A novel capillary electrophoresis method with pressure assisted field amplified sample injection in determination of thiol collectors in flotation process waters.

A new capillary electrophoresis method was developed for the quantification of diisobutyldithiophosphate (DTP), diisobutyldithiophosphinate (DTPI) and ethyl and isobutyl xanthates (EX, IBX) all of which are used as thiol collectors in froth flotation. This method uses pressure assisted field amplified sample injection (PA-FASI) to concentrate the analytes at the capillary inlet. The background electrolyte in electrophoretic separation was 60millimolar (mM) from 3-(cyclohexylamino)propane-1-sulfonic acid (CAPS) in 40mM NaOH solution. The similar CAPS electrolyte solution has earlier been used for screening for diuretics that contained sulphonamide and/or carboxylic groups. In this study, the functional groups are xanthate, phosphate and phosphinate. The method was developed using actual flotation process waters. The results showed that the water delivered from the plant did not contain significant amount of collectors; therefore, method development was accomplished by spiking analytes in these waters. Separation of analytes was achieved in 15min. The range of quantification was 0.27-66.6mg/L (R(2) 0.9991-0.9999) for all analytes other than ethyl xanthate, for which the range was 0.09-66.6mg/L (R(2) 0.9999). LOD (S/N=3) and LOQ (S/N=10) values for DTP, DTPI, IBX and EX were 0.05, 0.07, 0.06 and 0.01mg/L and 0.16, 0.25, 0.21 and 0.04mg/L, respectively. No interference from the matrices was observed, when the method was tested at a gold concentrator plant.

High-fluoride drinking water. A health problem in the Ethiopian Rift Valley 1. Assessment of lateritic soils as defluoridating agents.

PURPOSE: High-fluoride drinking water represents a health hazard to millions of people, not least in the East African Rift Valley. The aim of the present project was to establish a simple method for removing excessive fluoride from water. MATERIAL AND METHODS: Based on geological maps and previous experience, 22 soil samples were selected in mountainous areas in central Ethiopia. Two experiments were performed: 1. After sieving and drying, two portions of 50 g were prepared from each soil and subsequently mixed with solutions of NaF (500 mL). Aliquots (5 mL) of the solutions were taken at pre-set intervals of 1 hour to 30 days for fluoride analysis--using an F-selective electrode. 2. After the termination of the 30-days test, liquids were decanted and the two soil samples that had most effectively removed fluoride from the NaF solutions were dried, and subsequently exposed to 500 mL aqua destillata. The possible F-release into the distilled water was assessed regularly. RESULTS: Great variations in fluoride binding patterns were observed in the different soils. The percent change in F-concentration in the solutions, as compared to the original absolute value(F-), varied: at 1 hour from a decrease of 58% to an actual increase of 7.7%, while--at 30 days--all soil samples had caused a decrease in the F-concentration, varying from 0.5% to 98.5%. Only minute amounts of fluoride would leach from the fluoride-enriched soils. CONCLUSION: Lateritic soils may remove excessive fluoride from drinking water. Methods for practical application of this principle should be tested at household level.

Optimization of capillary electrophoretic-electrospray ionization-mass spectrometric analysis of catecholamines.

The capillary electrophoretic-mass spectrometric analysis (CE-MS) of catecholamines was optimized with coaxial sheath flow interface and electrospray ionization (ESI). The parameters studied included the sheath liquid composition and its flow rate, separation conditions in ammonium acetate buffer together with the ESI and cone voltages as mass spectrometric parameters. In addition, the effect of ESI voltage on injection as well as the siphoning effect were considered. The optimized conditions were a sheath liquid composition of methanol-water (80:20 v/v) with 0.5% acetic acid, with a flow rate of 6 microL/min. The capillary electrophoretic separation parameters were optimized with 50 mM ammonium acetate buffer, pH 4.0, to +25 kV separation voltage together with a pressure of 0.1 psi. The most intensive signals were obtained with an ESI voltage of +4.0 kV and a cone voltage of +20 V. The nonactive ESI voltage during injection as well as avoidance of the siphoning effect increased the sensitivity of the MS detection considerably. The use of ammonium hydroxide as the CE capillary conditioning solution instead of sodium hydroxide did not affect the CE-MS performance, but allowed the conditioning of the capillary between analyses to be performed in the MS without contaminating the ion source.

Determination of soluble anions and cations from waters of pulp and paper mills with on-line coupled capillary electrophoresis.

When the degree of closure of the paper machine wet end waters increases, wet end problems also become more difficult to control without specific and selective on-line measurements. The need to measure the concentrations of individual compounds in order to explain wet end phenomena is growing. This study was performed to set up a CE system to a paper machine and to determine soluble inorganic and organic ions in different locations of pulp and paper process waters with real time analyses by two on-line CE methods. A reconstructed commercial CE apparatus was connected to a papermaking machine via an apparatus, which was a combined sampling and sample pretreatment instrument, the role of which was to filter and dilute the samples before on-line determination by CE. The on-line procedures were optimized for simultaneous determination of anions as chloride, sulfate, oxalate, formate and acetate and for determination of cations as potassium, calcium, sodium, magnesium and traces of aluminium. The quantification was performed with external standard methods using the programs available in the commercial CE instrument. The concentrations of the ions were transferred by using a computerized transfer algorithm exporting the results from the analysis instrument to the process control unit. The developed on-line procedures were tested three times in paper and paperboard mills for 1 month at a time. Correlations were observed between the CE results and changes in the processes.

Determination of urinary catecholamines with capillary electrophoresis after solid-phase extraction.

The stabilities of 3,4-dihydroxybenzylamine (DHBA), dopamine, 3-methoxytyramine, normetanephrine and metanephrine standards under acid, base and enzymatic hydrolysis conditions were studied. Basic incubation media were not suitable for 3,4-dihydroxy compounds, but acid and enzymatic hydrolysis conditions were applicable to all the compounds. The results of acid and enzymatic hydrolysis were comparable and the enzymatic hydrolysis was applied to a urine matrix. A method including solid-phase extraction (SPE) with a copolymer sorbent was developed for purification of the urine samples. Due to poor recovery of DHBA, the most frequently used internal standard in catecholamine analysis, this compound was replaced with the 3-O-methoxy structure. The recoveries of the compounds in spiked urine samples in SPE were between 96.4 and 124.4%. The repeatability of the combination of enzymatic hydrolysis and SPE pretreatment was good for all the compounds, except for dopamine and 3-methoxytyramine due to some matrix compounds still interfering with the separation. The analyses were performed with capillary electrophoresis in an ammonium acetate buffer with UV detection. The validation data for the compounds including limit of detection, limit of quantification, linearity and repeatability of the method are presented.

Extraction of iridoid glycosides and their determination by micellar electrokinetic capillary chromatography.

Several methods for the extraction of two iridoid glycosides, catalpol and aucubin, from the plant matrix (Veronica longifolia leaves) were compared. Pressurized hot water extraction and hot water extraction were the most efficient isolation techniques for both. Pressurized liquid extraction and maceration with various organic solvents were also tested. Relative to the amounts extracted with hot water, ethanol extracted only 22% of catalpol and 25% of aucubin and pressurized hot water extracted 83% of catalpol and 92% of aucubin. The lowest relative standard deviations, 22% for catalpol and 8% for aucubin, were achieved with hot water extraction (13 repetitions), and the highest relative standard deviations, 76% for catalpol and 73% for aucubin, with pressurized liquid extraction (five repetitions). A fast capillary electrophoretic method was developed for the quantitative determination of catalpol and aucubin.

Quantification of anions and cations in environmental water samples. Measurements with capillary electrophoresis and indirect-UV detection.

The aim of this study was to validate two separation methods for determination of inorganic anions and cations from natural waters with capillary electrophoresis (CE) by using indirect-UV detection. The research is related to method development for screening of groundwater samples obtained in site investigations for spent fuel of the Finnish nuclear industry. In CE analysis, anions were separated in pyromellitic acid (pH 7.7) in the order bromide, chloride, sulphate, nitrite, nitrate, fluoride and dihydrogenphosphate. Cations were separated at pH 3.6 after anions using an 18-crown-6-ether solution. In these analyses, ammonium migrated first followed by potassium, calcium, sodium and magnesium. The concentrations of the ions in the natural water samples were calculated by using two or three calibration curves made using reference solutions at concentration levels of 0.5-250 mg/l. The repeatabilities of the anion and cation methods were tested using laboratory-made reference sample mixtures with high and low salt concentrations. The limits of quantification in the analyses were between 0.02 and 0.1 mg/l, depending on the ion. Concentrations of ions tested in natural waters varied from a few milligrams to tens of grams per litre.

Study of catecholamines in patient urine samples by capillary electrophoresis.

Capillary zone electrophoresis with photodiode array detection at 220 nm was used for analysis of catechol compounds in human urine. The method was optimized with reference compounds 3,4-dihydroxybenzylamine, adrenaline, noradrenaline, normetanephrine, dopamine, dopac (homogensitic acid), methanephrine, vanillyl-mandelic acid, 5-hydroxyindoleacetic acid (5-HIAA), homovanillic acid and 3-methoxytyramic acid at pH 4.0 and 8.0 for their electrophoretic separation. The UV spectra of the catechols were detected at a concentration of 20 microM. Repeatability of the method calculated using the absolute migration times of the catechols was below 1.5% and using the peak areas below 5%. The patient samples were hydrolyzed by 0.5 M acid or base solutions. In the studies, a few patient samples were analyzed using 3,4-dihydroxybenzylamine as an internal standard. In the hydrolysis steps needed for their detection in urine, all the other catecholamines, except 5-HIAA, did not decompose to detectable species at 220 or 254 nm. The concentrations of the catecholamines observed in real samples were at nM levels.

Validated method for the quantitation of quercetin from human plasma using high-performance liquid chromatography with electrochemical detection.

A validated method for the quantitation of trace levels of quercetin from human plasma to be used in pharmacokinetic and biomarker studies is presented. Quercetin conjugates were hydrolysed enzymatically, plasma proteins were removed using a Bond Elut C18 extraction column and additional interferences were removed by extracting them into a toluene-dichloromethane mixture. The HPLC system consisted of an Inertsil ODS-3 column (250 x 4.0 mm) and a mobile phase with 59% methanol in phosphate buffer (pH 2.4). High selectivity and a low quantitation limit (0.63 microg/l) were achieved by using electrochemical detection at a low potential. The method has excellent reproducibility: R.S.D. values of peak-heights were 2% and 7.9%, respectively, for within-day and between-day precision. The method was applied to a small scale study of quercetin pharmacokinetics and quercetin was shown to be absorbed from a 20 mg dose. No free quercetin was detected in plasma and no evidence of significant amounts of quercetin glycosides in plasma was found.

Determination of association constants of dansyl-amino acids and beta-cyclodextrin in N-methylformamide by capillary electrophoresis.

The use of nonaqueous background electrolytes in capillary electrophoresis (CE) is a promising new trend which should widen the scope of this technique. We demonstrate the chiral separation of dansyl-amino acids (Dns-AAs) in N-methylformamide (NMF) using beta-cyclodextrin (beta-CD) as chiral selector. The solubility of beta-CD is much better in NMF than in water, allowing high concentration of the chiral selector and successful enantioseparation despite the weak host-guest interaction between the Dns-AAs and beta-CD. The association constants for the complexation between Dns-AAs and beta-CD could be calculated from the electrophoretic mobilities, with attention paid to the change in viscosity of the electrolyte upon addition of beta-CD. The association constants ranged between 2 and 13 M(-1).

Characterization of flavonoids in extracts from four species of Epimedium by micellar electrokinetic capillary chromatography with diode-array detection.

A micellar electrokinetic capillary chromatographic (MEKC) method with diode-array detection is developed for the characterization of pharmacologically active flavonoids in extracts prepared from Epimedium brevicornum, E. humanense, E. coactum, and E. truncatum. The pK(a) values of icarin, epimedin B, and epimedin C are determined by spectrophotometry. Optimal separation of icarin, epimedin B and C, and eight other compounds is achieved by determining pK(a) values and by systematically optimizing electrolytic and instrumental parameters. The repeatability of analyses and the reliability of identification are evaluated by the marker technique. Calculated for relative migration times of flavonoids in the extracts, the repeatability of the analyses varies from 0.7 to 6.4% (nine replicates). For migration indices calculated with two markers, however, the repeatability almost falls below 0.5%. The distribution of the flavonoids is found to differ both qualitatively and quantitatively among the four species. The MEKC technique appears to provide a powerful tool for the identification and quality control of plant drugs and for phytotaxonomic investigations.

Selectivity in capillary electrophoresis in the presence of micelles, chiral selectors and non-aqueous media.

In addition to high efficiency, short analysis times and small sample volumes, a further attractive feature of capillary electrophoretic techniques is the possibility to achieve, high selectivities. Usually, selectivity control also allows improvement in the resolution. A simple way to enhance the selectivity of capillary electrophoretic separations is to add one or more surfactants above their critical micelle concentration, or in the case of chiral separations to add a chiral selector to the background electrolyte. Because of the dynamic structure of micelles, the aggregation of monomers and size of the micelles can be easily adjusted. This review describes the various type of surfactants used in micellar electrokinetic capillary chromatography, and the chiral selectors employed in enantiomeric separations by capillary electrophoresis. Factors affecting the selectivity are noted. A brief discussion is included of the selectivity enhancement obtainable in non-aqueous media.

Determination of serum corticosteroids by mixed micellar electrokinetic capillary chromatography with sodium dodecyl sulfate and sodium cholate.

The application of mixed micellar electrokinetic capillary electrochromatography to the determination of the corticosteroid hormones cortisone, cortisol (hydrocortisone), and dexamethasone in serum samples is demonstrated. The serum samples were enzymatically hydrolyzed, precipitated, solid-phase extracted, and analyzed by capillary electrophoresis. A micellar system of sodium dodecyl sulfate and sodium cholate buffered with an organic compound provided the electrolyte solution. Spiked blank serum samples were used for the linearity testing and limits of detection and quantitation were measured. Patient samples were analyzed and the concentrations of cortisol and cortisone were measured. The method, which was fast and easy to perform, was confirmed to be useful for the determination of corticosteroids in serum.

Determination of morphine analogues, caffeine and amphetamine in biological fluids by capillary electrophoresis with the marker technique.

A reliable method was sought for the fast screening and simultaneous determination of amphetamine, morphine, heroin (acetomorphine), codeine (methylmorphine) and caffeine in biological fluids and drug seizures. Capillary zone electrophoresis (CZE) and micellar electrokinetic capillary chromatography (MEKC), with detection at 200 and 220 nm, were investigated for analytes in human serum and urine. When adequate separation was not achieved in preliminary studies with CZE, further development was focused on the MEKC method. Glycine buffer containing sodium lauryl sulfate (pH 10.5) was used for the MEKC separations. The analytes and carboxylic acids used as marker compounds could be screened by a short-capillary method in less than 2 min. In the simultaneous determination of the drugs in urine and serum a longer separation of 18 min was preferred so that all the compounds, the markers and the endogenous compounds absorbing at the detection wavelength could be adequately separated in a single run. The migration times of the compounds increased in the order caffeine, morphine, heroin, codeine and amphetamine. The repeatability of the separation was tested by using two carboxylic acids as marker compounds in the determination of the migration indices of the analytes. The relative standard deviations for the migration indices were less than 1%, which is accurate enough for the determination of the drugs in biological fluids.

Structure-retention relationships of steroid hormones in reversed-phase liquid chromatography and micellar electrokinetic capillary chromatography.

Quantitative structure-retention relationship (QSRR) studies are useful in retention prediction, finding the relevant structural descriptors for analytes and estimating the relative biological activities of a series of analytes. Most of the studies have been conducted by RP-HPLC and a few by micellar electrokinetic capillary chromatography (MECC). The aim of this work was to find structural parameters and characteristics related to the RP retention and electrophoretic migration of steroids in order to predict the retention/migration of steroid hormones on the basis of their molecular structure. Retention data were obtained with an ODS column using a mobile phases aqueous methanol-acetonitrile (mobile phase A) and methanol-tetrahydrofuran (mobile phase B). MECC was conducted with a sodium dodecyl sulfate (SDS)-borate system and with a mixed micellar solution of SDS and sodium cholate. Several topological indices, such as the connectivity indices, chi, were used as structural parameters. Steric factors seem to have a great effect on the retention of steroid hormones, especially with the MeCN-containing mobile phase. Retention in mobile phase B could be predicted more accurately. Topological indices can be used in the modelling and prediction of the retention/migration of steroid hormones when the solutes form a congeneric series and stereochemical properties do not govern the separation process.

Ion-pair chromatography and micellar electrokinetic capillary chromatography in analyzing beta-adrenergic blocking agents from human biological fluids.

Ion-pair chromatography (IPC) and micellar electrokinetic capillary chromatography (MECC) were used for the separation and determination of parent beta-blockers from human biological fluids. In both these techniques, N-cetyl-N,N,N-trimethylammonium bromide (CTAB) was used as a buffer additive. In IPC, CTAB was an ion-pair former, and in MECC it was a micelle-forming surfactant. The effectiveness of the IPC method using methanol-gradient elution and that of MECC were compared for drug-spiked serum and urine samples. Detection was performed with a diode-array detector in the IPC method and with a 214-nm filter in the MECC technique. In both methods a phosphate buffer (pH 7.0) was used. In MECC the buffer solution contained 10 mM CTAB, while in IPC the CTAB concentration was decreased from 7 to 4 mM during the separation when a methanol gradient was used. The study showed that the IPC technique performed better for bioanalyses than the high-performance MECC technique, since in MECC UV detection presented a problem because of the low sample concentration. However, in MECC sample preparation was less time-consuming, using hydrolyzation and protein precipitation and, unlike the IPC technique, it did not require any liquid-liquid extraction step.

Screening and determination of beta-blockers, narcotic analgesics and stimulants in urine by high-performance liquid chromatography with column switching.

A fast method is described for the screening of eleven beta-blockers, two narcotic analgesics and two stimulants in urine by HPLC with column switching. The urine sample (100 microliters), buffered to pH 9-9.5, is injected onto a short extraction column packed with CN stationary phase. The extraction column is flushed with water for 2.5 min to elute polar matrix components to waste. The retained components are then backflushed by means of a six-port valve onto the ODS analytical column where they are separated. Phosphate buffer pH 3.0 and acetonitrile were used as mobile phase. Gradient elution was applied in the screening method to improve separation. Detection was performed with a diode-array detector at 220, 235 and 300 nm. Recoveries were near 100%, precision was excellent and sensitivity about 0.25 micrograms/l. To speed up the quantitative analysis, the same method but with isocratic elution was successfully applied to the determination of acebutolol and metoprolol in urine samples collected 4 h after administration of the compounds as single doses.

Optimized separation of seven corticosteroids by micellar electrokinetic chromatography.

The corticosteroids studied can be effectively separated by employing micellar electrokinetic capillary chromatography (MECC). The effect of pH, borate concentration and the sodium dodecyl sulfate (SDS) concentration on both the resolution and the selectivity was studied under 15 different experimental conditions. The experimental design was similar to the central composite design approach. Empirical quadratic regression models were derived for analyte migration time, band broadening and analyte velocity. Satisfactory regression fits and coefficients of determination for prediction were obtained with cross-validated models. The models for analyte migration time and analyte velocity were in good agreement with theory. Modeling of the band broadening seemed to be somewhat more complicated. Optimum conditions for resolution and selectivity were different. This is due to the fact that selectivity studies ignore the electroosmotic and band broadening properties of different electrolyte solutions. However, the study of the selectivity yielded good information about the suitability of the electrolyte systems for the particular separation problem. Although a high solubilizing power of SDS caused the corticosteroids to partition strongly into the SDS micelles, a good separation could be achieved at low SDS concentrations.

Introduction of migration indices for identification: chiral separation of some beta-blockers by using cyclodextrins in micellar electrokinetic capillary chromatography.

Because of the different physiological impact that stereoisomers may have, it is often vital to separate these forms from one another. Because of their structural similarity, the separation is usually difficult to achieve and zones may elute very close to each other. This is a particular problem in capillary electrophoresis, where the repeatability of absolute migration times is fairly poor, mainly due to the irreproducibility of the electroosmotic flow. The separation is usually repeatable, however, and when the disturbing effects are eliminated by using a migration index system incorporating two marker compounds the identification of the enantiomers becomes extremely good. Relative standard deviation (RSD) values less than 0.1% for the migration index of each enantiomer were obtained in both intra-day and day-to-day (6 days) studies. The best separation was achieved with the electrolyte solution made of 40 mM borate, 32 mM sodium dodecyl sulfate (SDS), 12 mM beta-cyclodextrin (beta-CD), and 6 mM alpha-cyclodextrin (alpha-CD) at pH 9.3.

Screening for diuretics in urine and blood serum by capillary zone electrophoresis.

Diuretics are therapeutic agents used to promote the excretion of bodily fluids and salts. They are also misused by some athletes to decrease body mass or to mask the use of anabolic steroids and other drugs. We have developed a method that screens for diuretics in urine and blood serum. Two successive runs were required because of the heterogeneity of this group of compounds. Screening for diuretics that contained sulphonamide and/or carboxylic groups was done at pH 10.6 with 3-(cyclohexylamino)-1-propane-sulphonic acid (0.06 M) as buffer. Diuretics that contained primary, secondary or tertiary amine groups were investigated at pH 4.5 with an acetate (0.07 M)-betaine (0.5 M) buffer system. Hydrostatic injection mode for 5 s gave the best efficiency. Longer injection times were acceptable but efficiency was then somewhat reduced. Detection limits at the low femtomole level are achievable for most compounds with a UV-Vis detector operating at 220 and 215 nm. Temperature affected the separation, and 20 degrees C proved best. All compounds were separated in less than 30 mins. A confirmation analysis of all compounds was done by GC-MS.