On-line solid-phase extraction of pharmaceutical compounds from wastewater treatment plant samples using restricted access media in column-switching liquid chromatography-tandem mass spectrometry.

An on-line solid phase extraction using a lab-made restricted access media (RAM) was developed as sample preparation procedure for determination of the pharmaceutical compounds caffeine (CAF), carbamazepine (CBZ), norfloxacin (NOR), ciprofloxacin (CIP), fluoxetine (FLX) and venlafaxine in wastewater treatment plant samples by liquid chromatography-tandem mass spectrometry (LC-MS/MS). This method is suitable for use in routine of analysis, avoiding cross-contamination and requiring only a small sample volume (50 µL), with minimal handling. The method was validated according to international guidelines. The chromatographic efficiency was evaluated using peak resolution and asymmetry parameters. Carryover was also evaluated, in order to ensure reliability of the analysis and the ability to reuse the cartridge. Satisfactory linearity (r2 > 0.99) was obtained for all the compounds. The intra- and inter-day precision values were lower than 5.79 and 14.1%, respectively. The limits of detection ranged from 0.01 to 3 µg L-1 and the limits of quantification were from 0.1 to 5 µg L-1. The method was applied to 20 environmental wastewater samples, with caffeine being the most widely detected compound, at the highest concentration of 392 µg L-1, while other compounds were detected in fewer samples at lower concentrations (up to 9.60 µg L-1). The lab-made modification is a cheaper option for on-line sample preparation, compared to commercially available on-line SPE cartridges and RAM columns. Moreover, a high-throughput procedure was achieved, with an analysis time of 16 min including sample preparation and chromatographic separation. The same RAM column was applied over 200 injections including method optimization, validation and application in wastewater samples without loss of analytical response.

Lab-made solid phase microextraction phases for off line extraction and direct mass spectrometry analysis: Evaluating the extraction parameters.

The analyses of drugs and metabolites in complex matrices have been widely studied in recent years. However, due to high levels endogenous compounds and matrix complexity, these analyses require a sample pre-treatment step. To this aim, two lab-made extractive phases were integrated to probe electrospray ionization mass spectrometry (PESI-MS) technique for direct analysis of illicit drugs in biological fluids and phorbol esters in Jatropha curcas extract. The polypyrrole (PPy) phase was electropolymerized onto a platinum wire surface by cyclic voltammetry. The molecularly imprinted polymer (MIP) was synthesized and adhered onto a stainless-steel needle with epoxy resin. The PPy-PESI-MS method showed to be linear in a concentration range from 1 to 500 µg L-1, with accuracy values between -2.1 and 14%, and precision values between 0.8 and 10.8%. The MIP-PESI-MS method showed to be linear in a concentration range from 0.9 to 30 mg L-1, with accuracy values between -1.6 and -15.3%, and precision values between 4.1 and 13.5%.

Occurrence of PPCPs in a Brazilian water reservoir and their removal efficiency by ecological filtration.

The presence of PPCPs (Pharmaceuticals and Personal Care Products) in water sources and drinking water has concerned researchers in recent times. This study was carried out to evaluate the occurrence of 6 PPCPs (namely paracetamol, diclofenac, naproxen, ibuprofen, benzophenone-3 and methylparaben) in the Lobo reservoir, their degradation products, and how efficiently they were removed by 22 ecological filters, considering individual and mixture of compounds. There were 3 spiking events of PPCPs (2 µg L-1) in the ecological filter influents conducted with a lag period of 15 days between spikes. Water samples were collected from the influent and effluent of the filters at 3, 6 and 24 h after each spiking event. All target PPCPs were identified in the Lobo reservoir water in the range of µg L-1. The personal care products were detected with 100% frequency in the samples, and in higher concentrations compared to the pharmaceuticals. Degradation products of diclofenac and benzophenone-3 were identified in the water samples. Results of this investigation show that ecological filtration was an effective process (70-99%) to remove 2 µg L-1 of the selected PPCPs, and demonstrated that the filters were resilient to individual compounds and to their mixtures.

Removal of pharmaceutically active compounds from synthetic and real aqueous mixtures and simultaneous disinfection by supported TiO2/UV-A, H2O2/UV-A, and TiO2/H2O2/UV-A processes.

Pharmaceutically active compounds are carried into aquatic bodies along with domestic sewage, industrial and agricultural wastewater discharges. Psychotropic drugs, which can be toxic to the biota, have been detected in natural waters in different parts of the world. Conventional water treatments, such as activated sludge, do not properly remove these recalcitrant substances, so the development of processes able to eliminate these compounds becomes very important. Advanced oxidation processes are considered clean technologies, capable of achieving high rates of organic compounds degradation, and can be an efficient alternative to conventional treatments. In this study, the degradation of alprazolam, clonazepam, diazepam, lorazepam, and carbamazepine was evaluated through TiO2/UV-A, H2O2/UV-A, and TiO2/H2O2/UV-A, using sunlight and artificial irradiation. While using TiO2 in suspension, best results were found at [TiO2] = 0.1 g L-1. H2O2/UV-A displayed better results under acidic conditions, achieving from 60 to 80% of removal. When WWTP was used, degradation decreased around 50% for both processes, TiO2/UV-A and H2O2/UV-A, indicating a strong matrix effect. The combination of both processes was shown to be an adequate approach, since removal increased up to 90%. H2O2/UV-A was used for disinfecting the aqueous matrices, while mineralization was obtained by TiO2-photocatalysis.

Expanding the applicability of cork as extraction phase for disposable pipette extraction in multiresidue analysis of pharmaceuticals in urine samples.

In this paper, cork is proposed as a natural and renewable material for the extraction phase in disposable pipette extraction to be applied in the multiresidue determination of pharmaceuticals in human urine by HPLC with diode array detection. The compounds carbamazepine, losartan, ketoprofen, 17­ß­estradiol, naproxen, diazepam, 17­α­ethinylestradiol, estrone, diclofenac and ibuprofen were studied. A known amount (5 mg) of cork, with a size of 200 mesh, was inserted into a pipette tip. The method optimization was carried out with univariate and multivariate approaches. The optimized conditions were sample pH adjusted to 3, urine dilution factor of 40 (158 µL of urine diluted in 6.142 mL of ultrapure water), 9 extraction cycles each performed with 700 µL of sample, and extraction time of 10 s per cycle. Desorption was performed with 85 µL of methanol applying 6 cycles of 10 s each using the same solvent aliquot. For the clean-up step, 4 cycles were carried out each with 200 µL of methanol. The limits of quantification varied from 5 to 10 µg L-1 with determination coefficients higher than 0.9919 for the calibration curves for all the analytes. Intra-day and inter-day precision and relative recovery from urine samples donated by two volunteers were assessed based on three spiked concentrations. The analyte relative recoveries ranged from 65 to 117% (n = 3) for the two samples. Intra-day precision ranged from 1.2 to 17% (n = 3) and inter-day precision varied from 11 to 21% (n = 9).

Functional polyaniline/multiwalled carbon nanotube composite as an efficient adsorbent material for removing pharmaceuticals from aqueous media.

The composite polyaniline/multiwalled carbon nanotube (PAni/MWCNT, 1:0.1 w/w) was developed with the intention of binding the adsorbent properties of two materials and using it to adsorb pharmaceuticals from aqueous media. PAni/MWCNT was characterized by scanning electron microscopy, thermogravimetry, infrared spectroscopy, pH at the point of zero charge, and the effect on the surface wettability of the material. As proof of concept, adsorption studies were carried out using meloxicam (MLX) as the pharmaceutical and it was evaluated as a function of pH, temperature, ionic strength, contact time and variation in concentration. Kinetics and isothermal models were applied to evaluate the mechanism of the adsorption process. The best MLX adsorption result was at pH 2 with 6 min of contact with PAni/MWCNT. The kinetics models that fitted the experimental data were pseudo-second order and Elovich and the kinetics model was the dual-site Langmuir-Freundlich. Both models suggest that the adsorption occurs by the chemical nature of the surface and in the pores of the energetically heterogeneous composite. The PAni/MWCNT presented an adsorption capacity of 221.2 mg g-1, a very good value when compared with the literature and can be used to remove pharmaceuticals from aqueous environments.

Enantiomeric Mixtures in Natural Product Chemistry: Separation and Absolute Configuration Assignment.

Chiral natural product molecules are generally assumed to be biosynthesized in an enantiomerically pure or enriched fashion. Nevertheless, a significant amount of racemates or enantiomerically enriched mixtures has been reported from natural sources. This number is estimated to be even larger since the enantiomeric purity of secondary metabolites is rarely checked in the natural product isolation pipeline. This latter fact may have drastic effects on the evaluation of the biological activity of chiral natural products. A second bottleneck is the determination of their absolute configurations. Despite the widespread use of optical rotation and electronic circular dichroism, most of the stereochemical assignments are based on empirical correlations with similar compounds reported in the literature. As an alternative, the combination of vibrational circular dichroism and quantum chemical calculations has emerged as a powerful and reliable tool for both conformational and configurational analysis of natural products, even for those lacking UV-Vis chromophores. In this review, we aim to provide the reader with a critical overview of the occurrence of enantiomeric mixtures of secondary metabolites in nature as well the best practices for their detection, enantioselective separation using liquid chromatography, and determination of absolute configuration by means of vibrational circular dichroism and density functional theory calculations.

A Simple Method for the Simultaneous Determination of Pharmaceuticals and Personal Care Products in River Sediment by Ultrasound-Assisted Extraction Followed by Solid-Phase Microextraction Coupled with Gas Chromatography-Mass Spectrometry.

A simple method was developed using ultrasound-assisted extraction (UAE) combined with solid-phase microextraction (SPME) coupled with gas chromatography-mass spectrometry (GC-MS) for the simultaneous determination of eight different pharmaceuticals and personal care products (PPCPs) (ibuprofen, 2-benzyl-4-chlorophenol, naproxen, triclosan, ketoprofen, diclofenac, bisphenol A and estrone) in river sediment. UAE conditions were optimized involving extraction variables such as extraction solvent, extraction time, sample amount, extraction temperature, pH and salt addition. A 100 mg of sediment was extracted by optimized UAE process using 7 mL deionized water (pH 3) + 1% methanol as solvent, room temperature and 1 min extraction at 70% of amplitude. A 5 mL of supernatant was subsequently extracted by SPME; the extracted analytes were derivatized on fiber in head-space mode with N-methyl-N-(tertbutyldimethylsilyl) trifluoroacetamide and then analyzed by GC-MS. The developed method was evaluated by testing: precision (CV < 8%), linearity (R  > 0.98), recoveries (56-108%), limits of detection (<0.25 ng/g) and quantitation (<0.8 ng/g). Finally, the method was successfully applied to sediment samples collected from Tula River, Hidalgo, Mexico (one of the most polluted river) in which naproxen and triclosan were identified at ng/g concentration levels. Quantitative results were compared with Tula River water samples, showing PPCPs distribution rate between water and sediment. The method is robust, simple and environmentally friendly, and provides straightforward analyses of these trace organic pollutants in sediment samples.

Adsorption of pharmaceuticals in water through lignocellulosic fibers synergism.

The contamination of water from disposal of drugs is an emerging problem due to their consequences on trophic webs. This study evaluated the ability of sugarcane and coconut fiber to reduce water toxicity contaminated by pharmaceuticals. The toxicity of solutions containing pharmaceuticals was studied by bioassay using Allium cepa, before and after filtration of contaminated water. The coconut and sugarcane fiber have not been satisfactory in reducing toxicity when tested separately. Despite no induction of chromosomal aberrations, our study found a reduction of the mitotic index. The mixture of fibers showed better results providing total reduction of toxicity, in addition to maintenance in the mitotic index and induction of chromosome aberrations. The interaction between fibers and drugs was confirmed by Thermogravimetry and Differential Thermal Analyses (TG/DTA) which presented differences in profile between the fibers before and after adsorption. The mixture of coconut and sugarcane proved viable for reduction of toxicity in contaminated water by a mixture of pharmaceuticals.

Multi-residue method for determination of 58 pesticides, pharmaceuticals and personal care products in water using solvent demulsification dispersive liquid-liquid microextraction combined with liquid chromatography-tandem mass spectrometry.

A rapid and efficient sample pretreatment using solvent-based de-emulsification dispersive liquid-liquid microextraction (SD-DLLME) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) was studied for the extraction of 58 pharmaceuticals and personal care products (PPCPs) and pesticides from water samples. Type and volume of extraction and disperser solvents, pH, salt addition, amount of salt and type of demulsification solvent were evaluated. Limits of quantification (LOQ) in the range from 0.0125 to 1.25 µg L(-1) were reached, and linearity was in the range from the LOQ of each compound to 25 µg L(-1). Recoveries ranged from 60% to 120% for 84% of the compounds, with relative standard deviations lower than 29%. The proposed method demonstrated, for the first time, that sample preparation by SD-DLLME with determination by LC-MS/MS can be successfully used for the simultaneous extraction of 32 pesticides and 26 PPCPs from water samples. The entire procedure, including the extraction of 58 organic compounds from the aqueous sample solution and the breaking up of the emulsion after extraction with water, rather than with an organic solvent, was environmentally friendly. In addition, this technique was less expensive and faster than traditional techniques. Finally, the analytical method under study was successfully applied to the analysis of all 58 pesticides and PPCPs in surface water samples.

Study of vortex-assisted MSPD and LC-MS/MS using alternative solid supports for pharmaceutical extraction from marketed fish.

A procedure based on vortex-assisted matrix solid-phase dispersion (MSPD) for the extraction of 15 pharmaceuticals from fish samples with determination by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was validated. Florisil, C18, diatomaceous earth, chitin, and chitosan were evaluated as solid supports. Best results were obtained with 0.5 g of diatomaceous earth, 0.5 g of sodium sulfate, and 5 mL of methanol. Analytical recoveries ranged from 58 to 128 % with relative standard deviation (RSD) lower than 15 %. Limit of quantification (LOQ) values for the 15 compounds ranged from 5 to 1000 ng g(-1). The method under investigation has shown to be a simple and fast extraction tool with minimum instrumentation and low amount of reagent, resulting in method low cost. Besides, alternative materials, such as chitin and chitosan, which were applied to the dispersion step for the first time, were found to be interesting alternatives.

Novel 3-hydroxypropyl-bonded phase by direct hydrosilylation of allyl alcohol on amorphous hydride silica.

A novel 3-hydroxypropyl (propanol)-bonded silica phase has been prepared by hydrosilylation of allyl alcohol on a hydride silica intermediate, in the presence of platinum (0)-divinyltetramethyldisiloxane (Karstedt's catalyst). The regio-selectivity of this synthetic approach had been correctly predicted by previous reports involving octakis(dimethylsiloxy)octasilsesquioxane (Q8 M8 (H) ) and hydrogen silsesquioxane (T8 H8 ), as molecular analogs of hydride amorphous silica. Thus, C-silylation predominated (∼94%) over O-silylation, and high surface coverages of propanol groups (5 ± 1 µmol/m(2) ) were typically obtained in this work. The propanol-bonded phase was characterized by spectroscopic (infrared (IR) and solid-state NMR on silica microparticles), contact angle (on fused-silica wafers) and CE (on fused-silica tubes) techniques. CE studies of the migration behavior of pyridine, caffeine, Tris(2,2'-bipyridine)Ru(II) chloride and lysozyme on propanol-modified capillaries were carried out. The adsorption properties of these select silanol-sensitive solutes were compared to those on the unmodified and hydride-modified tubes. It was found that hydrolysis of the SiH species underlying the immobilized propanol moieties leads mainly to strong ion-exchange-based interactions with the basic solutes at pH 4, particularly with lysozyme. Interestingly, and in agreement with water contact angle and electroosmotic mobility figures, the silanol-probe interactions on the buffer-exposed (hydrolyzed) hydride surface are quite different from those of the original unmodified tube.

Evaluation of the QuEChERS method for the extraction of pharmaceuticals and personal care products from drinking-water treatment sludge with determination by UPLC-ESI-MS/MS.

A modified version of the QuEChERS method has been evaluated for the determination of 21 pharmaceuticals and 6 personal care products (PPCPs) in drinking-water sludge samples by employing ultra high liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The performance of the method was evaluated through linearity, recovery, precision (intra-day), method detection and quantification limits (MDL and MQL) and matrix effect. The calibration curves prepared in acetonitrile and in the matrix extract showed a correlation coefficient ranging from 0.98 to 0.99. MQLs values were on the ng g(-1) order of magnitude for most compounds. Recoveries between 50% and 93% were reached with RSDs lower than 10% for most compounds. Matrix effect was almost absent with values lower than 16% for 93% of the compounds. By coupling a quick and simple extraction called QuEChERS with the UPLC-MS/MS analysis, a method that is both selective and sensitive was obtained. This methodology was successfully applied to real samples and caffeine and benzophenone-3 were detected in ng g(-1) levels.

Establishment of a design space for biopharmaceutical purification processes using DoE.

Recent trends in the pharmaceutical sector are changing the way protein purification processes are designed and executed, moving from operating the process in a fixed point to allowing a permissible region in the operating space known as design space. This trend is driving product development to design quality into the manufacturing process (Quality by Design) and not to rely exclusively on testing quality in the product. A typical purification step has numerous operating parameters that can impact its performance. Therefore, optimization and robustness analysis in purification processes can be time-consuming since they are mainly grounded on experimental work. A valuable approach consists in the combination of an adequate risk analysis technique for selecting the relevant factors influencing process performance and the design of experiment methodology. The latter allows for many process variables which can be studied at the same time; thus, the number of tests will be reduced in comparison with the conventional approach based on trial and error. These multivariate studies permit a detailed exploration in the experimental range and lay the foundation of Quality by Design principles application. This article outlines a recommended sequence of activities toward the establishment of an expanded design space for a purification process.

Nano-liquid chromatography in pharmaceutical and biomedical research.

Miniaturized separation techniques have emerged as environmentally friendly alternatives to available separation methods. Nano-liquid chromatography (nano-LC), microchip devices and nano-capillary electrophoresis are miniaturized methods that minimize reagent consumption and waste generation. Furthermore, the low levels of analytes, especially in biological samples, promote the search for more highly sensitive techniques; coupled to mass spectrometry, nano-LC has great potential to become an indispensable tool for routine analysis of biomolecules. This short review presents the fundamental aspects of nano-LC analytical instrumentation, discussing practical considerations and the primary differences between miniaturized and conventional instrumentation. Some theoretical aspects are discussed to better explain both the potential and the principal limitations of nano-LC. Recent pharmaceutical and biomedical applications of this separation technique are also presented to indicate the satisfactory performance for complex matrices, especially for proteomic analysis, that is obtained with nano-LC.

Selective silica-based sorbent materials synthesized by molecular imprinting for adsorption of pharmaceuticals in aqueous matrices.

The presence of pharmaceuticals in aqueous environmental matrices often requires efficient and selective preconcentration procedures. Thus, silicas (SILs) were synthesized by a molecular imprinting technique using an acid-catalyzed sol-gel process and the following drugs as templates: fluoxetine, gentamicin, lidocaine, morphine, nifedipine, paracetamol, and tetracycline. The materials were subjected to sorbent extraction assisted by ultrasonic treatment to remove the drugs and the consequent formation of molecular imprinted cavities. The surface area of the resulting materials ranged from 290 to 960 m(2)/g. Adsorption tests were performed with the molecular imprinting phases. In terms of the potential selectivity, the SILs were subjected to the adsorption of drugs from samples such as potable and surface water. The adsorption capacity remained in the range between 55 and 65% for both matrices, while for the nonimprinted SIL it remained between 15 and 20%.

Determination of some organic contaminants in water samples by solid-phase extraction and liquid chromatography-tandem mass spectrometry.

Independent methods for determination of organic contaminants such as pharmaceuticals and pesticides in drinking water samples, using SPE as the extraction technique and LC-MS/MS in the MRM mode with electrospray ionization, were developed and validated. Different SPE sorbents were evaluated, including lab-made fluorinated and phenyl and commercial Oasis HLB and C18, with the commercial phases being more suitable for the target compounds. Recoveries in the range of 70-120% were obtained for all target compounds, with the exception for paracetamol (acetaminophen), and precision values (inter-day and intra-day), expressed in terms of relative standard deviations (RSD), lower than 20% were obtained for all target compounds. Quantification limits were in the range of 0.006-0.208 µg L(-1) and the methods developed were successfully applied for the analysis of drinking water samples, detecting some pharmaceuticals and pesticides, but at concentration levels lower than the MRL.

Monitoring of phenolic compounds for the quality control of Melissa officinalis products by capillary electrophoresis.

INTRODUCTION: Official assays for the quality control of Melissa officinalis L. (Lamiaceae) leaves establish the quantification of total hydroxycinnamic derivatives expressed as rosmarinic acid. OBJECTIVE: The goal of this work was to develop a simple, fast and reliable method for monitoring the phenolic composition in herbs from the Lamiaceae family and for rapidly detecting M. officinalis adulteration or substitution in commercial medicinal samples in Argentina. METHODOLOGY: A capillary zone electrophoresis (CZE) method was performed under the following conditions: the background electrolyte (BGE) consisted of 20 m m sodium tetraborate buffer, pH 9.2; the applied voltage was 25 kV; the capillary and sample temperatures were kept at 25 °C; the hydrodynamic mode was selected for the sample injection (3.45 kPa during 5 s). RESULTS: A CZE method that achieved the separation and simultaneous determination of eight related phenolic compounds in less than 11 min was optimised for application to control quality analysis of M. officinalis-based products. The method was validated according to the US Federal Drug Agency requirements and offers advantages in terms of analysis time, cost and operation. CONCLUSIONS: The proposed methodology can be applied to the standardisation and quality control of plant material and phytopharmaceutical products derived from the Lamiaceae family, as indicated by the results obtained in the analysis of commercial medicinal products in Argentina.

Biocompatible in-tube solid phase microextraction coupled with liquid chromatography-fluorescence detection for determination of interferon α in plasma samples.

The present work demonstrates the successful application of automated biocompatible in-tube solid-phase microextraction coupled with liquid chromatography (in-tube SPME/LC) for determination of interferon alpha(2a) (IFN α(2a)) in plasma samples for therapeutic drug monitoring. A restricted access material (RAM, protein-coated silica) was employed for preparation of a lab-made biocompatible in-tube SPME capillary that enables the direct injection of biological fluids as well as the simultaneous exclusion of macromolecules by chemical diffusion barrier and drug pre-concentration. The in-tube SPME variables, such as sample volume, draw/eject volume, number of draw-eject cycles, and desorption mode were optimized, to improve the sensitivity of the proposed method. The IFN α(2a) analyses in plasma sample were carried out within 25min (sample preparation and LC analyses). The response of the proposed method was linear over a dynamic range, from 0.06 to 3.0MIUmL(-1), with correlation coefficient equal to 0.998. The interday precision of the method presented coefficient of variation lower than 8%. The proposed automated method has adequate analytical sensitivity and selectivity for determination of IFN α(2a) in plasma samples for therapeutic drug monitoring.

Challenges and rewards of research in marine natural products chemistry in Brazil.

Brazil is blessed with a great biodiversity, which constitutes one of the most important sources of biologically active compounds, even if it has been largely underexplored. As is the case of the Amazon and Atlantic rainforests, the Brazilian marine fauna remains practically unexplored in the search for new biologically active natural products. Considering that marine organisms have been shown to be one of the most promising sources of new bioactive compounds for the treatment of different human diseases, the 8000 km of the Brazilian coastline represents a great potential for finding new pharmacologically active secondary metabolites. This review presents the status of marine natural products chemistry in Brazil, including results reported by different research groups with emphasis on the isolation, structure elucidation, and evaluation of biological activities of natural products isolated from sponges, ascidians, octocorals, and Opistobranch mollusks. A brief overview of the first Brazilian program on the isolation of marine bacteria and fungi, directed toward the production of biologically active compounds, is also discussed. The current multidisciplinary collaborative program under development at the Universidade de São Paulo proposes to establish a new paradigm toward the management of the Brazilian marine biodiversity, integrating research on the species diversity, ecology, taxonomy, and biogeography of marine invertebrates and microorganisms. This program also includes a broad screening program of Brazilian marine bioresources, to search for active compounds that may be of interest for the development of new drug leads.