Synthesis, In Silico and In Vivo Toxicity Assessment of Functionalized Pyridophenanthridinones via Sequential MW-Assisted Intramolecular Friedel-Crafts Alkylation and Direct C-H Arylation.

A rapid, efficient, and original synthesis of novel pyrido[3,2,1-de]phenanthridin-6-ones is reported. First, the key cinnamamide intermediates 8a-f were easily prepared from commercial substituted anilines, cinnamic acid, and 2-bromobenzylbromide in a tandem amidation and N-alkylation protocol. Then, these N-aryl-N-(2-bromobenzyl) cinnamamides 8a-f were subjected to a TFA-mediated intramolecular Friedel-Crafts alkylation followed by a Pd-catalyzed direct C-H arylation to obtain a series of potentially bioactive 4-phenyl-4,5-dihydro-6H,8H-pyrido[3,2,1-de]phenanthridin-6-one derivatives 4a-f in good yields. Finally, the toxicological profile of the prepared final compounds, including their corresponding intermediates, was explored through in silico computational methods, while the acute toxicity toward zebrafish embryos (96 hpf-LC50, 50% lethal concentration) was also determined in the present study.

N,N'-Diaryldihydrophenazines as a Sustainable and Cost-Effective Alternative to Precious Metal Complexes in the Photoredox-Catalyzed Alkylation of Aryl Alkyl Ketones.

An inexpensive and highly efficient metal-free alternative to commonly used Ru- and Ir-based catalysts was proposed. It was shown that the new 2,7-di-tert-butyl-5,10-bis(4-trifluoromethylphenyl)-5,10-dihydrophenazine outcompeted the iridium phenylpyridyl complex in photoredox activity in the alkylation of silyl enol ethers yielding aryl alkyl ketones. The reaction occurred under visible light irradiation at room temperature and was also applicable to drug derivatives (ibuprofen and naproxen). In-depth photophysical, electrochemical, and quantum chemical studies showed that the aforementioned N,N-diaryldihydrophenazine exhibited enhanced properties that were essential for the photoredox catalysis (a long-lived triplet excited state, strong reducing ability, high stability of the radical cations formed in single-electron-transfer event, and chemical inertness of the catalyst with respect to reactants). Importantly, the substituted N,N'-diaryldihydrophenazines could be obtained directly from diaryl amines; a facile, easily handled and scaled-up one-pot synthetic procedure was elaborated.

Source, transportation, bioaccumulation, distribution and food risk assessment of perfluorinated alkyl substances in vegetables: A review.

Contamination of perfluoroalkyl substances (PFASs) in agricultural products have attracted more and more attentions recently. In this review, relationship between PFASs and vegetables is summarized comprehensively. PFASs could transfer to cultivation soils by irrigation water, bio-amended soil, and atmospheric deposition mainly from industrial emissions. Carbon chain length of PFASs, species of vegetables and so on are key factors for PFASs migration and bioaccumulation in soils, plants and vegetables. Studies on food risk assessment of PFOA and PFOS show low consumption risk for most vegetables, however researches on other substances are lacking. In the future, we need to pay more attention on novel pollution pathway in cultivation, traceability research for considerable contamination, dietary exposure levels for different vegetables and more substances, as well as more exact and scientific food risk assessments. Additionally, effective means for PFASs adsorption in soil and removal from soil are also expected.

Polycyclic aromatic hydrocarbons: bioaccumulation in dragonfly nymphs (Anisoptera), and determination of alkylated forms in sediment for an improved environmental assessment.

Road runoff carries a mixture of contaminants that threatens the quality of natural water bodies and the health of aquatic organisms. The use of sedimentation ponds is a nature-based solution for the treatment of road runoff. This study assessed the concentration of polycyclic aromatic hydrocarbons (PAHs) and their alkylated homologues in sediment from seven highway sedimentation ponds and three natural urban ponds. In addition, the study explored the bioaccumulation of PAHs in dragonfly nymphs (Anisoptera). Finally, biota-sediment accumulation factors (BSAFs) were estimated. The results revealed a significant difference in the concentrations of 16 priority PAHs in sediment, with overall higher levels in sedimentation ponds (2,911 µg/kg on average) compared to natural urban ponds (606 µg/kg on average). PAH levels increased substantially once alkylated homologues were considered, with alkylated comprising between 42 and 87% of the total PAH in sediment samples. These results demonstrate the importance of alkylated forms in the environmental assessment of PAHs. The bioaccumulation assessment indicates that dragonfly nymphs bioaccumulate PAHs to a certain degree. It is not clear, however, whether they metabolize PAHs. BSAF results ranged from approx. 0.006 to 10 and indicate that BSAFs can be a powerful tool to determine the functionality of sedimentation ponds.

Biocatalysis in drug discovery and development.

Enzyme catalysis, enabled by advances in protein engineering and directed evolution, is beginning to transform chemical synthesis in the pharmaceutical industry. This review presents recent examples of the creative use of biocatalysis to enable drug discovery and development. We illustrate how increased access to novel biotransformations and the rise of cascade biocatalysis allowed fundamentally new syntheses of novel medicines, representing progress toward more sustainable pharmaceutical manufacturing. Finally, we describe the opportunities and challenges the industry must address to ensure the reduction to practice of biotechnological innovations to develop new therapies in a faster, more economical, and environmentally benign way.

Mechanistic Studies of Bioorthogonal ATP Analogues for Assessment of Histidine Kinase Autophosphorylation.

Phosphorylation is an essential protein modification and is most commonly associated with hydroxyl-containing amino acids via an adenosine triphosphate (ATP) substrate. The last decades have brought greater appreciation to the roles that phosphorylation of myriad amino acids plays in biological signaling, metabolism, and gene transcription. Histidine phosphorylation occurs in both eukaryotes and prokaryotes but has been shown to dominate signaling networks in the latter due to its role in microbial two-component systems. Methods to investigate histidine phosphorylation have lagged behind those to study serine, threonine, and tyrosine modifications due to its inherent instability and the historical view that this protein modification was rare. An important strategy to overcome the reactivity of phosphohistidine is the development of substrate-based probes with altered chemical properties that improve modification longevity but that do not suffer from poor recognition or transfer by the protein. Here, we present combined experimental and computational studies to better understand the molecular requirements for efficient histidine phosphorylation by comparison of the native kinase substrate, ATP, and alkylated ATP derivatives. While recognition of the substrates by the histidine kinases is an important parameter for the formation of phosphohistidine derivatives, reaction sterics also affect the outcome. In addition, we found that stability of the resulting phosphohistidine moieties correlates with the stability of their hydrolysis products, specifically with their free energy in solution. Interestingly, alkylation dramatically affects the stability of the phosphohistidine derivatives at very acidic pH values. These results provide critical mechanistic insights into histidine phosphorylation and will facilitate the design of future probes to study enzymatic histidine phosphorylation.

Assessment of native and alkylated polycyclic aromatic hydrocarbons (PAHs) in sediments and mussels (Mytilus spp.) in the southern Baltic Sea.

Native and alkylated polycyclic aromatic hydrocarbons (PAHs) were investigated in whole and fine grained (F < 63 µm) sediment fractions and mussels in the southern Baltic Sea. Both sediments and mussels showed spatial differences in PAH levels and profiles. In whole sediments, mean concentrations of Σ16PAHs, ΣaPAHs, and ΣPAHs reached up to 1537, 415, and 1952 ng g-1 dry weight (dw), respectively, and in mussels they reached up to 235, 143, and 325 ng g-1 dw, respectively, depending on location. ΣaPAHs made up to 36% of ΣPAHs in whole sediments and up to 55% in some mussels in the Gulf of Gdansk. The association of PAHs with the sediment F < 63 µm differed depending on the PAH compound and season. Analysis of PAH sources was done using PCA and the ratios of specific compound concentrations. The sediment and mussel PAH levels evaluated against environmental quality criteria indicated no risk related to PAH occurrence in the study area.

Application of hydrolases and probiotic Pediococcus acidilactici BaltBio01 strain for cereal by-products conversion to bioproduct for food/feed.

The aim of this study was to apply the enzymatic treatment and fermentation by Pediococcus acidilactici BaltBio01 strain for industrial cereal by-products conversion to food/feed bioproducts with high amount of probiotic lactic acid bacteria (LAB). LAB propagated in potato media and spray-dried remained viable during 12 months (7.0 log10 cfu/g) of storage and was used as a starter for cereal by-products fermentation. The changes of microbial profile, biogenic amines (BAs), mycotoxins, lactic acid (L+/D-), lignans and alkylresorcinols (ARs) contents in fermented cereal by-product were analysed. Cereal by-products enzymatic hydrolysis before fermentation allows to obtain a higher count of LAB during fermentation. Fermentation with P. acidilactici reduce mycotoxins content in fermented cereal by-products. According to our results, P. acidilactici multiplied in potato juice could be used for cereal by-products fermentation, as a potential source to produce safer food/feed bioproduct with high amount of probiotic LAB for industrial production.

Enantioselective Catalysis Coupled with Stereodivergent Cyclization Strategies Enables Rapid Syntheses of (+)-Limaspermidine and (+)-Kopsihainanine†A.

Enantioselective Pd-catalyzed allylic alkylations of dihydropyrido[1,2-a]indolone (DHPI) substrates were used to construct the C20-quaternary stereocenters of multiple monoterpene indole alkaloids. Stereodivergent Pictet-Spengler and Bischler-Napieralski cyclization/reduction cascades furnish the cis- and trans-fused azadecalin subunits present in Aspidosperma and Kopsia alkaloids, respectively, en route to highly efficient syntheses of (+)-limaspermidine and (+)-kopsihainanine A.

Assessment of glutathione/glutathione disulphide ratio and S-glutathionylated proteins in human blood, solid tissues, and cultured cells.

Glutathione (GSH) is the major non-protein thiol in humans and other mammals, which is present in millimolar concentrations within cells, but at much lower concentrations in the blood plasma. GSH and GSH-related enzymes act both to prevent oxidative damage and to detoxify electrophiles. Under oxidative stress, two GSH molecules become linked by a disulphide bridge to form glutathione disulphide (GSSG). Therefore, assessment of the GSH/GSSG ratio may provide an estimation of cellular redox metabolism. Current evidence resulting from studies in human blood, solid tissues, and cultured cells suggests that GSH also plays a prominent role in protein redox regulation via S -glutathionylation, i.e., the conjugation of GSH to reactive protein cysteine residues. A number of methodologies that enable quantitative analysis of GSH/GSSG ratio and S-glutathionylated proteins (PSSG), as well as identification and visualization of PSSG in tissue sections or cultured cells are currently available. Here, we have considered the main methodologies applied for GSH, GSSG and PSSG detection in biological samples. This review paper provides an up-to-date critical overview of the application of the most relevant analytical, morphological, and proteomics approaches to detect and analyse GSH, GSSG and PSSG in mammalian samples as well as discusses their current limitations.

An alkylating immobilization linker for immunochemical epigenetic assessment.

A bifunctional linker molecule containing nitrogen mustard and a cyclic disulfide group has been developed for the covalent immobilization of intact DNA, which allows quantitative analysis of epigenomic modification in immobilized DNA using SPR-based immune sensing.

Safety Assessment of Alkyl PEG/PPG Ethers as Used in Cosmetics.

The Cosmetic Ingredient Review (CIR) Expert Panel assessed the safety of 131 alkyl polyethylene glycol (PEG)/polypropylene glycol ethers as used in cosmetics, concluding that these ingredients are safe in the present practices of use and concentration described in this safety assessment when formulated to be nonirritating. Most of the alkyl PEG/PPG ethers included in this review are reported to function in cosmetics as surfactants, skin-conditioning agents, and/or emulsifying agents. The alkyl PEG/PPG ethers share very similar physiochemical properties as the alkyl PEG ethers, which were reviewed previously by the CIR Expert Panel and found safe when formulated to be nonirritating. The alkyl PEG ethers differ by the inclusion of PPG repeat units, which are used to fine-tune the surfactant properties of this group. The Panel relied heavily on data on analogous ingredients, extracted from the alkyl PEG ethers and PPG reports, when making its determination of safety.

Assessment of electrophile damage in a human brain endothelial cell line utilizing a clickable alkyne analog of 2-chlorohexadecanal.

Peripheral leukocytes aggravate brain damage by releasing cytotoxic mediators that compromise blood-brain barrier function. One of the oxidants released by activated leukocytes is hypochlorous acid (HOCl) that is formed via the myeloperoxidase-H2O2-chloride system. The reaction of HOCl with the endogenous plasmalogen pool of brain endothelial cells results in the generation of 2-chlorohexadecanal (2-ClHDA), a toxic, lipid-derived electrophile that induces blood-brain barrier dysfunction in vivo. Here, we synthesized an alkynyl-analog of 2-ClHDA, 2-chlorohexadec-15-yn-1-al (2-ClHDyA) to identify potential protein targets in the human brain endothelial cell line hCMEC/D3. Similar to 2-ClHDA, 2-ClHDyA administration reduced cell viability/metabolic activity, induced processing of pro-caspase-3 and PARP, and led to endothelial barrier dysfunction at low micromolar concentrations. Protein-2-ClHDyA adducts were fluorescently labeled with tetramethylrhodamine azide (N3-TAMRA) by 1,3-dipolar cycloaddition in situ, which unveiled a preferential accumulation of 2-ClHDyA adducts in mitochondria, the Golgi, endoplasmic reticulum, and endosomes. Thirty-three proteins that are subject to 2-ClHDyA-modification in hCMEC/D3 cells were identified by mass spectrometry. Identified proteins include cytoskeletal components that are central to tight junction patterning, metabolic enzymes, induction of the oxidative stress response, and electrophile damage to the caveolar/endosomal Rab machinery. A subset of the targets was validated by a combination of N3-TAMRA click chemistry and specific antibodies by fluorescence microscopy. This novel alkyne analog is a valuable chemical tool to identify cellular organelles and protein targets of 2-ClHDA-mediated damage in settings where myeloperoxidase-derived oxidants may play a disease-propagating role.

Quantification of Polyfunctional Thiols in Wine by HS-SPME-GC-MS Following Extractive Alkylation.

Analyses of key odorous polyfunctional volatile thiols in wines (3-mercaptohexanol (3-MH), 3-mercaptohexylacetate (3-MHA), and 4-mercapto-4-methyl-2-pentanone (4-MMP)) are challenging due to their high reactivity and ultra-trace concentrations, especially when using conventional gas-chromatography electron impact mass spectrometry (GC-EI-MS). We describe a method in which thiols are converted to pentafluorobenzyl (PFB) derivatives by extractive alkylation and the organic layer is evaporated prior to headspace solid phase microextraction (HS-SPME) and GC-EI-MS analysis. Optimal parameters were determined by response surface area modeling. The addition of NaCl solution to the dried SPME vials prior to extraction resulted in up to less than fivefold improvement in detection limits. Using 40 mL wine samples, limits of detection for 4-MMP, 3-MH, and 3-MHA were 0.9 ng/L, 1 ng/L, and 17 ng/L, respectively. Good recovery (90%-109%) and precision (5%-11% RSD) were achieved in wine matrices. The new method was used to survey polyfunctional thiol concentrations in 61 commercial California and New York State wines produced from V. vinifera (Riesling, Gewürztraminer, Cabernet Sauvignon, Sauvignon blanc and non-varietal rosé wines), V. labruscana (Niagara), and Vitis spp. (Cayuga White). Mean 4-MMP concentrations in New York Niagara (17 ng/L) were not significantly different from concentrations in Sauvignon blanc, but were significantly higher than 4-MMP in other varietal wines.

Development of the adverse outcome pathway "alkylation of DNA in male premeiotic germ cells leading to heritable mutations" using the OECD's users' handbook supplement.

The Organisation for Economic Cooperation and Development's (OECD) Adverse Outcome Pathway (AOP) programme aims to develop a knowledgebase of all known pathways of toxicity that lead to adverse effects in humans and ecosystems. A Users' Handbook was recently released to provide supplementary guidance on AOP development. This article describes one AOP-alkylation of DNA in male premeiotic germ cells leading to heritable mutations. This outcome is an important regulatory endpoint. The AOP describes the biological plausibility and empirical evidence supporting that compounds capable of alkylating DNA cause germ cell mutations and subsequent mutations in the offspring of exposed males. Alkyl adducts are subject to DNA repair; however, at high doses the repair machinery becomes saturated. Lack of repair leads to replication of alkylated DNA and ensuing mutations in male premeiotic germ cells. Mutations that do not impair spermatogenesis persist and eventually are present in mature sperm. Thus, the mutations are transmitted to the offspring. Although there are some gaps in empirical support and evidence for essentiality of the key events for certain aspects of this AOP, the overall AOP is generally accepted as dogma and applies broadly to any species that produces sperm. The AOP was developed and used in an iterative process to test and refine the Users' Handbook, and is one of the first publicly available AOPs. It is our hope that this AOP will be leveraged to develop other AOPs in this field to advance method development, computational models to predict germ cell effects, and integrated testing strategies.

DNA-cellulose: an economical, fully recyclable and highly effective chiral biomaterial for asymmetric catalysis.

The challenge in DNA-based asymmetric catalysis is to perform a reaction in the vicinity of the helix by incorporating a small-molecule catalyst anchored to the DNA in a covalent, dative, or non-covalent yet stable fashion in order to ensure high levels of enantio-discrimination. Here, we report the first generation of a DNA-based catalyst bound to a cellulose matrix. The chiral biomaterial is commercially available, trivial to use, fully recyclable and produces high levels of enantioselectivity in various Cu(II)-catalyzed asymmetric reactions including Friedel-Crafts alkylations and Michael additions. A single-pass, continuous-flow process is also reported affording fast conversions and high enantioselectivities at low catalyst loadings thus offering a new benchmark in the field of DNA-based asymmetric catalysis.

Stereospecific cis- and trans-ring fusions arising from common intermediates.

Highly concise and stereospecific routes to cis and trans fusion, carrying various functionality at one of the bridgehead carbons, have been accomplished.

A direct and general method for the reductive alkylation of tertiary lactams/amides: application to the step economical synthesis of alkaloid (-)-morusimic acid D.

Full details of the direct and general method for the reductive alkylation of tertiary lactams and amides to give tertiary sec-alkylamines are presented. This one-pot method consists of in situ activation of a lactam or an amide with Tf2O/DTBMP, addition of a Grignard reagent, and reduction of the resulting iminium intermediates. Alkyl, benzyl, and aryl Grignard reagents and several reductants or reducing conditions (LiAlH4, NaBH4, Hantzsch ester, Bu3SnH, Pd(OH)2/C, H2) could be used effectively. Reductive alkylations of substituted lactams demonstrated good to excellent 1,3-asymmetric induction to provide the corresponding di- or trisubstituted pyrrolidine/piperidine in 6:1 (LiAlH4), 11:1 (Et3SiH), and 20:1 (catalytic hydrogenation) cis/trans diastereoselectivity, respectively. The versatility of this methodology was demonstrated by its application in the concise stereoselective synthesis of piperidine alkaloid (-)-morusimic acid.

Occurrence of perfluorinated alkylated substances in breast milk of French women and relation with socio-demographical and clinical parameters: results of the ELFE pilot study.

A previously developed and validated methodology based on liquid chromatography coupled to high resolution mass spectrometry was used for determine the concentration levels of 14 perfluoroalkylated substances (PFASs) in a set of 48 breast milk samples collected from French women in the frame of the ELFE pilot study. In accordance with other similar studies conducted at european and international levels, PFOS, PFOA, and PFHxS were detected and quantified in most of the analyzed samples (90%, 98% and 100%, respectively), and appeared as major contributors to the total PFAS exposure (38%, 37%, 25%, respectively), whereas the other targeted PFAS were very rarely, if not, found at the limits of detection of the method. Also in agreement with other published data, the concentration levels measured for the detected substances varied from <0.05 to 0.33µg/L for PFOS (median=0.079), from <0.05 to 0.22µg/L for PFOA (median=0.075), and from 0.04 to 0.07µg/L for PFHxS (median=0.050). On the basis of this relatively limited data set, no statistically significant relation was observed between these exposure levels and developmental outcomes, in particular the weight at birth. Similarly, no relation was observed between the measured PFAS levels and various socio-demographical parameters including the consumption of seafood, alcohol, smoking, or socio-economical level. These results suggest a need for further research and better knowledge regarding the sources, pharmacokinetics, and factors of exposure for other substances belonging to this class of emerging contaminants.

Safety assessment of alkyl PEG ethers as used in cosmetics.

The CIR Expert Panel assessed the safety of Alkyl PEG Ethers as used in cosmetics. These ingredients primarily function in cosmetics as surfactants, and some have additional functions as skin-conditioning agents, fragrance ingredients, and emulsion stabilizers. The Panel reviewed available relevant animal and clinical data, as well as information from previous CIR reports; when data were not available for individual ingredients, the Panel extrapolated from the existing data to support safety. The Panel concluded that the Alkyl PEG ethers are safe as used when formulated to be nonirritating, and the same applies to future alkyl PEG ether cosmetic ingredients that vary from those ingredients recited herein only by the number of ethylene glycol repeat units.