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Biomedical subjects

Salvatore Fanali

Publications and source records attributed to Salvatore Fanali.

At least 19 recordsLinked to original sources
PubMed2006

Synthesis and evaluation of polymeric continuous bed (monolithic) reversed-phase gradient stationary phases for capillary liquid chromatography and capillary electrochromatography.

There is a demand of novel high resolution separation media for separation of complex mixtures, particularly biological samples. One of the most flexible techniques for development of new separation media currently is synthesis of the continuous bed (monolithic) stationary phases. In this study the capillary format gradient stationary phases were formed using continuous bed (monolith) polymerization in situ. Different reversed-phase stationary phase gradients were tailored and their resolution using capillary liquid chromatography and capillary electrochromatography at isocratic mobile phase conditions was evaluated. It is demonstrated, that efficiency and resolution of the gradient stationary phases can be substantially increased comparing to the common (isotropic) stationary phases. The proposed formation approach of the gradient stationary phase is reproducible and compatible with the capillary format or microchip format separations. It can be easily automated for the separation optimizations or mass production of the capillary columns or chips.

Benzene Derivatives↗
PubMed2006

Separation of basic compounds of pharmaceutical interest by using nano-liquid chromatography coupled with mass spectrometry.

Nano-liquid chromatography-mass spectrometry (nano-LC-MS) was evaluated for the separation of basic compounds of pharmaceutical interest. The separation of selected beta-blockers, namely nadolol, oxprenolol, alprenolol and propranolol in the presence of terbutaline was performed using two 75 microm I.D. capillaries packed with two different RP18 stationary phases (SP). The best results concerning resolution and efficiency were achieved using the SP where free silanol groups were not present. As expected, this latter SP proved to be very efficient and symmetry factors were observed mainly in the case of the more retained analytes. Baseline resolution of all studied basic compounds was achieved with the Cogent bidentate C18 silica phase (CBC18) eluting analytes at 800 nL/min with a mobile phase containing 500 mM ammonium acetate pH 4.5-water-methanol (1:8:91, v/v/v). The separated basic compounds were revealed using on-column UV detector at 205 nm and electrospray-ion-trap mass spectrometer (ESI-MS). The packed capillary was connected to the MS through a commercial sheath liquid interface or a sheathless nano-spray interface and in both cases the sensitivity was studied and the results compared. Limit of detection (LOD) as low as 0.1 ng/mL was measured for nadolol using the sheathless nano-spray interface and the capillary column packed with the CBC18 stationary phase.

Chromatography, Liquid↗
PubMed2006

Enantiomeric separation of some demethylated analogues of clofibric acid by capillary zone electrophoresis and nano-liquid chromatography.

The enantiomeric separation of some demethylated analogues of clofibric acid, namely 2-(6-chloro-benzothiazol-2-ylsulfanyl)-, 2-(6-methoxy-benzothiazol-2-ylsulfanyl)-, 2-(quinolin-2-yloxy)-, 2-(6-chloro-quinolin-2-yloxy)-, 2-(7-chloro-quinolin-4-yloxy)-propionic acid (compounds A-E, respectively), has been studied by CZE and nano-LC using for the first technique two beta-CD derivatives and vancomycin added to the BGE and vancomycin-modified silica particles for the second one, with the aim to find the optimum experimental conditions for the baseline resolution. The type and the concentration of the chiral selector added to the BGE, the buffer pH, the type of organic modifier and its concentration, the capillary temperature and the applied voltage played a very important role in the enantioresolution of the analysed compounds. The use of 6-monodeoxy-6-monoamino-beta-CD allowed to achieve baseline resolution of four of five clofibric acid derivatives in less than 10 min while heptakis-(2,3,6-tri-O-methyl)-beta-CD partially resolved the same compounds in their enantiomers. Employing vancomycin as the chiral selector in CZE, the counter-current partial filling method was chosen achieving baseline resolution of four analytes. All the studied compounds were enantioresolved employing a capillary column packed with vancomycin stationary phase by nano-LC, and the resolution was strongly influenced by the concentration of the organic modifier and by the pH of the mobile phase. The best results were achieved at pH 4.5 in presence of 60% of methanol (MeOH). However, longer analysis times were observed in the experiments carried out by nano-LC.

Chromatography, Liquid↗
PubMed2006

Chiral MEKC-LIF of amino acids in foods: analysis of vinegars.

The formation of D-amino acids (D-aa's) in many fermented foods depends, among other factors, on the particular fermentation conditions, the action and autolysis of the microorganisms involved. In this sense, the analysis of chiral amino acids is an interesting analytical strategy for food scientists, since these compounds can be used as bacterial markers and can help, e.g., to detect adulterations, microbiological contaminations, etc. In this work, a fast and sensitive method based on MEKC-LIF has been developed to analyze and quantitate L-amino acid (L-aa) and D-aa in vinegars. The chiral MEKC-LIF procedure uses 100 mM sodium tetraborate, 30 mM SDS, and 20 mM beta-CD at pH 9.7 as running buffer, obtaining a good separation of the main vinegar L-/D-aa previously derivatized with fluorescein isothiocianate. Namely, L/D proline, alanine, arginine, glutamic, and aspartic acid, plus the nonchiral amino acid gamma-aminobutyric acid are separated in less than 20 min with high efficiency (up to 720,000 plates/m) and good sensitivity (LODs lower than 16.6 nM were achieved). Several D-aa's were detected and quantified in balsamic, sherry, white wine, and cider vinegars using this MEKC-LIF procedure, observing interesting differences in their L-aa and D-aa profiles and contents.

Acetic Acid↗
PubMed2006

On-line CE-MS using pressurized liquid junction nanoflow electrospray interface and surface-coated capillaries.

A simple and cost-effective laboratory-made liquid junction interface was used for coupling of CE with MS. In this device the capillary column and the spray tip were positioned in the electrode vessel containing appropriate spray liquid. The electrospray potential was applied on the electrode inside the liquid junction. A stable electrospray was produced at nanoliter per minute flow rates generated in the emitter tip without using an external pump. This arrangement provided high durability of the spray tip and independent optimization of the CE separation (use of coated capillaries) and ESI conditions. CE-MS analysis of mixtures of drugs, peptides, tryptic digests of proteins and biological fluids was optimized with respect to the effects of the distance between the separation capillary and electrospray tip and pressure applied on the liquid junction. The sensitivity of the system, in terms of the LOD (base peak monitoring) was below 10 ng/mL for the beta-blocker drugs and below 200 ng/mL for peptide analysis.

Adrenergic beta-Antagonists↗
PubMed2006

Simultaneous determination of aromatic and terpenic constituents of cloves by means of HPLC with diode array detection.

An HPLC method has been developed for the simultaneous determination of aromatic and terpenic constituents of cloves on a C8 RP column, with the mobile phase consisting of a pH 3.5 phosphate buffer-triethylamine (30%) and acetonitrile (70%); a flow rate of 1.2 mL/min and a diode-array detector were used. Complete separation of all analytes (eugenol (EUG), eugenol acetate (AEUG), beta-caryophyllene, a-humulene and caryophyllene oxide) was achieved within 7 min. Good linearity was found in the range 0.125-40.0 microg/mL for EUG and AEUG and in the range 0.250-20.0 microg/mL for the terpenic compounds. After validation, the method was successfully applied to the analysis of clove oil and clove extract samples. The results obtained indicate good accuracy (recovery percentage mean value corresponding to 99.9%) and satisfactory precision.

Calibration↗
PubMed2006

Use of tert-butylbenzoylated tartardiamide chiral stationary phase for the enantiomeric resolution of acidic compounds by nano-liquid chromatography.

Several racemic acidic compounds of pharmaceutical and environmental interest have been separated into their enantiomers by nano-liquid chromatography (nano-LC) employing a tert-butylbenzoylated tartardiamide chiral stationary phase (CHI-TBB). CHI-TBB was packed into a fused silica capillary of 100 microm id and retained by two frits made with a heated wire; detection was on-column at a window (about 0.5 cm) prepared by removing the polyimide layer. The normal phase mode was selected for eluting the studied acidic compounds and therefore n-hexane/2-propanol/acetic acid (89/10/1, v/v/v) was used as mobile phase. Working at a flow rate of 220 nL/min a good resolution was obtained for mecoprop, dichlorprop, diclofop, fenoxaprop (herbicides) and for DF 1738Y, DF 1770Y, DF 2008Y (drugs under evaluation). In order to optimize the chiral resolution we modified the polarity of the mobile phase by adding several polar additives such as ethyl acetate, dichloromethane, tert-butyl methyl ether. Better results were obtained for some herbicides on working with 2-propanol/CH2Cl2/n-hexane/acetic acid (8/4/87/1, v/v/v/v). The influence of the capillary temperature on chiral resolution was studied for two herbicides with different chemical structures, namely mecoprop and haloxyfop in the temperature range between 10 and 40 degrees C and with n-hexane/2-propanol/1% acetic acid (89/10/1, v/v/v) as the mobile phase. Linear correlation of ln k vs 1/T and In alpha vs 1/ T was observed; deltaH degrees values were negative, demonstrating that retention of analytes was an exothermic process. A decrease in resolution was observed with rising temperature, showing that enantioresolution was mainly influenced by selectivity factors.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed2005

Use of teicoplanin stationary phase for the enantiomeric resolution of atenolol in human urine by nano-liquid chromatography-mass spectrometry.

Nano-liquid chromatography (nano-LC) was used for the enantiomeric resolution of atenolol employing a teicoplanin modified silica stationary phase prepared in our laboratory. Experiments were carried out in a fused silica capillary of 75 microm i.d. packed with chiral modified silica particles of 5 microm diameter. Separated enantiomers were revealed by on-line UV detector at 205 nm or electrospray-ion-trap mass spectrometer (ESI-MS). Atenolol enantiomers were eluted utilizing a mobile phase with the following composition: 500 mM ammonium acetate pH 4.5/methanol/acetonitrile 1:60:39 (v/v/v) allowing to achieve good enantioresolution in a reasonable analysis time (about 8 min) with a flow rate of about 900 nL/min. After comparing the sensitivity of the nano-LC method using a conventional UV detector for capillary electrophoresis, a zeta cell (3 nL volume) employed in nano-LC and the ion-trap MS the method was validated with the MS detector offering the highest sensitivity (limit of detection (LOD)=50 ng/mL; limit of quantification (LOQ)=400 ng/mL for each atenolol enantiomer). (-)-Psi-Nor-ephedrine was used as the internal standard. The method was successfully applied to the analysis of atenolol enantiomers present in human urine samples of a patient under atenolol therapy.

Adrenergic beta-Antagonists↗
PubMed2005

Analysis and separation of enkephalin and dalargin analogues and fragments by capillary zone electrophoresis.

Capillary zone electrophoresis (CZE) has been applied to qualitative and quantitative analysis and separation of synthetic analogues and fragments of enkephalins ([Leu5]enkephalin, H-Tyr-Gly-Gly-Phe-Leu-OH, [Met5]enkephalin, H-Tyr-Gly-Gly-Phe-Met-OH), and dalargin (H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH), biologically active peptides with morphin-like effects acting as ligands for the opiate receptors in the brain. These oligopeptides (dipeptides to hexapeptides) were analyzed as cations in two acidic background electrolytes (BGEs), BGE I (100mM H3PO4, 50mM Tris, pH 2.25), BGE II (100mM iminodiacetic acid, pH 2.30), and both as cations and anions in alkaline BGE IV (40 mM Tris, 40 mM Tricine, pH 8.10). Purity degrees of peptides, expressed in three different ways (relative peak height, relative peak area and relative corrected peak area), were determined by their CZE analyses in the above BGEs, and their values were compared with respect to the peak shapes and migration times of the main synthetic products and their admixtures. Selected analogues and fragments of enkephalins and dalargin were successfully separated by CZE in acidic isoelectric buffers, 100 and 200 mM iminodiacetic acid, pH 2.30 and 2.32, respectively. The effective electrophoretic mobilities at standard temperature 25 degrees C, and effective and specific charges of all analyzed peptides in the above three BGEs were determined. Correlation between effective electrophoretic mobility of the analyzed peptides and their charge and size (relative molecular mass) was investigated, which revealed different molecular shape of analyzed peptides in acidic and alkaline BGEs. In addition, the selected characteristics of the UV-absorption detector (noise, signal to noise ratio, sensitivity, and limits of detection and quantification) were determined.

Electrolytes↗
PubMed2005

Separation and analysis of glycyrrhizin, 18beta-glycyrrhetic acid and 18alpha-glycyrrhetic acid in liquorice roots by means of capillary zone electrophoresis.

Glycyrrhizin is the main active compound of Glycyrrhiza glabra root extracts; according to recent studies, glycyrrhizin and its aglycon, glycyrrhetic acid, have interesting therapeutic properties. A new capillary electrophoretic method has been developed for the separation and quantification of glycyrrhizin, beta-glycyrrhetic acid and its isomer a-glycyrrhetic acid. Separation of the analytes was achieved in less than 3 min on a fused silica capillary, by injecting the samples at the short end of the capillary (effective length: 8.5 cm). The background electrolyte was composed of pH 10.0 carbonate buffer, methanol and ethylene glycol (80/10/10) and contained 0.4% beta-cyclodextrin; indomethacin was used as the internal standard. Diode array detection was used, with quantitative assays carried out at 254 nm. Linearity was found over the 5-200 and 2.5-100 microg mL(-1) concentration ranges for glycyrrhizin and glycyrrhetic acid, respectively. This method has been applied to the determination of the analytes in different matrices (liquorice roots and commercial confectionery products), and to the purity control of beta-glycyrrhetic acid obtained from the hydrolysis of glycyrrhizin. When analysing beta-glycyrrhetic acid and its epimer in roots, the samples were purified by means of a suitable solid-phase extraction (SPE) procedure with Oasis HLB cartridges, which granted good selectivity, eliminating matrix interference.

Candy↗
PubMed2005

Rapid assay of vitamin E in vegetable oils by reversed-phase capillary electrochromatography.

A rapid capillary electrochromatographic (CEC) method for the analysis of vitamin E in vegetable oils is reported. Vitamin E consists of a group of eight isomers, tocopherols (TOHs) and tocotrienols. The separation of four TOHs (alpha-, gamma-, delta-TOH), alpha-tocopherol acetate (alpha-TOH-Ac), and an antioxidant compound, butylated hydroxytoluene (BHT) used to prevent TOH autoxidation, was optimized. The CEC experiments were carried out in a 75 microm inner diameter (ID) fused-silica capillary, partially packed with 3 microm C(18 )stationary phase (33 cm total length, 8.4 cm and 7 cm effective and packed lengths, respectively). The optimum mobile phase was a polar organic phase composed of a mixture of methanol-acetonitrile in the ratio 50/50 v/v containing 0.01% ammonium acetate, applying a voltage and temperature set at -25 kV and 20 degrees C, respectively. The tocopherols and the BHT were successfully separated within 2.5 min using the short-end injection method. Under these experimental conditions, repeatability of retention time and peak area, analyte detection and quantitation limits, linearity, precision, and accuracy were studied. The CEC method was applied to determine the content of TOHs in different commercially available oils of virgin olive, hazelnut, sunflower, and soybean. The extraction of vitamin E isomers from oil samples was achieved using methanol and a methanol-isopropanol mixture.

Butylated Hydroxytoluene↗
PubMed2005

Separation and analysis of the major constituents of cloves by micellar electrokinetic chromatography.

A micellar electrokinetic chromatographic method was developed and applied for the first time to the separation and quantification of the main components of cloves, namely eugenol and beta-caryophyllene. The analytes were separated in less than ten minutes in an unmodified fused silica capillary (effective length: 40.0 cm) using a background electrolyte composed of 25 mM carbonate buffer (pH 10.0) and 60 mM sodium dodecylsulfate. Analyte concentrations were determined in clove oil and in methanolic clove extracts after simple sequential dilution with methanol and background electrolyte, and the resulting electropherograms showed no interference due to other components. Precision assays gave satisfactory results, with mean standard deviation of peak areas lower than 4.7% (lower than 1.5% for migration times). The eugenol content of a commercial clove oil was about 70% (RSD% = 3.2), while that of beta-caryophyllene was about 15% (RSD% = 4.2); in clove extracts, on the other hand, eugenol was approximately 15% (RSD% = 4.7) and beta-caryophyllene 4% (RSD% = 5.1). Accuracy was also good (mean recovery >98%).

Chromatography, Micellar Electrokinetic Capillary↗
PubMed2005

Enantiomeric separation of mirtazapine and its metabolites by nano-liquid chromatography with UV-absorption and mass spectrometric detection.

Mirtazapine (MIR) and two of its main metabolites, namely, 8-hydroxymirtazapine and N-desmethylmirtazapine, were separated in totheir enantiomers by nanoLC in a laboratory-made fused-silica capillary column (75 microm ID) packed with a vancomycin-modified silica stationary phase. The simultaneous separation of the three couples of the studied enantiomers was achieved in less than 33 min, using an experimentally optimized mobile phase delivered in the isocratic mode. Optimization of the mobile-phase composition was achieved by testing the influence of the buffer pH and concentration, the water concentration, the organic modifier type and concentration, and on the retention and resolution of the analytes. The optimum mobile-phase composition contained 500 mM ammonium acetate pH 4.5/water/MeOH/MeCN, 1:14:40:45 v/v/v/v. Using a UV detector at 205 nm, the method was validated studying several experimental parameters such as LOD and LOQ, intraday and interday repeatability, and linearity. Good results were achieved: LOD and LOQ were in the range 5-15 and 10-40 microg/mL, respectively (the highest value was obtained for the DEMIR enantiomers); correlation coefficients, 0.9993-0.9999; the intraday and interday precision was acceptable (RSD < 2%) using an internal standard. The method was tested for the separation of the studied enantiomers in an extracted (solid-phase) serum sample spiked with standard racemic mixture of MIR and its two metabolites. Finally, the nanoLC system was connected to a mass spectrometer through a nanoelectrospray interface and the MS, MS2, and MS3 spectra were acquired showing the potential of the system used for characterization and identification of the separated analytes.

Chromatography, Liquid↗
PubMed2005

Use of nano-liquid chromatography for the analysis of glycyrrhizin and glycyrrhetic acid in licorice roots and candies.

Glycyrrhizin (G) and glycyrrhetic acid (GA) were separated by using nano-liquid chromatography (nano-LC) in a fused silica capillary packed with RP18 stationary phase (75 microm ID, effective length 33 cm, packed 23 cm) eluting at 300 nL/min in a gradient mode. The mobile phase was a mixture of water-MeOH-MeCN-acetic acid (29:35:35:1, v/v/v/v) that was delivered for one minute and after this was modified by reducing the water content (14:42.5:42.5:1, v/v/v/v). The intra-day and inter-day relative standard deviations (of retention time and peak area) were satisfactory (lower than 2.9 and 4%, respectively). The linearity of the nano-LC method was assessed in the range 0.62-5.00 microg/mL and 80-200 microg/mL for GA and G, with R2 = 0.996 and 0.995, respectively. The licorice was extracted with a mixture of ethanol-water, diluted with the mobile phase, and injected for the analysis.

Chromatography, High Pressure Liquid↗
PubMed2005

Evaluation of cyclodextrins modified with dichloro-, dimethyl-, and chloromethylphenylcarbamate groups as chiral stationary phases for capillary electrochromatography.

Capillary electrochromatography using cyclodextrins modified with dichloro-, dimethyl-, and chloromethylcarbamate groups were used for the enantiomeric separation of standard analytes. The chiral selector was chemically bonded to aminopropylsilanized silica particles, and these chiral stationary phases (CSPs), mixed with aminopropylsilanized silica (1:1 wt:wt), were packed into 100-microm-i.d. fused-silica capillaries. The effect of the type of cyclodextrin, the nature and position of the substituents on the phenyl ring, and the binding mode of cyclodextrin phenylcarbamates onto the silica gel surface on the chiral recognition were studied. Experimental parameters such as organic solvent concentration were varied in order to better understand the mechanism contributing to the chiral recognition of these CSPs. Good enantioseparations were achieved for a racemic flavanone (FLA) and trans-cyclopropanedicarboxylic acid dianilide (CAD).

Carbamates↗
PubMed2004

A rapid HPLC-DAD method for the analysis of fluoxetine and norfluoxetine in plasma from overdose patients.

There is a need for fast, simple and reliable analytical methods for the analysis of fluoxetine and norfluoxetine in patients who voluntarily or involuntarily have taken an overdose of the drug. A new liquid chromatographic method with diode array detection is presented herein for the determination of fluoxetine and its main active metabolite in human plasma for toxicological purposes. A mobile phase composed of acetonitrile and aqueous tetramethylammonium perchlorate allows to obtain the complete separation of the analytes on a C18 reversed phase column. The fast and accurate sample pre-treatment step is carried out by means of solid-phase extraction using hydrophilic-lipophilic balance cartridges and loading 100 microL of plasma only. This procedure gives satisfactory extraction yield values, as well as good plasma sample purification from matrix interference. Linearity was obtained in the 150-3000 ng/mL range for both analytes. Selectivity with respect to other psychotropic drugs was satisfactory. The method seems to be suitable for the analysis of fluoxetine and its metabolite in human plasma for depressed patients in overdose.

Antidepressive Agents, Second-Generation↗
PubMed2004

Enantioselective determination of the novel antidepressant mirtazapine and its active demethylated metabolite in human plasma by means of capillary electrophoresis.

Mirtazapine is a recent noradrenergic and specific serotonergic antidepressant drug. A capillary electrophoretic method has been developed for the enantioseparation and analysis of mirtazapine and its main active metabolite, N-desmethylmirtazapine, in human plasma. For method optimisation several experimental parameters were investigated, such as type and concentration of the chiral selector, buffer pH and capillary temperature. Baseline enantioseparation of the analytes was achieved in 2.5 min in a fused silica capillary (50 microm i.d.; 48.5 cm total length; 8.5 cm effective length) using carboxymethyl-beta-cyclodextrin, dissolved in a background electrolyte consisting of 50 mM phosphate buffer at pH 2.5, as the chiral selector. UV detection was set at 205 nm. A careful pre-treatment of plasma samples was developed, using solid-phase extraction with hydrophilic-lipophilic balance cartridges (60 mg, 3 mL), eluting the sample with methanol, then concentrating it 37.5 times before injection. Extraction yield values are very satisfactory, being the average 89% for mirtazapine and 73% for N-desmethylmirtazapine. Application of the method to some human plasma samples has given satisfactory results.

Antidepressive Agents↗