Elevated Levels of Ultrashort- and Short-Chain Perfluoroalkyl Acids in US Homes and People.

Per- and polyfluoroalkyl substances (PFAS) make up a large group of fluorinated organic compounds extensively used in consumer products and industrial applications. Perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA), the two perfluoroalkyl acids (PFAAs) with 8 carbons in their structure, have been phased out on a global scale because of their high environmental persistence and toxicity. As a result, shorter-chain PFAAs with less than 8 carbons in their structure are being used as their replacements and are now widely detected in the environment, raising concerns about their effects on human health. In this study, 47 PFAAs and their precursors were measured in paired samples of dust and drinking water collected from residential homes in Indiana, United States, and in blood and urine samples collected from the residents of these homes. Ultrashort- (with 2 or 3 carbons [C2-C3]) and short-chain (with 4-7 carbons [C4-C7]) PFAAs were the most abundant in all four matrices and constituted on average 69-100% of the total PFAA concentrations. Specifically, trifluoroacetic acid (TFA, C2) and perfluoropropanoic acid (PFPrA, C3) were the predominant PFAAs in most of the samples. Significant positive correlations (n = 81; r = 0.23-0.42; p < 0.05) were found between TFA, perfluorobutanoic acid (PFBA, C4), and perfluoroheptanoic acid (PFHpA, C7) concentrations in dust or water and those in serum, suggesting dust ingestion and/or drinking water consumption as important exposure pathways for these compounds. This study demonstrates that ultrashort- and short-chain PFAAs are now abundant in the indoor environment and in humans and warrants further research on potential adverse health effects of these exposures.

Developed rapid and simple RP-HPLC method for simultaneous separation and quantification of bovine milk protein fractions and their genetic variants.

The aim was to develop a reliable rapid reversed-phase high-performance liquid chromatography (RP-HPLC) method to simultaneously determine the main bovine milk protein fractions, including their genetic variants. Compared to the previous studies, our method is able to separate the main protein fractions within 20 min of total run time. The method validation consisted of testing repeatability, reproducibility linearity, repeatability, and accuracy. The procedure was developed using raw individual, bulk, and commercially available heat-treated cow milk samples. The RSD of peak areas ranged from 1.43 to 3.16% within analytical day and from 3.29 to 6.70% across analytical days. The method can be applied to investigate both raw and heat-treated milk samples.

Rapid Identification between Two Fish Species Using UV-Vis Spectroscopy for Substitution Detection.

Fish species substitution and fraud has become a worldwide economic issue in the seafood industry. In this study, an ultraviolet-visible (UV-Vis) spectroscopy-based method was developed for the identification of fish samples. Sixty fish samples from twelve commonly consumed fish species in China were analyzed as models to testify the protocol. The obtained results show that UV-Vis spectroscopy combined with chemometric analysis, such as principal component analysis (PCA), can accurately distinguish two fish species by boiling fish tissue sample in trifluoroacetic acid (TFA) solution for 2 min and analyzing the resultant samples using a UV-Vis spectrometer. The developed strategy was successfully applied to the classification and identification of fish samples on the market. It is a promising strategy that can be applied to the classification and authenticity testing of closely related fish species in order to detect and recognize fish substitution.

Quantification of long-chain, short-chain, and ultrashort-chain liquid chromatography-amenable PFASs in water: Evaluation of approaches and tradeoffs for AFFF-impacted water.

The widespread use of aqueous film-forming foam (AFFF) for firefighting and firefighter training has led to extensive per- and polyfluoroalkyl substance (PFAS) contamination in the environment. Challenges remain in the analytical determination of PFASs via liquid chromatography-mass spectrometry (LC-MS), particularly when attempting to include ultrashort-chain perfluoroalkyl acids (PFAAs) and longer-chain anionic and zwitterionic PFASs in a single direct injection. In this study, we assessed the performance of three analytical LC columns (C18, JJ, and Acclaim columns) to separate targeted and suspect PFASs in AFFF-impacted water samples collected from five sites. The C18 column failed to retain ultrashort-chain PFAAs while the JJ and Acclaim columns were not suitable for hydrophobic PFASs. Ultrashort-chain PFAAs were detected at three sites and comprised 1.6-18% of the total perfluoroalkyl carboxylic and sulfonic acids. Semi-quantified concentrations of suspect PFASs comprised 0.70-13% of the total PFASs. When attempting to capture the entirety of the PFAS mass in a water sample, the C18 column captured the broadest suite of suspect PFASs, while the JJ column quantified the most total PFAS mass. Results of this study highlight the importance and tradeoffs of LC column choice to comprehensively determine the composition of PFASs and their concentrations in AFFF-impacted water samples.

Elucidation of the hydration pattern of trifluoroacetic acid in dilute solutions: FTIR and computational study.

The atmospheric partitioning of trifluoroacetic acid (TFA) in aerosol is a complex function of the size of suspended water droplets and their pH value. The unraveling of the affinity of TFA towards basic but not acidic conditions may be accomplished by providing an insight into the hydration pattern of undissociated TFA. Owing to rather scarce details on very dilute aqueous solutions of trifluoroacetic acid (TFA), we examined CF3COOD and CF3COONa solutions in D2O in the concentration range 0.001-0.1 mol dm-3 using transmission FTIR spectroscopy and computational methods. Besides detecting the signals originated from undissociated species in both CF3COOD (1787 cm-1 and 1766 cm-1 at c0 = 0.1 mol dm-3) and CF3COONa (1807 cm-1 at c0 = 0.1 mol dm-3) D2O solutions, through computational techniques we identified different TFA hydrates that contribute to the complexity of the spectral appearance. The combination of experimental and computational data suggested the concentration dependence of the predominant hydrogen bonding pattern of TFA. The results obtained in this work should help in understanding the partitioning of TFA into micron-size water droplets in the atmosphere in molecular and structural terms, i.e. the eventual stability of a hydrated complex for a particular TFA conformer.

Cardioprotective and hypotensive mechanistic insights of hydroethanolic extract of Cucumis melo L. kernels in isoprenaline-induced cardiotoxicity based on metabolomics and in silico electrophysiological models.

Background: Cardiovascular diseases (CVD) continue to threaten health worldwide, and account for a significant portion of deaths and illnesses. In both developing and industrialized nations, they challenge their health systems. There are several traditional uses of Cucurbitaceae seeds in Pakistan, India, Iran, and China, including treating cardiovascular, neurological, and urogenital diseases. Methods: In the present work, integrated techniques of metabolomics profiling and computational cardiomyocyte stimulation were used to investigate possible mechanisms of C. melo in isoprenaline (ISO)-induced myocardial infarction. In vitro, vasoconstrictions, paired atria, and in vivo invasive blood pressure measurement models were performed to explore the mechanism of action of C. melo hydroethanolic seed extract (Cm-EtOH). Results: Results showed that Cm-EtOH demonstrates NO-based endothelium-derived relaxing factor (EDRF) vasorelaxant response, negative chronotropic and inotropic response in the atrium, and hypotensive effects in normotensive rats. Results also revealed that Cm-EtOH decreases cardiomyocyte hypertrophy and reverts the altered gene expressions, biochemical, and metabolites in ISO-induced myocardial infarction (MI) rats. The extract additionally reversed ISO-induced MI-induced oxidative stress, energy consumption, and amino acid metabolism. Moreover, C. melo seeds increased EDRF function, energy production, and antioxidant capacity to treat myocardial and vascular disorders. In computational cardiomyocyte simulation, gallic acid reduced action potential duration, upstroke velocity (dV/dtmax), and effective refractory period. Conclusion: This study highlights the therapeutic potential of C. melo seeds to treat cardiovascular diseases and provides mechanistic insight into its antihypertensive and cardioprotective activities.

Inhibition of human cervical cancer development through p53-dependent pathways induced by the specified triple helical ß-glucan.

This study demonstrates that the purified ß-glucan (LNT) with a triple helix and relatively narrow molecular weight distribution, extracted and purified from artificially cultured Lentinus edodes, showed a significant cervical cancer inhibition with little cytotoxicity against normal cells in vitro and in vivo. From the in vitro data, the potential mechanism of anti-cervical cancer was preliminarily revealed as follows: LNT was firstly recognized by the human cervical cancer cell line of Hela and induced cell proliferation inhibition through p21 and apoptosis via a mitochondrion-dependent pathway by targeting the tumor suppressor of p53, indicated by an increase in reactive oxygen species (ROS) generation and a loss of mitochondrial membrane potential (Δψm), in a significant dosage-dependent manner. Meanwhile, LNT repressed tumor growth with an inhibition ratio of 61.2 % and induced tumor cell apoptosis through endogenous MDM2/p53/Bax/mitochondrion signal pathway by up-regulating the expression of p53, Bax, Cyt. c, caspase 9, and caspase 3, as well as down-regulating Bcl-2, MDM2, and PARP1 levels in Hela cells-transplanted BALB/c nude mice. This study provides a scientific basis for the clinical treatment of cervical cancer with LNT as a potential drug candidate characterized by the triple helix and specified molecular weight with a relatively narrow distribution.

Residue analytical method for the determination of trifluoroacetic acid and difluoroacetic acid in plant matrices by capillary electrophoresis tandem mass spectrometry (CE-MS/MS).

In the past years, the technology for trace residue analysis of plant protection compounds in plant and animal matrices, soil, and water has gradually changed to meet changing regulatory demands. Generally, from the '70s to the '90s of the last century, the active compounds and only a few major metabolites had to be determined in a typical "residue definition". Step by step and within the framework of product safety assessments of the enforcement of residues in dietary matrices and in the environment, further metabolites have come into the authorities focus. Many active substances were formerly determined via gas chromatography (GC) based detection techniques. The introduction of liquid chromatography tandem mass spectrometry (LC-MS/MS) technology in the '90s and the acceptance of this technique, by official bodies at the end of the '90s, has led to a major change for residue analytical laboratories all over the world. Most of the medium to non-polar active compounds as well as many of the more polar metabolites can be detected with this technique, and today LC-MS/MS is the "workhorse" in many residue analytical laboratories in the industry as well as official enforcement labs responsible for analyzing registration-related field studies. With the demand to analyze further breakdown products, more and more polar compounds, or even (permanently) charged target compounds, have now come into the focus of the registration authorities. This now brings standard LC-based techniques to their limits and requires the application of approaches such as hydrophilic interaction chromatography (HILIC) MS/MS or ion chromatography, however these techniques often incur related uncertainties and problems with matrix samples. The aim of this study was to develop a new CE-MS/MS-based approach to reduce the impact of matrix on the separation and detection of trifluoroacetic acid (TFA) and difluoroacetic acid (DFA) in agrochemical field trials. This project used 7 representative examples of fruit, grain and vegetables which had undergone homogenization and extraction with acetonitrile water and filtration before CE-MS/MS analysis. The CE-MS/MS developed reached the limit of quantitation (LOQ) requirement of current legislation for both TFA and DFA (0.01 mg/kg) in all 7 matrices tested. The mean relative standard deviation (RSD) obtained from the repeat analysis of control field trail samples in all matrices, for both TFA and DFA, was less than 10% meeting GLP guidelines. When compared with LC-MS/MS, using on column loading amounts, the CE-MS/MS was 17 - 43 times more sensitive than a standard method and less matrix effects were observed. The developed method was validated under GLP conditions to provide a GLP-validated residue analytical method for the charged metabolites TFA and DFA in matrix samples from GLP field residue trials.

Per- and polyfluoroalkyl substances and the contribution of unknown precursors and short-chain (C2-C3) perfluoroalkyl carboxylic acids at solid waste disposal facilities.

The emission of per- and polyfluoroalkyl substances (PFASs) from municipal solid wastes (MSW) disposal raises concerns for their potential of long-term release and risks. In this study, the occurrence of PFASs was investigated in ambient air and leachate from seven MSW disposal facilities including three landfills, two incineration plants, and two MSW transfer stations in Tianjin, China. Mass loads of PFASs (≥C4) released to the atmosphere were estimated at 0.007-0.97 kg/y/site, which were much lower than those to leachate (0.04-1.3 kg/y/site), while emission to the atmosphere at landfills was more considerable. With total oxidizable precursor (TOP) assay, unknown C4-C12 perfluoroalkyl acids (PFAAs)-precursors were found contributing 10-97 mol% in leachate and accounting for additional 15%-43% mass loads. Using IC-Ba/Ag/H cartridges, trifluoroacetic acid (C2) and perfluoropropionic acid (C3) were recovered in leachate for TOP assay (62%-78%) and determined at dominant levels of 19-81 µg/L, which accounted for mass loads of 0.08-2.6 kg/y/site. Unknown C2-C3 PFAA-precursors contributed 12-93 mol% with mass loads of 0.10-3.0 kg/y/site. Overall, unknown C2-C12 PFAA-precursors remained contributing 0.35-68 mol% in biochemically treated leachate. This study emphasizes that the profiles of unknown PFAA-precursors released during MSW disposal are to be identified, which is essential for their environmental risk assessment.

An improved LC-MS method to profile molecular diversity and quantify the six main bovine milk proteins, including genetic and splicing variants as well as post-translationally modified isoforms.

Here we describe a method based on Liquid Chromatography coupled with Mass Spectrometry (LC-MS) that provides an accurate determination of the six main bovine milk proteins, including allelic and splicing variants, as well as isoforms resulting from post-translational modifications, with an unprecedented level of resolution. Proteins are identified from observed molecular masses in comparison with theoretical masses of intact proteins indexed in an "in-house" database that includes nearly 3000 entries. Quantification was performed either from UV (214 nm) or mass signals. Thus, up to one hundred molecules, derived from the six major milk proteins, can be identified and quantified from an individual milk sample. This powerful and reliable method, initially developed as an anchoring method to estimate the composition of the six main bovine milk proteins from MIR spectra, is transferable to several mammalian species, including small ruminants, camels, equines, rabbits, etc., for which specific mass databases are available.

Review of the existing maximum residue levels for fluometuron according to Article 12 of Regulation (EC) No 396/2005.

According to Article 12 of Regulation (EC) No 396/2005, EFSA has reviewed the maximum residue levels (MRLs) currently established at European level for the pesticide active substance fluometuron. To assess the occurrence of fluometuron residues in plants, processed commodities, rotational crops and livestock, EFSA considered the conclusions derived in the framework of Commission Regulation (EC) No 33/2008 as well as the European authorisations reported by Member States (including the supporting residues data). Based on the assessment of the available data, an MRL proposal was derived and a consumer risk assessment was carried out. All information required by the regulatory framework was present and a risk to consumers was not identified. In addition, EFSA identified some data gaps which are not expected to impact on the validity of the MRL derived but which might have an impact on national authorisations.

Characterizing various monoclonal antibodies with milder reversed phase chromatography conditions.

Reversed phase liquid chromatography (RPLC) of therapeutic monoclonal antibodies (mAbs) is often performed at elevated temperatures (80-90 °C) and in the presence of relatively high concentrations of TFA (0.1%). Under such conditions, it is possible to achieve suitable performance in terms of peak shapes and recoveries. Yet, it is also possible, to cause on-column hydrolysis and the generation of artefact peaks. Interestingly, a wide-pore silica-based superficially porous (SPP) material with a high coverage phenyl bonding was recently introduced, and may offer a chance to perform protein RPLC with milder conditions (lower temperatures and lower TFA concentrations). To evaluate this possibility, 23 mAbs approved by Food and Drug Administration (FDA) and European Medicines Agency (EMA) were analysed on this new column, as well as a reference C4 SPP widepore silica-based column. Separations were performed at various temperatures ranging from 60 to 90 °C using various proportions of TFA and FA ranging from 0.1% TFA to 0.01% TFA/0.09% FA. It appears that temperature can be reduced down to 75 °C for intact mAbs and 65 °C for mAb subunits, using the high coverage phenyl bonded stationary phase. At such temperatures, desirable peak shape and >90% recovery can be observed for each of the studied mAbs. To achieve the same performance with the reference C4 bonded stationary phase, it was necessary to work at 90 °C and 85 °C, respectively. In addition, for mAb subunit analysis, it has been found that combining the phenyl bonded stationary phase with a 0.03% TFA/0.07% FA mobile phase can yield separations similar those obtained with 0.1% TFA, but with an increase of mass spectrometric sensitivity by about 40%. This work provides examples of milder conditions (65 °C and 0.03% TFA + 0.07% FA) being successfully employed in the RPLC(-MS) analysis of mAb subunits. A suitable stationary phase was selected, and milder conditions allow an improved MS sensitivity, while maintaining comparable elution profiles. Moreover, the use of milder conditions may reduce the risk of artefact peaks.

Separation of Peptides on HALO 2-Micron Particles.

Reversed-phase high-performance liquid chromatography (RP-HPLC) is of fundamental importance to the isolation and separation of peptides, proteins, and other biomolecules. Hence, there is a continuing high demand for the development of RP-HPLC stationary-phase materials with enhanced separation efficiency. HALO packing materials began the revolution in "core-shell" technology with the advantages of faster separations, higher resolution and peak capacity, high temperature stability, and rugged reliable performance compared to traditional HPLC and UHPLC. These materials are characterized by a solid core surrounded by a thin layer of porous material, and represent a technology for the future with continuing refinements. Such refinements are aided via the use of designed synthetic peptide standards during stationary-phase development. Concomitantly, such standards also enable the researcher to monitor RP-HPLC column performance and develop optimized separation protocols for peptides from a wide array of sources. © 2016 by John Wiley & Sons, Inc.

In vivo imaging of Aminopeptidase N (CD13) receptors in experimental renal tumors using the novel radiotracer (68)Ga-NOTA-c(NGR).

PURPOSE: Aminopeptidase N (APN/CD13) plays an important role in tumor neoangiogenic process and the development of metastases. Furthermore, it may serve as a potential target for cancer diagnosis and therapy. Previous studies have already shown that asparagine-glycine-arginine (NGR) peptides specifically bind to APN/CD13. The aim of the study was to synthesize and investigate the APN/CD13 specificity of a novel (68)Ga-labeled NOTA-c(NGR) molecule in vivo using miniPET. METHODS: c[KNGRE]-NH2 peptide was conjugated with p-SCN-Bn-NOTA and was labeled with Ga-68 ((68)Ga-NOTA-c(NGR)). Orthotopic and heterotopic transplanted mesoblastic nephroma (NeDe) bearing Fischer-344 rats were prepared, on which biodistribution studies and miniPET scans were performed for both (68)Ga-NOTA-c(NGR) and ανß3 integrin selective (68)Ga-NODAGA-[c(RGD)]2 tracers. APN/CD13 receptor expression of NeDe tumors and metastases was analyzed by western blot. RESULTS: (68)Ga-NOTA-c(NGR) was produced with high specific activity (5.13-5.92GBq/µmol) and with excellent radiochemical purity (95%<), at all cases. Biodistribution studies in normal rats showed that uptake of the (68)Ga-NOTA-c(NGR) was significantly (p⩽0.05) lower in abdominal organs in comparison with (68)Ga-NODAGA-[c(RGD)]2. Both radiotracers were mainly excreted from the kidney. In NeDe tumor bearing rats higher (68)Ga-NOTA-c(NGR) accumulation was found in the tumors than that of the (68)Ga-NODAGA-[c(RGD)]2. Using orthotopic transplantation, metastases were developed which showed specific (68)Ga-NOTA-c(NGR) uptake. Western blot analysis confirmed the presence of APN/CD13 expression in NeDe tumors and metastases. CONCLUSION: Our novel radiotracer (68)Ga-NOTA-c(NGR) showed specific binding to the APN/CD13 expressed ortho- and heterotopic transplanted NeDe tumors. Therefore, (68)Ga-NOTA-c(NGR) is a suitable tracer for the detection of APN/CD13 positive tumors and metastases in vivo.

Regioselective dehydrogenation of 3-keto-steroids to form conjugated enones using o-iodoxybenzoic acid and trifluoroacetic acid catalysis.

Mild and regioselective conversion of 3-keto-5α- and 3-keto-5ß-steroids (trans A/B- and cis A/B-ring juncture, respectively) to the corresponding enones (Δ(1)- and Δ(4)-3-ketones) by treatment with o-iodoxybenzoic acid (IBX) catalyzed by trifluoroacetic acid (TFA) in DMSO, is described. The IBX-mediated reaction involved dehydrogenation of the α- and ß-hydrogen atoms of the 3-ketones to give the enones regioselectively in good isolated yields without concomitant formation of related dienones and trienones.