Endothelial progenitor cells in patients with severe peripheral arterial disease.

The aims of this study were to investigate the interrelationships between endothelial progenitor cells (EPCs), peripheral arterial disease (PAD), and atherosclerotic risk factors, as only limited data are available regarding the EPCs in patients with PAD. The authors studied the number of EPCs by different methods in a carefully selected group of 45 patients with PAD along with 24 healthy subjects (HS). In patients with PAD, by utilizing the dual-binding method, the number of EPCs was significantly increased compared to HS (M +/- SD, PAD: 73 +/- 33, HS: 52 +/- 20 EPCs/high power field; p < .001). On the contrary, both CD34(+) cell count and CD133(+) cell count were significantly decreased compared to HS. Colony-forming units were significantly increased in PAD compared to HS (median and 25th and 75th percentiles, PAD: 7, 1, 9; HS: 1, 1, 4 CFU/well, respectively; Mann-Whitney, p = .006). In patients with PAD, the number and proliferative activity of circulating EPCs are increased with respect to HS even though EPC count by flourecence-activated cell sorting (FACS) analysis provided different results and this may explain the discrepancy in data collected using different methods. The regulation of the number and biological activity of EPCs in PAD remains unclear.

Carbohydrate-deficient transferrin (CDT): a reliable indicator of the risk of driving under the influence of alcohol when determined by capillary electrophoresis.

Carbohydrate-deficient transferrin (CDT) is a marker of chronic alcohol abuse, which has recently been introduced to evaluate the physical fitness for obtaining a driving license. The aim of the present study was to evaluate the prevalence of elevated CDT levels in subjects stopped while driving under the influence of alcohol by using a validated method based on capillary electrophoresis. The study was carried out on a group of 40 drunken drivers (group A) and on a control group (n=51) of subjects chosen from the general population (group B). CDT was directly determined by capillary electrophoresis in free solution and UV detection at 200 nm. CDT results from both groups were classified as "negative" or "positive" on the basis of the cut-off set at 2.00% (CDT index). The subjects classified as "positive" in group A were 24 (60%), whereas in group B were 2. The subjects classified as "negative" in group A were 16 (40%), whereas in group B was 49 (96.1%). The comparison of the observed percentages, evaluated with the chi(2)-test, was highly significant (p<0.001). The present study confirms the high prevalence of chronic alcohol abusers among drunken drivers and the usefulness of CDT as a predictor of the risk of drunk driving.

Direct analysis of bromide in human serum by capillary electrophoresis.

The purpose of the present work was the development and validation of a simple, rapid and reliable method for direct bromide quantification in serum based on capillary electrophoresis (CE). The analysis was carried out with an automated capillary electropherograph. Analytical conditions were as follows. Capillary: uncoated fused silica, effective length 50 cm, internal diameter 50 microm; voltage: 20 kV in reverse polarity mode; temperature: 25 degrees C; running buffer: 90 mmol/L sodium tetraborate decahydrate and 10 mmol/L NaCl, pH 9.24; detection: direct UV absorption at 200 nm; sample treatment: dilution of serum 1:10 with the internal standard solution (2 mmol/L thiocyanate). Under the described conditions, bromide ions and internal standard were baseline separated in 7 min. No interferences from other serum components were observed. The analytical sensitivity was characterized by a LOD: 0.05 mmol/L and a LOQ of 0.1 mmol/L. Excellent linearity was verified in the range from 2.5 to 60 mmol/L [y = 0.0746x - 0.0372; R2 = 0.9995 (x = bromide concentration; y = bromide peak area/internal standard (I.S.) peak area)]. Quantitative imprecision in intra-day (n = 7) and day-to-day (n = 7) experiments was always within R.S.D. values <2%. Recovery was quantitative throughout the range of linearity of the method. Clinical cases of infants undergoing potassium bromide therapy for refractory epilepsy were analyzed with results in agreement with literature data. On the basis of these considerations, capillary electrophoresis can be proposed as the method of choice for bromide analysis in serum samples, especially for therapeutic drug monitoring purposes.

Rapid and direct analysis of gamma-hydroxybutyric acid in urine by capillary electrophoresis-electrospray ionization ion-trap mass spectrometry.

The present work was aimed at the development of a capillary electrophoretic analysis of gamma-hydroxybutyric acid (GHB) using electrospray ion trap mass spectrometry to achieve the direct and unequivocal detection of this analyte in human urine. Optimized capillary electrophoretic conditions were: injection, 20 s at 0.5 psi (1 psi = 6894.76 Pa); buffer electrolyte, 12.5 mM ammonium formate adjusted to pH 8.35 with diethylamine; fused silicacapillary: 100 cm x 50 microm i.d.; separation voltage, 25 kV (forward polarity) + 0.5 psi; room temperature. Electrospray and mass spectrometric conditions were: drying gas and nebulizing gas (nitrogen) at flow rate 3 l/min, temperature 250 degrees C, nebulizer pressure: 10 psi; sheath liquid solution: methanol-water (90:10) containing 0.1% ammonia delivered at 3 microl/min; spray voltage 3.5 kV. Mass spetrometric detection was carried out in the selected ion monitoring mode of negative molecular ions at 103 m/z for GHB and 115 m/z for maleic acid (I.S.). Under these conditions the baseline separation of GHB and the I.S. was obtained. The selectivity of the analysis allowed for direct injection of unextracted urine, previously diluted 1:4 with water. Linearity was assessed in the GHB concentration range from 80 to 1280 microg/ml in urine. Analytical sensitivity (as limit of detection) resulted about 5 microg/ml in water and 20 microg/ml in original urine. Analytical precision was fairly acceptable with R.S.D. values lower than 5% for migration times and 18% for quantitation in real samples, in both intra day and day-to-day experiments. On these grounds, the developed method can be adopted for rapid identification of acute intoxications from GHB in humans.

Analysis of organic components of smokeless gunpowders: high-performance liquid chromatogaphy vs. micellar electrokinetic capillary chromatography.

The aim of the present study was to verify the analytical performances of high-performance liquid chromatography (HPLC) and micellar electrokinetic capillary chromatography (MEKC) for the separation and qualitative determination of a selected group of organic components of smokeless gunpowders. The HPLC method was based on a gradient reversed-phase elution with a mobile phase composed of 0.17 M H(3)PO(4)/methanol; detection was performed by UV absorption at the wavelengths of 220, 254, and 270 nm. The MEKC experiments were carried out by using uncoated fused-silica capillaries (50 microm inside diameter, 50 cm effective length) and a running buffer composed of 10 mM sodium tetraborate at pH 9.24 added with 25 mM sodium dodecyl sulfate (SDS); the applied voltage was 25 kV; detection was either at a fixed wavelength UV of 214 nm or with a diode-array detector operating in the wavelength range from 190 to 350 nm. Both reversed-phase HPLC and MEKC techniques succeeded in resolving the tested standard mixtures of organic components of smokeless powders. Although the sequence of elution of the different analytes was slightly different between HPLC and MEKC, a statistical analysis based on the Spearman's rank correlation test showed that the two separation patterns were highly correlated. HPLC and MEKC were comparable in terms of elution/migration time precision, whereas MEKC showed higher reproducibility of peak areas. The interfacing of capillary electrophoresis with diode array UV detection provided distinct UV spectra of the individual analytes, thus improving, on the detection side, the analytical selectivity and identification power of capillary electrophoresis.

Determination of carbohydrate deficient transferrin (CDT) with capillary electrophoresis: an inter laboratory comparison.

Carbohydrate deficient transferrin (CDT) is currently the most specific laboratory marker of chronic or sustained alcohol abuse. CDT is increasingly being used as a diagnostic tool in the areas employment, traffic safety and forensic medicine. In recent times, capillary electrophoresis (CE) has been proposed as a convenient tool for rapid, precise and accurate CDT determination, not only for research but also for routine analyses. Quite recently, commercial kits have been introduced which, reportedly, could simplify and standardize CDT analysis with capillary electrophoresis. The present work was aimed at testing the ruggedness of a capillary electrophoretic method based on a commercial kit (CEofix, Analis), by comparing the results obtained with different instruments in different laboratories, on a panel of sera randomly collected and exchanged. The results showed, notwithstanding few outliers, excellent correlation of the results obtained in the two laboratories (R=0.974). Also high concordance was found when results were classified as positive or negative on the basis of a cut-off (1.25%) established from a control group of teetotalers. In conclusion the present data support the usefulness of capillary electrophoresis for CDT determination for clinical, forensic and administrative diagnosis of chronic alcohol abuse.