Sustainable ionic liquids-based molecular platforms for designing acetylcholinesterase reactivators.

We report a green chemistry approach for preparation of oxime-functionalized ILs as AChE reactivators: amide/ester linked IL, l-alanine, and l-phenylalanine derived salts bearing pyridinium aldoxime moiety. The reactivation capacities of the novel oximes were evaluated towards AChE inhibited by typical toxic organophosphates, sarin (GB), VX, and paraoxon (PON). The studied compounds are mostly non-toxic up to the highest concentrations screened (2 mM) towards Gram-negative and Gram-positive bacteria cell lines and both filamentous fungi and yeasts in the in vitro screening experiments as well as towards the eukaryotic cell (CHO-K1 cell line). Introduction of the oxime moiety in initially biodegradable structure decreases its ability to biodegradation. The compound 3d was shown to reveal remarkable activity against the AChE inhibited by VX, exceeding conventional reactivators 2-PAM and obidoxime. The regularities on antidotal activity, cell viability, plasma stability, biodegradability as well as molecular docking study of the newly synthesized oximes will be used for further improvement of their structures.

2-Substituted quinazolines: Partial agonistic and antagonistic ligands of the constitutive androstane receptor (CAR).

Following the discovery of 2-(3-methoxyphenyl)-3,4-dihydroquinazoline-4-one and 2-(3-methoxyphenyl)quinazoline-4-thione as potent, but non-specific activators of the human Constitutive Androstane Receptor (CAR, NR1I3), a series of quinazolinones substituted at the C2 phenyl ring was prepared to examine their ability to selectively modulate human CAR activity. Employing cellular and in vitro TR-FRET assays with wild-type CAR or its variant 3 (CAR3) ligand binding domains (LBD), several novel partial human CAR agonists and antagonists were identified. 2-(3-Methylphenyl) quinazolinone derivatives 7d and 8d acted as partial agonists with the recombinant CAR LBD, the former in nanomolar units (EC50 = 0.055 µM and 10.6 µM, respectively). Moreover, 7d did not activate PXR, and did not show any signs of cytotoxicity. On the other hand, 2-(4-bromophenyl)quinazoline-4-thione 7l possessed significant CAR antagonistic activity, although the compound displayed no agonistic or inverse agonistic activities. A compound possessing purely antagonistic effect was thus identified for the first time. These and related compounds may serve as a remedy in xenobiotic intoxication or, conversely, in suppression of undesirable hepatic CAR activation.

Sustainable Phenylalanine-Derived SAILs for Solubilization of Polycyclic Aromatic Hydrocarbons.

The solubilization capacity of a series of sustainable phenylalanine-derived surface-active ionic liquids (SAILs) was evaluated towards polycyclic aromatic hydrocarbons-naphthalene, anthracene and pyrene. The key physico-chemical parameters of the studied systems (critical micelle concentration, spectral properties, solubilization parameters) were determined, analyzed and compared with conventional cationic surfactant, CTABr. For all studied PAH solubilization capacity increases with extension of alkyl chain length of PyPheOCn SAILs reaching the values comparable to CTABr for SAILs with n = 10-12. A remarkable advantage of the phenylalanine-derived SAILs PyPheOCn and PyPheNHCn is a possibility to cleave enzymatically ester and/or amide bonds under mild conditions, to separate polycyclic aromatic hydrocarbons in situ. A series of immobilized enzymes was tested to determine the most suitable candidates for tunable decomposition of SAILs. The decomposition pathway could be adjusted depending on the choice of the enzyme system, reaction conditions, and selection of SAILs type. The evaluated systems can provide selective cleavage of the ester and amide bond and help to choose the optimal decomposition method of SAILs for enzymatic recycling of SAILs transformation products or as a pretreatment towards biological mineralization. The concept of a possible practical application of studied systems for PAHs solubilization/separation was also discussed focusing on sustainability and a green chemistry approach.

Biobased natural deep eutectic system as versatile solvents: Structure, interaction and advanced applications.

The increasing emphasis on the development of green replacements to traditional organic solvents and ionic liquids (ILs) can be attributed to the rising concerns over human health and detrimental impacts of conventional solvents towards the environment. A new generation of solvents inspired by nature and extracted from plant bioresources has evolved over the last few years, and are referred to as natural deep eutectic solvents (NADES). NADES are mixtures of natural constituents like sugars, polyalcohols, sugar-based alcohols, amino acids and organic acids. Interest in NADES has exponentially grown over the last eight years, which is evident from an upsurge in the number of research projects undertaken. NADES are highly biocompatible as they can be biosynthesized and metabolized by nearly all living organisms. These solvents pose several noteworthy advantages, such as easy synthesis, tuneable physico-chemical properties, low toxicity, high biodegradability, solute sustainability and stabilization and low melting point. Research on the applicability of NADES in diverse areas is gaining momentum, which includes as - media for chemical and enzymatic reactions; extraction media for essential oils; anti-inflammatory and antimicrobial agent; extraction of bioactive composites; as chromatographic media; preservatives for labile compounds and in drug synthesis. This review gives a complete overview of the properties, biodegradability and toxicity of NADES which we propose can assist in further knowledge generation on their significance in biological systems and usage in green and sustainable chemistry. Information on applications of NADES in biomedical, therapeutic and pharma-biotechnology fields is also highlighted in the current article along with the recent progress and future perspectives in novel applications of NADES.

Role of Bacterial-Fungal Consortium for Enhancement in the Degradation of Industrial Dyes.

BACKGROUND: The presence of anthraquinone (Disperse blue 64) and azodyes (Acid yellow 17) in a waterbody are considered among the most dangerous pollutants. METHODS: In this study, two different isolated microbes, bacterium and fungus, were individually and as a co-culture applied for the degradation of Disperse Blue 64 (DB 64) and Acid Yellow 17 (AY 17) dyes. The isolates were genetically identified based upon 16S (for bacteria) and ITS/5.8S (for fungus) rRNA genes sequences as Pseudomoans aeruginosa and Aspergillus flavus, respectively. RESULTS: The fungal/bacterial consortium exhibited a higher percentage of dyes degradation than the individual strains, even at a high concentration of 300 mg/L. Azoreductase could be identified as the main catabolic enzyme and the consortium could induce azoreductase enzyme in the presence of both dyes. However, the specific substrate which achieved the highest azoreductase specific activity was Methyl red (MR) (3.5 U/mg protein). The tentatively proposed metabolites that were detected by HPLC/MS suggested that the reduction process catalyzed the degradation of dyes. The metabolites produced by the action consortium on two dyes were safe on Vicia faba and Triticum vulgaris germination and health of seedlings. Toxicity of the dyes and their degradation products on the plant was different according to the type and chemistry of these compounds as well as the type of irrigated seeds. CONCLUSION: We submit that the effective microbial degradation of DB64 and AY17 dyes will lead to safer metabolic products.

Ionic liquid based pretreatment of lignocellulosic biomass for enhanced bioconversion.

Lignocellulosic biomass is the most plentiful renewable biomolecule and an alternative bioresource for the production of biofuels and biochemicals in biorefineries. But biomass recalcitrance is a bottleneck in their usage, thus necessitating their pretreatment for hydrolysis. Most pretreatment technologies, result in toxic by-products or have lower yield. Ionic liquids (ILs) have successfully advanced as 'greener and recyclable' alternatives to volatile organic solvents for lignocellulosic biomass dissolution. This review covers recent developments made in usage of IL-based techniques with focus on biomass breakdown mechanism, process parameter design, impact of cation and anion groups, and the advantageous impact of ILs on the subsequent processing of the fractionated biomass. Progress and barriers for large-scale commercial usage of ILs in emerging biorefineries were critically evaluated using the principles of economies of scale and green chemistry in an environmentally sustainable way.

A new class of prophylactic metallo-antibiotic possessing potent anti-cancer and anti-microbial properties.

Immunocompromised cancer patients are often at high risk of developing infections. Standard infection control measures are required to prevent the onset of infection but, under some circumstances, antimicrobial prophylaxis is necessary. We have developed a family of innovative metallo-antibiotics of general formula [Cu(N,N)(CipA)Cl] where N,N represents a phenanthrene ligand and CipA stands for a derivative of the clinically used fluoroquinolone antibiotic ciprofloxacin. The X-ray crystal structure of one member from this family, [Cu(phen)(CipA)Cl] (where phen is 1,10-phenanthroline), is also reported. These complexes combine into one drug entity a Cu-N,N-framework with DNA binding and DNA oxidant properties and an antibiotic derivative with known anti-proliferative and anti-microbial activities. The complexes were all found to exhibit excellent DNA recognition with binding affinity of lead agents in the order of ∼107 M(bp)-1. Biophysical studies involving calf thymus DNA indicate the complexes intercalate or semi-intercalate DNA via the minor groove. All complexes exhibited excellent nuclease activity with DNA strand scission being mediated predominantly via superoxide and hydroxyl radicals. The complexes were found to have promising anti-proliferative effects against a human breast adenocarcinoma cell line (MCF-7) and a human prostate carcinoma cell line (DU145) with low micromolar and, in some cases, nanomolar cytotoxicities observed. Selective targeting of Gram positive bacteria was also identified by this complex class with one lead compound having an order of magnitude greater potency against Methicillin-resistant S. aureus (MRSA) as compared to the CipA ligand. Importantly, from a clinical stand point, these complexes were also found to be well tolerated in an in vivo Galleria mellonella larvae model, which has both functional and structural similarities to that of the innate immune system of mammals.

Site-Selective and Stereoselective C-H Functionalization of N-Cyclopropylamides via a Directed Remote Metalation Strategy.

A new methodology for site-selective and stereoselective C-H functionalization of aminocyclopropanes via directed remote lithiation has been developed. Treatment of N-directing group (DG = pivaloyl, tetramethylsuccinimidoyl) arylcyclopropanes with t-BuLi results in a clean ß-lithiation and, following quench with electrophiles, leads to a range of cyclopropane derivatives. Sequential double lithiation-methylation to give a dimethylated cyclopropane has been achieved. X-ray, NMR, and computational studies allow rationalization of syn-DG ß-deprotonation selectivity via a DG-lithium base coordinated complex.

Toxicity profiling of 24 l-phenylalanine derived ionic liquids based on pyridinium, imidazolium and cholinium cations and varying alkyl chains using rapid screening Vibrio fischeri bioassay.

A library of 24 pyridinium-, imidazolium-, and cholinium-based ionic liquids (ILs) with varying alkyl chain from C2 to C16 was toxicologically profiled using naturally luminescent marine bacteria Vibrio fischeri. The toxicity (30-min EC50) of studied ILs to Vibrio fischeri ranged from 7.82 µM (4.2 mg/L) (PyC12Phe) to 3096 µM (1227 mg/L) (ImidC2Phe), i.e. from "toxic" (EC50 1-10 mg/L) to "not harmful" (EC50 > 100 mg/L). Inhibition of the bacterial luminescence upon 30-min exposure to ILs correlated well with bacterial viability (exposure for 4 h). The toxicity of studied ILs was largely driven by the length of the alkyl chain (hydrophobicity) and not the type of cationic part of the IL: starting from C10 all the ILs irrespective of the cationic part proved "toxic". The toxicity of the studied ILs was increasing in parallel to their hydrophobicity up to log Kow = 1 (C8-C10) and then levelling up, being consistent with the previously obtained analogous data sets. The "cut-off" effect reported in this study for longer chain length members of the ILs series leads to the "limit" toxicity level for this type of ILs to be ca. 8 mM. Two open-access online tools (www.molinspiration.com and www.vcclab.org) have been applied for the calculation of the Kow values for the 24 ILs reported in this study and 21 ILs reported in the literature. This lead to plotting two nonlinear monotonic correlations between the values of experimental log (1/EC50) and calculated log Kow. The limitation of the online tools and an effect of the ILs structure on the "cut-off" effect have been discussed. The challenge of developing low microbial toxicity surface active ILs remains a significant task to overcome. Our results shed light on the new approaches for designing environmentally benign ILs and functional surfactants. As the hydrophobicity of the ILs significantly correlated with the toxicity, the Vibrio fischeri assay could be considered a powerful tool in providing toxicity data for building and evaluating the QSAR toxicity models for ILs.

Non-Toxic and Ultra-Small Biosilver Nanoclusters Trigger Apoptotic Cell Death in Fluconazole-Resistant Candida albicans via Ras Signaling.

Silver-based nanostructures are suitable for many biomedical applications, but to be useful therapeutic agents, the high toxicity of these nanomaterials must be eliminated. Here, we biosynthesize nontoxic and ultra-small silver nanoclusters (rsAg@NCs) using metabolites of usnioid lichen (a symbiotic association of algae and fungi) that exhibit excellent antimicrobial activity against fluconazole (FCZ)-resistant Candida albicans that is many times higher than chemically synthesized silver nanoparticles (AgNPs) and FCZ. The rsAg@NCs trigger apoptosis via reactive oxygen species accumulation that leads to the loss of mitochondrial membrane potential, DNA fragmentation, chromosomal condensation, and the activation of metacaspases. The proteomic analysis clearly demonstrates that rsAg@NCs exposure significantly alters protein expression. Most remarkable among the down-regulated proteins are those related to glycolysis, metabolism, free radical scavenging, anti-apoptosis, and mitochondrial function. In contrast, proteins involved in plasma membrane function, oxidative stress, cell death, and apoptosis were upregulated. Eventually, we also established that the apoptosis-inducing potential of rsAg@NCs is due to the activation of Ras signaling, which confirms their application in combating FCZ-resistant C. albicans infections.

Efficient DNA Condensation by a C3 -Symmetric Codeine Scaffold.

A novel tripodal codeine scaffold (CC3) was rationally designed using computational methods as a DNA condensing alkaloid. Separation of the piperidine nitrogen atoms in CC3 is considerably larger at 14.36 Šthan previously reported tripodal opioids allowing for enhanced aggregation of larger DNA plasmids (>4,000 bp). The scaffold undergoes protonation at physiological pH that allows for controlled compaction and release of nucleic acids. Condensation is inhibited under basic conditions and nucleic acid release can be achieved by modulating the ionic strength. Zeta potential experiments indicate stabilised DNA particles at low alkaloid loading with AFM measurements showing particles sizes with a height of 103 nm and diameter of 350 nm. Since condensation is a prerequisite for the cellular uptake of DNA, this new class of alkaloid represents a novel nucleic acid condensation agent with potential gene therapy applications.

Biotechnological Advances for Restoring Degraded Land for Sustainable Development.

Global land resources are under severe threat due to pollution and unsustainable land use practices. Restoring degraded land is imperative for regaining ecosystem services, such as biodiversity maintenance and nutrient and water cycling, and to meet the food, feed, fuel, and fibre requirements of present and future generations. While bioremediation is acknowledged as a promising technology for restoring polluted and degraded lands, its field potential is limited for various reasons. However, recent biotechnological advancements, including producing efficient microbial consortia, applying enzymes with higher degrees of specificity, and designing plants with specific microbial partners, are opening new prospects in remediation technology. This review provides insights into such promising ways to harness biotechnology as ecofriendly methods for remediation and restoration.

C 3-symmetric opioid scaffolds are pH-responsive DNA condensation agents.

Herein we report the synthesis of tripodal C3-symmetric opioid scaffolds as high-affinity condensation agents of duplex DNA. Condensation was achieved on both supercoiled and canonical B-DNA structures and identified by agarose electrophoresis, viscosity, turbidity and atomic force microscopy (AFM) measurements. Structurally, the requirement of a tris-opioid scaffold for condensation is demonstrated as both di- (C2-symmetric) and mono-substituted (C1-symmetric) mesitylene-linked opioid derivatives poorly coordinate dsDNA. Condensation, observed by toroidal and globule AFM aggregation, arises from surface-binding ionic interactions between protonated, cationic, tertiary amine groups on the opioid skeleton and the phosphate nucleic acid backbone. Indeed, by converting the 6-hydroxyl group of C3-morphine ( MC3: ) to methoxy substituents in C3-heterocodeine ( HC3: ) and C3-oripavine ( OC3: ) molecules, dsDNA compaction is retained thus negating the possibility of phosphate-hydroxyl surface-binding. Tripodal opioid condensation was identified as pH dependent and strongly influenced by ionic strength with further evidence of cationic amine-phosphate backbone coordination arising from thermal melting analysis and circular dichroism spectroscopy, with compaction also witnessed on synthetic dsDNA co-polymers poly[d(A-T)2] and poly[d(G-C)2]. On-chip microfluidic analysis of DNA condensed by C3-agents provided concentration-dependent protection (inhibition) to site-selective excision by type II restriction enzymes: BamHI, HindIII, SalI and EcoRI, but not to the endonuclease DNase I.

Biodegradation of ionic liquids--a critical review.

The importance of biodegradation data as part of the design of safer chemicals is presented using ionic liquids (ILs) as a model study. Structural features that promote/impede IL biodegradation, IL design strategies, methods of biodegradation analysis, properties of IL/surfactant derivatives and computational methods of predicting biodegradation are discussed. The importance of metabolite studies as part of biodegradation assays is highlighted. The relevance of applying the lessons learned developing biodegradable ILs to other chemical classes is proposed. A comprehensive appendix of IL biodegradation data published since 2010 (∼300 ILs) has been compiled.

Copper phenanthrene oxidative chemical nucleases.

Here we report the synthesis and isolation of a series of bis-chelate Cu(2+) phenanthroline-phenazine cationic complexes of [Cu(DPQ)(Phen)](2+), [Cu(DPPZ)(Phen)](2+), and [Cu(DPPN)(Phen)](2+) (where Phen = 1,10-phenanthroline, DPQ = dipyridoquinoxaline, DPPZ = dipyridophenazine, and DPPN = benzo[i]dipyridophenazine). These compounds have enhanced DNA recognition relative to the well-studied chemical nuclease, [Cu(Phen)2](2+) (bis-Phen), with calf thymus DNA binding constants of DPQ and DPPZ agents (∼10(7) M(bp)(-1)) being the highest currently known for Cu(2+) phenanthrene compounds. Complex DNA binding follows DPQ ≈ DPPZ > DPPN > bis-Phen, with fluorescence quenching and thermal melting experiments on poly[d(A-T)2] and poly[d(G-C)2] supporting intercalation at both the minor and major groove. Phenazine complexes, however, show enhanced targeting and oxidative cleavage on cytosine-phosphate-guanine-rich DNA and have comparable in vitro cytotoxicity toward the cisplatin-resistant ovarian cancer line, SKOV3, as the clinical oxidative DNA-damaging drug doxorubicin (Adriamycin). In this study we also describe how a novel "on-chip" method devised for the Bioanalyser 2100 was employed to quantify double-stranded DNA damage, with high precision, by the complex series on pUC19 DNA (49% A-T, 51% G-C). Both DPQ and bis-Phen complexes are highly efficient oxidizers of pUC19, with DPQ being the most active of the overall series. It is apparent, therefore, that oxidative chemical nuclease activity on homogeneous canonical DNA is not entirely dependent on dynamic nucleotide binding affinity or intercalation, and this observation is corroborated through catalytic interactions with the superoxide anion radical and Fenton breakdown of hydrogen peroxide.

Novel bronchodilatory quinazolines and quinoxalines: synthesis and biological evaluation.

A series of heterocyclic derivatives analogous to (-)vasicinone, in which the vasicinone C-ring was replaced with alkyl chain terminated by tertiary amine was prepared. N3, C4-O, C4-S or C4-N were used as the sites of attachment. The 4-[3-(1-piperidyl)propylsulfanyl]derivatives displayed bronchodilatory effect at low micromolar concentrations on isolated rat trachea, and low toxicity both on Balb/c 3T3 mouse fibroblast cells and in mice.

Regulating bioactivity of Cu2+ bis-1,10-phenanthroline artificial metallonucleases with sterically functionalized pendant carboxylates.

The synthetic chemical nuclease, [Cu(1,10-phenanthroline)2](2+), has stimulated research within metallonuclease development and in the area of cytotoxic metallodrug design. Our analysis reveals, however, that this agent is "promiscuous" as it binds both dsDNA and protein biomolecules, without specificity, and induces general toxicity to a diversity of cell lineages. Here, we describe the synthesis and characterization of small-molecule metallonucleases containing the redox-active cation, [Cu(RCOO)(1,10-phen)2](+), where 1,10-phen = 1,10-phenanthroline and R = -H, -CH3, -C2H5, -CH(CH3)2, and -C(CH3)3. The presence of coordinated carboxylate groups in the complex cation functions to enhance dsDNA recognition, reduce serum albumin binding, and offer control of toxicity toward human cancer cells, Gram positive and negative bacteria, and fungal pathogens. The induction of genomic dsDNA breaks (DSBs) were identified in ovarian adenocarcinoma cells using immunodetection of γ-H2AX. Formate, acetate, and pivalate functionalized complexes induced DSBs in a higher percentage of cells compared with [Cu(1,10-phen)2](2+), which supports the importance of inner-sphere modification toward enhancing targeted biological application.

Synthesis of polypeptide block copolymer hybrids by the combination of N-carboxyanhydride polymerization and RAFT.

The synthesis of hybrid bioconjugates via the ring-opening polymerization (ROP) of N-carboxyanhydrides (NCAs) using a synthetic macroinitiator is described. Poly(n-butyl acrylate), polystyrene, and poly(N-isopropyl acrylamide) are synthesized (polydisperity index, D < 1.1) using reversible addition-fragmentation chain transfer (RAFT) as the synthetic tool. A phthalimidomethyl trithiocarbonate RAFT chain transfer agent is used to prepare well-defined, end-functional polymers, which after deprotection result in amine terminal macroinitiators. The subsequent initiating systems could successfully be chain extended with ε-benzyloxycarbonyl-l-lysine or γ-benzyl-l-glutamate as the NCAs to produce a library of polymer-polypeptide conjugates. In doing so, a novel procedure for directly synthesizing bioconjugates via a non-modular route without the need for excessive purification and isolation steps is described.

The catalytic versatility of low toxicity dialkyltriazolium salts: in situ modification facilitates diametrically opposed catalysis modes in one pot.

The ability of triazolium salts to serve as a precatalyst for both an acid and a powerful base/nucleophile (controlled by additives) has been exploited in a process characterised by a unique in situ catalyst modification strategy.

A new phenanthroline-oxazine ligand: synthesis, coordination chemistry and atypical DNA binding interaction.

1,10-Phenanthroline-5,6-dione and l-tyrosine methyl ester react to form phenanthroline-oxazine (PDT) from which [Cu(PDT)(2)](ClO(4))(2) and [Ag(PDT)(2)]ClO(4)·2MeOH are obtained. Binding to calf-thymus DNA by Ag(I) and Cu(II) PDT complexes exceed bis-1,10-phenanthroline analogues and the minor groove binding drugs, pentamidine and netropsin. Furthermore, unlike the artificial metallonuclease, [Cu(phen)(2)](2+), the [Cu(PDT)(2)](2+) complex does not cleave DNA in the presence of added reductant indicating unique interaction with DNA.