Assessment of sulfonamide contamination in aquatic environments: A first report for Argentina and environmental risk assessment.

The global surge in pharmaceutical consumption, driven by increasing population and the demand for animal proteins, leads to the discharge of diverse pollutants, including antibiotic residues, into water bodies. Sulfonamides, being water-soluble compounds, can readily enter surface run-off, posing potential risks to non-target species despite their low environmental concentrations. Latin America has implemented intensive production systems highly dependent on antimicrobials for productivity and animal health, yet there is a paucity of information regarding their concentration in the region. The objective of this study was to evaluate the presence of sulfonamides in water and sediment samples and assess their potential ecological risks through an environmental risk assessment. The Río de la Plata basin collects the waters of the Paraguay, Paraná, and Uruguay rivers, together with their tributaries and various wetlands, passing through the provinces in Argentina known for their significant animal husbandry production. Two sampling campaigns were carried out for sediment, while only one campaign was conducted for surface waters. The samples were analyzed by high performance liquid chromatography tandem mass-spectrometry (HPLC-MS/MS). None of the examined sulfonamide antibiotics were detected in the sediment samples from both sampling campaigns. In contrast, sulfadiazine (95 %), sulfamethoxazole (91 %), and sulfathiazole (73 %) were detected in the water samples. Sulfadiazine was found in the concentration range of 8 to 128 ng/L, while sulfamethoxazole and sulfathiazole were observed at concentrations ranging from 3.0 to 32.5 ng/L and 2.9 to 8.1 ng/L, respectively. Based on the environmental risk assessment conducted using the sulfonamide concentrations, most samples indicated a medium risk for aquatic biota, with only one sample surpassing the high-risk threshold. This study represents the first report presenting data on the presence of sulfonamide antibiotics in the aquatic environment of Argentina.

Preconcentration of sulphonamides in bovine milk by the cloud point extraction method using smartphone-based digital images.

Sulphonamides are a group of synthetic antibiotics used specially in veterinary medicine. Among the procedures employed in the sample preparation for sulphonamide determination are liquid-liquid extraction (LLE) and solid-phase extraction (SPE) that use large volumes of organic solvents. Hence, a clean procedure was developed based on preconcentration and cloud point extraction (CPE) without using organic solvents to quantify total sulphonamides in bovine milk. The procedure was optimized as follows: 2 mL of pre-cleaned milk sample, 2 mL of reagent solution and 1 mL of Triton X-114 7% (m/v) were added to a tube, heated in a water bath at 40 °C for 10 minutes and centrifuged at 2950 rcf for 20 minutes. Digital image acquisition was employed directly at the tube without removing the supernatant/aqueous phase. The linear response was observed between 10 and 400 µg L-1 of total sulphonamides and described by the following equation: S = 2.50 + 0.0514C (µg L-1) and R = 0.999. The LOD and the CV (n = 11) were estimated to be 10 µg L-1 and 1.3%, respectively. The main interferents present at their usual concentrations in the sample did not interfere with the results. Spike and recovery tests of total sulphonamides were carried out in UHT and pasteurized milk with recovery values between 73 and 106% and the results obtained for this kind of sample were in agreement with those achieved by a high performance liquid chromatography (HPLC) procedure at the 95% confidence level. The analytical procedure presents an adequate sensitivity to determine total sulphonamides in bovine milk and does not require organic solvents, being aligned to the principles of green chemistry.

Hydrophilic interaction liquid chromatography coupled to quadrupole time-of-flight mass spectrometry as a potential combination for the determination of sulfonamide residues in complex infant formula matrices.

An accurate, sensitive and selective analytical method is proposed for sulfonamide residues analysis in infant formulas based on hydrophilic interaction liquid chromatography (HILIC) and quadrupole time-of-flight mass spectrometry in full scan mode. The sample preparation approach involves low-temperature lipid precipitation followed by dispersive solid-phase extraction with PSA and C18 sorbents, which was successfully optimized using Plackett-Burman design. In order to achieve high analytical sensitivity, the influence of HILIC conditions on sulfonamide ionization was investigated, such as the mobile phase composition, buffer concentration, and sample diluent for injection. The method performance characteristics, including linearity (range 5-120 µg kg-1), reliable limits of quantification (between 5 and 20 µg kg-1), recovery (72.9-109.2%) and precision (coefficient of variation values ≤ 19.8%) under repeatability and within-laboratory reproducibility conditions, were in accordance with the Codex Alimentarius Commission CAC/GL 71-2009 for quantitative analytical methods for veterinary drug residues in foods. Moreover, adequate identification of the compounds was provided with accurate mass measurement of both precursor and fragment ions in one single run. Finally, the developed method was applied to thirty-five powdered milk-based infant formula samples available in the Brazilian market.

Rapid monitoring of pesticides in tomatoes (Solanum lycopersicum L.) during pre-harvest intervals by paper spray ionization mass spectrometry.

A rapid analysis of acephate, chlorpyrifos, and cyazofamid in tomato peels during pre-harvest intervals using paper spray ionization mass spectrometry (PSI-MS) has been demonstrated. LODs of 0.01 ppm and LOQs of 0.03 ppm were achieved. Relative standard deviations were below 9%, and recoveries close to 100%. For pesticides monitoring, samples were separated into stored and field groups. Stored group contained fruits that were harvested and stored at low temperature during the pre-harvest interval, and the field group contained fruits that were on the agricultural field over the pre-harvest interval. The decrease in pesticide amounts was lower for the stored samples. At the end of the pre-harvest intervals, acephate, chlorpyrifos, and cyazofamid were determined at 0.86, 0.96, and 0.23 ppm in the stored group, and at 0.26, 0.37, and 0.09 ppm in the field group, respectively. This work demonstrates the utility of PSI-MS for performing rapid quality control of fruit harvesting.

Sistema microbiológico de cinco bioensayos para la detección de residuos de antibióticos / Five-assay microbiological system for the screeningf antibiotic residues

Abstract A novel microbiological system in microtiter plates consisting of five bioassays is presented for the detection and classification of antibiotic residues in milk. The bioassays were optimized for the detection of beta-lactams (Bioassay B: Geobacillus stearothermophilus), macrolides (Bioassay M: Bacillus megaterium with fusidic acid), tetracyclines (Bioassay T: B. megaterium with chloramphenicol), quinolones (Bioassay Q: Bacillus licheniformis) and sulfamides (Bioassay QS: B. licheniformis with trimethoprim) at levels near the maximum residue limits (MRL). The response of each bioassay was interpreted visually (positive or negative) after 4-5.5h of incubation. The system detects and classifies beta-lactams (5 pg/l of amoxicillin, 4 pg/l of ampicillin, 36 pg/l of cloxacillin, 22 pg/l of amoxicillin, 3 pg/l of penicillin, 114 pg/l of cephalexin, 89pg/l of cefoperazone and 116 pg/l of ceftiofur), tetracyclines (98 pg/l of chlortetracycline, 92 pg/l of oxytetracycline and 88 pg/l of tetracycline), macrolides (33 pg/l of erythromycin, 44 pg/l of tilmicosin and 50 pg/l of tylosin), sulfonamides (76 pg/l of sulfadiazine, 85 pg/l of sulfadimethoxine, 77 pg/l of sulfamethoxazole and 87pg/l of sulfathiazole) and quinolones (94 pg/l of ciprofloxacin, 98 pg/l of enrofloxacin and 79 pg/l marbofloxacin). In addition, the specificity values were high for B, T, Q (99.4%), M (98.8%) and QS (98.1%) bioassays. The control of antibiotics through this system can contribute to improving the quality and safety of dairy products.

Resumen Se presenta un novedoso sistema microbiológico en placas de microtitulación compuesto por 5 bioensayos para la detección y clasificación de residuos de antibióticos en leche. Los bioensayos fueron optimizados para la detección de betalactámicos (bioensayo B: Geobacillus stearothermophilus), macrólidos (bioensayo M: Bacillus megaterium con ácido fusídico), tetraciclinas (bioensayo T: Bacillus megaterium con cloranfenicol), quinolonas (bioensayo Q: Bacillus licheniformis) y sulfamidas (bioensayo QS: Bacillus licheniformis con trimetoprima), a niveles cercanos a los límites máximos de residuos (LMR). La respuesta de cada bioensayo se interpretó visualmente (positiva o negativa) después de 4 a 5,5 h de incubación. El sistema detecta y clasifica betalactámicos (5 pg/l de amoxicilina, 4 pg/l de ampicilina, 36 pg/l de cloxacilina, 22 pg/l de amoxicilina, 3 pg/l de penicilina, 114 pg/l de cefalexina, 89 pg/l de cefoperazona y 116 pg/l de ceftiofur), tetraciclinas (98 pg/l de clortetraciclina, 92 pg/l de oxitetraciclina y 88 pg/l de tetraciclina), macrólidos (33 pg/l de eritromicina, 44 pg/l de tilmi-cosina y 50 pg/l de tilosina), sulfamidas (76 pg/l de sulfadiacina, 85 pg/l de sulfadimetoxina, 77 pg/l de sulfametoxazol y 87 pg/l de sulfatiazol) y quinolonas (94 pg/l de ciprofloxacina, 98 pg/l de enrofloxacina y 79pg/l de marbofloxacina). Además, los valores de especificidad fueron altos para los bioensayos B, T, Q (99,4%), M (98,8%) y QS (98,1%). El control de residuos de antibióticos mediante este sistema puede contribuir a mejorar la calidad e inocuidad de los productos lácteos.

Robotic-assisted dynamic large drop microextraction.

By proper design of an innovative extraction device, a lab-made multipurpose autosampler was exploited in the automated performance of the dynamic large drops based microextraction. The pluses of this new analytical strategy were demonstrated in the determination of sulfonamides and fluoroquinolones in surface water samples, by direct immersion single drop microextraction (SDME) and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) analysis. Operational autosampler features and critical experimental factors influencing SDME, including the extraction mode (static or dynamic), extraction, stirring rate, salt addition, drop size, number of cycles and drop exposition time, were comprehensively investigated using both univariate and multivariate optimization. The lab-made autosampler allowed to performance challenging dynamic and static large drop based SDMEs in an automated and effortless way and with minimal requirements of hardware and software. Large stable drops provided high surface area, enhancing the phase ratio and in consequence increasing the analytes uptake. The best extraction efficiencies were obtained as a result of the synergic interaction between the use of large drops and the automated dynamic mode of extraction. The developed method proved to be a reliable, sensitive, and robust analytical tool, with intraday RSDs ranging between 4.0 and 7.6% (n = 6), and interday RSDs between 4.8 and 9.3% (n = 6), and, LOD and LOQ in the range of 15-50 and 35-100 ng L-1, respectively.

Behavioral response and dynamics of Eisenia fetida hemocytes exposed to environmentally relevant concentration of sulfentrazone.

Although the toxicity of the pesticide sulfentrazone in some aquatic organisms is known, its effects on edaphic organisms such as earthworms remain completely unknown. Thus, we aimed at evaluating the behavior and immune response of Eisenia fetida exposed to sulfentrazone at environmentally relevant concentrations (EC). E. fetida representatives exposed to this contaminant (for 48 h) were divided in the following groups: environmental concentration (EC1x: 318 ng sulfentrazone/g of dry weight soil) and EC100x (concentration 100 times higher than in EC1x). Based on the avoidance test results, earthworms responded to this pesticide and proved the toxicity of sulfentrazone. The observed immune response induction was expressed by increased granulocytes presenting phagocytic vacuoles and agglomerations/encapsulations, mainly in animals belonging to groups EC1x and EC100x. However, the reduced frequency of plasmocytes in these animals' hemolymphs suggested that the phagocytic immune response was not efficient to assure 100% survival. Our study is the first to report sulfentrazone toxicity in an edaphic organism, at environmental concentration.

An LC-ESI-MS/MS method for residues of fluoroquinolones, sulfonamides, tetracyclines and trimethoprim in feedingstuffs: validation and surveillance.

An increasing concern about food safety has been observed over the years. The presence of drugs residues in food is one of the major subjects of research in food safety. Feedingstuffs can be responsible for carryover into the food chain of residues of several drugs. This paper describes the development, validation and application of a fast and simple method for analysis of 24 antibiotic residues in feedingstuffs for cattle, pigs and poultry. Analytes include compounds from different antimicrobials classes, such as sulfonamides (sulfadiazine, sulfamethazine, sulfamethoxazole, sulfaquinoxaline, sulfachlorpyridazine, sulfadoxine, sulfadimethoxine, sulfizoxazole, sulfamerazine and sulfathiazole), fluoroquinolones (ciprofloxacin, enrofloxacin, norfloxacin, danofloxacin, difloxacin, sarafloxacin, flumequine, nalidixic acid and oxolinic acid), tetracyclines (tetracycline, doxycycline, oxytetracycline and chlortetracycline) and trimethoprim. Samples were extracted with methanol:water (70:30) 0.1% formic acid, followed by clean-up steps using centrifugation, low-temperature purification (LTP) and ultracentrifugation. Instrumental analysis was performed using liquid chromatography coupled to tandem mass spectrometry. Chromatographic separation was achieved using a C18 column and a mobile phase composed of acetonitrile and water, both with 0.1% formic acid. Validation parameters such as limit of detection (LOD), limit of quantification (LOQ), selectivity, linearity, accuracy, precision, decision limit (CCα) and detection capability (CCß) were determined and meet the adopted criteria. LOD and LOQ were set to 30 and 75 µg kg-1, respectively. Inter-day precision were in the range from 4.0 to 11.1%, and linearity provides values of r2 above 0.95 for all analytes. The optimised method was applied to the analysis of more than 1500 real samples within the period 2012-2017. Non-compliant results were discussed and classified in terms of analytes, feed types and target species. Multivariate analysis of the data was performed using principal component analysis.

Sulfluramid use in Brazilian agriculture: A source of per- and polyfluoroalkyl substances (PFASs) to the environment.

N-Ethyl perfluorooctane sulfonamide (EtFOSA) is a perfluorooctane sulfonate (PFOS) precursor and the active ingredient in sulfluramid, a pesticide which is used extensively in Brazil for management of leaf cutting ants. Here we investigate the occurrence of EtFOSA, PFOS, and other per- and polyfluoroalkyl substances (PFASs) in soil, eucalyptus leaves, water (ground, riverine, and coastal (estuarine/marine)) and coastal sediment from an agricultural region of Bahia State, Brazil. This area contains a larger number of eucalyptus plantations where sulfluramid is suspected to be applied. Soil, leaves, and coastal water (marine/estuarine) contained ∑PFAS concentrations of up to 5400 pg g-1, 979 pg g-1, and 1020 pg L-1, respectively, with PFAS profiles generally dominated by PFOS and perfluorooctane sulfonamide (FOSA). Coastal sediment contained ∑PFAS concentrations of up to 198 pg g-1, with PFOS, FOSA, and perfluorooctanoic acid (PFOA) being the most frequently observed PFASs. These substances are all potential EtFOSA transformation products, pointing to sulfluramid as a possible source. In riverine water, ∑PFAS concentrations of up to 8930 pg L-1 were observed. PFOS and PFOA were detected in all river water samples. Groundwater also exhibited PFAS contamination (5730 pg L-1 ∑PFASs), likely from sulfluramid use. The observation of other PFASs (e.g. perfluorobutanoic acid) in freshwater suggests that other PFAS sources (in addition to sulfluramid) may be important in this region. Overall, these data support the hypothesis that sulfluramid use contributes to the occurrence of PFASs in the Brazilian environment.

Determination of sulfachloropyridazine residue levels in feathers from broiler chickens after oral administration using liquid chromatography coupled to tandem mass spectrometry.

Several antimicrobials are routinely used by the poultry farming industry on their daily operations, however, researchers have found for some antimicrobials that their residues persist for longer periods in feathers than they do in edible tissues, and at higher concentrations, as well. But this information is not known for other classes of antimicrobials, such as the sulfonamides. Therefore, this work presents an accurate and reliable analytical method for the detection of sulfachloropyridazine (SCP) in feathers and edible tissues from broiler chickens. This method was also validated in-house and then used to study the depletion of sulfachloropyridazine in those matrices. The experimental group comprised 54 broiler chickens, who were raised under controlled conditions and then treated with a commercial formulation of 10% sulfachloropyridazine for 5 days. Samples were analyzed via LC-MS/MS, using 13C6-sulfamethazine (SMZ-13C6) as an internal standard. Aromatic sulfonic acid solid phase extraction (SPE) cartridges were used to clean up the samples. The Limit of Detection (LOD) for this method was set at 10 µg kg-1 on feathers and liver; and at 5 µg kg-1 on muscle. Within the range of 10-100 µg kg-1, the calibration curves for all matrices presented a determination coefficient greater than 0.96. Our results show, with a 95% confidence level, that sulfachloropyridazine persisted in feathers for up to 55 days after ceasing treatment, and its concentrations were higher than in edible tissues. In consequence, to avoid re-entry of antimicrobial residues into the food-chain, we recommend monitoring and inspecting animal diets that contain feather derivatives, such as feathers meals, because they could be sourced from birds that might have been medicated with sulfachloropyridazine.

Evaluation of ionic liquids supported on silica as a sorbent for fully automated online solid-phase extraction with LC-MS determination of sulfonamides in bovine milk samples.

Sulfonamides are antibiotics widely used in the treatment of diseases in dairy cattle. However, their indiscriminate use for disease control may lead to their presence in tissues and milk and their determination requires a sample preparation step as part of an analytical approach. Among the several sample preparation techniques available, those based upon the use of sorptive materials have been widely employed. Recently, the application of ionic liquids immobilized on silica surfaces or polymeric materials has been evaluated for such an application. This manuscript addresses the evaluation of silica-based ionic liquid obtained by a sol-gel synthesis process by basic catalysis as sorbent for online solid-phase extraction with liquid chromatography and electrospray ionization time-of-flight mass spectrometry for sulfonamides determination. Infrared vibrational spectroscopy confirmed the presence of the ionic liquid on the silica surface, suggesting that the ionic liquid was anchored on to the silica surface. Other sorbents varying the ionic liquid alkyl chain were also synthesized and evaluated by off-line solid-phase extraction in the sulfonamide extraction. As the length of the alkyl chain increased, the amount of extracted sulfonamides decreased, possibly due to a decrease in the electrostatic interaction caused by the reduction in the polarity, as well as the presence of a hexafluorophosphate anion that increases the hydrophobic character of the material. The use of 1-butyl-3-methylimidazolium hexafluorophosphate as a selective ionic liquid sorbent enabled the isolation and sulfonamide preconcentration in bovine milk by online solid-phase extraction with liquid chromatography and electrospray ionization time-of-flight mass spectrometry. The limit of quantification for the method developed was 5-7, 5 µg/mL, with extraction recoveries ranging between 74 and 93% and intra- and interassay between 1.5-12.5 and 2.3-13.1, respectively.

Direct Determination of Nimesulide in Natural Waters and Wastewater by Cathodic Stripping Voltammetry.

Pharmaceuticals are included in a group of compounds considered as contaminants of emerging concern (CECs) in environmental matrices. The quantification of CECs is typically accomplished using chromatographic methods that require several sample pretreatment steps. The current study proposes a voltammetric method to quantify directly nimesulide (NIM) in aqueous samples. The voltammetric parameters were optimized and chosen based on the profile of the voltammograms and peak current intensity. The adequacy of the analytical method was evaluated using validation criteria, such as accuracy, selectivity, linearity, detection, and quantification limits. Linearity was assessed by a standard addition curve at a concentration range of 0.5-130 µg L-1 of NIM. The limit of quantification was 0.50 µg L-1, with 60 s of preconcentration time. Accuracy was expressed as recovery percentages of NIM and ranged from 111.4 to 119.8%. The voltammetric method herein proposed has several advantages over others already used to determine NIM, such as fewer sample pretreatment steps, faster, and cheaper analyses. NIM was detected in wastewater samples at concentration levels ranging from 101.7 to 385.0 µg L-1. This result is the first evidence about NIM occurrences in environmental matrices in the area surrounding the Itaipu Lake reservoir in Brazil.

Monitoring of chlorsulfuron in biological fluids and water samples by molecular fluorescence using rhodamine B as fluorophore.

A new simple methodology is proposed for chlorsufuron (CS) traces quantification based upon enhancement of rhodamine B (RhB) fluorescent signal. Experimental variables that influence fluorimetric sensitivity have been studied and optimized. The zeroth order regression calibration was linear from 0.866 to 35.800µgL(-1) CS, with a correlation coefficient of 0.99. At optimal experimental conditions, a limit of detection of 0.259µgL(-1) and a limit of quantification of 0.866µgL(-1) were obtained. The method showed good sensitivity and adequate selectivity and was applied to the determination of trace amounts of CS in plasma, serum and water samples with satisfactory results analyzed by ANOVA test. The proposed methodology represents an alternative to traditional chromatographic techniques for CS monitoring in complex samples, using an accessible instrument in control laboratories.

On-line solid phase extraction-ultra high performance liquid chromatography-tandem mass spectrometry as a powerful technique for the determination of sulfonamide residues in soils.

Sulfonamides are antimicrobials used widely as veterinary drugs, and their residues have been detected in environmental matrices. An analytical method for determining sulfadiazine, sulfathiazole, sulfamethazine, sulfamethoxazole, sulfadimethoxine and sulfaquinoxaline residues in soils employing a solid phase extraction on-line technique coupled with ultra-high performance liquid chromatography and tandem mass spectrometry (SPE-UHPLC-MS/MS) was developed and validated in this study. SPE and chromatographic separation were performed using an Oasis HLB column and an Acquity UPLC BEH C18 analytical column, respectively, at 40°C. Samples were prepared by extracting sulfonamides from soil using a solid-liquid extraction method with water:acetonitrile, 1:1v/v (recovery of 70.2-99.9%). The following parameters were evaluated to optimize the on-line SPE process: sorbent type (Oasis and C8), sample volume (100-400µL), loading solvent (water and different proportions of water:methanol) and washing volume (0.19-0.66mL). The method produced linear results for all sulfonamides from 0.5 to 12.5ngg(-1) with a linearity greater than 0.99. The precision of the method was less than 15%, and the matrix effect was -27% to -87%. The accuracy was in the range of 77-112% for all sulfonamides. The limit of quantitation in the two soils (clay and sand) was 0.5ngg(-1). The SPE column allowed for the analysis of many (more than 2000) samples without decreasing the efficiency.

Is Ongoing Sulfluramid Use in South America a Significant Source of Perfluorooctanesulfonate (PFOS)? Production Inventories, Environmental Fate, and Local Occurrence.

Despite international phase-out initiatives, production and use of perfluorooctanesulfonate (PFOS) and related substances continues in some countries. In Brazil, the PFOS-precursor N-ethyl perfluorooctane sulfonamide (EtFOSA) is used in Sulfluramid, a pesticide for controlling leaf-cutting ants. New data on production, environmental fate, and occurrence of Brazilian Sulfluramid are reported herein. From 2003 to 2013, Brazilian Sulfluramid manufacturing increased from 30 to 60 tonnes yr(-1) EtFOSA. During this time <1.3 tonnes yr(-1) were imported, while exports increased from ∼0.3 to 2 tonnes yr(-1). From 2004 to 2015, most EtFOSA was exported to Argentina (7.2 tonnes), Colombia (2.07 tonnes), Costa Rica (1.13 tonnes), Equador (2.16 tonnes), and Venezuela (2.4 tonnes). Within Brazil, sales occurred primarily in the states of Minas Gerais, São Paulo, Mato Grosso do Sul, Espírito Santo, and Bahia. Model simulations predict EtFOSA will partition to soils, while transformation products perfluorooctane sulfonamide (FOSA) and PFOS are sufficiently mobile to leach into surface waters. In support of these predictions, up to 3400 pg L(-1) of FOSA and up to 1100 pg L(-1) of PFOS were measured in Brazilian surface water, while EtFOSA was not detected. The high FOSA/PFOS ratio observed here (up to 14:1) is unprecedented in the scientific literature to our knowledge. Depending on the extent of conversion of EtFOSA, cumulative Brazilian Sulfluramid production and import from 2004 to 2015 may contribute between 167 and 487 tonnes of PFOS/FOSA to the environment. These levels are clearly nontrivial and of concern since production is continuing unabated.

Synthesis of some novel enzyme inhibitors and antibacterial agents derived from 5-(1-(4-tosyl)piperidin-4-yl)-1, 3, 4-oxadiazol-2-thiol

ABSTRACT Keeping in mind the pharmacological importance of the 1,3,4-oxadiazole moiety, a series of new S-substituted derivatives, 5a-h, of 5-(1-(4-tosyl)piperidin-4-yl)-1,3,4-oxadiazol-2-thiol (3) were synthesized. The reaction of p-toluenesulfonyl chloride (a) and ethyl isonipecotate (b) produced ethyl 1-(4-tosyl)piperidin-4-carboxylate (1) which was further transformed into 1-(4-tosyl)piperidin-4-carbohydrazide (2) by hydrazine hydrate in methanol. Compound 2 was refluxed with CS2 in the presence of KOH to synthesize 5-(1-(4-tosyl)piperidin-4-yl)-1,3,4-oxadiazol-2-thiol (3). The desired compounds, 5a-h, were synthesized by stirring 3 with aralkyl halides, 4a-h, in DMF using NaH as an activator. The structures of synthesized compounds were elucidated by 1H-NMR, IR and EI-MS spectral studies. These compounds were further evaluated for enzyme inhibitory activity against lipoxygenase and alpha-glucosidase, along with antibacterial activity against Gram-negative and Gram-positive bacteria.

RESUMO Tendo em vista a importância farmacológica da porção 1,3,4-oxadiazol, sintetizou-se uma série de novos derivados S-substituídos, 5a-h, de 5-(1-(4-tosi)piperidin-4-il)-1,3,4-oxadiazol-2-tiol (3). A reação do cloreto de p-toluenossulfonila (a), com isonipecotato de (b) etila forneceu 1-(4-tosil)piperidin-4-carboxilato de metila (1), que foi, em seguida, transformado em 1-(4-tosil)piperidin-4-carbo-hidrazida (2) por reação com hidrato de hidrazina em metanol. O composto 2 foi submetido a refluxo com CS2 na presença de KOH para se obter 5-(1-(4-tosil)piperidin-4-il)-1,3,4-oxadiazol-2-tiol (3). Os compostos desejados, 5a-h, foram obtidos por agitação de 3 com haletos de aralquila, 4a-h, em DMF, na presença de NaH. As estruturas dos compostos sintetizados foram elucidadas através de análise dos espectros de 1H-MNR, IR e EI-MS. Estes compostos foram, ainda, avaliados quanto à inibição das enzimas lipoxigenase e alfa-glucosidase, juntamente com a atividade antibacteriana contra bactérias Gram positivas e Gram negativas.

Determination of quinolones and fluoroquinolones, tetracyclines and sulfonamides in bovine, swine and poultry liver using LC-MS/MS.

Antibacterials are widely used in veterinary medicine. Residues of these drugs can remain in food of animal origin, including bovine liver. This paper describes a fast and simple analytical method for the determination of quinolones and fluoroquinolones, tetracyclines and sulfonamides in bovine liver samples. Deuterated enrofloxacin, sulfapyridine and demeclocycline were used as internal standards. The homogenised liver samples were extracted with acidified acetonitrile. Steps of non-solid-phase extraction (SPE) clean-up and concentration were used in the presented method. The final extracts were analysed by sensitive and selective detection of all components in a single run using LC-MS/MS. Acceptable recoveries between 66% and 110% were obtained. Good linearity (r(2)) above 0.96, considering three different days, for all drugs was achieved in concentrations ranging from 0.0 to 2.0 × the maximum residue limit (MRL). Intraday precision with coefficient of variation (CV%) (n = 6) lower than 14.7% and inter-day precision lower than 18.8% in agreement with European Commission Decision 2002/657/EC were obtained in concentrations ranging from 0.5 to 1.5 MRL. Accuracy was between 86% and 110%. Limits of detection and quantitation, as well as decision limit (CCα) and detection capability (CCß), were also evaluated.

Screening and determination of sulphonamide residues in bovine milk samples using a flow injection system.

A new, simple, rapid and sensitive flow injection spectrophotometric method was developed for the screening and determination of sulphonamides in bovine milk samples. The method is based on the condensation of sulphathiazole, sulphamethazine, and sulphadimethoxine with p-dimethylaminobenzaldehyde (p-DAB) in acid medium, in the presence of sodium dodecyl sulphate (SDS), producing a yellow compound (λmax=465 nm). Optimisation of the experimental parameters was performed using a multivariate methodology. The linear range was 90-500 µg/L and the limits of detection and quantification were in the ranges 25-29 µg/L and 84-88 µg/L, respectively. The procedure was applied for the determination of sulphonamide antibiotics in bovine milk samples submitted to a prior extraction procedure based on QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) methodology. Recoveries of 60.5-70.5% were achieved for milk samples spiked with 0.09, 0.1, 0.2, and 0.3 µg/g of each sulphonamide.

On-line hyphenation of solid-phase extraction to chromatographic separation of sulfonamides with fused-core columns in sequential injection chromatography.

On-line sample pretreatment (clean-up and analyte preconcentration) is for the first time coupled to sequential injection chromatography. The approach combines anion-exchange solid-phase extraction and the highly effective pentafluorophenylpropyl (F5) fused-core particle column for separation of eight sulfonamide antibiotics with similar structures (sulfathiazole, sulfanilamide, sulfacetamide, sulfadiazine, sulfamerazine, sulfadimidine, sulfamethoxazole and sulfadimethoxine). The stationary phase was selected after a critical comparison of the performance achieved by three fused-core reversed phase columns (Ascentis(®) Express RP-Amide, Phenyl-Hexyl, and F5) and two monolithic columns (Chromolith(®) High Resolution RP-18 and CN). Acetonitrile and acetate buffer pH 5.0 at 0.60 mL min(-1) were used as mobile phase to perform the separations before spectrophotometric detection. The first mobile phase was successfully used as eluent from SPE column ensuring transfer of a narrow zone to the chromatographic column. Enrichment factors up to 39.2 were achieved with a 500 µL sample volume. The developed procedure showed analysis time <10.5 min, resolutions >1.83 with peak symmetry ≤1.52, LODs between 4.9 and 27 µg L(-1), linear response ranges from 30.0 to 1000.0 µg L(-1) (r(2)>0.996) and RSDs of peak heights <2.9% (n=6) at a 100 µg L(-1) level and enabled the screening control of freshwater samples contaminated at the 100 µg L(-1) level. The proposed approach expanded the analytical potentiality of SIC and avoided the time-consuming batch sample pretreatment step, thus minimizing risks of sample contamination and analyte losses.

A critical review of properties of darunavir and analytical methods for its determination.

Darunavir is a synthetic non-peptidic protease inhibitor that has been shown to be extremely potent against wild-type HIV, and it is an important component of highly active antiretroviral treatment (HAART), which is considered as one of the most significant advances in the field of HIV therapy. However, there are some concerns about darunavir quality control. Darunavir shows pseudo-polymorphism: in different ambient conditions one pseudo-polymorphic form can change to another. This behavior of darunavir is problematic because the dosage form is exposed to different ambient conditions around the world, since HIV/AIDS is prevalent globally. Issues around differences in the solubility and effects that different forms of darunavir can cause are of concern, and a more stable form is preferable. Important investigations of darunavir such as dissolution behavior, polymorphism, stability and degradation studies, and the impact of that on the quality of the product are being conducted by our working group. A cure for HIV/AIDS remains a long-term commitment, and there is much yet to achieve. This article discusses, by a critical review of the literature, the impact of the use of darunavir in the treatment of HIV-infected patients, its physical-chemical properties, the analytical methods to determine it, and challenges that remain in order to ensure the quality and stability of darunavir.