Effects of organic modifier and temperature on the enantiomeric separation of several azole drugs using supercritical fluid chromatography and the Chiralpak AD column.

The effects of organic modifier and temperature on the enantioseparation of 10 triazoles and eight imidazoles, using supercritical fluid chromatography with the Chiralpak AD column, have been investigated in this work. For this purpose four different organic modifiers (methanol, ethanol, 2-propanol and acetonitrile) were evaluated. Only in the case of two compounds could the enantiomeric separation not be achieved with any of the modifiers tested; the rest of compounds were baseline or partially resolved with at least one of the modifiers. The alcohol-type modifiers provided the best results in terms of retention time and resolution. In general, retention increased in the order methanol < ethanol < 2-propanol; moreover it was possible to establish a relationship between the retention and the number of aromatic rings and dioxolane groups in the molecule, that is, the higher the number is, the higher the retention time. From the study of the temperature effect, the enthalpy-entropy compensation was demonstrated for all the compounds, except for bifonazole using methanol and miconazole using acetonitrile. This suggested that both analytes are enantiomerically resolved through different mechanisms.

Study of the enantiomeric separation of an acetamide intermediate by using supercritical fluid chromatography and several polysaccharide based chiral stationary phases.

Four chiral stationary phases, based on the phenylcarbamate derivatives of amylose or cellulose: Chiralcel OD-H, Chiralpak AD, Lux Cellulose-2 and Lux Amylose-2, were evaluated for the enantiomeric separation of an acetamide chiral intermediate, the (4S-trans)-4-(ethylamino)-4-(N-acetamide)-5,6-dihydro-(6S)-methyl-4H-thieno-[2,3-b]thiopyran-7,7-dioxide, using SFC. The effect of the different modifiers and temperatures, on the separation, was also studied. The chiral separation could not be achieved using the Chiralpak AD column, nevertheless the other columns provided excellent results with analysis times close to 6 min and resolutions higher than 2. The highest enantioresolutions and retentions were obtained with the Lux Cellulose-2 column and 2-propanol as organic modifier. The isoelution temperatures were estimated from the van't Hoff plots, and in all the cases they were above the temperature range studied which means that the enantiomeric separation was enthalpy driven.

Overview of pesticide residues in stored pollen and their potential effect on bee colony (Apis mellifera) losses in Spain.

In the last decade, an increase in honey bee (Apis mellifera L.) colony losses has been reported in several countries. The causes of this decline are still not clear. This study was set out to evaluate the pesticide residues in stored pollen from honey bee colonies and their possible impact on honey bee losses in Spain. In total, 1,021 professional apiaries were randomly selected. All pollen samples were subjected to multiresidue analysis by gas chromatography-mass spectrometry (MS) and liquid chromatography-MS; moreover, specific methods were applied for neonicotinoids and fipronil. A palynological analysis also was carried out to confirm the type of foraging crop. Pesticide residues were detected in 42% of samples collected in spring, and only in 31% of samples collected in autumn. Fluvalinate and chlorfenvinphos were the most frequently detected pesticides in the analyzed samples. Fipronil was detected in 3.7% of all the spring samples but never in autumn samples, and neonicotinoid residues were not detected. More than 47.8% of stored pollen samples belonged to wild vegetation, and sunflower (Heliantus spp.) pollen was only detected in 10.4% of the samples. A direct relation between pesticide residues found in stored pollen samples and colony losses was not evident accordingly to the obtained results. Further studies are necessary to determine the possible role of the most frequent and abundant pesticides (such as acaricides) and the synergism among them and with other pathogens more prevalent in Spain.

Comparative study of sample preparation procedures to determine fipronil in pollen by gas chromatography with mass spectrometric and electron-capture detection.

Several sample preparation methods have been assayed to analyze residues of fipronil in pollen at trace concentrations. Extraction with organic solvents, solid-phase extraction on either commercial cartridges or a Florisil-packed column and a matrix solid-phase dispersion, also with Florisil as a dispersing agent, have been tested. Determination of fipronil in the extracts has been carried out by GC with electron-capture and mass spectrometric detection. An extraction with acetonitrile followed by a clean-up on ODS or polymeric cartridges was the most suitable procedure to obtain acceptable recoveries and relatively simple chromatograms. The matrix-effects observed in the quantification can be corrected with a matrix-matched calibration.

Semipreparative enantiomeric separation of omeprazole by supercritical fluid chromatography.

Omeprazole, a widely used antiulcer drug, has been enantiomerically separated at semipreparative scale on a polysaccharide based chiral stationary phase by supercritical fluid chromatography (SFC). For this work, a modular supercritical fluid chromatograph was adapted to operate at semipreparative scale and a Chiralpak AD (250 mm x 10 mm) column was used. The effect of two organic modifiers (ethanol and 2-propanol) was studied, and different injection volumes and concentrations of the omeprazole racemic mixture were evaluated in order to obtain high enantiomeric purities and production rates. Better results were achieved using concentration overloading instead of volume overloading. The recoveries decreased when the requirements of enantiomeric purity or the load increased, but it was possible to recover 100% of both enantiomers at an enantiomeric purity higher than 99.9% under some loading conditions, like injecting 1 and 2 ml of a solution of 3g/l. As far as production rates are concerned, the best result for S-(-)-omeprazole at that purity (27.2mg/h) was achieved with sample concentrations of 10 g/l and the injection of 2 ml, while a volume of 4 ml was better in the case of R-(+)-omeprazole (20.5mg/h).

Study of the enantiomeric separation of oxfendazole and cetirizine using subcritical fluid chromatography on an amylose-based column.

The enantiomeric separation of cetirizine and oxfendazole on a Chiralpak AD column using subcritical fluid chromatography has been studied in this work. The enantioseparation of cetirizine was only possible when 2-propanol was used as a modifier, obtaining better results in presence of the additives triethylamine (TEA) and trifluoroacetic acid (TFAA). On the contrary, 2-propanol provided the lowest enantioresolutions for oxfendazole, in this case the best results in terms of high resolution and short analysis time were obtained with ethanol. The study of the temperature effect revealed that in the case of cetirizine using 2-propanol, and oxfendazole using methanol, the separation was enthalpy-driven and the isoelution temperature was above the working range. Using ethanol or 2-propanol, the results showed that the oxfendazole enantioseparation was entropically driven and the isoelution temperatures were below the range studied.

[Cholesterol, alpha-tocopherol, and retinoid concentrations in silicone oil used as a vitreous substitute]. / concentraciones de colesterol, alfa-tocoferol y retinoides en aceite de silicona tras su utilización como sustitutivo vítreo.

OBJECTIVE: To verify the existence of organic lipophylic compounds in silicone oil extracted from human eyes following its use for previous retinal detachment, and to determine the intraocular permanence time of these substances in the oil. METHODS: Concentrations of retinoic acid, retinol, retinal, cholesterol and alpha-tocopherol were detected by HPLC in 23 samples of silicone oil extracted from patients with complicated retinal detachments. The time interval between the time of injection of the silicone oil and the subsequent assessment varied from 3 to 50 months (the permanence time). RESULTS: All tested compounds were found in the samples, but these were most commonly cholesterol and less frequently alpha-tocopherol. There was an inverse relationship between retinoic acid concentration and age (p=0.023), and a direct relationship between cholesterol concentration and permanence time (p=0.0008) at least up to 20 months. CONCLUSIONS: These findings confirm that silicone oil is not an inert substance but is capable of extracting lipophylic compounds from the intraocular tissues. There is a clear linear elevation of cholesterol levels with increased intraocular permanence time. This finding could be used to further establish a safe permanence time for intraocular silicone oil used in ophthalmologic surgery. More studies with larger samples are warranted to evaluate this further.

Comparative study of the enantioselective separation of several antiulcer drugs by high-performance liquid chromatography and supercritical fluid chromatography.

A comparative study of the enantiomeric separation of several antiulcer drugs such as omeprazole, lansoprazole, rabeprazole and pantoprazole using HPLC and supercritical fluid chromatography (SFC) on the Chrialpak AD column is presented in this work. The results show that employing the above mentioned column only two compounds (omeprazole and pantoprazole) could be enantiomerically resolved using HPLC, on the contrary SFC allowed the enantiomeric separation of all the compounds studied with higher resolutions and lower analysis times.

Chiral separation of some triazole pesticides by supercritical fluid chromatography.

The enantiomeric separation of six triazole pesticides: cyproconazole, propiconazole, diniconazole, hexaconazole, tebuconazole and tetraconazole, using supercritical fluid chromatography and the Chiralpak AD column, is presented in this work. The effect of different organic modifiers such as methanol, ethanol and 2-propanol on the retention and resolution was studied as well as the presence of additives in the mobile phase. The results obtained were highly successful, all the compounds were enantiomerically separated and in most of the cases the analysis time was close to 10 min. The type of organic modifier that provided the best results depended on the compound.

Separation of triadimefon and triadimenol enantiomers and diastereoisomers by supercritical fluid chromatography.

The enantiomeric separation of triadimenol and triadimefon on a Chiralpak AD column using supercritical fluid chromatography, was studied in this work. The effect of different modifiers (methanol, ethanol and 2-propanol) was tested, with methanol and ethanol providing the best results for the enantiomeric separation of the two compounds. The enantioseparation of a mixture of triadimenol and triadimefon (six stereoisomers) was achieved in only 15 min using a gradient of ethanol, 200 bar, 35 degrees C and a flow-rate of 2 ml/min. The separation of triadimenol diastereoisomers on different achiral columns (diol, silica and ODS) was also investigated. In this case, the type of organic modifier to be used depended on the stationary phase, the Spherex Diol being the column that gave the best separation. Using this column, resolutions higher than 3 were obtained in analysis times of 5 min with any of the modifiers checked.

[Glutathione-related enzymes activity during vitrectomy. Effect of BSS Plus(R) in retinal tissue]. / Actividad de los enzimas glutation-dependientes durante la vitrectomía. Efecto de la solución BSS Plus(R) en retina.

PURPOSE: To determine changes in the retinal activity of two enzymes related to the glutathione metabolism (Glutathione synthetase -GSHS- and glutathione reductase -GSSGR-) after vitrectomy using BSS Plus(R). MATERIAL AND METHODS: Forty-five pigmented rabbits were distributed in 6 groups: group I (GSHS control in retinal tissue); Group II (analysis of GSHS at 3 hours post-vitrectomy); Group III (determination of GSHS at 48 hours after vitrectomy); Group IV (GSSGR control in retinal tissue); Group V (measurement of GSSGR at 3 hours post-surgery) and Group VI (study of GSSGR at 48 hours post-vitrectomy). Statistical analysis was done by a parametric test (ANOVA of single factor) (p<0.05). RESULTS: The activity of these enzymes was: Group I (n=5, basal GSHS). 2785,63 D.S. 419,51 U/g; Group II (n=4, GSHS at 3 hours). 6053,50 D.S. 2788,84 U/g; Group III (n=4, GSHS at 48 hours). 7424,30 D.S. 997,47 U/g; Group IV (n=10, basal GSSGR). 150,86 D.S. 24,40 mU/mL; Group V (n=7, GSSGR at 3 hours). 212,03 D.S. 53,30 mU/mL; Group VI (n=8, GSSGR at 48 hours). 210,84 D.S. 46,03 mU/mL. CONCLUSIONS: At retinal tissue, BSS Plus(R) intraocular irrigating solution increases GSHS activity without modifications of GSSGR levels. It seems to be related to the <> synthesis of glutathione.

Capillary gas chromatography with mass spectrometric and atomic emission detection for characterization and monitoring chlordimeform degradation in honey.

Capillary gas chromatography-mass spectrometry and capillary gas chromatography-atomic emission detection have been successfully used to identify and monitor the main degradation products of chlordimeform when this compound is initially present in honey. The analysis of laboratory-spiked honey samples over 28 weeks revealed the occurrence of two degradation products: 4-chloro-o-toluidine (I) and N-formyl-4-chloro-o-toluidine (III). During this period the concentration of chlordimeform decreased to 7.5% of its initial value; the concentration of compound I increased gradually whereas compound III was present in a larger proportion and reached a maximum around the 14th week.

Separation of antifungal chiral drugs by SFC and HPLC: a comparative study.

The enantiomeric separation of several compounds, including an antifungal drug and several of its precursors, using HPLC and SFC is described in this work. The columns employed were based on polysaccharide derivatives and the results show that most of the separations obtained by SFC are better, in terms of high resolution and short analysis time, than those obtained by HPLC. Only one compound could not be resolved using SFC but, in this case, HPLC provided baseline resolution.

Separation of albendazole sulfoxide enantiomers by chiral supercritical-fluid chromatography.

The enantioseparation of albendazole sulfoxide (ABZSO) by chiral supercritical-fluid chromatography (SFC) on two columns, based on the polysaccharide derivatives Chiralpak AD and Chiralcel OD, was studied. The effect of different modifiers, methanol, ethanol, 2-propanol, and acetonitrile, was examined. The results showed that ABZSO can be separated on both columns, using an alcohol-type modifier. Using the Chiralpak AD column, the best results were obtained with 2-propanol and, in the case of the Chiralcel OD, with methanol.

[Inhibition of lipid peroxidation in retinal tissue during vitrectomy. Effect of intraocular irrigating solution BSS plus]. / Inhibición de la lipoperoxidación retiniana durante la vitrectomía. Efecto de la solución de irrigación intraocular BSS plus.

PURPOSE: Several antioxidant substances have been included in intraocular solutions for vitrectomy in order to reduce oxidative stress. The objective of this work is to evaluate the antioxidant effect of BBS plus(R) (with oxidized glutathione) by means of determining scores of Malondialdehyde (MDA) in retinal tissue at different post vitrectomy time-periods. MATERIAL AND METHODS: Thirty pigmented rabbits distributed in three groups (n=10). Group I: Control (no surgery); group II: Vitrectomy adding BSS plus(R), scoring of malondialdehyde at 3 hours post surgery; group III: Vitrectomy adding BBS plus(R), analysis of malondialdehide at 48 hours after vitrectomy. Retinal samples were used to determine the score of malondialdehyde using ion-pairing high performance liquid chromatography. Statistical analysis was performed using ANOVA-Scheffé test (p<0.05). RESULTS: Scores of free MDA (microgram of retinal tissue) were: group I (n=8): 1.080 S.D. 0.389 microgram group II (n=8): 1.273 S.D. 0.710 microgram and group III (n=10): 0.639 S.D. 0.374 microgram. CONCLUSIONS: BSS plus(R) showed antioxidant effects in retinal tissue which were observed better at a later rather than early stage after a vitrectomy.

Analysis of pesticide residues in wine by solid-phase extraction and gas chromatography with electron capture and nitrogen-phosphorus detection.

A feasible and reproducible method for multiresidue analysis of several common pesticides, of different polarities, in wine samples is proposed. The method combines a solid-phase extraction on polymeric cartridges eluted with ethyl acetate and a gas chromatographic determination using electron capture and nitrogen-phosphorus detection. To avoid the matrix effect, previous washing of the cartridges with a mixture of water-2-propanol (90:10) and further clean-up of the extract on Florisil cartridges, together with a calibration using spiked extracts, are recommended.

Applications of the Chiralpak AD and Chiralcel OD chiral columns in the enantiomeric separation of several dioxolane compounds by supercritical fluid chromatography.

Two chiral columns based on polysaccharide derivatives (Chiralpak AD and Chiralcel OD) have been tested for the chiral separation of several dioxolane compounds, using supercritical fluid chromatography. The compounds studied included ketoconazole and some of its precursors. The effect of the different modifiers and the pressure, on the chromatographic parameters was also evaluated. In general, the alcohol modifiers provided better results than acetonitrile, and all the compounds could be separated with these two columns, but the selection of the column depends on the kind of compound.

Use of deactivated fused-silica capillary precolumns in pesticide analysis by gas chromatography with electron-capture detection.

The advantages and disadvantages of coupling a retention gap of fused-silica between the injection port and the chromatographic column are discussed. The influence on the peak width and height of several factors such as the solvent (n-hexane, acetone, ethyl acetate and methanol), the gap (length, inner diameter, deactivation mode), the injection volume and the pesticide concentration has been examined. Those factors have very different incidences so, it is not possible to extract a general recommendation about the use of gaps. For this reason, checking its viability in each particular case is more advisable.

Determination of oxalate, sulfate and nitrate in honey and honeydew by ion-chromatography.

An ion chromatographic method for determining the anions oxalate, sulfate and nitrate in honey and honeydew samples is described. To prevent matrix interference and to isolate the anions a clean-up step using solid-phase extraction on anionic cartridges and eluting with a 0.01 M chromate solution is recommended. The anions are separated on an anionic column with a mobile phase of borate-gluconate buffer and using conductimetric detection. The method is applied to the analysis of samples from different botanical origin.

Gas chromatographic determination of acrinathrine and 3-phenoxybenzaldehyde residues in honey.

A procedure involving an extraction step and further gas chromatographic analysis with flame ionization detection to determine residues of acrinathrine and its main metabolite, 3-phenoxybenzaldehyde, in honey is proposed. Residues can be isolated from the matrix by means of liquid-liquid extraction with a mixture of benzene-isopropanol, by solid-phase extraction with octadecylsilane cartridges or Florisil packed columns, the latter method giving higher recoveries. Assays on spiked honey samples are carried out to test the procedures that are afterwards applied to honey samples from treated beehives.