Turn-on fluorescence of mirabegron for its sensitive detection in human plasma: Box-Behnken-Design for optimization.

Here, a novel fluorescence strategy was established for the detection of mirabegron (MBG) sensitively on the basis of hantzsch dihydropyridine synthesis. The developed method adopts turn-on fluorescence of MBG for the first time, permitting its selective determination in spiked human plasma at 486 nm after excitation at 410 nm. The developed method exhibited a good linear range from 0.5 µgmL-1 to 2.0 µgmL-1 with detection and quantification limits of 0.05 and 0.2 (µgmL-1), respectively. The profitable applicability of the developed method in spiked human plasma samples was demonstrated, achieving limit of detection below the previously levels reported by spectroscopic methods, allowing application of the developed method for selective determination of MBG in its tablets and spiked human plasma samples with good recovery.

Synthesis of MIL-88(Fe) coordinated to carboxymethyl cellulose fibers nanocomposite for dispersive solid phase microextraction of acetanilide herbicides from cereal and agricultural soil samples.

In this study, MIL-88(Fe) coordinated to carboxymethyl cellulose fibers was successfully synthesized, characterized, and utilized as a nanocomposite for the dispersive solid phase microextraction of butachlor and acetochlor. These analytes served as representative analytes for acetanilide herbicides (AHs) present in real samples. Effective parameters on the extraction efficiency were investigated to maximize the analytical performance of the developed method. Under optimized conditions, which encompassed sorbent amount of 12 mg, solution pH of 7.0, 4.0 min of the vortex time, 3.0 min of the extraction time, chloroform as desorption agent and no salt addition, the developed method exhibited remarkable figures of merit, such as high linearity (R2> 0.99), low limits of detection of 0.90 ng mL-1, substantial preconcentration factors (between 213 and 228), relative recoveries in the range of 90.8% to 109%, and good repeatability with relative standard deviations equal or below 7.2%. After validation, the developed method was applied to detect AHs in various cereal and agricultural soil samples.

Ultimate fate, transformation, and toxicological consequences of herbicide pretilachlor to biotic components and associated environment: An overview.

Herbicides are applied for effective weed management in order to increase the crop yield. In recent decades, the overuse of these chemicals has posed adverse effects on different biotic components of the environment. Pretilachlor has been widely used during last few decades for weed management in paddy crop. Its excessive use may prove fatal for environment, various organisms, and nontarget plants. Thus, it is pertinent to know the extent to which herbicide residues remain in environment. The potential mobility and the release rate of herbicide in the soil are important factors governing ecotoxicological impact and degradation rate. Therefore, several techniques are being investigated for its effective removal from the contaminated sites. Furthermore, efforts have also been made to study the degradation of pretilachlor by various physicochemical processes, resulting into the formation of different types of metabolites. This review summarizes the available information on environmental fate, various degradation processes, microbial biotransformation, metabolites formed, ecotoxicological effects, techniques for detection in environmental samples, effect of safener, and various control release formulations for sustained release of pretilachlor in applied fields. The information so obtained will be very advantageous in deciding the future policies for safe and judicious use of the herbicide by maintaining health and environmental sustainability.

Biodegradation of butachlor in rice intensified by a regulator of OsGT1.

The constant utilization of herbicide butachlor to prevent weeds in agronomic management is leading to its growing accumulation in environment and adverse impact on crop production and food security. Some technologies proposed for butachlor degradation in waters and farmland soils are available, but the catabolic mechanism in crops polluted with butachlor remains unknown. How plants cope with the ecotoxicity of butachlor is not only a fundamental scientific question but is also of critical importance for safe crop production and human health. This study developed a genetically improved rice genotype by overexpressing a novel glycosyltransferase gene named OsGT1 to accelerate removal of butachlor residues in rice crop and its growth environment. Both transcriptional expression and protein activates of OsGT1 are considerably induced under butachlor stress. The growth of the OsGT1 overexpression rice (OsOE) was significantly improved and butachlor-induced cellular damage was greatly attenuated compared to its wild-type (WT). The butachlor concentrations in shoots and roots of the hydroponically grown OsOE plants were reduced by 14.1-30.7 % and 37.8-47.7 %. In particular, the concentrations in the grain of OsOE lines were reduced to 54.6-85.6 % of those in wild-type. Using LC-Q-TOF-HRMS/MS, twenty-three butachlor derivatives including 16 metabolites and 7 conjugations with metabolic pathways were characterized, and it turns out that the OsOE lines accumulated more degradative products than wild-type, implying that more butachlor molecules were intensively catabolized. Taken together, the reduced residues of parent butachlor in rice and its growth media point out that OsGT1 plays a critical role in detoxifying and catabolizing the poisoning chemical in plants and its environment.

Synthesis, molecular modelling and QSAR study of new N-phenylacetamide-2-oxoindole benzensulfonamide conjugates as carbonic anhydrase inhibitors with antiproliferative activity.

In continuation of our previous studies to optimise potent carbonic anhydrase inhibitors, two new series of isatin N-phenylacetamide based sulphonamides were synthesised and screened for their human (h) carbonic anhydrase (EC 4.2.1.1) inhibitory activities against four isoforms hCA I, hCA II, hCA IX and hCA XII. The indole-2,3-dione derivative 2h showed the most effective inhibition profile against hCAI and hCA II (KI = 45.10, 5.87 nM) compared to acetazolamide (AAZ) as standard inhibitor. Moreover, 2h showed appreciable inhibition activity against the tumour-associated hCA XII, similar to AAZ showing KI of 7.91 and 5.70 nM, respectively. The analogs 3c and 3d showed good cytotoxicity effects, and 3c revealed promising selectivity towards lung cell line A549. Molecular docking was carried out for 2h and 3c to predict their binding conformations and affinities towards the hCA I, II, IX and XII isoforms.

Design, synthesis, and α-glucosidase-inhibitory activity of phenoxy-biscoumarin-N-phenylacetamide hybrids.

Thirteen new phenoxy-biscoumarin-N-phenylacetamide derivatives (7a-m) were designed based on a molecular hybridization approach as new α-glucosidase inhibitors. These compounds were synthesized with high yields and evaluated in vitro for their inhibitory activity against yeast α-glucosidase. The obtained results revealed that a significant proportion of the synthesized compounds showed considerable α-glucosidase-inhibitory activity in comparison to acarbose as a positive control. Representatively, 2-(4-(bis(4-hydroxy-2-oxo-2H-chromen-3-yl)methyl)phenoxy)-N-(4-bromophenyl)acetamide (7f), with IC50 = 41.73 ± 0.38 µM against α-glucosidase, was around 18 times more potent than acarbose (IC50 = 750.0 ± 10.0 µM). This compound was a competitive α-glucosidase inhibitor. Molecular modeling and dynamic simulation of these compounds confirmed the obtained results through in vitro experiments. Prediction of the druglikeness/ADME/toxicity of the compound 7f and comparison with the standard drug acarbose showed that the new compound 7f was probably better than the standard drug in terms of toxicity.

Cryo-EM structure of the ß3-adrenergic receptor reveals the molecular basis of subtype selectivity.

The ß3-adrenergic receptor (ß3AR) is predominantly expressed in adipose tissue and urinary bladder and has emerged as an attractive drug target for the treatment of type 2 diabetes, obesity, and overactive bladder (OAB). Here, we report the cryogenic electron microscopy structure of the ß3AR-Gs signaling complex with the selective agonist mirabegron, a first-in-class drug for OAB. Comparison of this structure with the previously reported ß1AR and ß2AR structures reveals a receptor activation mechanism upon mirabegron binding to the orthosteric site. Notably, the narrower exosite in ß3AR creates a perpendicular pocket for mirabegron. Mutational analyses suggest that a combination of both the exosite shape and the amino-acid-residue substitutions defines the drug selectivity of the ßAR agonists. Our findings provide a molecular basis for ßAR subtype selectivity, allowing the design of more-selective agents with fewer adverse effects.

Crystal Structure and Hirshfeld Surface Analysis of Acetoacetanilide Based Reaction Products.

We report an unprecedented multicomponent reaction of acetoacetanilide with malononitrile leading to a structurally novel bicyclic product (9) in a high yield. The structure has been confirmed by X-ray crystallography and comparative Hirshfeld surface analysis of 5-cyano-2-hydroxy-2-methyl-N-phenyl-4-(yridine-4-yl)-6-(thiophen-2-yl)-3,4-dihydro-2H-pyran-3-carboxamide 2, 5-cyano-2-hydroxy-2-methyl-6-oxo-N-phenyl-4-(thiophen-2-yl)piperidine-3-carboxamide 4 and 2-(8-amino-7,8a-dicyano-1-imino-4a-methyl-3-oxo-2-phenyl-1,3,4,4a,5,8a-hexahydroisoquinolin-6(2H)-ylidene)-N-phenylacetamide 9.

Effect of crop residue ashes on sorption behavior of herbicides used in the succeeding crop in Indian soils.

Effect of the wheat straw ash (WSA) on pretilachlor and the rice straw ash (RSA) on sulfosulfuron kinetics and adsorption behavior was studied. Kinetics study suggested that adsorption of herbicides in soil/soil + 0.2% ash mixture was best explained by the pseudo second order model. Ashes at 0.1%-0.5% levels increased adsorption of respective herbicide; but, effect varied with ash content and soil type. Effect of ash (0.2%) on herbicide's adsorption was more in the sandy loam soil (144%-188%) than in the clay loam soil (112%-122%) suggesting masking of ash particles. The Freundlich adsorption isotherm explained the adsorption of herbicides in the soils/soil + ash mixtures and sorption was highly nonlinear as 1/n (slope) values varied between 0.57 and 1.25 for pretilachlor and 0.32 and 0.77 for sulfosulfuron. Adsorption increased with increase in temperature. High surface area unburnt carbon in ashes was responsible for increase in adsorption and decrease in desorption of herbicides in ash mixed soils. The pH of soil/soil + ash mixtures affected herbicide adsorption, but effect was significant for pretilachlor. The negative free energy change (ΔG) values suggested that the sorption process was exothermic and spontaneous in nature. This study has implications in identifying the role of crop residue burning on fate of herbicides applied in succeeding crop.

Allosteric p97 Inhibitors Can Overcome Resistance to ATP-Competitive p97 Inhibitors for Potential Anticancer Therapy.

A major challenge of targeted cancer therapy is the selection for drug-resistant mutations in tumor cells leading to loss of treatment effectiveness. p97/VCP is central regulator of protein homeostasis and a promising anticancer target because of its vital role in cell growth and survival. One ATP-competitive p97 inhibitor, CB-5083, has entered clinical trials. Selective pressure on HCT116 cells dosed with CB-5083 identified five different resistant mutants. Identification of p97 inhibitors with different mechanisms of action would offer the potential to overcome this class of resistance mutations. Our results demonstrate that two CB-5083 resistant p97 mutants, N660 K and T688 A, were also resistant to several other ATP-competitive p97 inhibitors, whereas inhibition by two allosteric p97 inhibitors NMS-873 and UPCDC-30245 were unaffected by these mutations. We also established a CB-5083 resistant cell line that harbors a new p97 double mutation (D649 A/T688 A). While CB-5083, NMS-873, and UPCDC-30245 all effectively inhibited proliferation of the parental HCT116 cell line, NMS-873 and UPCDC-30245 were 30-fold more potent in inhibiting the CB-5083 resistant D649 A/T688 A double mutant than CB-5083. Our results suggest that allosteric p97 inhibitors are promising alternatives when resistance to ATP-competitive p97 inhibitors arises during anticancer treatment.

Discovery of 8-Methyl-pyrrolo[1,2-a]pyrazin-1(2H)-one Derivatives as Highly Potent and Selective Bromodomain and Extra-Terminal (BET) Bromodomain Inhibitors.

The bromodomain and extra-terminal (BET) family proteins have recently emerged as promising drug targets for cancer therapy. In this study, identification of an 8-methyl-pyrrolo[1,2-a]pyrazin-1(2H)-one fragment (47) as a new binder to the BET bromodomains and the subsequent incorporation of fragment 47 to the scaffold of ABBV-075, which recently entered Phase I clinical trials, enabled the generation of a series of highly potent BET bromodomain inhibitors. Further druggability optimization led to the discovery of compound 38 as a potential preclinical candidate. Significantly, compared with ABBV-075, which exhibits a 63-fold selectivity for BRD4(1) over EP300, compound 38 demonstrates an excellent selectivity for the BET bromodomain family over other bromodomains, with an ∼1500-fold selectivity for BRD4(1) over EP300. Orally administered 38 achieves a complete inhibition of tumor growth with a tumor growth inhibition (TGI) of 99.7% accompanied by good tolerability.

Pharmacophore-fusing design of pyrimidine sulfonylacetanilides as potent non-nucleoside inhibitors of HIV-1 reverse transcriptase.

Twenty-seven derivatives (40-66) were generated by pharmacophore fusing of sulfonylacetanilide-diarylpyrimidine (1) with rilpivirine or biphenyl-diarylpyrimidines. They displayed up to single-digit nanomolar activity against wild-type (WT) virus and various drug-resistant mutant strains in HIV-1-infected MT-4 cells, thereby targeting the reverse transcriptase (RT) enzyme. Compound 51 displayed exceptionally potent activity against WT virus (EC50 = 6 nM) and several mutant strains (L100I, EC50 = 8 nM, K103N, EC50 = 6 nM, Y181C, EC50 = 26 nM, Y188L, EC50 = 122 nM, E138K, EC50 = 26 nM). The structure-activity relationships of the newly obtained pyrimidine sulfonylacetanilides were also elucidated. Molecular docking analysis explained the activity and provided a structural insight for follow-up research.

Biodegradation of propanil by Acinetobacter baumannii DT in a biofilm-batch reactor and effects of butachlor on the degradation process.

The herbicide, propanil, has been extensively applied in weed control, which causes serious environmental pollution. Acinetobacter baumannii DT isolated from soil has been used to determine the degradation rates of propanil and 3,4-dichloroaniline by freely suspended and biofilm cells. The results showed that the bacterial isolate could utilize both compounds as sole carbon and nitrogen sources. Edwards's model could be fitted well to the degradation kinetics of propanil, with the maximum degradation of 0.027 ± 0.003 mM h-1. The investigation of the degradation pathway showed that A. baumannii DT transformed propanil to 3,4-dichloroaniline before being completely degraded via the ortho-cleavage pathway. In addition, A. baumannii DT showed high tolerance to butachlor, a herbicide usually mixed with propanil to enhance weed control. The presence of propanil and butachlor in the liquid media increased the cell surface hydrophobicity and biofilm formation. Moreover, the biofilm reactor showed increased degradation rates of propanil and butachlor and high tolerance of bacteria to these chemicals. The obtained results showed that A. baumannii DT has a high potential in the degradation of propanil.

Purity determination of a new antifungal drug candidate using quantitative 1 H NMR spectroscopy: Method validation and comparison of calibration approaches.

Quantitative nuclear magnetic resonance (qNMR) is an analytical technique that offers numerous advantages in pharmaceutical applications including minimum sample preparation and rapid data collection times with no need for response factor corrections, being a powerful tool for assaying drug content in both drug discovery and early drug development. In the present work, we have applied qNMR, using both the internal standard and the electronic reference to access in vivo concentrations 2 calibration methods, to assess the purity of RI76, a novel antifungal drug candidate. NMR acquisition and processing parameters were optimized in order to obtain spectra with intense, well-resolved signals of completely relaxed nuclei. The analytical method was validated following current guidelines, demonstrating selectivity, linearity, accuracy, precision, and robustness. The calibration approaches were statistically compared, and no significant difference was observed when comparing the obtained results and their dispersion in terms of relative standard deviation. The proposed qNMR method may, therefore, be used for both qualitative and quantitative assessments of RI76 in early drug development and for characterization of this compound.

Bimodal Molecule as NIR-CT Contrast Agent for Hepatoma Specific Imaging.

With currently available molecular imaging techniques, hepatocellular carcinoma (HCC), a liver cancer with high mortality rates and poor treatment responses, is mostly diagnosed at its late stage. This is largely due to the lack of highly sensitive contrast agents with high liver specificity. Herein, we report a novel bimodal contrast agent molecule CNCI-1 for the effective detection of HCC at its early stage both in vitro and in vivo. The agent has high liver specificity with effective X-ray computed tomography (CT)/near-infrared (NIR) imaging functions. It has been successfully applied to in vivo NIR imaging with high sensitivity and high selectivity to the HCC region of the HepG2 tumor-xenografted mice model and LM3 orthotopic hepatoma mice model. Moreover, the agent was found to be noninvasive and hepatocarcinoma cells preferential. Furthermore, it also enhanced the tumor imaging by revealing the blood vessels nearby for the CT image acquisition in the VX2 orthotopic hepatoma rabbit model. Our design strategy provides a new avenue to develop the medical relevant bimodal contrast agents for diagnosis of HCC at its early stage.

Cosolvent Analysis Toolkit (CAT): a robust hotspot identification platform for cosolvent simulations of proteins to expand the druggable proteome.

Cosolvent Molecular Dynamics (MD) simulations are increasingly popular techniques developed for prediction and characterization of allosteric and cryptic binding sites, which can be rendered "druggable" by small molecule ligands. Despite their conceptual simplicity and effectiveness, the analysis of cosolvent MD trajectories relies on pocket volume data, which requires a high level of manual investigation and may introduce a bias. In this work, we present CAT (Cosolvent Analysis Toolkit): an open-source, freely accessible analytical tool, suitable for automated analysis of cosolvent MD trajectories. CAT is compatible with commonly used molecular graphics software packages such as UCSF Chimera and VMD. Using a novel hybrid empirical force field scoring function, CAT accurately ranks the dynamic interactions between the macromolecular target and cosolvent molecules. To benchmark, CAT was used for three validated protein targets with allosteric and orthosteric binding sites, using five chemically distinct cosolvent molecules. For all systems, CAT has accurately identified all known sites. CAT can thus assist in computational studies aiming at identification of protein "hotspots" in a wide range of systems. As an easy-to-use computational tool, we expect that CAT will contribute to an increase in the size of the potentially 'druggable' human proteome.

Extraction of acetanilide herbicides in naked oat (Avena nuda L.) by using ionic-liquid-based matrix solid-phase dispersion-foam flotation solid-phase extraction.

The herbicides in naked oat (Avena nuda L.) samples were extracted, separated, and determined by using ionic-liquid-based matrix solid-phase dispersion-solvent flotation coupled with high-performance liquid chromatography. The experimental parameters were optimized and evaluated by a univariate method and orthogonal experiment. A good linear relationship was obtained in the range of 5-5000 µg/kg, and the linear correlation coefficient are between 0.9989∼0.9993. The quantification limits for alachlor, metazachlor, propanil, acetochlor, pretilachlor, metolachlor, and butachlor are 5.03, 2.62, 2.73, 4.58, 7.28, 5.05, 5.78 µg/kg, respectively. The average recoveries of the acetanilide herbicides at spiked concentrations of 10, 100, and 500 µg/kg ranged from 92.1 to 104.7%, and relative standard deviations were equal to or lower than 2.9%.

Extraction and separation of acetanilide herbicides in beans based on metal-organic framework MIL-101 (Zn) as sorbent.

An extraction method based on metal-organic framework has been developed and applied to acetanilide herbicides, including metazachlor, propanil, pretilachlor, and butachlor, in black beans, red beans, and kidney beans. The acetanilide herbicides are extracted with a mixture of ethyl acetate and n-hexane. The extract solution is absorbed and purified with metal-organic framework MIL-101 (Zn). The separation and determination of four acetanilide herbicides were implemented by high-performance liquid chromatography. The experimental parameters were evaluated by a univariate method and orthogonal experiments. The presented method can obtain effective extraction and purification. The detection limits for metazachlor, propanil, pretilachlor, and butachlor were 0.58, 0.90, 1.78 and 1.18 µg/kg, respectively. The average recoveries of the acetanilide herbicides at spiked concentrations of 10, 50, and 100 µg/kg ranged from 86.9% to 119.0%, and relative standard deviations were equal to or lower than 2.80%.

Multi-Armed 1,2,3-Selenadiazole and 1,2,3-Thiadiazole Benzene Derivatives as Novel Glyoxalase-I Inhibitors.

Glyoxalase-I (Glo-I) enzyme was established to be a valid target for anticancer drug design. It performs the essential detoxification step of harmful byproducts, especially methylglyoxal. A robust computer-aided drug design approach was used to design and validate a series of compounds with selenium or sulfur based heterorings. A series of in-house multi-armed 1,2,3-selenadiazole and 1,2,3-thiadiazole benzene derivatives were tested for their Glo-I inhibitory activity. Results showed that these compounds bind Glo-I active sites competitively with strong potential to inhibit this enzyme with IC50 values in micro-molar concentration. Docking poses revealed that these compounds interact with the zinc atom at the bottom of the active site, which plays an essential role in its viability.

Physicochemical properties and drug-release mechanisms of dual-release bilayer tablet containing mirabegron and fesoterodine fumarate.

Introduction: In this study, a dual release bi-layer tablet containing Fesoterodine fumarate (Fst) 5 mg and Mirabegron (Mrb) 50 mg was prepared to investigate the different release behavior of each drug in bilayer tablet. The bilayer tablet was prepared based on monolayer-tablet formulation of each drug. Methods: The optimized bi-layer tablet showed an in vitro dissolution profile similar to commercial reference tablets Toviaz and Betmiga, based on a satisfactory similarity factor. Drug-release kinetics of each drug in the bilayer tablet were evaluated based on dissolution profiles. Drug-release behavior was evaluated by observing the surface of each layer by scanning electron microscopy and measuring the changes in weight and volume of the tablet during dissolution. Drug transfer between each layer was also investigated by Fourier -transform infrared spectroscopic imaging by observing the cross-section of the bilayer tablet cut vertically during dissolution. Results: The release of Fst was well suited for the Higuchi model, and the release of Mrb was well suited for the Hixson-crowell model. Compared with dissolution rate of each monolayer tablet, that of Fst in the bilayer tablet was slightly reduced (5%), but the dissolution rate of Mrb in bilayer tablet was dramatically decreased (20%). Also, a drug-release study confirmed that polymer swelling was dominant in Fst layer compared with polymer erosion, and degradation was dominant in MRB layer. Fourier-transform infrared imaging and 3-D image reconstruction showed that drug transfer in the bilayer tablet correlates with the results of drug-release behavior. Conclusion: These findings are expected to provide scientific insights in the development of a dual-release bilayer drug-delivery system for Fst and Mrb.