Determination of selected neurotransmitter metabolites using monolithic column chromatography coupled with chemiluminescence detection

The oxidation of selected clinically important neurotransmitter metabolites with acidic potassium permanganate in the presence of polyphosphates evokes chemiluminescence of sufficient intensity to enable the sensitive determination of these species. Limits of detection for 5-hydroxyindole-3-acetic acid (5-HIAA), vanilmandelic acid (VMA; α,4-dihydroxy-3-methoxybenzeneacetic acid), 4-hydroxy-3-methoxyphenylglycol (MHPG), homovanillic acid (HVA, 4-hydroxy-3-methoxyphenylacetic acid) and 3,4-dihydroxyphenylacetic acid (DOPAC) were between 5 × 10⁻⁹ and 4 × 10⁻⁸ M, using flow-injection analysis methodology. In addition, we demonstrate the rapid determination of homovanillic acid and 5-hydroxyindole-3-acetic acid in human urine - without the need for extraction procedures - using monolithic column chromatography with chemiluminescence detection.     

Separation of six purine bases by capillary electrophoresis with electrochemical detection

Capillary zone electrophoresis with electrochemical detection (ED) has been employed for the separation and determination of adenine (A), guanine (G), theophylline (Thp), hypoxanthine (HX), xanthine (Xan) and uric acid (UA). Effects of several important factors such as the acidity and concentration of running buffer, separation voltage, injection time and detection potential were investigated to acquire the optimum conditions. The detection electrode was a 300 μm carbon disc electrode at a working potential of +0.95 V (versus saturated calomel electrode (SCE)). The six purine bases can be well separated within 14 min in a 40 cm length fused-silica capillary at a separation voltage of 10 kV in a 100 mmol/l borate buffer (BB, pH 10.0). The current response was linear over about three orders of magnitude with detection limits (S/N=3) ranging from 0.157×10⁻⁶ to 0.767×10⁻⁶ mol/l for all compounds. The proposed method was successfully applied to determine Thp in tea and aminophylline tablets, UA in human urine, and two purine bases in DNA.     

Study of uptake kinetics of vincristine for human erythrocytes by capillary zone electrophoresis with electrochemical detection

Capillary zone electrophoresis (CZE) was employed for the determination of vincristine using electrochemical detection with a carbon fiber microdisk bundle electrode at a constant potential of 1.0 V versus saturated calomel electrode (SCE). The optimum conditions of separation and detection are 1.7×10⁻² Na₂HPO₄− 3.2×10⁻³ mol/l NaH₂PO₄ (pH 7.5) for the buffer solution, 20 kV for the separation voltage. The limit of detection is 5.0×10⁻⁷ mol/l or 2.2 fmol (S/N=3) for the injection voltage of 5 kV and the injection time of 10 s. The recovery of the method is between 95 and 101% for the vincristine taken by human erythrocytes. The method was applied to investigate uptake and accumulation behavior of vincristine for human erythrocytes. The advantages of the method are the small sample volume of CZE and the high selectivity and sensitivity of electrochemical detection.     

Separation and determination of l-tyrosine and its metabolites by capillary zone electrophoresis with a wall-jet amperometric detection

A method of capillary electrophoresis with wall-jet amperometric detection (AD) has been developed for separation and determination of l-tyrosine (Tyr) and its metabolites, such as Tyramine (TA), p-hydroxyphenylpyruvic (pHPP), homogentisic acid (HGA) and some dipeptides containing Tyr, such as Tyr-Gly-Gly (YGG), Tyr-Arg (YR) and Tyr-d-Arg (Y-d-R). A carbon disk electrode was used as the working electrode and the optimal detection potential was 1.00 V (versus Ag/AgCl). At 18 kV of applied voltage, the seven compounds were completely separated within 20 min in 110 × 10⁻³ mol/L Na₂HPO₄-NaH₂PO₄ buffer (pH 7.10) containing 3 × 10⁻³ mol/L β-cyclodextrin (β-CD). Good linear relationship was obtained for all analytes and the detection limits of seven analytes were in the range of 0.95-4.25 ng/mL. The proposed method has been applied to examine the metabolic process of l-tyrosine in rabbit's urine.     

Study on the co-luminescence effect of Tb-Gd-epinephrine system and its application to the sensitive determination of epinephrine at nanomol level

A new fluorimetric method for the determination of epinephrine is established by rare earth co-luminescence effect of Tb-Gd-E-Tris system. Under optimum conditions, a linear relationship has been obtained between the fluorescence intensity and the concentration of epinephrine in the range of 1.8 × 10⁻⁸ to 2.5 × 10⁻⁶ mol/l, and the detection limit is 4.5 × 10⁻⁹ mol/l (S/N = 3). The method is applied for the determination of epinephrine in injection and the recovery test in urine by standard addition method, and the results obtained are satisfactory. The mechanism of the co-luminescence in the system was also discussed.     

Preparation of silver hexacyanoferrate nanoparticles and its application for the simultaneous determination of ascorbic acid, dopamine and uric acid

A silver hexacyanoferrate nanoparticles/carbon nanotubes modified glassy carbon electrode was fabricated and then successfully used for the simultaneous determination of ascorbic acid, dopamine and uric acid by cyclic voltammetry. A detailed investigation by transmission electron microscopy (TEM) and electrochemistry was performed in order to elucidate the preparation process and properties of the nanocomposites. The size of silver hexacyanoferrate nanoparticles was examined by TEM around 27 nm. Linear calibration plots were obtained over the range of 4.0 × 10⁻⁶-7.8 × 10⁻⁵, 2.4 × 10⁻⁶-1.3 × 10⁻⁴ and 2.0 × 10⁻⁶-1.5 × 10⁻⁴ mol L⁻¹ with detection limits of 4.2 × 10⁻⁷,1.4 × 10⁻⁷ and 6.0 × 10⁻⁸ mol L⁻¹ for ascorbic acid, dopamine and uric acid, respectively. The practical analytical utilities of the modified electrode were demonstrated by the determination of ascorbic acid, dopamine and uric acid in urine and human blood serum samples.     

Square wave adsorptive stripping voltametric determination of the mixture of nalidixic acid and its main metabolite (7-hydroxymethylnalidixic acid) by multivariate methods and artificial neural network

Nalidixic acid (NA) and its main metabolite, 7-hydroximethylnalidixic acid (OHNA), are quinolones antibacterial used as agents used for the treatment of urinary tract infection. For both compounds an adsorption process on a hanging mercury electrode (HMDE). On this basis, a square wave adsorptive stripping voltammetry (SWadSV) method has been developed for the individual and simultaneous determination of NA and OHNA. The variables that affect to accumulation process, such as concentration of perchloric acid, accumulation potential and accumulation time have been optimised by using an experimental design (concretely a Box-Behnken design with three levels) together with the response surface methodology (RSM). Calibration curves were linear in the range (0-1.38) × 10⁻⁷ mol L⁻¹ for NA and (0-3.23) × 10⁻⁸ mol L⁻¹ for OHNA, in the optimized conditions, with detection limits of 9.48 × 10⁻⁹ mol L⁻¹ and 8.06 × 10⁻¹⁰ mol L⁻¹ for NA and OHNA, respectively. The method was applied to urine samples containing only one of the analytes with satisfactory recoveries. As the voltammetric signals of these compounds show a high overlapping, different chemometric methods, such as classical least squares (CLS), partial least squares (PLS), principal component regression (PCR) and artificial neural network (ANN) have been used for the resolution of the mixture. The analysis of these compounds in urine samples were carried out using the different chemometric tools and the best recoveries were obtained by using ANN. No pre-treatment of the sample was necessary.     

Biosensor for chlorogenic acid based on an ionic liquid containing iridium nanoparticles and polyphenol oxidase

A biosensor based on the ionic liquid, 1-n-butyl-3-methylimidazolium hexafluorophosphate containing dispersed iridium nanoparticles (Ir-BMI.PF₆) and polyphenol oxidase was constructed. This enzyme was obtained from the sugar apple (Annona squamosa), immobilized in chitosan ionically crosslinked with oxalate. The biosensor was used for determination of chlorogenic acid by square wave voltammetry. The polyphenol oxidase catalyzes the oxidation of chlorogenic acid to the corresponding o-quinone, which is electrochemically reduced back to this substance at +0.25 V vs. Ag/AgCl. Under optimized operational conditions the chlorogenic acid concentration was linear in the range of 3.48 × 10⁻⁶ to 4.95 × 10⁻⁵ mol L⁻¹ with a detection limit of 9.15 × 10⁻⁷ mol L⁻¹. The biosensor was applied in the determination of chlorogenic acid in organic and decaffeinated coffee and the results compared with those obtained using the capillary electrophoresis method. The recovery study for chlorogenic acid in these samples gave values of 93.2-105.7%.     

Determination of puerarin and daidzein in Puerariae radix and its medicinal preparations by micellar electrokinetic capillary chromatography with electrochemical detection

The use of micellar electrokinetic capillary chromatography (MECC) with electrochemical detection is described for the determination of puerarin and daidzein in Puerariae radix and its medicinal preparations. Operated in a wall-jet configuration, a 300 μm diameter carbon-disk electrode was used as the working electrode, which exhibits good responses at +900 mV (versus SCE) for the two analytes. Under the optimum conditions, the analytes were base-line separated within 11 min in a sodium dodecyl sulphate—borax (pH 7.8) running buffer, and excellent linearity was obtained in the concentration range from 5.0×10⁻⁴ to 5.0×10⁻⁶ mol/l. The detection limit (S/N=3) was 6×10⁻⁷ and 1.1×10⁻⁶ mol/l for puerarin and daidzein, respectively. This work provides a useful method for the analysis of traditional Chinese medicines.     

A novel capillary microliter droplet sample injection-chemiluminescence detector and its application to the determination of benzoyl peroxide in wheat flour

A novel capillary microliter order droplet injection-chemiluminescence (CL) system is proposed. In this system, the liquid sample microdrops, automatically formed at the end of a capillary tip by the effect of the gravity and the gas pressure, repeatedly drop into the miniature reaction cell and reacts with CL reagent to generate CL signal. The phenomena of sample zone dilution and spreading are eliminated as the capillary is used as the sample channel and gas pressure is used as driving force without the liquid carrier stream. Therefore, a high sensitivity is obtained. To evaluate the applicability of the proposed method, a determination of benzoyl peroxide (BP) is examined. The system shows that the benzoyl peroxide is detected linearly in the concentration range from 5×10⁻¹⁰ to 1×10⁻⁶ g ml⁻¹. The detection limit (signal-to-noise ratio=3) is 1.4×10⁻¹⁰ g ml⁻¹ for benzoyl peroxide (mass concentration is 1.1 pg, i.e., 4.5 fmol), which is the best result reported so far. The relative standard deviation (n=11) is 1.5% for 2.0×10⁻⁸ g ml⁻¹ BP. The proposed detector for the detection of benzoyl peroxide offers the advantages of sensitivity, simplicity, rapidity, automation and miniaturization. The proposed method has been applied satisfactorily to the determination of benzoyl peroxide in wheat flour.     

Ultratrace analysis for trans,trans-muconic acid by electrophoric derivatization and capillary gas chromatography

A highly sensitive method is described for the determination of trans,trans-muconic acid (MA), a biomarker of benzene exposure. The method is based on the derivatization of MA with an electrophoric reagent, 2-(pentafluorophenoxy)ethyl-2-(piperidino)ethanesulfonate, using potassium carbonate and 18-crown-6 ether as reaction activators. The resulting pentafluorophenoxy derivative of MA was analyzed by capillary GC with an electron-capture detector (ECD). The lower quantitation limit of the method is attainable at 0.3 μM of MA with a detection limit of about 60 nM (S/N=3) (60 fmol per 1.0 μl injection). Application of the method to the analysis of MA in urine proved feasible.     

Simultaneous determination of two active ingredients in Flos daturae by capillary electrophoresis with electrochemiluminescence detection

The coupling of Ru(bpy)₃²⁺ based electrochemiluminescence (ECL) detection with capillary electrophoresis (CE) was developed for the simultaneous determination of the two major active ingredients (atropine and scopolamine) in Flos daturae. Parameters related to the separation and detection were discussed and optimized. It was proved that 20 mM phosphate buffer at pH 8.48 could achieve the most favorable resolution, and the high sensitivity of detection was obtained by maintaining the detection potential at 1.2 V. Under the optimized conditions: ECL detection at 1.2 V, 20 mM phosphate buffer at pH 8.48, 5 mM Ru(bpy)₃²⁺ and 50 mM phosphate buffer at pH 7.48 in the detection reservoir, detection limits of 5 × 10⁻⁸ mol/l for atropine and 1 × 10⁻⁶ mol/l for scopolamine were obtained. Relative standard derivations of the ECL intensity and the migration time were 5.16 and 0.71% for atropine and 5.07 and 1.22% for scopolamine, respectively. Developed method was successfully applied to determine the amounts of both alkaloids in Flos daturae. A baseline separation for atropine and scopolamine was achieved within 11 min.     

A quick method for the simultaneous determination of ascorbic acid and sorbic acid in fruit juices by capillary zone electrophoresis

A method of quickly determining ascorbic acid and sorbic acid by capillary zone electrophoresis with ultraviolet detection was developed. The choice of background electrolyte, wavelength, injection time and applied voltage were discussed. Ascorbic acid and sorbic acid were well separated in 80 mmol L⁻¹ boric acid-5 mmol L⁻¹borax (pH = 8.0) in 5 min at the detecting wavelength of 270 nm. Under the optimum condition, the method has linear ranges of 2.54-352.00 mg L⁻¹ for ascorbic acid and 1.08-336.39 mg L⁻¹ for sorbic acid with the detection limit of 1.70 mg L⁻¹ for ascorbic acid and 0.54 mg L⁻¹ for sorbic acid, respectively. Other organic acids in fruit juices have no effect on the detection. This method is very feasible and simple and can be used to detect ascorbic acid and sorbic acid in fruit juices.     

A flow-injection chemiluminescence method for the determination of some estrogens by enhancement of luminol-hydrogen peroxide-tetrasulfonated manganese phthalocyanine reaction

A novel flow-injection chemiluminescence (FI-CL) method for the determination of estrogens is proposed, based upon its enhancing effect on the CL reaction of luminol with hydrogen peroxide catalyzed by tetrasulfonated manganese phthalocyanine (MnTSPc) in alkaline solution. Under the selected experimental conditions, a linear relationship was obtained between the CL intensity and the concentration of estrone in the range of 1.0 × 10⁻⁷ to 1.0 × 10⁻⁶ mol/l, estradiol in the range of 9.0 × 10⁻⁸ to 1.0 × 10⁻⁶ mol/l and estriol in the range of 3.0 × 10⁻⁷ to 2.0 × 10⁻⁶ mol/l, respectively. The detection limits were 5.1 × 10⁻⁸ mol/l for estrone, 7.2 × 10⁻⁹ mol/l for estradiol and 6.5 × 10⁻⁸ mol/l for estriol with a relative standard deviation of 2.8% for 5.0 × 10⁻⁷ mol/l estrone, 2.4% for 1.0 × 10⁻⁷ mol/l estradiol, and 3.1% for 7.0 × 10⁻⁷ mol/l estriol (n = 11). This method has been applied to the determination of estrogen in pharmaceutical injections and tap water with satisfactory results.     

A flow-based procedure with solenoid micro-pumps for the spectrophotometric determination of uric acid in urine

The determination of uric acid in urine shows clinical importance, once it can be related to human organism dysfunctions, such as gout. An analytical procedure employing a multicommuted flow system was developed for the determination of uric acid in urine samples. Cu(II) ions are reduced by uric acid to Cu(I) that can be quantified by spectrophotometry in the presence of 2,2′-biquinoline 4,4′-dicarboxylic acid (BCA). The analytical response was linear between 10 and 100 μmol L^− 1 uric acid with a detection limit of 3.0 μmol L^− 1 (99.7% confidence level). Coefficient of variation of 1.2% and sampling rate of 150 determinations per hour were achieved. Per determination, 32 μg of CuSO₄ and 200 μg of BCA were consumed, generating 2.0 mL of waste. Recoveries from 91 to 112% were estimated and the results for 7 urine samples agreed with those obtained by the commercially available enzymatic kit for determination of uric acid. The procedure required 100-fold dilution of urine samples, minimizing sample consumption and interfering effects. In order to avoid the manual dilution step, on-line sample dilution was achieved by a simple system reconfiguration attaining a sampling rate of 95 h^− 1.     

Determination of ethylenediaminetetraacetic acid, ethylenediaminedisuccinic acid and iminodisuccinic acid in cosmetic products by capillary electrophoresis and high performance liquid chromatography

A capillary electrophoresis (CE) and a high performance liquid chromatography (HPLC) method are described for the simultaneous determination of ethylenediaminetetraacetic acid (EDTA), S,S′-ethylenediaminedisuccinic acid (EDDS) and R,S-iminodisuccinic acid (IDS) complexing agents as their Fe(III) complexes in cosmetics like shower cream and foam bath. The non-biodegradable EDTA is used in combination with biodegradable analogues like EDDS and IDS in many commercial products. The HPLC method involves separation by reversed-phase ion pair chromatography on a C₁₈ column using methanol-formate buffer (20 mM tetrabutylammonium hydrogen sulfate, 15 mM sodium formate adjusted to pH 4.0 with formic acid) (10:90, v/v) as mobile solvent at a flow rate of 0.8 mL min⁻¹ at 24 °C using UV detection at 240 nm. The CE separation was performed in a fused silica capillary of 50 μm i.d. with the total length of 50 cm with a 10 mM MES and MOPSO (pH 5.5) at an applied voltage of −25 kV. The samples were introduced by applying a 50 mbar pressure for 2 s. Absorbances at 215 and 225 nm were monitored for the detection of the complexes. The methodology performance of the two methods was evaluated in terms of linearity, limit of detection (LOD), limit of quantitation (LOQ) and reproducibility. The LOD values obtained from HPLC are low when compared with CE. The applicability of both the methods was demonstrated for the analysis of cosmetic products such as shower cream and foam bath. The results obtained by both CE and HPLC were found to be comparable and in good agreement.     

Simultaneous determination of epinephrine, uric acid and xanthine in the presence of ascorbic acid using an ultrathin polymer film of 5-amino-1,3,4-thiadiazole-2-thiol modified electrode

This paper describes the simultaneous determination of epinephrine (EP), uric acid (UA) and xanthine (XN) in the presence of ascorbic acid (AA) using electropolymerized ultrathin film of 5-amino-1,3,4-thiadiazole-2-thiol (p-ATT) modified glassy carbon (GC) electrode in 0.2 M phosphate buffer solution (pH 5). Although bare GC electrode resolves the voltammetric signals of AA and XN, it fails to resolve the voltammetric signals of EP and UA in a mixture. However, the p-ATT modified electrode not only separates the voltammetric signals of AA, EP, UA and XN with potential difference of 150, 120 and 400 mV between AA-EP, EP-UA and UA-XN, respectively but also shows higher oxidation current for these molecules. The p-ATT modified electrode exhibits excellent selectivity towards the oxidation of EP, UA and XN in the presence of 40-fold higher concentration of AA. Further, the p-ATT modified electrode was also used for the selective determination of EP in the presence of 40-fold higher concentrations of AA, UA and XN. Using amperometric method, we achieved the lowest detection of 40 nM EP and 60 nM each UA and XN. The amperometric current response was increased linearly with increasing EP concentration in the range of 4.0 × 10⁻⁸ to 4.0 × 10⁻⁵ M and the detection limit was found to be 27 × 10⁻¹¹ M (S/N = 3). The practical application of the present modified electrode was demonstrated by determining the concentration of EP in epinephrine tartrate injection and XN in human urine samples.     

Rosmarinic acid determination using biomimetic sensor based on purple acid phosphatase mimetic

A new heterodinuclear Fe(III)Zn(II) complex which mimics the active site of the hydrolytic enzyme red kidney bean purple acid phosphatase was synthesized and characterized by IR, CHN and X-ray crystallographic analyses. This complex, [Fe^IIIZn^II(μ-OH)bpbpmp-CH₃](ClO₄)₂, containing the ligand (H₂bpbpmp-CH₃ = {2-[bis(2-pyridylmethyl)aminomethyl]-6-[(2-hydroxy-5-methylbenzyl) (2-pyridyl-methyl) aminomethyl]-4-methyl-phenol}) was employed in the construction of a biomimetic sensor and used in the determination of rosmarinic acid in plant extract samples. The response parameters and optimization of the biomimetic sensor design were evaluated. The best performance of this sensor was obtained for 75:15:10% (w/w/w) of the graphite powder:nujol:Fe(III)Zn(II) complex, 0.1 mol L⁻¹ phosphate buffer solution (pH 7.5), 1.19 × 10⁻⁴ mol L⁻¹ hydrogen peroxide with frequency, pulse amplitude, and scan increment at 30 Hz, 100 mV, and 0.6 mV, respectively. The rosmarinic acid concentration was linear in the range of 2.98 × 10⁻⁵ to 3.83 × 10⁻⁴ mol L⁻¹ (r = 0.9991) with a detection limit of 2.30 × 10⁻⁶ mol L⁻¹. This biomimetic sensor demonstrated long-term stability (300 days; 900 determinations) and reproducibility, with a relative standard deviation of 12.0%. The recovery study of rosmarinic acid in plant extract samples gave values from 90.3 to 98.3% and the concentrations determined showed agreement when compared with those obtained using capillary electrophoresis at the 95% confidence level.     

Simultaneous determination of flumequine and oxolinic acid in chicken tissues by solid phase extraction and capillary electrophoresis

This paper describes capillary electrophoresis (CE) methodology for simultaneous determination of oxolinic acid (OXO) and flumequine (FLU) in spiked chicken tissue using norfloxacin (NOR) as internal standard (IS), with diode array detection. The analytes were extracted using dichlorometane and NaOH and pre-concentrated by solid phase extraction (SPE).The recoveries obtained were 94 and 84% for oxolinic acid and flumequine , respectively. The detection and quantification limits achieved were 15 and 48 μg kg⁻¹ for oxolinic acid, respectively, and 10 and 30 μg kg⁻¹ for flumequine. The sensitivity of the method proposed allows the determination of these drugs at a residue level far below their maximum residue limit (MRL) established by the European Union (EU).     

Determination of three Tumor Biomarkers (Homovanillic Acid,Vanillylmandelic Acid,and 5‐Hydroxyindole‐3‐Acetic Acid) Using Flow Injection Analysis with Amperometric Detection

Flow injection analysis with amperometric detection (FIA‐AD) at screen‐printed carbon electrodes (SPCEs) in optimum medium of Britton‐Robinson buffer (0.04 mol ? L^?1, pH 2.0) was used for the determination of three tumor biomarkers (homovanillic acid (HVA), vanillylmandelic acid (VMA), and 5‐hydroxyindole‐3‐acetic acid (5‐HIAA)). Dependences of the peak current on the concentration of biomarkers were linear in the whole tested concentration range from 0.05 to 100 μmol ? L^?1, with limits of detection (LODs) of 0.065 μmol ? L^?1 for HVA, 0.053 μmol ? L^?1 for VMA, and 0.033 μmol ? L^?1 for 5‐HIAA (calculated from peak heights), and 0.024 μmol ? L^?1 for HVA, 0.020 μmol ? L^?1 for VMA, and 0.012 μmol ? L^?1 for 5‐HIAA (calculated from peak areas), respectively.