Ion mobility spectrometry in liquid analysis

Ion mobility spectrometry (IMS) is a straightforward, low cost method for fast and sensitive determination of organic and inorganic analytes. Originally this portable technique was applied to the determination of gas phase compounds in security and military use. Nowadays, IMS has received increasing attention in environmental and biological analysis, and in food quality determination. This thesis consists of literature review of suitable sample preparation and introduction methods for liquid matrices applicable to IMS from its early development stages to date. Thermal desorption, solid phase microextraction (SPME) and membrane extraction were examined in experimental investigations of hazardous aquatic pollutants and potential pollutants. Also the effect of different natural waters on the extraction efficiency was studied, and the utilised IMS data processing methods are discussed. Parameters such as extraction and desorption temperatures, extraction time, SPME fibre depth, SPME fibre type and salt addition were examined for the studied sample preparation and introduction methods. The observed critical parameters were extracting material and temperature. The extraction methods showed time and cost effectiveness because sampling could be performed in single step procedures and from different natural water matrices within a few minutes. Based on these experimental and theoretical studies, the most suitable method to test in the automated monitoring system is membrane extraction. In future an IMS based early warning system for monitoring water pollutants could ensure the safe supply of drinking water. IMS can also be utilised for monitoring natural waters in cases of environmental leakage or chemical accidents. When combined with sophisticated sample introduction methods, IMS possesses the potential for both on-line and on-site identification of analytes in different water matrices.

Design, construction, and performance of a real-time holographic refractometry prototype for liquid analysis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Hybrid materials for biomedical applications

The increased longevity of humans and the demand for a better quality of life have led to a continuous search for new implant materials. Scientific development coupled with a growing multidisciplinarity between materials science and life sciences has given rise to new approaches such as regenerative medicine and tissue engineering. The search for a material with mechanical properties close to those of human bone produced a new family of hybrid materials that take advantage of the synergy between inorganic silica (SiO4) domains, based on sol-gel bioactive glass compositions, and organic polydimethylsiloxane, PDMS ((CH3)2.SiO2)n, domains. Several studies have shown that hybrid materials based on the system PDMS-SiO2 constitute a promising group of biomaterials with several potential applications from bone tissue regeneration to brain tissue recovery, passing by bioactive coatings and drug delivery systems. The objective of the present work was to prepare hybrid materials for biomedical applications based on the PDMS-SiO2 system and to achieve a better understanding of the relationship among the sol-gel processing conditions, the chemical structures, the microstructure and the macroscopic properties. For that, different characterization techniques were used: Fourier transform infrared spectrometry, liquid and solid state nuclear magnetic resonance techniques, X-ray diffraction, small-angle X-ray scattering, smallangle neutron scattering, surface area analysis by Brunauer–Emmett–Teller method, scanning electron microscopy and transmission electron microscopy. Surface roughness and wettability were analyzed by 3D optical profilometry and by contact angle measurements respectively. Bioactivity was evaluated in vitro by immersion of the materials in Kokubos’s simulated body fluid and posterior surface analysis by different techniques as well as supernatant liquid analysis by inductively coupled plasma spectroscopy. Biocompatibility was assessed using MG63 osteoblastic cells. PDMS-SiO2-CaO materials were first prepared using nitrate as a calcium source. To avoid the presence of nitrate residues in the final product due to its potential toxicity, a heat-treatment step (above 400 °C) is required. In order to enhance the thermal stability of the materials subjected to high temperatures titanium was added to the hybrid system, and a material containing calcium, with no traces of nitrate and the preservation of a significant amount of methyl groups was successfully obtained. The difficulty in eliminating all nitrates from bulk PDMS-SiO2-CaO samples obtained by sol-gel synthesis and subsequent heat-treatment created a new goal which was the search for alternative sources of calcium. New calcium sources were evaluated in order to substitute the nitrate and calcium acetate was chosen due to its good solubility in water. Preparation solgel protocols were tested and homogeneous monolithic samples were obtained. Besides their ability to improve the bioactivity, titanium and zirconium influence the structural and microstructural features of the SiO2-TiO2 and SiO2-ZrO2 binary systems, and also of the PDMS-TiO2 and PDMS-ZrO2 systems. Detailed studies with different sol-gel conditions allowed the understanding of the roles of titanium and zirconium as additives in the PDMS-SiO2 system. It was concluded that titanium and zirconium influence the kinetics of the sol-gel process due to their different alkoxide reactivity leading to hybrid xerogels with dissimilar characteristics and morphologies. Titanium isopropoxide, less reactive than zirconium propoxide, was chosen as source of titanium, used as an additive to the system PDMS-SiO2-CaO. Two different sol-gel preparation routes were followed, using the same base composition and calcium acetate as calcium source. Different microstructures with high hydrophobicit were obtained and both proved to be biocompatible after tested with MG63 osteoblastic cells. Finally, the role of strontium (typically known in bioglasses to promote bone formation and reduce bone resorption) was studied in the PDMS-SiO2-CaOTiO2 hybrid system. A biocompatible material, tested with MG63 osteoblastic cells, was obtained with the ability to release strontium within the values reported as suitable for bone tissue regeneration.

Sequential injection analysis implementing multiple standard additions for As speciation by liquid chromatography and atomic fluorescence spectrometry (SIA-HPLC-AFS)

An analytical procedure for multiple standard additions of arsenic species using sequential injection analysis (SIA) is proposed for their quantification in seafood extracts. SIA presented flexibility for generating multiple specie standards at the ng mL(-1) concentration level by adding different volumes of As(III), As(V), monomethylarsonic (MMA) and dimethylarsinic (DMA) to the sample. The mixed sample plus standard solutions were delivered from SIA to fill the HPLC injection loop. Subsequently, As species were separated by HPLC and analyzed by atomic fluorescence spectrometry (AFS). The proposed system comprised two independently controlled modules, with the HPLC loop acting as the intermediary device. The analytical frequency was enhanced by combining the actions of both modules. While the added sample was flowing through the chromatographic column towards the detection system, the SIA program started performing the standard additions to another sample. The proposed method was applied to spoiled seafood extracts. Detection limits based on 3 sigma for As(III), As(V), MMA and DMA were 0.023, 0.39, 0.45 and 1.0 ng mL(-1), respectively. (C) 2011 Elsevier B.V. All rights reserved.

Liquid-phase microextraction combined with high-performance liquid chromatography for the enantioselective analysis of mefloquine in plasma samples

A simple and rapid method, which involves liquid-phase microextraction (LPME) followed by HPLC analysis using Chiralpak AD column and UV detection, was developed for the enantioselective determination of mefloquine in plasma samples. Several factors that influence the efficiency of three-phase LPME were investigated and optimized. Under the optimal extraction conditions, the mean recoveries were 33.2 and 35.0% for (-)-(SR-)-mefloquine and (+)-(RS)-mefloquine, respectively. The method was linear over 50-1500 ng/ml range. Within-day and between-day assay precision and accuracy were below 15% for both enantiomers at concentrations of 150, 600 and 1200 ng/ml. Furthermore, no racemization or degradation were seen with the method described. (C) 2007 Elsevier B.V. All rights reserved.

Enantioselective analysis of zopiclone and its metabolites in plasma by liquid chromatography/tandem mass spectrometry

A high-performance liquid chromatographic method with triple-quadrupole mass spectrometry detection (LC-MS-MS) was developed and validated for the first time for the simultaneous quantification of zopiclone and its metabolites in rat plasma samples. The analytes were isolated from rat plasma by liquid-liquid extraction and separated using a chiral stationary phase based on an amylose derivative, Chiralpak ADR-H column, and ethanol-methanol-acetonitrile (50:45:5, v/v/v) plus 0.025% diethylamine as the mobile phase, at a flow-rate of 1.0 mL min(-1). Moclobemide was used as the internal standard. The developed method was linear over the concentration range of 7.5-500 ng mL(-1). The mean absolute recoveries were 74.6 and 75.7; 61.6 and 56.9; 72.5, and 70.7 for zopiclone enantiomers, for N-desmethyl zopiclone enantiomers and for zopiclone-N-oxide enantiomers, respectively, and 75.9 for the internal standard. Precision and accuracy were within acceptable levels of confidence (<15%). The method application in a pilot study of zopiclone kinetic disposition in rats showed that the levels of (+)-(S)-zopiclone were always higher than those of (-)-R-zopiclone. Higher concentrations were also observed for (+)-(S)-N-desmethyl zopiclone and (+)-(S)-N-oxide zopiclone, confirming the stereoselective disposition of zopiclone.

Enantioselective analysis of mirtazapine and its two major metabolites in human plasma by liquid chromatography-mass spectrometry after three-phase liquid-phase microextraction

A three-phase liquid-phase microextraction (LPME) method using porous polypropylene hollow fibre membrane with a sealed end was developed for the extraction of mirtazapine (MRT) and its two major metabolites, 8-hydroxymirtazapine (8-OHM) and demethylmirtazapine (DMR), from human plasma. The analytes were extracted from 1.0 mL of plasma, previously diluted and alkalinized with 3.0 mL 0.5 mol L-1 pH 8 phosphate buffer solution and supplemented with 15% sodium chloride (NaCl), using n-hexyl ether as organic solvent and 0.01 moL L-1 acetic acid solution as the acceptor phase. Haloperidol was used as internal standard. The chromatographic analyses were carried out on a chiral column, using acetonitrile-methanol-ethanol (98:1:1, v/v/v) plus 0.2% diethylamine as mobile phase, at a flow rate of 1.0 mL min(-1). Multi-reaction monitoring (MRM) detection was performed by mass spectrometry (MS-MS) using a triple-stage quadrupole and electrospray ionization interface operating in the positive ion mode. The mean recoveries were in 18.3-45.5% range with linear responses over the 1.25-125 ng mL(-1) concentration range for all enantiomers evaluated. The quantification limit (LOQ) was 1.25 ng mL(-1). Within-day and between-day assay precision and accuracy (2.5, 50 and 100 ng mL(-1)) showed relative standard deviation and the relative error lower than 11.9% for all enantiomers evaluated. Finally, the method was successfully used for the determination of mirtazapine and its metabolite enantiomers in plasma samples obtained after single drug administration of mirtazapine to a healthy volunteer. (c) 2007 Elsevier B.V. All rights reserved.

Hollow fiber-based liquid-phase microextraction (HF-LPME) of isradipine and its main metabolite followed by chiral HPLC analysis: application to an in vitro biotransformation study

An enantioselective liquid chromatographic method using two-phase hollow fiber liquid-phase microextraction (HF-LPME-HPLC) was developed for the determination of isradipine (ISR) enantiomers and its main metabolite (pyridine derivative of isradipine, PDI) in microsomal fractions isolated from rat liver. The analytes were extracted from 1 mL of microsomal medium using a two-phase HF-LPME procedure with hexyl acetate as the acceptor phase, 30 min of extraction, and sample agitation at 1,500 rpm. For the first time, ISR enantiomers and PDI were resolved. For this separation, a ChiralpakA (R) AD column with hexane/2-propanol/ethanol (94:04:02, v/v/v) as the mobile phase at a flow rate of 1.5 mL min(-1) was used. The column was kept at 23 A +/- 2 A degrees C. The drug and metabolite detection was performed at 325 nm and the internal standard oxybutynin was detected at 225 nm. The recoveries were 23% for PDI and 19% for each ISR enantiomer. The method presented quantification limits (LOQ) of 50 ng mL(-1) and was linear over the concentration range of 50-5,000 and 50-2,500 ng mL(-1) for PDI and each ISR enantiomer, respectively. The validated method was employed to an in vitro biotransformation study of ISR using rat liver microsomal fraction showing that (+)-(S)-ISR is preferentially biotransformed.

Determination of beta-artemether and its main metabolite dihydroartemisinin in plasma employing liquid-phase microextraction prior to liquid chromatographic-tandem mass spectrometric analysis

A method for the determination of artemether (ART) and its main metabolite dihydroartemisinin (DHA) in plasma employing liquid-phase microextraction (LPME) for sample preparation prior to liquid chromatography-tandem mass spectrometry (LC-MS-MS) was developed. The analytes were extracted from 1 nil, of plasma utilizing a two-phase LPME procedure with artemisinin as internal standard. Using the optimized LPME conditions, mean absolute recovery rates of 25 and 32% for DHA and ART, respectively, were achieved using toluene-n-octanol (1:1, viv) as organic phase with an extraction time of 30 min. After extraction, the analytes were resolved within 5 min using a mobile phase consisting of methanol-ammonium acetate (10 mmol L(-1) pH 5.0, 80:20. v/v) on a laboratory-made column based on poly(methyltetradecylsiloxane) attached to a zirconized-silica support. MS-MS detection was employed using an electrospray interface in the positive ion mode. The method developed was linear over the range of 5-1000 ng mL(-1) for both analytes. Precision and accuracy were within acceptable levels of confidence (<15%). The assay was applied to the determination of these analytes in plasma from rats treated with ART. The two-phase LPME procedure is affordable and the solvent consumption was very low compared to the traditional methods of sample preparation. (C) 2010 Elsevier B.V. All rights reserved.

Two-step liquid-phase microextraction and high-performance liquid chromatography for the simultaneous analysis of the enantiomers of mefloquine and its main metabolite carboxymefloquine in plasma

A method for the simultaneous analysis of the enantiomers of mefloquine (MQ) and its main metabolite carboxymefloquine (CMQ) in plasma is described for the first time. The assay involves two-step liquid-phase micro-extraction (LPME) and enantioselective high-performance liquid chromatography. In the first LPME step, the enantiomers of MQ were extracted from an alkalinized sample through a thin layer of di-n-hexyl ether immobilized in the pores of the hollow fiber and into 0.01 M perchloric acid as acceptor solution. In the second LPME step, the same sample was acidified to enable the extraction of CMQ using the same organic solvent and 0.05 M sodium hydroxide as acceptor phase. The analytes were resolved on a Chirobiotic T column in the polar-organic mode of elution and detected at 285 nm. The recovery rates from 1 mL of plasma were in the range 35-38%. The method presented limits of quantification of 50 ng/mL for all analytes and was linear up to 1,500 and 3,000 ng/mL for the enantiomers of MQ and CMQ, respectively. The plasmatic concentrations of (+)-(RS)-MQ were higher than those of (-)-(SR)-MQ after oral administration of the racemic drug to rats.

Simultaneous Analysis of Tramadol, O-Desmethyltramadol, and N-Desmethyltramadol Enantiomers in Rat Plasma by High-Performance Liquid Chromatography-Tandem Mass Spectrometry: Application to Pharmacokinetics

Tramadol (T) is available as a racemic mixture of (+)-trans-T and (-)-trans-T. The main metabolic pathways are O-demethylation and N-demethylation, producing trans-O-desmethyltramadol (M1) and trans-N-desmethyltramadol (M2) enantiomers, respectively. The analgesic effect of T is related to the opioid activity of (+)-trans-T and (+)-M1 and to the monoaminergic action of (+/-)-trans-T. This is the first study using tandem mass spectrometry as a detection system for the simultaneous analysis of trans-T, M1, and M2 enantiomers. The analytes were resolved on a Chiralpak (R) AD column using hexane: ethanol (95.5:4.5, v/v) plus 0.1% diethylamine as the mobile phase. The quantitation limits were 0.5 ng/ml for trans-T and M1 and 0.1 ng/ml for M2. The method developed and validated here was applied to a pharmacokinetic study in rats. Male Wistar rats (n = 6 at each time point) received a single oral dose of 20 mg/kg racemic trans-T. Blood samples were collected up to 12 h after drug administration. The kinetic disposition of trans-T and M2 was enantioselective (AUC((+)/(-)) ratio = 4.16 and 6.36, respectively). The direction and extent of enantioselectivity in the pharmacokinetics of trans-T and M2 in rats were comparable to data previously reported for healthy volunteers, suggesting that rats are a suitable model for enantioselective studies of trans-T pharmacokinetics. Chirality 23: 287-293, 2011. (C) 2010 Wiley-Liss, Inc.

Analysis of Liquid Crystalline Nanoparticles by Small Angle X-Ray Diffraction: Evaluation of Drug and Pharmaceutical Additives Influence on the Internal Structure

The goal of this work was to study the liquid crystalline structure of a nanodispersion delivery system intended to be used in photodynamic therapy after loading with photosensitizers (PSs) and additives such as preservatives and thickening polymers. Polarized light microscopy and light scattering were performed on a standard nanodispersion in order to determine the anisotropy of the liquid crystalline structure and the mean diameter of the nanoparticles, respectively. Small angle X-ray diffraction (SAXRD) was used to verify the influence of drug loading and additives on the liquid crystalline structure of the nanodispersions. The samples, before and after the addition of PSs and additives, were stable over 90 days, as verified by dynamic light scattering. SAXRD revealed that despite the alteration observed in some of the samples analyzed in the presence of photosensitizing drugs and additives, the hexagonal phase still remained in the crystalline phase. (C) 2011 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 100: 2849-2857, 2011

Solid-phase extraction method with high-performance liquid chromatography and electrochemical detection for the quantitative analysis of oxycodone in human plasma

A sensitive and reproducible solid-phase extraction (SPE) method for the quantification of oxycodone in human plasma was developed. Varian Certify SPE cartridges containing both C-8 and benzoic acid functional groups were the most suitable for the extraction of oxycodone and codeine (internal standard), with consistently high (greater than or equal to 80%) and reproducible recoveries. The elution mobile phase consisted of 1.2 ml of butyl chloride-isopropanol (80:20, v/v) containing 2% ammonia. The quantification limit for oxycodone was 5.3 pmol on-column. Within-day and inter-day coefficients of variation were 1.2% and 6.8% respectively for 284 nM oxycodone and 9.5% and 6.2% respectively for 28.4 nM oxycodone using 0.5-ml plasma aliquots. (C) 1998 Elsevier Science BN. All rights reserved.

Solid-phase microextraction using poly(pyrrole) film and liquid chromatography with UV detection for analysis of antidepressants in plasma samples

Poly(pyrrole) (PPY) coating was prepared on a stainless-steel (SS) wire for solid-phase microextraction (SPME) by electrochemical deposition (cyclic voltammetric). The PPY was evaluated by analyzing new-generation antidepressants (mirtazapine, citalopram, paroxetine, duloxetine, fluoxetine, and sertraline) in plasma sample by SPME and liquid chromatography with UV detection (LC-UV). The effect of electrolyte Solution (lithium perchlorate or tetrabutylammonium perchlorate) and the number of cycles (50, 100 or 200) applied during the polymerization process on the SPME performance was evaluated. Important factors in the optimization of SPME efficiency such as extraction time, temperature, pH, influence of plasma proteins on sorption mechanisms, and desorption conditions are discussed. The SPME-PPY/LC method showed to be linear in concentrations ranging from the limit of quantification (LOQ) to 1200 ng mL(-1). The LOQ values range from 16 to 25 ng mL-1. The inter-day precision of the SPME-PPY/LC method presented coefficient of variation (CV) lower than 15%. Based on analytical validation results, the SPME-PPY/LC methodology showed to be adequate for antidepressant analysis, from therapeutic to toxic levels. In order to evaluate the proposed method for clinical use, the SPME-PPY/LC method was applied to the analysis of plasma samples from elderly depressed patients. (c) 2009 Elsevier B.V. All rights reserved,