Discovery of SARS-CoV-2 antiviral drugs through large-scale compound repurposing.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 2019 has triggered an ongoing global pandemic of the severe pneumonia-like disease coronavirus disease 2019 (COVID-19)1. The development of a vaccine is likely to take at least 12-18 months, and the typical timeline for approval of a new antiviral therapeutic agent can exceed 10 years. Thus, repurposing of known drugs could substantially accelerate the deployment of new therapies for COVID-19. Here we profiled a library of drugs encompassing approximately 12,000 clinical-stage or Food and Drug Administration (FDA)-approved small molecules to identify candidate therapeutic drugs for COVID-19. We report the identification of 100 molecules that inhibit viral replication of SARS-CoV-2, including 21 drugs that exhibit dose-response relationships. Of these, thirteen were found to harbour effective concentrations commensurate with probable achievable therapeutic doses in patients, including the PIKfyve kinase inhibitor apilimod2-4 and the cysteine protease inhibitors MDL-28170, Z LVG CHN2, VBY-825 and ONO 5334. Notably, MDL-28170, ONO 5334 and apilimod were found to antagonize viral replication in human pneumocyte-like cells derived from induced pluripotent stem cells, and apilimod also demonstrated antiviral efficacy in a primary human lung explant model. Since most of the molecules identified in this study have already advanced into the clinic, their known pharmacological and human safety profiles will enable accelerated preclinical and clinical evaluation of these drugs for the treatment of COVID-19.

Dissipation and residues of fluazinam and dimethomorph in potatoes, potato plants, and soil, determined by QuEChERS ultra-performance liquid chromatography tandem mass spectrometry.

Fluazinam and dimethomorph 35% suspension concentrate (SC) is a new combined fungicide formulation introduced in China to improve fungicidal efficacy and decrease the risk of resistance in potatoes. Fluazinam and dimethomorph dissipation and residues in potatoes, potato plants, and soil under field conditions were determined by ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). Fluazinam and dimethomorph 35% SC was applied at two doses to potatoes and soil in Ningxia Autonomous Region and Anhui Province, China. Fluazinam and dimethomorph dissipation fitted first-order kinetics, and the fluazinam half-lives in potato plants and soil were 3.3-5.4 and 9.4-9.5 days, respectively. The dimethomorph half-lives in potato plants and soil were 2.1-2.6 and 5.9-8.6 days, respectively. Fluazinam and dimethomorph 35% SC was sprayed onto potato plants three or four times at application rates of 420 and 630 g a.i. ha-1 with 7 days between applications. Potato and soil samples were collected at 3, 7, and 14 days after the last application. Potatoes and soil had fluazinam concentrations of < 0.01 and < 0.05-0.183 mg kg-1, respectively, and dimethomorph concentrations of < 0.01 and 0.129-0.677 mg kg-1, respectively. The final fluazinam and dimethomorph concentrations in potatoes were below the EU maximum residue limits (0.02 and 0.05 mg kg-1, respectively) 3 days after application. Fluazinam and dimethomorph can therefore be applied to potatoes at the recommended doses.

Residual dynamic and risk assessment of dimethomorph in Swiss chard grown at two different sites.

Residue analysis of dimethomorph in Swiss chard cultivated at two different locations under greenhouse conditions was conducted using high-performance liquid chromatography-ultraviolet detection and confirmed by tandem mass spectrometry. The randomly collected samples (over 14 days) were extracted with acetonitrile and purified using a Florisil solid-phase extraction cartridge. Linearity over a concentration range of 0.05-50.0 mg/L had an excellent coefficient of determination of 0.9996. Recovery rate ranged from 82.98 to 95.43% with relative standard deviations ≤5.12% and limits of detection and quantification of 0.003 and 0.01 mg/kg, respectively. The initial deposits [day 0 (2 h post-application)] were considerably lower (7.57 and 8.55 mg/kg for sites 1 and 2, respectively) than the maximum residue limit (30 mg/kg) set by the Korean Ministry of Food and Drug Safety. The dissipation half-life was approximately the same, being 5.0 and 5.1 days for sites 1 and 2, respectively. Risk assessment estimated as acceptable daily intake revealed a value of 0.084 or 0.094% (day 0) and 0.014% (10 days post-application), for sites 1 and 2, respectively. The values indicated that dimethomorph can be safely used on Swiss chard, with no hazardous effects expected for Korean consumers.

A molecularly imprinted polymer for the selective solid-phase extraction of dimethomorph from ginseng samples.

A molecularly imprinted polymer (MIP) was synthesized and evaluated to selectively extract dimethomorph from ginseng samples. Dimethomorph molecularly imprinted polymers with template to monomer molar ratios were contrived and developed via precipitation polymerization employing methacrylic acid as functional monomer, ethylene dimethacrylate as cross-linker and butanone:N-heptane (7:3, v:v)as porogen. The LOD (limit of detection) of this method was 0.002 mg kg(-1), and the LOQ (limit of quantification) was 0.005 mg kg(-1). The different spiked level of ginseng was 0.1 mg kg(-1), 1.0 mg kg(-1), 5.0 mg kg(-1), and the average recovery of dimethomrph was 89.2-91.6%. Under the optimized condition, good linearity was obtained from 0.01 to 5 mg kg(-1) (r(2) ≥ 0.9997) with the relative standard deviations of less than 3.20%. This proposed MISPE-GC procedure eliminated the effect of template leakage on quantitative analysis and could be applied to direct determination of dimethomrph in ginseng samples.

Mass transfer ways of ultraviolet printing ink ingredients into foodstuffs.

The case of isopropylthioxanthone (ITX) showed conclusively that the ingredients of ultraviolet printing inks may migrate into packaged foodstuffs. For multilayered materials like beverage cartons, the only way that mass transfer can occur is by the so-called set-off effect. In contrast, in the case of rigid plastics like yoghurt cups, two other methods of mass transfer, permeation and gas phase, have to be considered. In cooperation with producers of ink, plastic cups and yoghurt, a project was conducted in order to elucidate the mass transfer of ink ingredients. In addition, the influence of storage time and the age of ultraviolet lamps on the migration level was examined. The suitability of 50% ethanol as a simulant for yoghurt was also tested. ITX was chosen as a model migrant, as it is easily detectable. Furthermore, the migration of two other substances, the photo-initiator 2-methyl-4'-(methylthio)-2-morpholinopropiophenone (MTMP) and the amine synergist ethyl-4-(dimethylamino)benzoate (EDAB), which may be used in combination with ITX, was studied. Before being filled with yoghurt or 50% ethanol, the printed cups were stored under different contact conditions, with and without contact between the inner layer and the printed surfaces, in order to distinguish between the possible mass transfer ways. All analyses were performed by means of high performance liquid chromatography with diode array and fluorescence detection (HPLC-DAD/FLD). It was shown that contamination with ITX and EDAB occurs via set-off and that the degree of migration increases with lamp age and storage time of the unfilled cups. Migration of MTMP was not detectable. The results show that besides the careful selection of the appropriate raw materials for printing ink, a close monitoring of the process also plays a major role in migration control. In addition, the results proved that 50% ethanol is a suitable simulant for yoghurt.

Variations in amino acid composition of antisense peptide-phosphorodiamidate morpholino oligomer affect potency against Escherichia coli in vitro and in vivo.

The potency of antisense peptide-phosphorodiamidate morpholino oligomers (PPMOs) was improved by varying the peptide composition. An antisense phosphorodiamidate morpholino oligomer (PMO) complementary to the mRNA of the essential gene acpP (which encodes the acyl carrier protein required for lipid biosynthesis) in Escherichia coli was conjugated to the 5' ends of various cationic membrane-penetrating peptides. Each peptide had one of three repeating sequence motifs: C-N-N (motif 1), C-N (motif 2), or C-N-C (motif 3), where C is a cationic residue and N is a nonpolar residue. Variations in the cationic residues included arginine, lysine, and ornithine (O). Variations in the nonpolar residues included phenylalanine, valine, beta-alanine (B), and 6-aminohexanoic acid (X). The MICs of the PPMOs varied from 0.625 to >80 microM (about 3 to 480 microg/ml). Three of the most potent were the (RX)(6)B-, (RXR)(4)XB-, and (RFR)(4)XB-AcpP PMOs, which were further tested in mice infected with E. coli. The (RXR)(4)XB-AcpP PMO was the most potent of the three conjugates tested in mice. The administration of 30 microg (1.5 mg/kg of body weight) (RXR)(4)XB-AcpP PMO at 15 min postinfection reduced CFU/ml in blood by 10(2) to 10(3) within 2 to 12 h compared to the numbers in water-treated controls. All mice treated with 30 microg/dose of (RXR)(4)XB-AcpP PMO survived infection, whereas all water-treated mice died 12 h postinfection. The reduction in CFU/ml in blood was proportional to the dose of PPMO from 30 to 300 microg/ml. In summary, the C-N-C motif was more effective than the other two motifs, arginine was more effective than lysine or ornithine, phenylalanine was more effective than 6-aminohexanoic acid in vitro but not necessarily in vivo, and (RXR)(4)XB-AcpP PMO reduced bacterial infection and promoted survival at clinically relevant doses.

Volatilization of the pesticides chlorpyrifos and fenpropimorph from a potato crop.

Volatilization of pesticides from crops in the field can be an important emission pathway. In a field experiment with characterization of meteorological conditions, the pesticides chlorpyrifos and fenpropimorph were sprayed onto a potato crop, after which concentrations in the air and on/in the plants were measured. Rates of volatilization were estimated with the aerodynamic profile (ADP), energy balance (EB), relaxed eddy accumulation (REA), and plume dispersion (PD) methods. The volatilization rates obtained with the ADP and EB methods were similar, while some rates obtained with the REA and PD methods in the initial period were lower. Cumulative volatilization of chlorpyrifos during daylight hours (ADP and EB methods) was estimated to be about 65% of the dosage. By far the majority of this volatilization occurred in the first few days. Competing processes at the plant surface had a considerable effect on the dissipation of fenpropimorph, so cumulative volatilization during daylight hours was estimated to be only 7% of the dosage. Plant surface residues were higher than would correspond with the volatilization rate, indicating that penetration into the leaves had occurred.

Measured and computed volatilisation of the fungicide fenpropimorph from a sugar beet crop.

Depending on their vapour pressure, volatilisation can be one of the main pathways of emission of pesticides into the environment. The volatilisation of fenpropimorph was studied in a field experiment in which the fungicide was sprayed onto a sugar beet crop. Volatilisation rates were calculated by measuring the concentration gradient in air, using the Aerodynamic and Bowen Ratio methods. A simplified computation model was used to simulate pesticide volatilisation, together with the concurrent processes of penetration into the plant leaves and phototransformation. Input data for the model had already been obtained by carrying out a wind-tunnel study with fenpropimorph, whereby field conditions were imitated. The computations yielded a reasonable description of the level and rate of decline of fenpropimorph volatilisation in the first 4 h after spraying. The continued volatilisation 2 and 3 days after spraying could be described by assuming that a fraction of the deposit was poorly exposed with comparatively low rates of the decline processes. In the first 3 days, penetration of fenpropimorph into the plant leaves was computed to be the main route for the pesticide (52% of the dosage), with substantial contributions from volatilisation (12%) and phototransformation (11%). The computation model can be developed further as a tool for extrapolating results on volatilisation from small-scale experiments to field conditions, but this requires more information on the effect of environmental conditions on the model parameters.

Identification of aminoethylcysteine ketimine decarboxylated dimer, a natural antioxidant, in dietary vegetables.

Aminoethylcysteine ketimine decarboxylated dimer (simply named dimer) is a natural sulfur-containing tricyclic compound detected, until now, in human urine, bovine cerebellum, and human plasma. Recently, the antioxidant properties of this compound have been demonstrated. In this investigation, the presence of aminoethylcysteine ketimine decarboxylated dimer was identified in garlic, spinach, tomato, asparagus, aubergine, onion, pepper, and courgette. Identification of this compound in dietary vegetables was performed using gas chromatography, high-performance liquid chromatography, and gas chromatography-mass spectrometry. Results from GC analysis range in the order of 10(-4) micromol of dimer/g for all the tested vegetables. These results and the lack of a demonstrated biosynthetic pathway in humans might account for a dietary supply of this molecule.

[Application of derivative spectrophotometric signal multiplier method in multicomponent mixture--determination of moroxydine hydrochloride in Gan Mao Qing capsules].

This paper provides a basic principle and experimental technique of derivative signal multiplier spectrophotometry in multicomponent mixture. A microcomputer was used to process the spectral data measured on a manual spectrophotometer (UV-7520) for the determination of moroxydine hydrochloride in Gan Mao Qing capsules. Quantitative analysis of multicomponent mixture can be done without sample separation. The selection of optimal wavelength pairs is performed through the program with a computer. The method needs no special spectrophotometer and is simple, rapid and easy to operate. The mean recovery was 99.98 +/- 0.53% (n = 12).