Use of a Novel Sub‐2 µm Silica Hydride Vancomycin Stationary Phase in Nano‐Liquid Chromatography. II. Separation of Derivatized Amino Acid Enantiomers

A novel vancomycin silica hydride stationary phase was synthesized and the particles of 1.8 µm were packed into fused silica capillaries of 75 µm internal diameter (I.D.). The chiral stationary phase (CSP) was tested for the separation of some derivatized amino acid enantiomers by using nano‐liquid chromatography (nano‐LC). Some experimental parameters such as the type and the content of organic modifier, the pH, and the concentration of the buffer added to the mobile phase were modified and the effect on enantioselectivity, retention time, and enantioresolution factor was studied. The separation of selected dansyl amino acids (Dns‐AAs), e.g., Asp, Glu, Leu, and Phe in their enantiomers was initially achieved utilizing a mobile phase containing 85% (v/v) methanol (MeOH) and formate buffer measuring the enantioresolution factor and enantioselectivity in the range 1.74–4.17 and 1.39–1.59, respectively. Better results were obtained employing a more polar organic solvent as acetonitrile (ACN) in the mobile phase. Optimum results (Rs 1.41–6.09 and α 1.28–2.36) were obtained using a mobile phase containing formate buffer pH 2.5/water/MeOH/ACN 6:19:12.5:62.5 (v/v/v/v) in isocratic elution mode at flow rate of 130 nL/min. Chirality 27:767–772, 2015. © 2015 Wiley Periodicals, Inc.     

Analysis of Oxcarbazepine and the 10‐Hydroxycarbazepine Enantiomers in Plasma by LC‐MS/MS: Application in a Pharmacokinetic Study

Oxcarbazepine is a second‐generation antiepileptic drug indicated as monotherapy or adjunctive therapy in the treatment of partial seizures or generalized tonic–clonic seizures in adults and children. It undergoes rapid presystemic reduction with formation of the active metabolite 10‐hydroxycarbazepine (MHD), which has a chiral center at position 10, with the enantiomers (S)‐(+)‐ and R‐(?)‐MHD showing similar antiepileptic effects. This study presents the development and validation of a method of sequential analysis of oxcarbazepine and MHD enantiomers in plasma using liquid chromatography with tandem mass spectrometry (LC‐MS/MS). Aliquots of 100 μL of plasma were extracted with a mixture of methyl tert‐butyl ether: dichloromethane (2:1). The separation of oxcarbazepine and the MHD enantiomers was obtained on a chiral phase Chiralcel OD‐H column, using a mixture of hexane:ethanol:isopropanol (80:15:5, v/v/v) as mobile phase at a flow rate of 1.3 mL/min with a split ratio of 1:5, and quantification was performed by LC‐MS/MS. The limit of quantification was 12.5 ng oxcarbazepine and 31.25 ng of each MHD enantiomer/mL of plasma. The method was applied in the study of kinetic disposition of oxcarbazepine and the MHD enantiomers in the steady state after oral administration of 300 mg/12 h oxcarbazepine in a healthy volunteer. The maximum plasma concentration of oxcarbazepine was 1.2 µg/mL at 0.75 h. The kinetic disposition of MHD is enantioselective, with a higher proportion of the S‐(+)‐MHD enantiomer compared to R‐(?)‐MHD and an AUC^0‐12 _{S‐(+)/R‐(?)} ratio of 5.44. Chirality 25:897–903, 2013. © 2013 Wiley Periodicals, Inc.     

A novel colorimetric assay of β‐D‐glucans in basidiomycete strains by alcian blue dye in a 96‐well microtiter plate

Basidiomycete strains synthesize several types of β‐d ‐glucans, which play a major role in the medicinal properties of mushrooms. Therefore, the specific quantification of these β‐d ‐glucans in mushroom strains is of great biochemical importance. Because published assay methods for these β‐d ‐glucans present some disadvantages, a novel colorimetric assay method for β‐d ‐glucan with alcian blue dye was developed. The complex formation was detected by following the decrease in absorbance in the range of 620 nm and by hypsochromic shift from 620 to 606 nm (～14 nm) in UV‐Vis spectrophotometer. Analysis of variance was used for optimization of the slope of the calibration curve by using the assay mixture containing 0.017% (w/v) alcian blue in 2% (v/v) acetic acid at pH 3.0. The high‐throughput colorimetric assay method on microtiter plates was used for quantification of β‐d ‐glucans in the range of 0–0.8 μg, with a slope of 44.15 × 10^?2 and a limit of detection of 0.017 μg/well. Recovery experiments were carried out by using a sample of Hericium erinaceus, which exhibited a recovery of 95.8% for β‐1,3‐d ‐glucan. The present assay method exhibited a 10‐fold higher sensitivity and a 59‐fold lower limit of detection compared with the published method with congo red. β‐d ‐glucans of several mushrooms strains were isolated from fruiting bodies and mycelia, and they were quantified by this assay method. This assay method is fast, specific, simple, and it can be used to quantify β‐d ‐glucans from other biological sources. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:1526–1535, 2015     

Enantiospecific Analysis of 8‐Prenylnaringenin in Biological Fluids by Liquid‐Chromatography‐Electrospray Ionization Mass Spectrometry: Application to Preclinical Pharmacokinetic Investigations

8‐Prenylnaringenin (8PN) is a naturally occurring bioactive chiral prenylflavonoid found most commonly in the female flowers of hops (Humulus lupulus L.). A stereospecific method of analysis for 8PN in biological fluids is necessary to study the pharmacokinetic disposition of each enantiomer. A novel and simple liquid chromatographic‐electrospray ionization‐mass spectrometry (LC‐ESI‐MS) method was developed for the simultaneous determination of R‐ and S‐8PN in rat serum and urine. Carbamazepine was used as the internal standard (IS). Enantiomeric resolution of 8PN was achieved on a Chiralpak^® AD‐RH column with an isocratic mobile phase consisting of 2‐propanol and 10 mM ammonium formate (pH 8.5) (40:60, v/v) and a flow rate of 0.7 mL/min. Detection was achieved using negative selective ion monitoring (SIM) of 8PN at m/z 339.15 for both enantiomers and positive SIM m/z at 237.15 for the IS. The calibration curves for urine were linear over a range of 0.01–75 µg/mL and 0.05–75 µg/mL for serum with a limit of quantification of 0.05 µg/mL in serum and 0.01 µg/mL in urine. The method was successfully validated showing that it was sensitive, reproducible, and accurate for enantiospecific quantification of 8PN in biological matrices. The assay was successfully applied to a preliminary study of 8PN enantiomers in rat. Chirality 26:419–426, 2014. © 2014 Wiley Periodicals, Inc.     

Identification of N‐octylnortadalafil and its Stereoisomers in Dietary Supplements with Chiral Liquid Chromatography–Circular Dichroism

A direct chiral liquid chromatography–circular dichroism (LC‐CD) method was developed for the simple and rapid identification of N‐octylnortadalafil [(6R, 12aR)‐6‐(1,3‐benzodioxol‐5‐yl)‐2‐octyl‐2,3,6,7,12,12a‐hexahydropyrazino[1’,2’:1,6]pyrido[3,4‐b]indole‐1,4‐dione; RR‐OTDF] and its stereoisomers in dietary supplements. Samples were extracted with methanol. Compounds were then separated by chiral LC‐CD using Chiralcel OD‐RH (4.6 × 1 50 mm, 5 µm) with 5 mM ammonium formate (pH 3)/0.1% formic acid in acetonitrile (95:5, v/v) mixture solution (mobile phase A) and 0.1% formic acid in acetonitrile (mobile phase B). The isocratic elution used was mobile phase A / mobile phase B (3:7, v/v) at a flow rate of 1 ml/min. The column temperature was held at 30°C. RR‐OTDF and its stereoisomers were separated within 20 min with the resolution factors being over 2.0. Using this method, RR‐OTDF and (6R, 12aS)‐6‐(1,3‐benzodioxol‐5‐yl)‐2‐octyl‐2,3,6,7,12,12a‐hexahydropyrazino[1’,2’:1,6]pyrido[3,4‐b]indole‐1,4‐dione were detected in a dietary supplement. Chirality 28:204–208, 2016. © 2016 Wiley Periodicals, Inc.     

Determination of carbamazepine in human urine and serum samples by high‐performance liquid chromatography with post‐column Ru(bipy)‐Ce(SO4)2 chemiluminescence detection

A novel, sensitive and rapid CL method coupled with high‐performance liquid chromatography separation for the determination of carbamazepine is described. The method was based on the fact that carbamazepine could significantly enhance the chemiluminescence of the reaction of cerium sulfate and tris(2,2‐bipyridyl) ruthenium(II) in the presence of acid. The chromatographic separation was performed on a Kromasil® (Sigma‐Aldrich) TM RP‐C18 column (id: 150 mm × 4.6 mm, particle size: 5 µm, pore size: 100 Å) with a mobile phase consisting of methanol–water‐glacial acetic acid (70:29:1, v/v/v) at a flowrate of 1.0 mL/min, the total analysis time was within 650 s. Under optimal conditions, CL intensity was linear for carbamazepine in the range 2.0 × 10^?8 ~ 4.0 × 10^?5 g/mL, with a detection limit of 6.0 × 10^?9 g/mL (S/N = 3) and the relative standard detection was 2.5% for 2.0 × 10^?6 g/mL (n = 11). This method was successfully applied to the analysis of carbamazepine in human urine and serum samples. The possible mechanism of the CL reaction is also discussed briefly. Copyright © 2012 John Wiley & Sons, Ltd.     

A validated enantiospecific method for determination and purity assay of clopridogrel

A new and accurate HPLC method using β‐cyclodextrin chemically bonded to spherical silica particles as chiral stationary phase (CSP) was developed and validated for determination of S‐clopidogrel and its impurities R‐enantiomer and S‐acid as a hydrolytic product. The effects of acetonitrile and methanol content in the mobile phase and temperature on the resolution and retention of enantiomers were investigated. A satisfactory resolution of S‐clopidogrel active form and its impurities was achieved on ChiraDex® column (5 μm, 4 × 250 mm) at a flow rate of 1.0 ml/min and 17°C using acetonitrile, methanol and 0.01 M potassium dihydrogen phosphate solution (15:5:80 v/v/v) as mobile phase. The detection wavelength was set at 220 nm. The method was validated in terms of accuracy, precision, linearity, and robustness. The limit of detection for R‐enantiomer and S‐acid were 0.75 and 0.09 μg/ml, respectively, injection volume being 20 μl. Finally, the molecular modeling of the inclusion complexes between the analytes and β‐cyclodextrin was performed to investigate the mechanism of the enantiorecognition and to study the quantitative structure–retention relationships. Chirality, 2009. © 2008 Wiley‐Liss, Inc.     

Stereoselective Determination of Metoprolol and its Metabolite α‐Hydroxymetoprolol in Plasma by LC‐MS/MS: Application to Pharmacokinetics during Pregnancy

Metoprolol is available for clinical use as a racemic mixture. The S‐(?)‐metoprolol enantiomer is the one expressing higher activity in the blockade of the β₁‐adrenergic receptor. The α‐hydroxymetoprolol metabolite also has activity in the blockade of the β₁‐adrenergic receptor. The present study describes the development and validation of a stereoselective method for sequential analysis of metoprolol and of α‐hydroxymetoprolol in plasma using high‐performance liquid chromatography with tandem mass spectrometry (LC‐MS/MS). 1‐ml aliquots of plasma were extracted with dichloromethane : diisopropyl ether (1:1, v/v). Metoprolol enantiomers and α‐hydroxymetoprolol isomers were separated on a Chiralpak AD column (Daicel Chemical Industries, New York, NY, USA) and quantitated by LC‐MS/MS. The limit of quantitation obtained was 0.2 ng of each metoprolol enantiomer/ml plasma and 0.1 ng/ml of each α‐hydroxymetoprolol isomer/ml plasma. The method was applied to the study of kinetic disposition of metoprolol in plasma samples collected up to 24 h after the administration of a single oral dose of 100‐mg metoprolol tartrate to a hypertensive parturient with a gestational age of 42 weeks. The clinical study showed that the metoprolol pharmakokinetics is enantioselective, with the observation of higher area under the curve (AUC)^0?∞ values for S‐(?)‐metoprolol (AUC_S‐(?)/AUC_R‐(+) = 1.81) and the favoring of the formation of the new chiral center 1′R of α‐hydroxymetoprolol (AUC^0?∞_1′R/1′S = 2.78). Chirality, 25:1–7, 2013. © 2012 Wiley Periodicals, Inc.     

Chromatographic profiles of tryptophan and kynurenine enantiomers derivatized with (S)‐4‐(3‐isothiocyanatopyrrolidin‐1‐yl)‐7‐(N,N‐dimethylaminosulfonyl)‐2,1,3‐benzoxadiazole using LC–MS/MS on a triazole‐bonded column

d ‐ and l ‐Tryptophan (Trp) and d ‐ and l ‐kynurenine (KYN) were derivatized with a chiral reagent, (S )‐4‐(3‐isothiocyanatopyrrolidin‐1‐yl)‐7‐(N,N‐dimethylaminosulfonyl)‐2,1,3‐benzoxadiazole (DBD‐PyNCS), and were separated enantiomerically by high‐performance liquid chromatography (HPLC) equipped with a triazole‐bonded column (Cosmosil HILIC) using tandem mass spectrometric (MS/MS) detection. Effects of column temperature, salt (HCO₂NH₄) concentration, and pH of the mobile phase in the enantiomeric separation, followed by MS detection of (S )‐DBD‐PyNCS‐d ,l ‐Trp and ‐d ,l ‐KYN, were investigated. The mobile phase consisting of CH₃CN/10 mM ammonium formate in H₂O (pH 5.0) (90/10) with a column temperature of 50–60 °C gave satisfactory resolution (R s) and mass‐spectrometric detection. The enantiomeric separation of d ,l ‐Trp and d ,l ‐KYN produced R s values of 2.22 and 2.13, and separation factors (α) of 1.08 and 1.08, for the Trp and KYN enantiomers, respectively. The proposed LC–MS/MS method provided excellent detection sensitivity of both enantiomers of Trp and KYN (5.1–19 nM).     

Validated spectrofluorimetric method for the determination of carbamazepine in pharmaceutical dosage forms after reaction with 4‐chloro‐7‐‐nitrobenzo‐2‐oxa‐1,3‐diazole (NBD‐Cl)

A sensitive and simple spectrofluorimetric method has been developed and validated for the determination of the anti‐epileptic drug carbamazepine (CBZ) in its dosage forms. The method was based on a nucleophilic substitution reaction of CBZ with 4‐chloro‐7‐nitrobenzo‐2‐ oxa‐1,3‐diazole (NBD‐Cl) in borate buffer (pH 9) to form a highly fluorescent derivative that was measured at 530 nm after excitation at 460 nm. Factors affecting the formation of the reaction product were studied and optimized, and the reaction mechanism was postulated. The fluorescence–concentration plot is rectilinear over the range of 0.6–8 µg/mL with limit of detection of 0.06 µg/mL and limit of quantitation of 0.19 µg/mL. The method was applied to the analysis of commercial tablets and the results were in good agreement with those obtained using the reference method. Validation of the analytical procedures was evaluated according to ICH guidelines. Copyright © 2015 John Wiley & Sons, Ltd.     

Spectrofluorimetric determination of atenolol from human urine using high‐affinity molecularly imprinted solid‐phase extraction sorbent

This study presents a novel, sensitive and selective molecularly imprinted solid‐phase extraction (MISPE)–spectrofluorimetric method for the removal and determination of atenolol from human urine. Molecularly imprinted and non‐imprinted polymers were synthesized thermally using a radical chain polymerization technique and used as solid‐phase extraction sorbents. Acrylic acid ethylene glycol dimethacrylate, dibenzoyl peroxide and dichloroethane were used as a functional monomer, cross‐linker, initiator and porogen, respectively. The calibration curve was in the range of 0.10–2.0 μg/ml for the developed method. Limit of detection and limit of quantification values were 0.032 and 0.099 μg/ml, respectively. Owing to the selectivity of the MISPE technique and the sensitivity of spectrofluorimetry, trace levels of atenolol have been successfully determined from both organic and aqueous media. Relatively high imprinting factor (4.18) and recovery results (74.5–75.3%) were obtained. In addition, intra‐ and interday precision values were 0.38–1.03% and 0.47–2.05%, respectively, proving the precision of the proposed method. Thus, a selective, sensitive and simple MISPE–spectrofluorimetric method has been developed and applied to the direct determination of atenolol from human urine.     

Separation of Ofloxacin and Its Six Related Substances Enantiomers by Chiral Ligand‐Exchange Chromatography

A chiral ligand‐exchange high‐performance liquid chromatography method was developed for the enantioseparation of ofloxacin and its six related substances termed impurities A, B, C, D, E, and F. The separation was performed on a conventional C₁₈ column. Different organic modifiers, copper salts, amino acids, the ratio of Cu²⁺ to amino acid, pH of aqueous phase, and column temperature were optimized. The optimal mobile phase conditions were methanol‐water systems consisting of 5 mmol/L copper sulfate and 10 mmol/L L‐isoleucine (L‐Ile). Under such conditions, good enantioseparation of ofloxacin and impurities A, C, E, and F could be observed with resolutions (R_S) of 3.54, 1.97, 3.21, 3.50, and 2.12, respectively. On the relationship between the thermodynamic parameters and structures of analytes, the mechanism of chiral recognition was investigated. It was concluded that ofloxacin and impurities A, C, E, and F were all enthalpically driven enantioseparation and that low column temperature was beneficial to enantioseparation. Furthermore, the structure–separation relationship of these analytes is also discussed. Chirality 27:843–849, 2015. © 2015 Wiley Periodicals, Inc.     

Simultaneous Enantiomeric Determination of Propranolol,Metoprolol, Pindolol,and Atenolol in Natural Waters by HPLC on New Polysaccharide‐Based Stationary Phase using a Highly Selective Molecularly Imprinted Polymer Extraction

A simple high performance liquid chromatography method HPLC‐UV for simultaneous enantiomeric determination of propranolol, metoprolol, pindolol, and atenolol in natural water samples was developed and validated, using a molecularly imprinted polymer solid‐phase extraction. To achieve this purpose, Lux® Cellulose‐1/Sepapak‐1 (cellulose tris‐(3,5‐dymethylphenylcarbamate)) (Phenomenex, Madrid, Spain) chiral stationary phase was used in gradient elution and normal phase mode at ambient temperature. The gradient elution program optimized consisted of a progressive change of the mobile phase polarity from n‐hex/EtOH/DEA 90/10/0.5 (v/v/v) to 60/40/0.5 (v/v/v) in 13 min, delivered at a flow rate of 1.3 ml/min and a sudden change of flow rate to 2.3 ml/min in 1 min. Critical steps in any molecularly imprinted polymer extraction protocol such as the flow rate to load the water sample in the cartridges and the breakthrough volume were optimized to obtain the higher extraction recoveries for all compounds. In optimal conditions (100 ml breakthrough volume loaded at 2.0 ml/min), extraction recoveries for the four pairs of β‐blockers were near 100%. The MIP‐SPE‐HPLC‐UV method developed demonstrates good linearity (R² ≥ 0.99), precision, selectivity, and sensitivity. Method limit detection was 3.0 µg/l for propranolol and pindolol enantiomers and 20.0 and 22.0 µg/l for metoprolol and atenolol enantiomers, respectively. The proposed methodology should be suitable for routine control of these emerging pollutants in natural waters for a better understanding of the environmental impact and fate. Chirality 24:860–866, 2012. © 2012 Wiley Periodicals, Inc.     

Indirect Chiral Separation of New Recreational Drugs by Gas Chromatography‐Mass Spectrometry Using Trifluoroacetyl‐L‐Prolyl Chloride as Chiral Derivatization Reagent

New recreational drugs such as amphetamine‐, cathinone, and benzofury derivatives gained high popularity on the drug market in recent years. They can be purchased via the Internet from different providers and online portals. Most of these compounds are chiral, which makes the development of chiral separation methods necessary. Besides this, it is useful to find out if the compounds were sold as racemic mixtures. Also, it is important to check whether the new psychoactive compounds contain further ingredients or impurities. The aim of this research was the continuation of the application of a method for indirect chiral separation of 24 new psychoactive compounds recently purchased via the Internet. After derivatization with the chiral derivatization reagent trifluoroacetyl‐L‐prolyl chloride, chromatographic separation of diastereomers was achieved using a 30 m HP5‐MS capillary column. As carrier gas, helium was used with a constant flow of 1.0 ml/min. Three different column temperature programs were tested. Under optimum conditions 13 out of 24 compounds were successfully resolved into their enantiomers obtaining Rs values up to 7.0. The use of a single quadrupole mass spectrometer as the detector allowed the identification of the compounds in multicomponent samples. Chirality 27:211–215, 2015. © 2014 Wiley Periodicals, Inc.     

Spectrofluorimetric determination of aluminium using 2‐hydroxy‐1‐naphthylidene‐(8‐aminoquinoline)

A sensitive and selective spectrofluorimetric method has been developed for the rapid determination of aluminium. This method is based on the complex formation between aluminium and 2‐hydroxy‐1‐naphthylidene‐(8‐aminoquinoline) (HNAQ). The optimum conditions for the complex formation were a metal‐to‐ligand (M : L) stoichiometric ratio of 1:1, a pH of 5.5 and a 0.20 m acetate buffer. The fluorescence of the complex was monitored at an emission wavelength of 502 nm with excitation at 438 nm. Under these conditions, linear calibration curves were obtained in the ranges 0.05–1 and 1–5 ppm. The detection limit was 3.4 ppb for the former and 13.5 ppb for the latter. The maximum relative standard deviation of the method for an aluminium standard of 200 ppb was 1.5% (n = 5). This method was successfully applied for the determination of aluminium in drinking water, pharmaceutical antacid tablets and suspension samples. Copyright © 2010 John Wiley & Sons, Ltd.     

Enantioselective analysis of N‐hydroxymexiletine glucuronide in human plasma for pharmacokinetic studies

Enzymatic hydrolysis with β‐glucuronidase/sulfatase was used for the enantioselective determination of N‐hydroxymexiletine glucuronide in plasma for pharmacokinetic studies. N‐Hydroxymexiletine glucuronide was determined as the quantity of mexiletine released by hydrolysis (difference between the enantiomeric concentrations of mexiletine obtained with and without hydrolysis). Plasma samples (100 μl) were treated at pH 5.0 with 10 mg of the enzyme (Limpet Acetone Powder type I) for 16 hr at 37°C and extracted at pH 10.4 with diisopropyl ether. Chiral mexiletine discrimination was obtained by reaction with o‐phthalaldehyde/N‐acetyl‐L ‐cysteine, separation of the resulting diastereomers on a C‐18 reversed‐phase column with a mobile phase of methanol–0.05 N acetate buffer, pH 5.5 (6.5:3.5, v/v), and fluorescence detection (λ_ex 350 nm, λ_em 455 nm). The performance characteristics for the enantioselective analysis of mexiletine preceded by enzymatic hydrolysis were recovery ∼90%, quantification limit 1 ng/ml, and linearity up to 1000 ng/ml plasma for both enantiomers. The coefficients of variation obtained in the study of intra‐ and inter‐day precision were respectively 5% and 7% for both enantiomers. The assay was shown to be suitable for a pharmacokinetic study performed in a patient with the arrhythmic form of chronic Chagas' heart disease treated with 200 mg t.i.d. of racemic mexiletine hydrochloride. The high sensitivity of the method allows analysis of only 100 μl plasma. Chirality 11:85–90, 1999. © 1999 Wiley‐Liss, Inc.     

Determination of Trantinterol Enantiomers in Human Plasma by High‐Performance Liquid Chromatography – Tandem Mass Spectrometry Using Vancomycin Chiral Stationary Phase and Solid Phase Extraction and Stereoselective Pharmacokinetic Application

A sensitive and enantioselective vancomycin chiral stationary phase high‐performance liquid chromatography–tandem mass spectrometry method was developed for the determination of trantinterol enantiomers in human plasma. Baseline resolution was achieved using the vancomycin chiral stationary phase known as Chirobiotic V with polar ionic mobile phase consisting of acetonitrile–methanol (60:40, v/v) containing 0.01% ammonia and 0.02% acetic acid at a flow rate of 1.0 mL/min. Waters Oasis HLB C18 solid phase extraction cartridges were used in the sample preparation of trantinterol samples from plasma. The detection was performed on a triple‐quadrupole tandem mass spectrometer by multiple reaction monitoring mode via electrospray ionization. The calibration curve was linear in a concentration range from 0.0606 to 30.3 ng/mL in plasma, with the lower limit of quantification of 0.0606 ng/mL. The intra‐ and interday precision (relative standard deviation) values were within 9.7% and the accuracy (relative error) was from ?6.6 to 7.2% at all quality control levels. The method was successfully applied to a study of stereoselective pharmacokinetics in human. Chirality 27:327–331, 2015.© 2015 Wiley Periodicals, Inc.     

Enantiomeric Separation and Simulation Studies of Pheniramine,Oxybutynin, Cetirizine,and Brinzolamide Chiral Drugs on Amylose‐Based Columns

Solid phase extraction ( SPE)‐chiral separation of the important drugs pheniramine, oxybutynin, cetirizine, and brinzolamide was achieved on the C₁₈ cartridge and AmyCoat (150 x 46 mm) and Chiralpak AD (25 cm x 0.46 cm id) chiral columns in human plasma. Pheniramine, oxybutynin, cetirizine, and brinzolamide were resolved using n‐hexane‐2‐PrOH‐DEA (85:15:0.1, v/v), n‐hexane‐2‐PrOH‐DEA (80:20:0.1, v/v), n‐hexane‐2‐PrOH‐DEA (70:30:0.2, v/v), and n‐hexane‐2‐propanol (90:10, v/v) as mobile phases. The separation was carried out at 25 ± 1 ºC temperature with detection at 225 nm for cetirizine and oxybutynin and 220 nm for pheniramine and brinzolamide. The flow rates of the mobile phases were 0.5 mLmin^‐1. The retention factors of pheniramine, oxybutynin, cetirizine and brinzolamide were 3.25 and 4.34, 4.76 and 5.64, 6.10 and 6.60, and 1.64 and 2.01, respectively. The separation factors of these drugs were 1.33, 1.18, 1.09 and 1.20 while their resolutions factors were 1.09, 1.45, 1.63 and 1.25, and 1.15, respectively. The absolute configurations of the eluted enantiomers of the reported drugs were determined by simulation studies. It was observed that the order of enantiomers elution of the reported drugs was S‐pheniramine > R‐pheniramine; R‐oxybutynin > S‐oxybutynin; S‐cetirizine > R‐cetirizine; and S‐brinzolamide > R‐brinzolamide. The mechanism of separation was also determined at the supramolecular level by considering interactions and modeling results. The reported SPE‐chiral high‐performance liquid chromatography ( HPLC) methods are suitable for the enantiomeric analyses of these drugs in any biological sample. In addition, simulation studies may be used to determine the absolute configuration of the first and second eluted enantiomers. Chirality 26:136–143, 2014. © 2014 Wiley Periodicals, Inc.     

Highly sensitive and selective electrochemiluminescence determination of cholesterol utilizing a functional electrode with a core–shell nanostructure

A highly sensitive and selective method for the determination of cholesterol is required to evaluate trace amounts of cholesterol in test samples. In this work, selected gold nanoparticles (AuNPs) and 5‐amino‐2‐mercapto‐1,3,4‐thiadiazole (AMT) were used and a thin film of three‐dimensional gold–AMT core–shell nanoparticles (p‐AMT–AuNPs) was prepared using an electrochemical method. Cholesterol oxidase was then bonded to the film surface to give a functional electrode. Based on catalysis by the electrode functionalized for cholesterol and a luminol–H₂O₂ electrochemiluminescence (ECL) system, a highly sensitive and selective ECL method was developed for the determination of cholesterol. Under optimized conditions, ECL intensity showed a good linear relationship with cholesterol over the concentration range 0.05–11.0 µg/ml, with a correlation coefficient of 0.999 and a limit of detection of 0.02 µg/ml. The proposed method was used to determine cholesterol in dairy products with a relative standard deviation of < 1.8% and recovery rates of 98.1–104%. Copyright © 2015 John Wiley & Sons, Ltd.     

Supramolecular interaction of 18‐crown‐6 ether with mesalazine and spectrofluorimetric determination of mesalazine in pharmaceutical formulations

The supramolecular interaction of protonated mesalazine (MSZ) and 18‐crown‐6 ether (18C6) has been examined by Ultraviolet–visible, FT‐IR and fluorescence spectroscopy. The formation of the inclusion complex has been confirmed based on the changes of the spectral properties. The MSZ–18C6 host–guest complex formed in (1:1) stoichiometry and the inclusion constant (K = 1.411 × 10² L mol^–1) was ascertained by the typical double reciprocal plots. Furthermore, the thermodynamic parameters (ΔG°, ΔH° and ΔS°) of (MSZ‐18C6) were obtained. Based on the remarkable enhancement of the fluorescence intensity of MSZ produced through complex formation, a simple, accurate, rapid and highly sensitive spectrofluorometric method for the determination of MSZ in aqueous solution in the presence of 18C6 was developed. The measurement of relative fluorescence intensity was carried with excitation at 298 nm, emission 410 nm. All variables affecting the reactions were studied and optimized. Beer's law was obeyed in the concentration range of 0.1–0.9 µg/mL. The absorbance was found to increase linearly with increasing concentration of MSZ. The molar absorptivity, Sandell sensitivity, limit of detection (LOD) and limit of quantification (LOQ) were calculated. The validity of the described method was assessed, and the method was successfully applied to the determination of MSZ in its pharmaceutical formulation. In addition, a solid inclusion complex was synthesized by the coprecipitation method. Copyright © 2015 John Wiley & Sons, Ltd.