Use of Sugar Cane Bagasse as Solid Extraction Phase Sorbent to Analyze Hormones from Industrial Effluent.

Sugar cane bagasse (SCB) is an abundant byproduct of sugar and bioethanol mills. It has been effectively used as a low-cost biosorbent to remove hazardous chemical compounds from a variety of effluent sources. Herein, we report on the preparation of SCB and its use as a solid phase extraction (SPE) sorbent to retain synthetic hormones (ethinylestradiol, drospirenone, and levonorgestrel) from industrial pharmaceutical plant effluent samples prior to LC-MS/MS quantitative analysis. We evaluated the reproducibility and recoveries and accuracy data analyses were compared with that of commercial SPE (cSPE) cartridges. The results from the evaluated parameters indicated that the SCB bed had an efficiency of >99%, comparable to that of cSPE cartridges, demonstrating the applicability and feasibility of this material as an effective and green chemistry alternative, as well as its biosorbent potential to remove hormones from industrial pharmaceutical effluent.

Prospective Prediction of Dapaconazole Clinical Drug-Drug Interactions Using an In Vitro to In Vivo Extrapolation Equation and PBPK Modeling.

This study predicted dapaconazole clinical drug−drug interactions (DDIs) over the main Cytochrome P450 (CYP) isoenzymes using static (in vitro to in vivo extrapolation equation, IVIVE) and dynamic (PBPK model) approaches. The in vitro inhibition of main CYP450 isoenzymes by dapaconazole in a human liver microsome incubation medium was evaluated. A dapaconazole PBPK model (Simcyp version 20) in dogs was developed and qualified using observed data and was scaled up for humans. Static and dynamic models to predict DDIs following current FDA guidelines were applied. The in vitro dapaconazole inhibition was observed for all isoforms investigated, including CYP1A2 (IC50 of 3.68 µM), CYP2A6 (20.7 µM), 2C8 (104.1 µM), 2C9 (0.22 µM), 2C19 (0.05 µM), 2D6 (0.87 µM), and 3A4 (0.008−0.03 µM). The dynamic (PBPK) and static DDI mechanistic model-based analyses suggest that dapaconazole is a weak inhibitor (AUCR > 1.25 and <2) of CYP1A2 and CYP2C9, a moderate inhibitor (AUCR > 2 and <5) of CYP2C8 and CYP2D6, and a strong inhibitor (AUCR ≥ 5) of CYP2C19 and CYP3A, considering a clinical scenario. The results presented may be a useful guide for future in vivo and clinical dapaconazole studies.

Mussels get higher: A study on the occurrence of cocaine and benzoylecgonine in seawater, sediment and mussels from a subtropical ecosystem (Santos Bay, Brazil).

Data on the occurrence of cocaine (COC) and benzoylecgonine (BE) in marine environmental compartments are still limited, with few studies reporting superficial water contamination, mainly in tropical zones. In this sense, environmental data of these substances are essential to identify potential polluting sources, as well as their impact in costal ecosystems. The aim of this study was to evaluate the occurrence of COC and BE in seawater, sediment and mussels from a subtropical coastal zone (Santos Bay, São Paulo, Brazil), as well as to determine a field measured Bioaccumulation Factor (BAF). COC and BE were detected in all water samples in concentrations ranging from 1.91 ng·L-1 to 12.52 ng·L-1 and 9.88 ng·L-1 to 28.53 ng·L-1, respectively. In sediments, only COC was quantified in concentrations ranging from 0.94 ng·g-1 to 46.85 ng·g-1. Similarly, only COC was detected in tissues of mussels 0.914 µg·kg-1 to 4.58 µg·kg-1 (ww). The field-measured BAF ranged from 163 to 1454 (L·kg-1). Our results pointed out a widespread contamination by cocaine and its main human metabolite benzoylecgonine in Santos Bay. Mussels were able to accumulate COC in areas used by residents and tourists for bathing, fishing, and harvest, denoting concern to human health. Therefore, our data can be considered a preliminary assessment, which indicates the need to evaluate drugs (including illicit as COC) in environmental and seafood monitoring programs, in order to understand their risks on the ecosystem and human health.

Detection of anti-cancer drugs and metabolites in the effluents from a large Brazilian cancer hospital and an evaluation of ecotoxicology.

The use of chemotherapy agents has been growing worldwide, due to the increase number of cancer cases. In several countries, mainly in Europe countries, these drugs have been detected in hospitals and municipal wastewaters. In Brazil this issue is poorly explored. The main goal of this study was to assess the presence of three anti-cancer drugs, 5-fluorouracil (5-FU), gemcitabine (GEM) and cyclophosphamide (CP), and two metabolites, alpha-fluoro-beta-alanine (3-NH2-F) and 2'-deoxy-2',2'-difluorouridine (2-DOH-DiF), in effluents from a large cancer hospital, in the municipal wastewater treatment plant (WWTP) influent and effluent, and also to evaluate toxicity of the mixtures of these compounds by ecotoxicological testing in zebrafish. The sample collections were performed in Barretos Cancer Hospital of the large cancer center in Brazil. After each collection, the samples were filtered for subsequent Liquid Chromatography Mass Spectrometry analysis. The presence of CP, GEM, and both metabolites (3-NH2-F and 2-DOH-DiF) were detected in the hospital wastewater and the WWTP influent. Three drugs, GEM, 2-DOH-DiF and CP, were detected in the WWTP effluent. Two drugs were detected below the limit of quantification, 2-DOH-DiF:

Determination of Organochlorines in Soil of a Suburban Area of São Paulo Brazil.

Technological advances have promoted improvements in several science fields, especially related to environmental and analytical areas with the improvement of detection and development of environmentally friendly extraction techniques. This study applied Quick, Easy, Cheap, Effective, Rugged and Safe method (QuEChERS) for soil extraction and assessed its performance through a validation study using samples from the soil of a contaminated area in Caieiras, SP, Brazil. Nine organochlorine pesticides, including the isomers alpha, beta, gamma and delta- hexachlorocyclohexane; cis- and trans-heptachlor epoxide; cis- and trans-chlordane and heptachlor were analyzed by gas chromatography coupled to electron capture detector. The method was validated according to ISO 5725-4 (2020), EURACHEM (2014) and DOQ-CGCRE-008 (2016). The limits of detection and quantification of the method for the nine organochlorines were α-HCH (1.2 and 12.6 µg kg-1), ß-HCH (1.7 and 12.0 µg kg-1), γ-HCH (1.5 and 11.6 µg kg-1), δ-HCH (0.8 and 11.6 µg kg-1), heptachlor (1.0 and 10.8 µg kg-1), cis-heptachlor epoxide (0.9 and 11.5 µg kg-1), trans-heptachlor epoxide (0.9 and 11.5 µg kg-1), cis-chlordane (0.4 and 7.9 µg kg-1) and trans-chlordane (0.5 and 10.9 µg kg-1), respectively, and all of them were within the maximum limits recommended by the EPA for the compounds α-HCH (86.0 and 360.0 µg kg-1), ß-HCH (300.0 and 1.3 × 103 µg kg-1), γ-HCH (570.0 and 2.5 × 103 µg kg-1), δ-HCH (not defined), heptachlor (130.0 and 630.0 µg kg-1), cis-/trans-heptachlor epoxide (7.0 and 330.0 µg kg-1), cis-/trans-chlordane (1.77 × 103 and 7.7 × 103 µg kg-1) in residential and industrial soil, respectively. Recovery results were between 65% and 105% for almost all compounds, which is an optimum result for multi-residue analytical methods, considering the complexity of the matrix used in the study. Caieiras presented contamination levels of α-HCH in the range of 2.0 to 66.0 µg g-1, which was higher than the limits established by EPA, corresponding to 0.077 µg g-1 for residential soil and 0.27 µg g-1 for industrial soil. According to the validation study, the analytical method proposed was reliable for organochlorine quantification, and the QuEChERS was considered efficient for organochlorine extraction from soil.

Extending matrix-assisted laser desorption/ionization triple quadrupole mass spectrometry enzyme screening assays to targets with small molecule substrates.

Mass spectrometry (MS)-based high-throughput screening (HTS) has tremendous potential as an alternative to current screening methods due to its speed, sensitivity, reproducibility and label-free readout. We recently reported that a new generation matrix-assisted laser desorption/ionization triple quadrupole (MALDI-QqQ) mass spectrometer is ideally suited for a variety of enzyme assays and screening protocols. However, all the targets measured to date had peptide substrates that were easily monitored by selected ion monitoring (SIM) without interference from the MALDI matrix. To further extend the application to enzymes with small molecule, non-peptide substrates, we evaluated this method for measuring enzyme activity and inhibition of acetylcholinesterase (AChE). Due to the potential of MALDI matrix interference, multiple reaction monitoring (MRM) was investigated for selective MS/MS transitions and to accurately measure the conversion of acetylcholine into choline. Importantly, ionization, detection and MRM transition efficiency differences between the substrate and product can be overcome by pre-balancing the MRM transitions during method development, thus allowing for a direct readout of the enzyme activity using the ratio of the substrate and product signals. Further validation of the assay showed accurate concentration-dependent inhibition measurements of AChE with several known inhibitors. Finally, a small library of 1008 drug-like compounds was screened at a single dose (10 microM) and the top 10 inhibitors from this primary screen were validated in a secondary screen to determine the rank order of inhibitory potency for each compound. Collectively, these data demonstrate that a MALDI-QqQMS-based readout platform is amenable to measuring small molecule substrates and products and offers significant advantages over current HTS methods in terms of speed, sensitivity, reproducibility and reagent costs.

Seasonal monitoring of cocaine and benzoylecgonine in a subtropical coastal zone (Santos Bay, Brazil).

Illicit drugs and their metabolites represent a new class of emerging contaminants. These substances are continuously discharged into wastewater which have been detected in the aquatic environment in concentrations ranging from ng.L-1 to µg.L-1. Our study detected the occurrence of cocaine (COC) and benzoylecgonine (BE) in a subtropical coastal zone (Santos Bay, SP, Brazil) within one year. Water samples (surface and bottom) were collected from the Santos Submarine Sewage Outfall (SSOS) area. COC and BE were measured in the samples using ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-ESI-MS/MS). Concentrations ranged from 12.18 to 203.6 ng.L-1 (COC) and 8.20 to 38.59 ng.L-1 (BE). Higher concentrations of COC were observed during the end of spring, following the population increase at summer season. COC and its metabolite occurrence in this coastal zone represent a threat to coastal organisms.

Detoxification, oxidative stress, and cytogenotoxicity of crack cocaine in the brown mussel Perna perna.

The presence of cocaine and its metabolites and by-products has been identified in different aquatic matrices, making crack cocaine the target of recent studies. The aim of this study was to evaluate the sublethal effects of crack on the brown mussel Perna perna. Mussels were exposed to three concentrations of crack cocaine (0.5, 5.0, and 50.0 µg L-1) for 168 h. Gills, digestive glands, and hemolymph were extracted and analyzed after three different exposure times using a suite of biomarkers (EROD, DBF, GST, GPX, LPO, DNA damage, ChE, and lysosomal membrane stability [LMS]). After 48 and 96 h of exposure, EROD, DBF, GST, GPX activities and DNA strand breaks in the gills increased significantly after 48 and 96 h of exposure. Alterations in LMS were also observed in the mussels exposed to all crack concentrations after 96 and 168 h. Our results demonstrated that crack cocaine is metabolized by CYP-like and GST activities in the gills. GPX was not able to prevent primary genetic damage, and cytotoxic effects in the hemocytes were also observed in a dose- and time-dependent response. Our study shows that the introduction of illicit drugs into coastal ecosystems must be considered a threat to marine organisms.

A tiered approach to assess effects of diclofenac on the brown mussel Perna perna: A contribution to characterize the hazard.

Pharmaceutical discharges into the aquatic ecosystem are of environmental concern and sewage treatment plants (STPs) have been pointed out as the major source of these compounds to coastal zones, where oceanic disposal of sewage occurs through submarine outfalls. Diclofenac (DCF) is one of the most frequently detected pharmaceuticals in water, but little is known about the effects on marine organisms. In this study, we employed a tiered approach involving the determination of environmental concentrations of DCF in marine water and the adverse biological effects for fertilization, embryo-larval development and biomarker responses of the mussel Perna perna. Results indicate that effects in fertilization rate and embryo-larval development were found in the order of mg·L-1. However, low concentrations of DCF (ng·L-1) significantly decreased the lysosomal membrane stability and COX activity, as well as triggered DNA damage, oxidative stress and changes in antioxidant defenses. Our results point to an environmental hazard at coastal ecosystems and suggest the need for improvements in the treatment of domestic wastewater aiming to reduce DCF concentrations, as well as regulation on current environmental legislation and monitoring of aquatic ecosystems.

An alternative and fast method for determination of glyphosate and aminomethylphosphonic acid (AMPA) residues in soybean using liquid chromatography coupled with tandem mass spectrometry.

A simple and specific method using reversed-phase liquid chromatography coupled with electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) was investigated, which allowed the determination of residues of glyphosate and its metabolite, aminomethylphosphonic acid (AMPA), in soybean samples. An aqueous extraction with liquid-liquid partition followed by protein precipitation was performed before the LC/MS/MS determination. The quantitation of glyphosate and AMPA was performed in positive and negative ESI mode, respectively, using the multiple reaction monitoring (MRM) mode with three transitions for each analyte to enhance the specificity of the method and avoid false positives. The methodology reported in this work is capable of detecting residues of glyphosate and AMPA in soybean samples with limits of quantification of 0.30 and 0.34 mg kg(-1), respectively. This alternative method has throughput advantages such as simpler sample preparation and faster chromatographic analysis.

An electrospray ionization tandem mass spectrometric study of p-tert-butylcalix[6]arene complexation with ammonium hydroxide, and ammonium and sodium ions.

The formation of complexes involving p-tert-butylcalix[6]arene with neutral and charged species has been investigated by tandem mass spectrometry combined with electrospray ionization. Complexes of p-tert-butylcalix[6]arene with NH4+ ions were observed in the ratios 1:1, 2:1, and 3:1, together with the complexes of p-tert-butylcalix[6]arene with NH4OH and Na+ ions in the ratios 1:1:1, 2:1:1, and 3:1:1. A single 1:1 complex of p-tert-butylcalix[6]arene with Na+ ions was observed. In addition, a doubly charged complex of p-tert-butylcalix[6]arene with NH4OH, Na+, and NH4+ ions in the ratio 6:1:1:1 was observed. The identity of each complex was determined by mass analysis of product ions formed by the application of a declustering potential over the range 20-220 V and by observation of product ion mass spectra wherein the collision energy was varied from 5 to 50 eV. Fragmentation of the complexes is characterized by the facile loss of the ammonia molecule, sodium and ammonium ions, loss of neutral p-tert-butylcalix[6]arene, and successive neutral losses of C4H8 from the six tert-butyl groups in each p-tert-butylcalix[6]arene molecule.