ALBERT

Description: Guandu River is the main water source for 9 million inhabitants in Rio de Janeiro city and some others included in the metropolitan region of the Rio de Janeiro State, Brazil. Here, the development of a chromatographic method and its application to assess the occurrence of 4-nonylphenol (4NP), benzophenone (BP), bisphenol A (BPA) and diethyl-phthalate (DEP), known as endocrine disruptors (EDs), is reported. Sample were prepared by solid phase extraction (SPE) with C18 cartridge and methanol as elution solvent. Validation of analytical method followed the United States Environmental Protection Agency protocol (USEPA 8000D guide) and selectivity, matrix effect, linearity, precision, accuracy, robustness, limit of detection (LOD) and limit of quantification (LOQ) were evaluated. The recovery was greater than 90%, accuracy was found between 80% and 115% and relative standard deviation (RSD) below 11.03%. LOQ ranged from 10.0 to 50.0 ng L−1, while the LOD ranged from 0.87 to 5.72 ng L−1. The coefficients of determination (R2) were greater than 0.99 for all compounds within a linear ranges of 10.0 to 500 ng L−1 for 4NP and BP and 50.0 to 500 ng L−1 for BPA and DEP. The method was therefore considered selective and robust for all micropollutants. Matrix effect was observed for BP, 4NP and DEP. The developed method was applied to analyze five samples collected monthly during 2018 at a selected sampling point of a river in Rio de Janeiro State. The maximum concentrations found for BPA, BP, DEP and 4NP were 182.04, 286.20, 2.56×103 and 13.48 ng L−1 respectively. These values are high enough to justify an investigation on the presence of these micropollutants in drinking water as well as to extend the monitoring for the search of similar pollutants and their metabolites.

Description: In this work, we demonstrate a simple spectrophotometry approach to more accurately quantify and measure paclitaxel (PTX) concentrations. PTX cannot be precisely quantified when mixed with an aqueous solvent, and carries the risk of undergoing crystal precipitation. It is likely that PTX undergoes numerous interactions with aqueous solvents and enters a supersaturated state due to its low solubility. Therefore, a quantitative method is required to measure PTX for quality control before clinical use. Although several high-performance liquid chromatography (HPLC) methods have been reported to date, not all medical facilities have a clinical laboratory with such HPLC devices and analysis techniques. Spectroscopy is a simple and convenient method; however, calibration standards are prepared with an organic solvent, such as methanol and acetonitrile, which, when mixed with PTX, can cause solvent effects that lead to inaccurate results. We generated a calibration curve of PTX at various concentrations (40%, 50%, 60%, 70%, 80%, 90% and 100%) of methanol and evaluated the relative error from HPLC results. The optimum methanol concentration for quantification of PTX was 65.8%, which corresponded to the minimum relative error. The detection limit and quantification limit were 0.030 μg/mL and 0.092 μg/mL, respectively. It was possible to predict the PTX concentration even when polyoxyethylene castor oil and anhydrous ethanol were added, as in the commercially available PTX formulation, by diluting 32-fold with saline after mixing. Our findings show that PTX can be more accurately quantified using a calibration curve when prepared in a methanol/water mixture without the need for special devices or techniques.

Description: The study tested the efficiency and reproducibility of a method for optimal separation of low and high abundant proteins in blood plasma. Firstly, three methods for the separation and concentration of eluted (E: low abundance), or bound (B: high abundance) proteins were investigated: TCA protein precipitation, the ReadyPrep™ 2-D cleanup Kit and Vivaspin Turbo 4, 5 kDa ultrafiltration units. Secondly, the efficiency and reproducibility of a Seppro column or a ProteoExtract Albumin/IgG column were assessed by quantification of E and B proteins. Thirdly, the efficiency of two elution buffers, containing either 25% or 10% glycerol for elution of the bound protein, was assessed by measuring the remaining eluted volume and the final protein concentration. Compared to the samples treated with TCA protein precipitation and the ReadyPrep™ 2-D cleanup Kit, the E and B proteins concentrated by the Vivaspin4, 5 kDa ultrafiltration unit were separated well in both 1-D and 2-D gels. The depletion efficiency of abundant protein in the Seppro column was reduced after 15 cycles of sample processing and regeneration and the average ratio of E/(B + E) × 100% was 37 ± 11(%) with a poor sample reproducibility as shown by a high coefficient of variation (CV = 30%). However, when the ProteoExtract Albumin/IgG column was used, the ratio of E/(B + E) × 100% was 43 ± 3.1% (n = 6) and its CV was 7.1%, showing good reproducibility. Furthermore, the elution buffer containing 10% (w/v) glycerol increased the rate of B protein elution from the ProteoExtract Albumin/IgG column, and an appropriate protein concentration (3.5 µg/µl) for a 2-D gel assay could also be obtained when it was concentrated with Vivaspin Turbo 4, 5 kDa ultrafiltration unit. In conclusion, the ProteoExtract Albumin/IgG column shows good reproducibility of preparation of low and high abundance blood plasma proteins when using the elution buffer containing 10% (w/v) glycerol. The optimized method of preparation of low/high abundance plasma proteins was when plasma was eluted through a ProteoExtract Albumin/IgG removal column, the column was further washed with elution buffer containing 10% glycerol. The first and second elution containing the low and high abundance plasma proteins, respectively, were further concentrated using Vivaspin® Turbo 4, 5 kDa ultrafiltration units for 1 or 2-D gel electrophoresis.

Description: Water has been described as a universal solvent, and this is perhaps the strength behind its many uses. Despite this unique property, anthropogenic activities along its course and natural factors often determine the composition of water. In the current research, the portion of River Nworie having past Owerri town was sampled in the dry season 2017 to determine its ionic composition at predestinated points and to relate such properties to its physicochemical characteristics. Studies relating physicochemical properties and dissolved toxic ions in water could develop a body of knowledge that could enable detection and quantification of potential risk of ions such as heavy metals from natural water to aquatic ecosystem, animal and human health without actually involving aquatic organism, animal and human. Clean sterile plastic bottles were used for collecting surface water. A total of 30 sub-samples from five points at 300 m apart were sampled in the morning. Physicochemical properties were determined using standard methods and ionic composition of water was determined according methods of APHA. Results revealed that Ca2+ had a mean 23.60 ± 0.67 mg/l and was the highest while K+ with a mean 0.72 ± 0.30 was the least amongst major cations. Amongst the major anions Cl− had mean of 31.58 ± 4.47 mg/l while mean of PO43− was 1.42 ± 0.13 mg/l. The ionic balance calculate as % balance error showed high values for all sampling sites ranging from 30 to 39.42% indicating that there is massive input from anthropogenic activities. The computed relationships for selected heavy metals, cations and anions revealed that R2 values were ranging between  ± 0.012 to 1 indicating some form of relationship existing. The water pH weakly correlated with dissolved cations and anions while moderate with pH only, due to the pH level (5.2–6.2). The cations and anions were more influenced by the water temperature than the heavy metals. Therefore, high temperature ranges of 31–32.4 °C will favour more dissolution of cations and anions in natural water. Cations showed stronger relationship with EC while only heavy metals showed no relationship with DO (Dissolved oxygen). Dissolved oxygen relationship with cations and anions was in the order; K+ 〉 Mg2+ 〉 Ca2+ 〉 Na+ while anions was SO42− 〉 NO3− 〉 Cl− 〉 PO43−, respectively. Information here could be used to predict the effects of using this water for various purposes including water for agricultural purposes, in the management of ion polluted waters, and also to inform on the mitigation process to be taken.

Description: Background Extracellular vesicles (EVs) are released by most cell types and are involved in multiple basic biological processes. Medium/large EVs (m/lEVs), which are of a different size from exosomes, play an important role in the coagulation in blood, and are secreted from cancer cells, etc., suggesting functions related to malignant transformation. The m/lEVs levels in blood or urine may help unravel pathophysiological findings in many diseases. However, it remains unclear how many naturally-occurring m/lEV subtypes exist as well as how their characteristics and functions differ from one another. Methods We used the blood and urinal sample from each 10 healthy donors for analysis. Using a flow cytometer, we focus on characterization of EVs with large sizes (〉200 nm) that are different from exosomes. We also searched for a membrane protein for characterization with a flow cytometer using shotgun proteomics. We then identified m/lEVs pelleted from plasma and urine samples by differential centrifugation and characterized by flow cytometry. Results Using proteomic profiling, we identified several proteins involved in m/lEV biogenesis including adhesion molecules, peptidases and exocytosis regulatory proteins. In healthy human plasma, we could distinguish m/lEVs derived from platelets, erythrocytes, monocytes/macrophages, T and B cells, and vascular endothelial cells with more than two positive surface antigens. The ratio of phosphatidylserine appearing on the membrane surface differed depending on the cell-derived m/lEVs. In urine, 50% of m/lEVs were Annexin V negative but contained various membrane peptidases derived from renal tubular villi. Urinary m/lEVs, but not plasma m/lEVs, showed peptidase activity. The knowledge of the new characteristics is considered to be useful as a diagnostic material and the newly developed method suggests the possibility of clinical application.

Description: Fizzy extraction (FE) facilitates analysis of volatile solutes by promoting their transfer from the liquid to the gas phase. A carrier gas is dissolved in the sample under moderate pressure (Δp ≈ 150 kPa), followed by an abrupt decompression, what leads to effervescence. The released gaseous analytes are directed to an on-line detector due to a small pressure difference. FE is advantageous in chemical analysis because the volatile species are released in a short time interval, allowing for pulsed injection, and leading to high signal-to-noise ratios. To shed light on the mechanism of FE, we have investigated various factors that could potentially contribute to the extraction efficiency, including: instrument-related factors, method-related factors, sample-related factors, and analyte-related factors. In particular, we have evaluated the properties of volatile solutes, which make them amenable to FE. The results suggest that the organic solutes may diffuse to the bubble lumen, especially in the presence of salt. The high signal intensities in FE coupled with mass spectrometry are partly due to the high sample introduction rate (upon decompression) to a mass-sensitive detector. However, the analytes with different properties (molecular weight, polarity) reveal distinct temporal profiles, pointing to the effect of bubble exposure to the sample matrix. A sufficient extraction time (~12 s) is required to extract less volatile solutes. The results presented in this report can help analysts to predict the occurrence of matrix effects when analyzing real samples. They also provide a basis for increasing extraction efficiency to detect low-abundance analytes.

Description: Background There exists great interest in using dried bloodspots across the clinical, public health, and nutritional sciences to characterize circulating levels of essential elements yet current methods face several challenges related to instrumentation, quality control, and matrix effects. Elemental analysis via total X-ray fluorescence (TXRF) may help overcome these challenges. The objective of this study was to develop and apply a novel TXRF-based analytical method to quantify essential elements (copper, selenium, zinc) in dried bloodspots. Methods Analytical methods were developed with human whole blood standard reference materials from the Institut National de Santé Publique du Québec (INSPQ). The method was developed in careful consideration of several quality control parameters (e.g., analytical accuracy, precision, linearity, and assay range) which were iteratively investigated to help refine and realize a robust method. The developed method was then applied to a quantitative descriptive survey of punches (n = 675) taken from residual dried bloodspots from a newborn screening biobank program (Michigan BioTrust for Health). Results The analytical method developed to quantify the three target elements in dried bloodspots fared well against a priori quality control criteria (i.e., analytical accuracy, precision, linearity and range). In applying this new method, the average (±SD) blood copper, selenium, and zinc levels in the newborn samples were 1,117.0 ± 627.1 µg/L, 193.1 ± 49.1 µg/L, and 4,485 ± 2,275 µg/L respectively. All the elements were normally distributed in the sample population, and the measured concentrations fall within an expected range. Conclusions This study developed and applied a novel and robust method to simultaneously quantify three essential elements. The method helps overcome challenges in the field concerning elemental analysis in dried bloodspots and the findings help increase understanding of nutritional status in newborns.