Determination of hydrazines by chip electrophoresis with contactless conductivity detection

In this report, a new approach for the fast determination of hydrazine compounds (hy) in complex matrices is presented. The experimental protocol is based on poly(methylmethacrylate) (PMMA) microchip separations with contactless conductivity detection using a compact portable device, which integrates all separation and detection components. Three hy (hydrozine (Hy), methylhydrazine (MH), and 1,1-dimethylhydrazine (UDMH)) were separated within < 30 s at a separation voltage of 3.8 kV using a L(-)-histidine/2-(N-morpholinoethanesulfonic acid) (His/MES) buffer (25:50 mM, pH 5.87). The contactless conductivity detection enables detection limits for Hy, MH, and UDMH of 11.9, 35.5, and 337.8 ng/mL, respectively, with linear concentration dependence up to 10 μg/mL. In complex matrices such as soil samples or river water, interferences were eliminated by implementing ultrasound-assisted headspace single-drop microextraction of hy under strongly alkaline conditions, using an aqueous drop of His/MES buffer as the extractant phase. The incorporation of this miniaturized sample preparation step led to improved limits of detection for Hy, MH, and UDMH of 6.5, 15.3, and 11.4 ng/mL, respectively. The overall protocol demonstrates a promising approach for interfacing chip electrophoresis with real-world applications.     

Determination of enkephalin peptides by nonaqueous capillary electrophoresis with electrochemical detection

Nonaqueous capillary electrophoresis with electrochemical detection (NACE-ED) was applied to the analysis of enkephalin peptides. The effect of different buffer compositions on the electrophoretic behavior of methionine enkephalin, leucine enkephalin, and [D-Ala2]-leucine enkephalin was studied. Separation of the protonated and the deprotonated peptides was obtained using ACN/methanol-based electrolyte systems. The electrochemical behavior of the enkephalins was studied by the capillary batch injection analysis technique. NACE-ED yielded well-defined signals in the oxidation mode only for the negatively charged analytes. The optimized BGE for the counterelectroosmotic separation consisted of 10 mM ammonium acetate in ACN/methanol (3:1 v/v). Using a platinum microdisk electrode set to an actual potential of +0.65 V detection limits in the submicromolar range were observed which are about one order of magnitude lower compared to UV detection. Problems concerning EOF instability and electrode fouling caused by water and other neutral sample impurities transported by the EOF can be avoided in the EOF-inverted mode using poly(ethylene glycol)-coated capillaries and an actual working electrode potential of +1.0 V. For the quantification of the enkephalins [D-Ala2]leucine enkephalin was used as internal standard. The practical utility for the determination of enkephalins in spiked plasma samples after SPE was demonstrated.     

Polycations as displacer in high-performance bioseparation

Non-aqueous capillary electrophoresis with electrochemical detection (NACE-ED) was applied to the determination of nicotine. The measurements were performed using an acetonitrile-based buffer. Nicotine was shown to yield well defined voltammetric signals suitable for oxidative detection. The precision of NACE-ED regarding migration time and peak height for samples containing 8 micrograms/ml nicotine is expressed by relative standard deviations of 0.1% and 1.6%, respectively (n = 8). The limit of detection for nicotine was 13 ng/ml (286 fg). For nicotine determination in tobacco samples various solutions were studied regarding the extraction efficiency in an ultrasonic bath. The highest extraction efficiency was obtained using a solvent mixture consisting of acetonitrile-acetic acid-water (20:5:75, v/v). The results for nicotine determination in tobacco were evaluated using tobacco reference material with certified nicotine content. Analytical aspects such as accuracy, reproducibility and selectivity were addressed in this work. The measurements were based on the use of a newly developed electrochemical detector cell which was found to enable user-friendly operation of NACE-ED measurements.     

Determination of 1,1-dimethylhydrazine and its decomposition products using ion-pair chromatography

The influence of the nature of the ion-pair reagent (IPR) using sodium octylsulfate, octylsulfonate, and dodecylsulfate as an example on the separation of asymmetric dimethylhydrazine (ADMH) and its decomposition products has been investigated using a Synergi Hydro RP column. It has been shown that the use of sodium octylsulfate as an IRP is preferable. The approach to simultaneous determination of hydrazine, methylhydrazine, ADMH, N-nitrosodimethylamine, and tetramethyl-2-tetrazene with detection limits in water matrices of 0.3, 0.7, 0.8, 7, and 1.2 μg/L respectively (sample volume of 100 μL) has been proposed.     

Determination of cannabinoids in hair using high-pH* non-aqueous electrolytes and electrochemical detection. Some aspects of sensitivity and selectivity

Non-aqueous capillary electrophoresis with electrochemical detection (NACE-ED) was applied to the determination of cannabinoids in hair. The effect of different electrolyte compositions on the selectivity of the separation of tetrahydrocannabinol (THC), cannabinol (CBN), cannabidiol (CBD) and tetrahydrocannabinol carboxylic acid (THCA) was studied. Complete electrophoretic resolution was obtained using a strongly basic background electrolyte consisting of 5 mM sodium hydroxide dissolved in acetonitrile-methanol (1:1). Electrochemical detection yielded well defined signals in the oxidation mode. In order to obtain low limits of detection experimental parameters, which determine the sensitivity and the noise level, were optimized. A crucial parameter for sensitive measurements using a wall-tube flow cell as end-column detector is the distance between the capillary outlet and the working electrode. The highest signal-to-noise ratio using a 50 microm I.D. capillary was obtained at a distance of 25 microm. When the capillary outlet was moved away from the working electrode, thus reducing the strength of the separation field present at the working electrode, a large low frequency noise developed. This rise was attributed to disturbances of the hydrodynamic pattern in the flow cell. Analytical aspects such as sensitivity, reproducibility and selectivity were addressed in this work. The precision of NACE-ED regarding migration time and peak height for a sample containing 1 microg/ml THC was 0.4% and 1.1% (RSD), respectively (n=5). The calibration curve was linear for concentrations ranging between 0.1 and 10 microg/ml (r=0.998). The limit of detection for THC was 37 ng/ml, which is almost two orders of magnitude lower when compared with on-column UV detection. The method was evaluated using hair samples containing cannabinoids as sample material.     

Simultaneous determination of celecoxib, meloxicam, and rofecoxib using capillary electrophoresis with surfactant and application in drug formulations

A simple and selective CE using surfactant with UV detection is described for the simultaneous determination of selective cyclooxygenase-2 inhibitors, celecoxib, meloxicam, and rofecoxib. The simultaneous analysis of celecoxib, meloxicam, and rofecoxib was performed in Tris buffer (10 mM; pH 11) with 60 mM sodium octane-sulfonate and 20% ACN as an anionic surfactant and organic modifier, respectively. Under this condition, good separation with high efficiency and the required short analysis time is achieved. The linear ranges of the method for the determination of celecoxib, meloxicam, and rofecoxib were over 5-100 microg/mL; the detection limits at 200 nm (S/N = 3; injection 3.45 kPa, 5 s) were 2, 1, and 1 microg/mL, respectively. The small amount of sample required and the expeditiousness of the procedure allow content uniformity to be determined in individual pharmaceutical products.     

Analysis of ecdysteroids by micellar electrokinetic chromatography with on-line preconcentration

In order to enhance the UV detection sensitivity, an application study of an on-line preconcentration technique for micellar electrokinetic chromatographic (MEKC) was carried out. The simultaneous determination of four test ecdysteroids, 20-hydroxyecdysone, ajugasterone C, polypodine B and ponasterone A has been investigated by using the normal stacking mode in MEKC with UV detection. The effects of anionic surfactant composition and concentration, the applied voltage, the pH buffer, the kind and the amount of organic solvent and the injection time on the analyte resolution were evaluated. The optimised conditions for the separation involved the use of a 50 mM borate as the running buffer containing 50 mM of a mixture of sodium dodecyl sulphate (SDS) and sodium cholate (SC) in the ratio of 1:1 together with a concentration of 10% (v/v) of 2-PrOH at pH 9.0. Hydrodynamic injection of 12 s at 50 mbar and separation voltage of 20 kV at temperature of 20 degrees C were employed. These conditions allowed a repeatability separation within 21 min. Concentration detection limit for the neutral analytes studied improve about an order of magnitude. The method was also applied to the determination of ecdysteroids in a real sample.     

Direct determination of hydrazine,methylhydrazine, and 1,1-dimethylhydrazine by zwitterionic hydrophilic interaction liquid chromatography with amperometric detection

A promising alternative to ion-chromatographic methods currently used for the direct determination of hydrazines is provided by hydrophilic interaction liquid chromatography (HILIC). In this work, we propose a method for the simultaneous determination of hydrazine, methylhydrazine and 1,1-dimethylhydrazine in natural waters and soils based on a combination of chromatographic separation on a zwitterionic sulfobetaine stationary phase (Nucleodur HILIC) in the HILIC mode with amperometric detection.

Effects of different factors on the retention of analytes were studied and the optimum conditions of analysis were found. We recommend a mixture of acetonitrile with an aqueous phosphate buffer solution of pH 2.5 (78:22 v/v) with an ionic strength of 20 mM as a mobile phase. Detection in the direct current mode was performed at a working electrode potential of 1.1 V.

The advantages of the method are the high efficiency of separation, rapidity, high sensitivity and a wide dynamic range of analyte concentrations, covering four orders of magnitude. The attained LOD values for analytes lie in the range 0.07–0.13 μg L^–1, which is two orders of magnitude lower than those in currently used methods of ion chromatography with electrochemical and mass spectrometric detection.

The method was validated on samples of natural waters of different origin using the added–found technique. It was found that the error of analysis did not exceed 10% for river and ground waters and increased to 20–30% for peat bog surface waters.

The possibility of application of the developed method to the analysis of soils was shown on samples of peat bog soils selected at places of impact of the first steps of carrier rockets and polluted by rocket fuel based on 1,1-dimethylhydrazine.     

Determination of aluminium,copper, iron and manganese in biological and other samples as 8-quinolinol complexes by high-performance liquid chromatography with electrochemical and spectrophotometric detection

A range of methods based on reverse-phase high-performance liquid chromatography is described for determining Al, Cu, Fe and Mn. The simplest method for determining Fe, Cu and Al involves direct formation and separation of the 8-quinolinol complexes on the column, with 1:1 acetonitrile/water containing 5 × 10^?3 M 8-quinolinol, 0.4 M potassium nitrate and 0.02 M acetate buffer (pH 6.0) as the mobile phase, followed by electrochemical detection at ?0.5 V vs. Ag/AgCl and/or spectrophotometric detection at 400 nm. Electrochemical detection enables < 2 ng Cu and < 1 ng Fe to be quantified for injection volumes of 20 μ1. Spectrophotometric detection allows simultaneous determinations of Al, Cu and Fe with lower sensitivity. The method is applied to the determination of Cu, Fe and Al in bovine liver and oyster tissue. Down to 1 ng of manganese (in the 20 μ1 injected) can be determined in biological samples by liquid chromatography with a mobile phase containing 1 mM Tris buffer (pH 8.8) after injection of an externally prepared 8-quinolinol complex. Preconcentration on Sep-Pak cartridges after dichloromethane extraction is used for the determination of low concentrations of iron in water. A sensitive determination of aluminium based on detection at 254 nm is also reported.     

Optimized Separation of cis-trans Isomers and Enantiomers of Sertraline Using Cyclodextrin-Modified Micellar Electrokinetic Chromatography

Micellar electrokinetic chromatography has been investigated for the separation of cis-trans isomers and enantiomers of sertraline. The effects of various separating factors were studied. Optimum separation was achieved using a buffer (pH 11.5) of 35 mM sodium borate containing 30 mM sodium deoxycholate and 20 mM hydroxypropyl –-cyclodextrin; the optimum voltage and temperature were 25 kV and 20 °C, respectively. A detection wavelength of 210 nm was used. The analytical performance of the method was discussed in terms of linearity response, precision, detection limits, quantification limits and recoveries. The method developed was successfully applied to the determination of sertraline in bulk drug, tablets and capsules.Revised: 1 April and 18 May 2004     

Separation and determination of flavonoids using microemulsion EKC with electrochemical detection

A selective and sensitive method of microemulsion EKC (MEEKC) with electrochemical detection (ED) was developed for separation and determination of 14 flavonoids. In order to obtain the better stability for the studied flavonoids, oil (ethyl acetate) with low interfacial surface tension was employed as organic solvent. A running buffer composed of 0.9% (w/v, 30 mM) SDS, 0.9% (w/v, 21 mM) sodium cholate (SC), 0.9% (w/v, 121 mM) butan-1-ol, 0.6% (w/v, 68 mM) ethyl acetate, and 98.2% v/v 10 mM Na(2)B(4)O(7)-20 mM H(3)BO(3) buffer (pH 7.5) was applied for the separation of flavonoids. Under the optimum conditions, the relationship between peak currents and analyte concentrations was linear over about 1.3 and 1.7 orders of magnitude with detection limits (defined as S/N = 3) ranging from 0.02 to 0.5 microg/mL for all analytes. This method was applied for the determination of flavonoids in real samples with simple extraction procedures, and the assay results were satisfactory.     

Separating stereoisomers of di-, tri-, and tetrapeptides using capillary electrophoresis with contactless conductivity detection

The separation and detection of small oligopeptides in CE with contactless conductivity detection were demonstrated. A strongly acidic separation buffer (0.5 M acetic acid) was employed in order to render the species cationic. Separation of the stereoisomers was achieved in typically 10-15 min by using either dimethyl-beta-CD (DM-beta-CD), (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid (18C(6)H(4)), a combination of the two substances, or of histidine, as buffer additives. Calibration curves were determined for isomers of Gly-Asp and H-Pro-Asp-NH(2), in the range of 0.05-0.5 mM and 0.1-1 mM, respectively, and were found to be linear. LODs were determined to be in the order of 1.0 microM. The determination of isomeric impurities down to about 1% was found possible. Species showing good separation could also be successfully determined on an electrophoretic lab-on-chip device, with analysis times of a few minutes.     

Separation of perfluorocarboxylic acids using capillary electrophoresis with UV detection

A capillary electrophoretic method with UV detection for separation and quantitation of perfluorocarboxylic acids (PFCAs) from C6-PFCA to C12-PFCA has been developed. The optimization of measurement conditions included the choice of the most appropriate type and concentration of buffer in the background electrolyte (BGE), as well as the type and the content of an organic modifier. The optimal separation of investigated PFCAs was achieved with 50 mM phosphate buffer and 40% isopropanol in the BGE using direct UV detection. The optimum wavelength for direct UV detection was optimized at 190 nm. For indirect detection, several chromophores were studied. Five mM 3,5-Dinitrobenzoic acid (3,5-DNBA) in 20 mM phosphate buffer BGE and indirect UV detection at 280 nm gave the optimal detection and separation performance for the investigated PFCAs. The possibility of on-line preconcentration of solutes by stacking has been examined for indirect detection. The detection limits (LODs) determined for direct UV detection ranged from 2 microg/mL for C6-PFCA to 33 microg/mL for C12-PFCA. The LODs obtained for indirect UV detection were comparable to those obtained for direct UV detection.     

Temperature optimization for improved determination of phosphatidylserine species by micro liquid chromatography with electrospray tandem mass spectrometric detection

A sensitive method for determination of disaturated phosphatidylserine species in the presence of their monounsaturated analogs has been developed, using micro liquid chromatography coupled to electrospray ionization tandem mass spectrometry. The hydrophobic nature of the phosphatidylserine species required a combination of low-eluting sample solvents and sub-ambient temperatures in order to focus large sample volumes up to 20 microL. The samples were dissolved in 2-propanol:hexane:water (20:10:4, v/v/v) prior to 1:9 dilution with ammonium formate buffer:2-propanol:tetrahydrofuran (30:55:15, v/v/v) and final 1:4 dilution with ammonium formate buffer (10 mM):2-propanol: tetrahydrofuran (55:37.5:7.5, v/v/v). The analytical column was a 0.5 x 150 mm stainless steel column packed with 5 microm C30 particles, while the mobile phase contained ammonium formate buffer (10 mM): 2-propanol:tetrahydrofuran (30:55:15, v/v/v). A temperature program from 5 degrees C (hold for 3 minutes) to 75 degrees C at 8 K/min provided separation of the disaturated phosphatidylserine species from their monounsaturated analogs, making available a sensitive determination of the isobaric species. The mass limit of detection for dipalmitoyl phosphatidylserine was 100 pg, corresponding to a concentration limit of detection of 5 pg/microL when using an injection volume of 20 microL. This is an improvement by a factor of 20 as compared to previously reported numbers obtained with conventional LC columns. The within-assay precision of dipalmitoyl phosphatidylserine was 11.9% RSD (n = 3), while the retention time precision was 4.1% RSD (n = 6).     

Determination of gamma-hydroxybutyric acid in clinical samples using capillary electrophoresis with contactless conductivity detection

A method for the determination of gamma-hydroxybutyric acid (GHB) in urine and serum samples with capillary electrophoresis using capacitively coupled contactless conductivity detection (CE-C(4)D) was developed. The optimized separation buffer consisting of 20 mM of arginine, 10 mM of maleic acid and 30 microM of cetyltrimethylammonium bromide (CTAB) contained 5 mM vancomycin to facilitate the separation of gamma-hydroxybutyric acid from beta-hydroxybutyric acid (BHB), which is also present in clinical samples. The detection limits in the clinical samples were found to be about 2 microg/ml, which is well below the required sensitivity for the recognition of overdosage and adequate for the determination of endogenous concentrations in urine. The determination of GHB in both types of samples was carried out directly after a fourfold dilution without requiring any derivatization or extraction procedures.     

Separation and determination of closely related lantibiotics by micellar electrokinetic chromatography

A sensitive micellar electrokinetic chromatography (MEKC) method was developed for the separation and determination of four closely related lantibiotics: gallidermin, cinnamycin, duramycin and nisin. Factors affecting the separation of the lantibiotics such as pH, phosphate buffer concentration, SDS concentration and wavelength for UV detection were investigated. By optimizing these experimental conditions, successful separation was achieved between class 1A lantibiotics (nisin and gallidermin) and class 1B lantibiotics (duramycin and cinnamycin). The four lantibiotics were separated within 12 min in 50 mM phosphate buffer at pH 3.95 ± 0.1 containing 80 mM SDS with UV detection of 214 nm. The LOD (S/N = 3) were 61 ng/mL for gallidermin, 57 ng/mL for cinnamycin, 55 ng/mL for duramycin and 58 ng/mL for nisin. The method was successfully applied to real samples such as fermentation broth, bovine colostrum and predrop beer. This method yielded satisfactory results, with quantitative recoveries of spiked lantibiotics in the three samples ranging from 86.1 to 99.6%.     

Ion chromatography as a tool for the investigation of unsymmetrical hydrazine degradation in soils

A new procedure for the determination of 1,1-dimethylhydrazine (UDMH) in soil samples was developed. This involves the distillation of UDMH from an alkaline suspension of soil and ion chromatographic analysis of the distillate. The separation was performed on a silica cation-exchanger column with ammonium acetate buffer solution as mobile phase and amperometric detection at +1.2?V. Hydrazine (Hy) and methylhydrazine (MH), which are decomposition products of UDMH, can be determined simultaneously. The limits of detection in aqueous solutions were 0.2, 0.5 and 1?µg?L^?1 for Hy, MH and UDMH, respectively. The developed technique was used for investigating the behaviour of UDMH in spills of rocket fuels on soils. It was found that the addition of 4?kg?m^?2 UDMH resulted in a 0.02% residue one year after the soil treatment. The vertical migration of UDMH in soil was less than 50?cm.     

Capillary electrophoretic-ultraviolet method for the separation and estimation of zineb, maneb, and ferbam in food samples

A simple and sensitive capillary electrophoretic method with ultraviolet detection has been developed for the separation and determination of ferbam [iron(III)-dimethyldithiocarbamate], maneb [manganese(II)-ethylenebisdithiocarbamate] and zineb [zinc(II)-ethylenebisdithiocarbamate], in borate buffer, after their acidic decomposition and complexation with CDTA (trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid monohydrate), as CDTA-metal complexes of Fe+3, Mn +2, and Zn+2. The determination is dependent on the pH and the nature of the buffer solutions. In this method, the detection limit (signal-to-noise ratio = 3) is 0.0013, 0.0022, and 0.0023 mM for ferbam, maneb, and zineb, respectively. The relative standard deviation for the analysis of 1 mM of each was found to be 1.5 +/- 0.2%. The method was successfully applied for the analysis of red beans and grain samples spiked with ferbam, maneb, and zineb. The applicability of capillary electrophoresis as a useful tool for the simultaneous determination and analysis of ferbam, maneb, and zineb is demonstrated.     

Determination of copper and iron using [S,S']-ethylenediaminedisuccinic acid as a chelating agent in wood pulp by capillary electrophoresis.

A capillary electrophoresis (CE) method is described for the simultaneous determination of copper and iron after complexation with a readily biodegradable chelating agent, [S,S']-ethylenediaminedisuccinic acid (EDDS), in wood pulp. CE separation was performed in a fused-silica capillary (50 microm i.d.; total length, 65 cm) with an electrolyte containing 25 mM borate buffer and 0.5 mM CTAB at pH 7.0 and an applied voltage of -25 kV. The samples were introduced by applying a 50 mbar pressure for 2 s, and detection of the complexes was monitored at 245 nm. The methodology performance of the methods was evaluated in terms of the linearity, limit of detection (LOD), limit of quantitation (LOQ) and reproducibility. The applicability of the method was demonstrated for the analysis of copper and iron in wood pulp.     

Rapid and selective micellar electrokinetic chromatography for simultaneous determination of amikacin, kanamycin A, and tobramycin with UV detection and application in drug formulations

A simple and selective micellar electrokinetic chromatography (MEKC) with UV detection is described for simultaneous determination of amikacin, tobramycin, and kanamycin A, performed in Tris buffer (180 mM; pH 9.1) with 300 mM sodium pentanesulfonate (SPS) as an anionic surfactant. Under this condition, good separation with high efficiency and the required short analysis time is achieved. The linear ranges of the method for the determination of amikacin, tobramycin, and kanamycin A were 0.1-0.5 mg / mL, 0.4-2.0 mg / mL, and 0.4-2.0 mg / mL, respectively; the detection limits (signal-to-noise ratio = 3; injection, 0.5 psi 5 s) were 0.08, 0.2, and 0.2 mg / mL, respectively. The small amount of sample required and the expeditiousness of the procedure allow content uniformity to be determined in individual commercial products.