycotoxin detection through the immobilization of OTA–bovine serum albumin (OTA–BSA) conjugate on gold-coated quartz crystals (AT-cut/5 MHz). Immunoassays were performed in a flow-injection system through frequency decreases in a quartz–crystal microbalance (QCM) because of a mass increasing during immunoreaction with anti-OTA antibodies. Three immobilization procedures for OTA–BSA (direct adsorption and covalent attachment to two alkane thiol self-assembled monolayers) were characterized with QCM in real time. Covalent attachment of the OTA–BSA conjugates through gold nanoparticles was also tested for amplifying the signal. Binding of the excess of antibodies to the immobilized OTA in an indirect competitive analysis decreased linearly the resonant frequency in the range of the OTA concentration from 10 to 128 ng/mL, with a detection limit of 8 ng/mL (signal/noise ratio of 3). A pepsin 2 mg/mL (pH = 2.1) solution was used to release antigen–antibody complexes, regenerating the biorecognition surface.     

Electrochemical behavior of isoproterenol in the presence of uric acid and folic acid at a carbon paste electrode modified with 2,7-bis(ferrocenyl ethyl)fluoren-9-one and carbon nanotubes

This paper reports the selective determination of isoproterenol (IP) in the presence of uric acid (UA) and folic acid (FA) using 2,7-bis(ferrocenyl ethyl)fluoren-9-one modified carbon nanotube paste electrode (2,7-BFCNPE) in 0.1 M phosphate buffer solution (PBS) (pH 7.0). The bare carbon paste electrode does not separate the voltammetric signals of IP, UA, and FA. However, 2,7-BFCNPE not only resolved the voltammetric signals of IP, UA, and FA with potential differences of 150, 325, and 475 mV between IP–UA, UA–FA, and IP–FA, respectively, but also dramatically enhanced the oxidation peak currents of them when compared to bare carbon paste electrode. In PBS of pH 7.0, the oxidation current increased linearly with two concentration intervals of IP, one is 0.08 to 17.5 μM and the other is 17.50 to 700.0 μM. The detection limit (3σ) obtained by DPV was 26.0 ± 2 nM. The practical application of the modified electrode was demonstrated by determining IP in IP injection, urine, and human blood serum.     

Photofragmentation of nitro-based explosives with chemiluminescence detection

A simple, fast, reliable, sensitive and potentially portable explosive detection device was developed employing laser photofragmentation (PF) followed by heterogeneous chemiluminescence (CL) detection. The PF process involves the release of NO_{x(x = 1,2)} moieties from explosive compounds such as TNT, RDX, and PETN through a stepwise excitation–dissociation process using a 193 nm ArF laser. The NO_{x(x = 1,2)} produced upon PF is subsequently detected by its CL reaction with basic luminol solution. The intensity of the CL signal was detected by a thermoelectrically cooled photomultiplier tube with high quantum efficiency and negligible dark current counts. The system was able to detect trace amounts of explosives in various forms in real time under ambient conditions. Detection limits of 3 ppbv for PETN, 2 ppbv for RDX, and 34 ppbv for TNT were obtained. It was also demonstrated that the presence of PETN residue within the range of 61 to 186 ng/cm² can be detected at a given signal-to-background ratio of 10 using a few microjoules of laser energy. The technique also demonstrated its potential for the direct analysis of trace explosive in soil. An LOD range of 0.5–4.3 ppm for PETN was established, which is comparable to currently available techniques. Figure Photofragmentation–chemiluminescence detector     

Ultrasensitive Assay of Rhein in Medicine Based on its Enhanced Luminol-K3Fe(CN)6 Chemiluminescence Reaction Using the Flow Injection Technique

Determination of the effective components in traditional Chinese medicine is one of the key steps for its identification. In this paper a novel and sensitive chemiluminescence (CL) method for the determination of rhein coupled with flow-injection analysis (FIA) is developed. It is based on the strong sensitizing effect on the weak CL reaction between luminol and ferricyanide in alkaline solution. Under optimal experimental conditions, the relative CL intensity is proportional to the concentration of rhein in the range of 7.0 × 10⁻¹²–7.0 × 10⁻¹⁰ mol L⁻¹ and 1.0 × 10⁻⁹–4.0 × 10⁻⁵ mol L⁻¹, the detection limit is 1.478 × 10⁻¹³ mol L⁻¹, and the relative standard deviation (RSD) for 9 parallel measurements of 1.408 × 10⁻⁷ mol L⁻¹ rhein is 3.4%. The method was successfully applied to the determination of rhein in pharmaceutical preparations. The possible mechanism of CL is also briefly discussed.     

Catecholamine analysis with strong cation exchange column liquid chromatography–peroxyoxalate chemiluminescence reaction detection

A liquid chromatography–chemiluminescence detection method was developed and validated for the determination of catecholamines (norepinephrine, epinephrine, and dopamine) in mouse brains. Chromatography was performed on a strong cation exchange column (150 × 2.0-mm id) using an isocratic mobile phase of 65 mM potassium acetate/75 mM potassium phosphate (95:5, pH 3.5) at a flow rate of 0.2 mL/min following post-column fluorescence derivatization of catecholamines with ethylenediamine and peroxyoxalate chemiluminescence reaction detection. The recovery of catecholamines added to mouse brain samples was more than 95.0%, while intra- and inter-day precision of the assay were <4.8%. The validated method was used to determine norepinephrine and dopamine concentrations in mouse brains without prior sample purification.     

Applications of nanostructured Pt-Au hybrid film for the simultaneous determination of catecholamines in the presence of ascorbic acid

Nanostructured platinum-gold (Pt–Au) hybrid film modified glassy carbon electrode (GCE) was fabricated by electro-deposition method in the presence of 2 × 10⁻⁴ mol l⁻¹ l-cysteine. To examine the surface morphological analysis, the (Pt–Au) hybrid film were electrochemically deposited on transparent semiconductor indium tin oxide (ITO) electrodes for scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS) studies. From the SEM analysis, it was observed that the deposited nanoplatinum (250–400 nm) was formed as a cauliflower-shaped structure with the gold nanoparticles (30–90 nm). The concentration variation of additive l-cysteine results in the formation of cauliflower-shaped platinum nanoparticles. Further, the Pt–Au hybrid film modified GCE could be used for the detection of catecholamine neurotransmitters epinephrine (EP), norepinephrine (NEP) individually and in the presence of ascorbic acid (AA) in pH 7 phosphate-buffered solutions (PBS). Furthermore, the proposed Pt–Au hybrid film could be applied for the detection of epinephrine in injection solution and ascorbic acid from commercially available vitamin C tablets.     

Determination of some sulfonylurea herbicides in soil by a novel liquid-phase microextraction combined with sweeping micellar electrokinetic chromatography

A novel method for the determination of five sulfonylurea herbicides in soil was developed by a dispersive solid-phase extraction (DSPE) clean-up followed by dispersive liquid–liquid microextraction (DLLME), prior to sweeping micellar electrokinetic chromatography (MEKC). In the DSPE-DLLME, 10 g of soil sample was first extracted with 10 mL of acetonitrile containing 5% formic acid (pH 3.0). The extract was then cleaned-up by a DSPE with C₁₈ as sorbent. A 1 mL aliquot of the resulting extract was then added into a centrifuge tube containing 5 mL of water adjusted to pH 2.0 and 60.0 μL chlorobenzene (as extraction solvent) for DLLME procedure. Then, the organic sample extraction solution was evaporated to dryness, and reconstituted with 20.0 μL of 1.0 mmol L⁻¹ Na₂HPO₄ (pH 10.0) for sweeping-MEKC analysis after DLLME. Under optimized conditions, the method provided as high as 3,000- to 5,000-fold enrichments factors. The linearity of the method was in the range of 3.3–200 ng g⁻¹ for chlorimuron ethyl and bensulfuron methyl, and in the range of 1.7–200 ng g⁻¹ for tribenuron methyl, chlorsulfuron and metsulfuron methyl, with the correlation coefficients (r) ranging from 0.9965 to 0.9983, respectively. The limits of detection (LODs) ranged from 0.5 to 1.0 ng g⁻¹. The intraday relative standard deviations (RSDs, n = 5) were below 5.3% and interday RSDs (n = 15) within 6.8%. The recoveries of the method for the five sulfonylureas from soil samples at spiking levels of 5.0, 20.0, and 100.0 ng g⁻¹ were 76.0–93.5%, respectively. The developed method has been successfully applied to the analysis of the target sulfonylurea herbicide residues in soil samples with a satisfactory result.     

In-line solid-phase extraction–capillary electrophoresis coupled with mass spectrometry for determination of drugs of abuse in human urine

In-line solid-phase extraction–capillary electrophoresis coupled with mass spectrometric detection (SPE–CE–MS) has been used for determination of 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), codeine (COD), hydrocodeine (HCOD), and 6-acetylmorphine (6AM) in urine. The preconcentration system consists of a small capillary filled with Oasis HLB sorbent and inserted into the inlet section of the electrophoresis capillary. The SPE–CE–MS experimental conditions were optimized as follows: the sample (adjusted to pH 6.0) was loaded at 930 mbar for 60 min, elution was performed with methanol at 50 mbar for 35 s, 60 mmol L⁻¹ ammonium acetate at pH 3.8 was used as running buffer, the separation voltage was 30 kV, and the sheath liquid at a flow rate of 5.0 μL min⁻¹ was isopropanol–water 50:50 (v/v) containing 0.5% acetic acid. Analysis of urine samples spiked with the four drugs and diluted 1:1 (v/v) was studied in the linear range 0.08–10 ng mL⁻¹. Detection limits (LODs) (S/N = 3) were between 0.013 and 0.210 ng mL⁻¹. Repeatability (expressed as relative standard deviation) was below 7.2%. The method developed enables simple and effective determination of these drugs of abuse in urine samples at the levels encountered in toxicology and doping.     

Development of a sensitive micro-magnetic chemiluminescence enzyme immunoassay for the determination of carcinoembryonic antigen

A micro-magnetic chemiluminescence (CL) enzyme immunoassay with high sensitivity, selectivity, and reproducibility was developed for the determination of the tumor marker, carcinoembryonic antigen (CEA) in human serum. A sandwich scheme assay has been utilized with fluorescein isothiocyanate antibody (FITC)-labeled anti-CEA antibody and alkaline phosphate (ALP)-labeled anti-CEA antibody being used in the CL detection. The CL signal produced by the emission of photons from 4-methoxy-4-(3-phosphate-phenyl)-spiro-(1,2-dioxetane-3,2′-adamantane) (AMPPD) was directly proportional to the amount of analyte present in a sample solution. The influences of the reaction time of antigen with antibody, the reaction time of substrate with label, the dilution ratio of ALP-labeled anti-CEA antibody, the concentration of FITC-labeled anti-CEA antibody, and other relevant variables upon the CL signal were examined and optimized. The CL responses depended linearly on the CEA concentration over the range from 2 to 162 ng mL⁻¹ in a logarithmic plot. Assay sensitivity as low as 0.69 ng mL⁻¹ was achieved. A coefficient of variance of less than 13% was obtained for intra- and inter-assay precision. This method has been successfully applied to the analysis of CEA in human serum. According to the procedure based on spiked standards, the recoveries obtained were 80–110%. Comparison experiments were carried out with the commercially available CEA chemiluminescence immunoassay. Satisfactory results were obtained according to a paired t-test method (t value < t _critical at the 95% confidence level).     

Multi-mycotoxin analysis of maize silage by LC-MS/MS

This paper describes a method for determination of 27 mycotoxins and other secondary metabolites in maize silage. The method focuses on analytes which are known to be produced by common maize and maize-silage contaminants. A simple pH-buffered sample extraction was developed on the basis of a very fast and simple method for analysis of multiple pesticide residues in food known as QuEChERS. The buffering effectively ensured a stable pH in samples of both well-ensiled maize (pH < 4) and of hot spots with fungal infection (pH > 7). No further clean-up was performed before analysis using liquid chromatography–tandem mass spectrometry. The method was successfully validated for determination of eight analytes qualitatively and 19 quantitatively. Matrix-matched calibration standards were used giving recoveries ranging from 37% to 201% with the majority between 60% and 115%. Repeatability (5–27% RSD_r) and intra-laboratory reproducibility (7–35% RSD_IR) was determined. The limit of detection (LOD) for the quantitatively validated analytes ranged from 1 to 739 μg kg⁻¹. Validation results for citrinin, fumonisin B₁ and fumonisin B₂ were unsatisfying. The method was applied to 20 selected silage samples and alternariol monomethyl ether, andrastin A, alternariol, citreoisocoumarin, deoxynivalenol, enniatin B, fumigaclavine A, gliotoxin, marcfortine A and B, mycophenolic acid, nivalenol, roquefortine A and C and zearalenone were detected.     

Electrocatalytic reduction of hydrogen peroxide at Prussian blue modified electrodes: a RDE study

Electrocatalytic reduction of hydrogen peroxide at Prussian blue modified electrode has been studied with rotating disk electrode in pH 5.5 and 7.3 solutions. It has been shown that the electrocatalytic cathodic reduction obeys Koutecky–Levich relationship at electrode potentials ranging from 0.1 to −0.4 V vs. Ag/AgCl for low concentrations of peroxide not exceeding 0.3 mM. Within this potential window, the calculated kinetic cathodic current ranges within the limits of 2.15–6.09 and 1.00–3.60 mA cm⁻² mM⁻¹ for pH 5.5 and 7.3, respectively. For pH 5.5 and 7.3 solutions, a linear slope of the dependence of kinetic current on electrode potential of −10.8 and −2.89 mA cm⁻² mM⁻¹ V⁻¹, respectively, has been obtained. At a higher concentration of peroxide, exceeding 0.6 mM, deviations from Koutecky–Levich relationship have been observed. These deviations appear more expressed at higher potentials and higher solution pH. The results obtained have been interpreted within the frame of two-step reaction mechanism, including (1) dissociative adsorption of hydrogen peroxide with the formation of OH radicals and (2) one-electron reduction of these radicals to OH⁻ anions. At a higher concentration of peroxide, and especially at a higher pH, the second process becomes rate limiting.     

Solid state electrochemical of the erbium hexacyanoferrate-modified carbon ceramic electrode and its electrocatalytic oxidation of <Emphasis Type="SmallCaps">l</Emphasis>-cysteine

A kind of erbium hexacyanoferrate (ErHCF)-modified carbon ceramic electrodes (CCEs) fabricated by mechanically attaching ErHCF samples to the surface of CCEs derived from sol–gel technique was proposed. The resulting modified electrodes exhibit well-defined redox responses with the formal potential of +0.215 V [vs saturated calomel electrode (SCE)] at a scan rate of 20 mV s⁻¹ in 0.5 M KCl (pH 7) solution. The voltammetric characteristics of the ErHCF-modified CCEs were investigated by voltammetry. Attractively, the ErHCF-modified CCEs presented good electrocatalytic activity with a marked decrease in the overvoltage about 400 mV for l-cysteine oxidation. The calibration plot for l-cysteine determination was linear at 5.0 × 10⁻⁶–1.3 × 10⁻⁴ M with a linear regression equation of I(A) = 0.558 + 0.148c (μM) (R ² = 0.9989, n = 20), and the detection limit was 2 × 10⁻⁶ M (S/N = 3). At last, the ErHCF-modified CCEs were used for amperometric detection of l-cysteine in real samples.     

The effect of chloride concentration and pH on pitting corrosion of AA7075 aluminum alloy coated with phenyltrimethoxysilane

The effect of chloride ion concentration and pH of solution on the corrosion behavior of aluminum alloy AA7075 coated with phenyltrimethoxysilane (PTMS) immersed in aqueous solutions of NaCl is reported. Potentiodynamic polarization, linear polarization, open circuit potential, and weight loss measurements were performed. The surface of samples was examined using SEM and optical microscopy. Elemental characterization of the coating by secondary ion mass spectrometry indicates an intermediate layer between coating and aluminum alloy surface. The corrosion behavior of the aluminum alloy AA7075 depends on chloride concentration and pH of solution. In acidic or neutral solutions, general and pitting corrosion occur simultaneously. On the contrary, exposure to alkaline solutions results in general corrosion only. Results further reveal that aluminum alloy AA7075 is susceptible to pitting corrosion in all chloride solutions with concentrations between 0.05 M and 2 M NaCl; an increase in the chloride concentration slightly shifted both the pitting and corrosion potentials to more active values. Linear polarization resistance measurements show a substantially improved corrosion resistance value in case of samples coated with PTMS as compared to uncoated samples in both neutral (pH = 7), acidic (pH = 0.85 and 3), and alkaline chloride solutions (pH = 10 and 12.85). The higher corrosion resistance of the aluminum alloy coated with PTMS can be attributed to the hydrophobic coating which acts as a barrier and prevents chloride ion penetration and subsequent reaction with the aluminum alloy.     

Glucose Oxidase-dextran Conjugates with Enhanced Stabilities Against Temperature and pH

Multipoint covalent bonding of glucose oxidase (EC 1.1.3.4) to hydrophilic natural polymer dextran and optimization of procedures to obtain, with enhanced temperature and pH stabilities, were studied. Purified enzyme was conjugated with various molecular weight dextrans (17.5, 75, and188 kD) in a ratio of 20:1, 10:1, 1:1, 1:5, 1:10, 1:15, and 1:20. After 1 h of incubation at pH 7, the activities of purified enzyme and conjugates were determined at different temperatures (25°C, 30°C, 35°C, 40°C, 50°C, 60°C, 70°C, and 80°C), and the results were evaluated for thermal resistance. Increases in temperature from 25°C to 50°C did not change the activities of the conjugates. The conjugate, which was prepared with 75 kDa dextran in a molar ratio of 1:5, showed the highest thermal resistance and even the activity still remains at 80°C at pH 7.0. This conjugate also displayed activity in a wide pH range (pH 4.0–7.0) at high temperatures. Conjugate, which was synthesized with 75 kDa dextran in a molar ratio of 1:5, appears to be feasible and useful for biotechnological applications.     

1,3,5,7-Tetramethyl-8-(<Emphasis Type="Italic">N</Emphasis>-hydroxysuccinimidyl butyric ester)difluoroboradiaza-<Emphasis Type="Italic">s</Emphasis>-indacene as a new fluorescent labeling reagent for HPLC determination of amino acid neurotransmitters in the cerebral cortex of mice

1,3,5,7-Tetramethyl-8-(N-hydroxysuccinimidyl butyric ester)difluoroboradiaza-s-indacene (TMBB-Su), a new BODIPY-based fluorescent probe, was designed and synthesized for the labeling of amino compounds. It was used as a pre-column derivatizing reagent for determination of amino acid neurotransmitters by high-performance liquid chromatography (HPLC). The fluorescence quantum yield in acetonitrile increased from 0.84 to 0.95 when it reacted with amino acid neurotransmitters. Derivatization of TMBB-Su with seven amino acid neurotransmitters was completed within 30 min at 25 °C in 24.0 mmol L⁻¹ pH 7.8 boric acid buffer. The separation was performed on a C₁₈ column with methanol–water–buffer 55:35:10 (v/v) as mobile phase (buffer: 0.10 mol L⁻¹ H₃Cit–0.10 mol L⁻¹ NaOH). Interference from the other concomitant amino acids was eliminated successfully by means of pH gradient elution. With fluorescence detection at 494 and 504 nm for excitation and emission, respectively, the limits of detection (signal-to-noise ratio = 3) were from 2.1 to 12.0 nmol L⁻¹. The proposed method has been used to determine amino acid neurotransmitters in the cerebral cortex of mice with cerebral ischemia at the convalescence stage with satisfactory recoveries varying from 94.9 to 105.2%.