Electrophoretic Parameters Research Articles

Coupling sweeping-micellar electrokinetic chromatography with tandem mass spectrometry for the therapeutic monitoring of benzimidazoles in animal urine by dilute and shoot

A new method based on micellar electrokinetic chromatography-tandem mass spectrometry (MEKC-MS/MS) has been developed for the identification and simultaneous quantification of thirteen benzimidazoles in animal urine. In order to obtain an appropriate separation with the highest sensitivity, different electrophoretic parameters were evaluated. Under optimum conditions, the separation was performed using ammonium perfluorooctanoate as volatile surfactant and electrophoretic buffer (50mM, pH 9). To increase the sensitivity, a stacking mode named sweeping was applied, using water as injection solvent at 50mbar for 75s, obtaining sensitivity enhancement factors from 50 to 181. The method was applied to different animal urine samples, including sheep, cow and goat. The sample treatment consisted of a 1:10 (v/v) dilution with water. Calibration using sheep urine samples can be used for both goat and cow urine samples with a relative bias below 25% and relative standard deviations lower than 8%. The limits of detection were below 70µgL−1. As a result, the applicability of this rapid, simple, sensitive, and environmentally friendly method for therapeutic drug monitoring of benzimidazoles based on the analysis of animal urine has been demonstrated.

Escherichia coli adhesive coating as a chiral stationary phase for open tubular capillary electrochromatography enantioseparation

Bacteria, the microorganism with intrinsic chirality, have numerous fascinating chiral phenomena such as various chirality-triggered biological processes and behaviors. Herein, bacteria were firstly explored as novel chiral stationary phases in open-tubular capillary electrochromatography (OT-CEC) for enantioseparation of fluoroquinolone enantiomers and simultaneous separation of six fluoroquinolone antibiotics. The model strain, i.e. non-pathogenic Escherichia coli (E. coli) DH5α, was adhered onto the inner surface of positively charged polyethyleneimine (PEI) modified capillaries based on the bacterial adhesion characteristics and strong electrostatic interaction. The morphology and thickness of the bacteria adhesive coatings in the capillary were characterized by field emission scanning electron microscopy (FESEM) and atomic force microscopy (AFM). Baseline separation of ofloxacin and partial separation of lomefloxacin enantiomers could be achieved by the E. coli coated columns. The preparation parameters including the coating time and concentration of bacteria that affecting the chiral resolution were intensively investigated. The electrophoretic parameters, including pH, buffer concentration and applied voltage, were also optimized. The developed method was validated (linearity, LOD, LOQ, intra-day, inter-day and column-to-column repeatability and recovery) and successfully utilized for the quantitative analysis of ofloxacin enantiomers in the ofloxacin tablets. Moreover, only a slight decrease in the separation efficiency was observed after 90 consecutive runs on the E. coli@capillary. These results demonstrated that bacteria are promising stationary phases for chiral separation in CEC.

Simultaneous determination of amlodipine and telmisartan from pharmaceutical products by way of capillary electrophoresis

Abstract A rapid, simple and sensitive capillary zone electrophoresis method was developed for the simultaneous determination of amlodipine besylate and telmisartan, in fixed-dose combinations, through the utilization of a UV photodiode array detector. Electrophoretic parameters such as buffer concentration and pH, system temperature, applied voltage and injection parameters, were optimized in order to improve the efficiency of the separation. The best results were obtained when employing fused silica capillary (48 cm length X 50 μm ID) and 50 mM phosphate buffer electrolyte at pH 4.50, + 25 kV applied voltage, as well as 25°C system temperature. The separation was achieved in approximately 3 minutes, with a resolution of 4.90, while the order of migration was amlodipine followed by telmisartan. The analytical performance of the method was verified with regard to linearity, precision and robustness. In addition, the limit of detection and the quantification were calculated.

Separation and determination of optical isomers of phenylephrine by chiral ligand exchange capillary elcctrophoresis coupling with the promoting effect of ionic liquid

A method for the separation and determination of optical isomers of phenylephrine was developed based on the promoting effect of non-chiral ionic liquid on chiral ligand-exchange capillary electrophoresis after the electrophoretic parameters were optimized systematically. R-phenylephrine and S-phenylephrine can be separated and determined effectively in 20 mmol/L Tris-H3PO4 buffer solution (pH 5.4) composed of 4.0 mmol/L Cu(II), 8.0 mmol/L L-proline (L-Pro) and 15 mmol/L 1-butyl-3-methylimidazolium chloride ([BMIM] Cl) with the applied voltage of 20 kV, capillary temperature of 25 °C , detection wavelength of 254 nm, and injection of 5 s at 3,447 Pa. The resolution of R- and S-phenylephrines was 1. 42. The linear ranges for the determination of R-phenylephrine and S-phenylephrine were 12. 5 - 150 mg/L and 15. 0-150 mg/L, respectively. The method has been satisfactorily used for the determination of R-phenylephrine and S-phenylephrine in the spiked blood and urine samples. The spiked recoveries in the urine sample were in the range of 93. 7% -108. 2% with the RSDs lower than 3. 18% (n= 3) , and the spiked recoveries in the blood sample were in the range of 91. 4% and 113. 1% with the RSDs lower than 4. 82% (n =3).

Fully automated electrophoretically mediated microanalysis for CYP1A1 activity monitoring optimized by multivariate approach.

In this study, a fully automated incapillary system was developed to monitor the activity of CYP1A1 (Cytochrome P450, family 1, subfamily A, polypeptide 1) in physiological conditions. Ethoxycoumarin, the selected substrate, undergoes an inline bioreaction in the presence of CYP1A1 supersomes and Nicotinamide adenine dinucleotide phosphate reduced as cofactor, giving rise to hydroxycoumarin, the product that was assayed. The optimization of the experimental conditions was supported by the application of a design of experiment, providing a better understanding of electrophoretic mixing parameters that influence the metabolic reactions. The results obtained in optimal conditions were compared not only to those achieved after offline metabolization but also with liver microsomes. Finally, inhibition studies were conducted showing an important decrease of hydroxycoumarin formation using apigenin as CYP1A1 potent inhibitor. This study demonstrates the usefulness of our inline system for the fully automated in vitro metabolism studies and the screening of new CYP1A1 inhibitors.

Polymorphism of alpha-amylase and conjugation in common wheat enzyme types with quantitative traits of plants

Using polymorphism of alpha-amylase in the winter common wheat studied inheritance isoenzymes and its conjugation enzyme types with germinating grain on the "vine", grain productivity, plant height and time of ear formation. It is shown that the polymorphism isoenzyme of alpha-amylase wheat is limited by the presence of different loci whose products are similar in electrophoretic parameters. In this regard, one component of the enzyme can be controlling at one or two or three genes. Identification of a locus controlling alpha-amylase isoenzyme in the fast moving part of the electrophoretogram, designated as α-Amy-B7. Determine the distance of the locus to factor α-Amy-B6.

Electrophoretic Deposition of CuIn1–xGaxSe2 Thin Films Using Solvothermal Synthesized Nanoparticles for Solar Cell Application

CuIn1–xGaxSe2 (CIGS) thin films are successfully prepared by convenient electrophoretic deposition, using colloidal nanoparticles. This is the first report which focuses on the electrophoretic deposition (EPD) of presynthesized CIGS nanoparticles directly from their colloid for solar cell application. In this research, CIGS nanoparticles are first synthesized via solvothermal process and then dispersed in a media containing a mixture of ethanol as the solvent and triethylenetetramine as the additive to be used for the film deposition via the electrophoretic method. By simple adjustment of the electrophoretic parameters, including applied voltage, pH, deposition time, and composition of nanoparticles, CIGS thin films with controlled thickness and optoelectronic properties can be fabricated. The highest photovoltaic efficiency of 5.57% is obtained in the CuIn0.75Ga0.25Se2 sample. It is believed that this fabrication approach may open up a new gate to reduce the production cost of a highly demanding CIGS abs...

Different Training Schedules Influence Serum Electrophoretic Protein Profile in the Athletic Horse

Physical exercise induces various physiological responses and metabolic adaptations that have not been completely elucidated. The monitoring of alterations in blood parameters related to exercise is required to detect subclinical conditions. In this study, changes in serum total proteins and globulin fractions (albumin, α1, α2, β1, β2, and γ-globulins) were investigated in 15 clinically healthy horses subjected to different physical exercise for 5 weeks. Animals were divided in three equal groups: group A performed an intense training schedule, group B performed a light training schedule, and group C included sedentary subjects. After 5 weeks, group B was subjected to the same training schedule used for group A and it was indicated as group B1. Blood samples were collected from all animals at rest conditions. During the first week, three blood samples were collected at first, third, and fifth day and afterward, once a week for additional 4 weeks. Two-way repeated measures analysis of variance showed a statistical effect of sampling time and of different training schedules on total proteins and albumin levels in groups A and B as compared with groups C and B1. Based on these results, it is evident that the interpretation of changes in electrophoretic parameters in athletic horses cannot be limited to the comparison with a static normal range, but must consider their dynamic evolution with the progression of training.

Effect of Electrophoretic Deposition Parameters on the Corrosion Behavior of Hydroxyapatite-Coated Cobalt–Chromium Using Response Surface Methodology

Cobalt–chromium (Co–Cr)-based alloys have been used extensively as medical implants, but the ion release and the corrosion products can affect their mechanical integrity and biocompatibility. One of the solutions is to surface coat the substrate with hydroxyapatite via electrophoretic deposition technique. Two variables—pH of electrolyte and current density—were used to examine the electrochemical behavior of the coated sample. An experimental strategy was developed based on the response surface methodology together with the analysis of variance to verify the precision of the mathematical models and their relative parameters. Close agreement was observed between the predicted models and the experimental results. The pH value of electrolyte was a more significant factor than current density in increasing the corrosion potential (Ecorr) of the substrate. The maximum Ecorr was obtained with a current density of 12 mA cm−2 and a pH value of 4.71.

Hexafluoroisopropanol-modified cetyltrimethylammonium bromide/sodium dodecyl sulfate vesicles as a pseudostationary phase in electrokinetic chromatography

A novel catanionic surfactant vesicle system, formulated from hexafluoroisopropanol (HFIP), cetyltrimethylammonium bromide (CTAB) and sodium dodecyl sulfate (SDS), was developed as pseudostationary phase (PSP) in electrokinetic chromatography (EKC). HFIP, as an organic modifier with the prominent properties of ionization, hydrogen bond donor and hydrophobicity, was used to effectively promote the spontaneous vesicle formation from CTAB/SDS mixed aqueous solutions, where precipitates are easy to occur due to long carbon chains, and adjust the performance of CTAB/SDS vesicles. The physical features (size and viscosity) and electrophoretic parameters (electroosmotic mobility, electrophoretic mobility and elution range) of HFIP-modified CTAB/SDS vesicles were characterized as HFIP volume content (0–4%, v/v), CTAB/SDS molar ratio (2:8–7:3mol/mol) and total surfactant concentration (10–50mM) varying, respectively. The 3% v/v HFIP-modified CTAB/SDS (3:7mol/mol, 50mM) vesicle system proves to have the largest mean diameter (288.20nm) and the widest elution range (12.41), which is also much wider than that of the corresponding other four PSP systems including trifluoroethanol (TFE)-modified CTAB/SDS vesicles (5.69), isopropanol-modified CTAB/SDS micelles (2.03), HFIP-modified SDS micelles (4.86) and unmodified SDS micelles (3.12). The chromatographic performance of the HFIP-modified vesicle system was evaluated by separating eight polycyclic aromatic hydrocarbons, nitrotoluene positional isomers, five positively charged and five negatively charged/neutral drugs, respectively. Baseline or near-baseline separation was achieved for each series of solutes. Compared with the TFE-modified vesicle system, as well as the HFIP-modified and unmodified SDS micelle systems, the HFIP-modified vesicle system shows the best separation selectivity, the highest or comparable efficiency, and the lowest retention.

Rapid Determination of Uric Acid in Human Urine by CE with an Improved Electromagnetic Induction Detector

A novel and rapid method was developed for the determination of uric acid in human urine by capillary electrophoresis with an improved electromagnetic induction detector. Electrophoretic parameters affecting the separation efficiency, such as buffer composition, buffer pH, buffer concentration, and electroosmotic flow modifier, were systematically investigated. An electrolyte solution consisting of 8.5 mmol L−1 tris(hydroxymethyl)aminomethane (Tris), 1.5 mmol L−1 citric acid, and 0.2 mmol L−1 cetyltrimethylammonium bromide, pH 8.0, was found to be suitable for sample determination. Uric acid was separated and detected within 2.3 min, with a linear response range from 5 to 400 µg mL−1 and a correlation coefficient of 0.9998. Intraday and interday precision were 1.6 and 2.5 % (n = 6), respectively. The recoveries were between 98.3 and 100.5 %. This simple, effective, and stable method is a good alternative to existing methods for uric acid determination. Also, the improved electromagnetic induction detector holds great promise in clinical analysis.

Development of a Capillary Electrophoresis Method for the Separation of Fluoroquinolone Derivatives in Acidic Background Electrolyte

Abstract Introduction: Antibacterial quinolones class comprises a series of synthetic antibacterial agents, following the model of nalidixic acid. Because of their common 6-fl uorosubtituent on the quinolone ring, fluroquinolones are the most potent analogues with extended spectrum of activity and great pharmacokinetic properties. The applicability of capillary zone electrophoresis for the separation of fl uoroquinolones in acidic background electrolyte has been studied, our aim being the development of a capillary zone electrophoresis method for the simultaneous separation of six fl uoroquinolones and also to optimize the analytical conditions. The six studied fl uoroquinolones were ciprofloxacin, enoxacin, enrofloxacin, moxifloxacin, ofloxacin and sarafloxacin. Material and methods: Preliminary, we studied the electrophoretic behavior of six fluoroquinolones in an acidic pH, which highlighted the possibility of developing a separation method in this area of pH. Electrophoretic parameters infl uencing separation performance were studied and optimized. Results: A fast and reliable method has been developed, using a background electrolyte containing 100 mM phosphoric acid and the following conditions: applied voltage: + 25 kV, temperature: 20°C, injection pressure 30 mbar - 5 sec, UV detection at 280 nm, capillary: 60 cm (52 cm effective length) x 50 μm, analyte concentration: 167 μg/ml. The separation of the studied fl uoroquinolones was achieved in less than 8 minutes. Conclusions: Capillary zone electrophoresis using an acidic background electrolyte proved to be an efficient tool in the separation of fluoroquinolones from different generations. Also the proposed methods are particular environment-friendly replacement and improvement of a common high performance liquid chromatography determination with rapid analysis time without using any organic solvents.

Analysis of cerebrospinal fluid γ‐aminobutyric acid by capillary electrophoresis with laser‐induced fluorescence detection

The measurement of γ-aminobutyric acid (GABA) is suitable for investigating various neurological disorders. In this study, a sensitive and selective method for free GABA quantification in cerebrospinal fluid (CSF) has been standardised. This method is based on CE with LIF detection using 4-fluoro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-F) as a derivatisating agent. The reaction conditions (NBD-F concentration, pH, temperature and reaction time) and the electrophoretic parameters (run buffer composition and pH and separation voltage) were optimised to obtain the maximum derivatisation efficiency and electrophoretic resolution. The best resolution was obtained using 200 mM sodium borate, 10 mM SDS, 8.5 mM β-CD, pH 10 and 20 kV voltage. The method was linear in the concentration range of 2.5-1000 nM with good inter- and intra-assay precision values. The effects of CSF handling on free GABA concentrations were also evaluated. Our results show that the time delay between CSF collection and freezing strongly increases the CSF GABA values. Age-related reference values were established in 55 paediatric controls. The influence of antiepileptic therapy on free CSF GABA was studied in 38 neuropaediatric patients. Significantly, higher GABA values were obtained in patients taking valproic acid or vigabatrin therapy, which are antiepileptic drugs that modulate GABA metabolism.

Reference intervals of some electrophoretic and haematological parameters in Italian goats: comparison between Girgentana and Aspromontana breeds

The aim of this study was to establish and compare reference intervals of some haematological and electrophoretic parameters at different ages (1–2, 3–4 and over 5 years) in two Italian goat breeds: Aspromontana and Girgentana. A total of 90 female Aspromontana goats and 95 female Girgentana goats, ranging from 1 to 11 years of age, clinically healthy, were used. Blood samples were collected from each animal and analysed for haematological parameters and serum proteins profile with routine methods. Two-way analysis of variance showed a significant effect of age on white blood cell (WBC, P < 0.0001) in Aspromontana goats and on haematocrit (Hct, P < 0.009) in Girgentana goats. Statistical analysis showed a significant effect of age on concentrations of total protein (P < 0.05), albumin (P < 0.01) and on the albumin: globulins (A/G) ratio (P < 0.05) in Aspromontana goats. In Girgentana goats, statistical analysis showed a significant effect of age on concentrations of total protein (P < 0.01), albumin (P < 0.01) α-globulins (P < 0.05), γ-globulins (P < 0.01) and on the A/G ratio (P < 0.01). Age-related effects were not detected in β1-globulins and β2-globulins fractions in booth breed. Statistical analysis showed a significant effect of breed in the following haematological and electrophoretic parameters: WBC (P < 0.005), Hct (P < 0.0001), mean corpuscular volume (P < 0.0001), mean corpuscular haemoglobin (P < 0.01), mean corpuscular haemoglobin concentrations (P < 0.0001), concentrations of total protein (P < 0.03), α-globulins (P < 0.001), β1-globulins (P < 0.05), β2-globulins (P < 0.01) fractions and on the A/G ratio (P < 0.05).

Anodic Electrophoretic Deposition of TiO&lt;sub&gt;2&lt;/sub&gt; Nanoparticles Synthesized Using Sol Gel Method

Many researches on electrophoretic deposition of TiO2 nanoparticles (NPs) use commercial TiO2 nanoparticles from Degussa. TiO2 from Degussa is not use in this research because in order to make the TiO2 nanoparticle electronically charged and move under an applied constant voltage, a small amount of iodine and acetylacetone needs to be added to the suspension. It is suggested that the H+ ions generated by the reaction is absorbed on the suspended particles making them positively charged. For anodic EPD, negatively charged nanoparticles are required. In this research TiO2 nanoparticles were prepared by adding 1 ml of titanium isopropoxide dilute with 1 ml ethanol to 10 ml DI water and stirrer under 0°C for 4 hr. As prepared TiO2 nanoparticle were centrifuged at 5500 rpm for 10 min to isolate the particles from the solution. Then the TiO2 nanoparticles were resuspended in ethanol or water. TiO2 nanoparticle with particle diameter of 4.66 nm and anatase phase were produced. Important electrophoretic deposition process parameters, such as apparent pH of the TiO2 NPs, applied voltage, deposition time and solid loading, were studied during electrophoretic deposition of TiO2 nanoparticles.

Optimization of Pulse-Field Gel Electrophoresis for Borrelia burgdorferi Subtyping

ObjectiveTo optimize the performance of Pulsed-Field Gel Electrophoresis (PFGE) for the comparison of inter-laboratory results and information exchange of Borrelia burgdorferi subtyping. MethodsA panel of 34 strains of B. burgdorferi were used to optimize PFGE for subtyping. In order to optimize the electrophoretic parameters (EPs), all 34 strains of B. burgdorferi were analyzed using four EPs, yielding different Simpson diversity index (D) values and the epidemiological concordance was also evaluated. ResultsThe EP of a switch time of 1 s to 25 s for 13 h and 1 s to 10 s for 6 h produced the highest D value and was declared to be optimal for MluI and SmaI PFGE of B. burgdorferi. MluI and SmaI were selected as the first and second restriction enzymes for PFGE subtyping of B. burgdorferi according to discrimination and consistency with epidemiological data. ConclusionPFGE can be used as a valuable test for routine genospecies identification of B. burgdorferi.

Enantiomeric Separation of 13 New Amphetamine‐Like Designer Drugs by Capillary Electrophoresis, Using Modified‐‐Cyclodextrins

An easy-to-prepare chiral CE method for the enantiomeric separation of 13 new amphetamine-like designer drugs, using CDs as chiral selectors, was developed. Sulfated-β-CD was found to be the best chiral selector among the three used (sulfated-β-CD, caroboxymethyl-β-CD, dimethyl-β-CD). The separation of the analytes was achieved in a fused-silica gel capillary at 20 °C using an applied voltage of +25 kV. The optimized background electrolyte consisted of 63.5 mM H3 PO4 and 46.9 mM NaOH in water. Several electrophoretic parameters such as CD type, CD concentration (1 - 40 mg/mL), buffer pH (2.6, 3.6, 5.0, 6.0), length of the capillary (70 - 40 cm total length), amount of the organic solvent (methanol and acetonitrile) were investigated and optimized.

Analysis of genomic methylation level using micellar electrokinetic chromatography with UV detection

Analytical methods for quantification of 5'-methylcytosine in genomes are important tools to investigate epigenetic changes in gene expression during development, differentiation, aging, or cancer. Here, we report a novel genomic methylation content assay based on enzymatic hydrolysis of DNA and MEKC separation of 5'-deoxyribonucleoside monophosphates (dNMP) using the cationic surfactant CTAB as pseudostationary phase. Calf Thymus DNA was used during method development to determine electrophoretic parameters and electrolyte composition for a complete separation between 2'-deoxycytosine-5'-monophosphate and 2'-deoxy-5'-methylcytosine 5'-monophosphate (d5mCMP). Methylated and not methylated oligonucleotides were used to confirm the identity of each peak and evaluate analytical parameters of the method. The LOD of the method was found to be 12.5 pmol/μL for d5mCMP.

Establishment and Comparison of Pulsed-field Gel Electrophoresis, Multiple-locus Variable Number Tandem Repeat Analysis and Automated Ribotyping Methods for Subtyping of Citrobacter Strains

ObjectiveTo establish and compare the pulsed-field gel electrophoresis (PFGE), multiple-locus variable number tandem repeat analysis (MLVA) and automated ribotyping for subtyping of Citrobacter strains. MethodsPFGE protocol was optimized in terms of plug preparation procedure, restriction enzymes and configuration of electrophoretic parameters. MLVA method was evaluated by finding variable number tandem repeats in two genomes of Citrobacter strains. The ribotyping was performed by using the automated RiboPrinter system. ResultsWe optimized the plug preparation procedure, focused on the cell suspension concentration (turbidity of 2.5 to 3.5), SDS addition (no SDS needed) and lysis time (1 h), and selected the appropriate restriction enzyme (XbaI) and the electrophoretic parameters (1.0 s-20.0 s for 19 h) of PFGE. There was nearly no discriminatory power of MLVA between Citrobacter strains. For 51 Citrobacter strains, automated ribotyping gave a D-value of 0.9945, while PFGE gave a D-value of 0.9969. Both PFGE and automated ribotyping clustered strains from the same sources (with the same species from the same place at the same time identified as the same source) and divided strains from different sources (from different years, places and hosts) into different subtypes. ConclusionPFGE protocol established in this paper and automated ribotyping are suitable for application in Citrobacter subtyping.

Selective Deposition of TiO&lt;sub&gt;2 &lt;/sub&gt;during Monolayer Formation of TiO&lt;sub&gt;2&lt;/sub&gt; and Iron Oxide Nanocrystals by Electrophoretic Deposition in Non-Polar Solvents

The electrophoretic deposition of TiO2 iron oxide nanocrystal monolayers from stable, mixed colloidal suspensions by electrophoretic deposition using non polar solvents is reported. The selective deposition of TiO2 was accomplished by controlling the mobility of the nanocrystals and the voltage during electrophoretic deposition. The effect of the electrophoretic deposition processing parameters (voltage and particle mobility) on the promotion and suppression of nanocrystal deposition was investigated.