Free energy perturbation (FEP)-guided scaffold hopping

Scaffold hopping refers to computer-aided screening for active compounds with different structures against the same receptor to enrich privileged scaffolds, which is a topic of high interest in organic and medicinal chemistry. However, most approaches cannot efficiently predict the potency level of candidates after scaffold hopping. Herein, we identified potent PDE5 inhibitors with a novel scaffold via a free energy perturbation (FEP)-guided scaffold-hopping strategy, and FEP shows great advantages to precisely predict the theoretical binding potencies ΔG FEP between ligands and their target, which were more consistent with the experimental binding potencies ΔG EXP (the mean absolute deviations | Δ G FEP - Δ G EXP |  < 2 kcal/mol) than those ΔG MM-PBSA or ΔG MM-GBSA predicted by the MM-PBSA or MM-GBSA method. Lead L12 had an IC50 of 8.7 nmol/L and exhibited a different binding pattern in its crystal structure with PDE5 from the famous starting drug tadalafil. Our work provides the first report via the FEP-guided scaffold hopping strategy for potent inhibitor discovery with a novel scaffold, implying that it will have a variety of future applications in rational molecular design and drug discovery.

Discovery of novel diarylamides as orally active diuretics targeting urea transporters

Urea transporters (UT) play a vital role in the mechanism of urine concentration and are recognized as novel targets for the development of salt-sparing diuretics. Thus, UT inhibitors are promising for development as novel diuretics. In the present study, a novel UT inhibitor with a diarylamide scaffold was discovered by high-throughput screening. Optimization of the inhibitor led to the identification of a promising preclinical candidate,

Thermodynamic analysis and preferential solvation of gatifloxacin in DMF + methanol cosolvent mixtures / Análisis termodinâmico y solvatación preferencial de gatifloxacina en mezclas cosolventes DMF + metanol

SUMMARY This paper presents the thermodynamic analysis of solubility of gatifloxacin in the N,N-Dimethylformamide (DMF) + methanol (MeOH) cosolvent system at 10 temperatures. From the solubility data, the thermodynamic functions of solution, mixing, and transfers are calculated and analyzed using the Perlovich graphical method. On the other hand, an enthalpy-entropy compensation analysis is performed and the preferential solvation parameters are calculated using the inverse Kirkwood-Buff integral (IKBI) method. The result of the performed calculations indicates that the gatifloxacin solution process is endothermic with entropic favor, where the addition of DMF has a positive cosolvent effect in all cases. Regarding preferential solvation, the results are not entirely conclusive, since in all cases the values of the preferential solvation parameter are less than 0.01, so that, negligible preferential solvation takes place.

RESUMEN Este artículo presenta el análisis termodinâmico de la solubilidad de gatifloxacina en el sistema cosolvente de A,A-Dimetilformamida (DMF) + metanol (MeOH) a 10 temperaturas. A partir de los datos de solubilidad se calculan las funciones termodinámicas de solución, mezcla y transferencia. Para el análisis además se utiliza el método gráfico Perlovich. Por otro lado, se realiza un análisis de compensación entalpía-entropía y se calculan los parámetros de solvatación preferencial utilizando el método de las integrales inversas de Kirkwood-BufF (IIKB). Los resultados del análisis termodinámico indican que el proceso de solución de gatifloxacina es endotérmica con favorecimiento entrópico, donde la adición de DMF tiene un efecto cosolvente positivo en todos los casos. En cuanto a la solvatación preferencial, los resultados no son del todo concluyentes, debido a que en todos los casos los valores del parámetro de solvatación preferencial son menores a 0,01 indicando una solvatación insignificante.

Identificación de biomarcadores saturados en un carbón bituminoso tratado con ácido y un solvente no convencional / Identification of biomarkers in a bituminous coal treated with acid and an unconventional solvent / Identificação de biomarcadores saturados em carvão betuminoso tratado com ácido e solvente não convencional

Resumen Este artículo describe los biomarcadores saturados (n־alcanos, hopanos y esteranos) identificados en los extractos obtenidos de un carbón bituminoso, tratado con ácido y un solvente no convencional. Los extractos de carbones bituminosos venezolanos se obtuvieron a través de cloroformo (solvente convencional) y N,N־dimetilformamida ־DMF־ como solvente de alto poder extractivo, con y sin tratamiento ácido previo, con el fin de evaluar el rendimiento y distribución de biomarcadores característicos. El rendimiento de extracción con DMF alcanzó 44% p/p, mientras que con CHCl3, este valor no superó el 2% p/p. La desmineralización previa con HCl incrementó el rendimiento de extracción sin afectar la distribución de los biomarcadores, salvo en la relación de hidrocarburos ligeros respecto a los pesados. Se propone DMF como solvente de elección para obtener información geoquímica en carbones meteorizados.

Abstract This article describes saturated biomarkers (n־alkanes, hopanes and steranes) identified in the extracts obtained from a bituminous coal treated with acid and an unconventional solvent. Extracts of Venezuelan bituminous coals were obtained through chloroform (conventional solvent) and N,N־ dimethylformamide ־DMF־ as high extractive solvent, with and without previous acid treatment, to evaluate the performance and distribution of characteristic biomarkers. The extraction yield with DMF reached 44% w/w, while with CHCl3, this value did not exceed 2% w/w. Previous demineralization with HCl increased the extraction yield without affecting the distribution of the biomarkers, except in the ratio of light hydrocarbons to the heavy ones. DMF, therefore, is proposed as the solvent of choice to obtain geochemical information on weathered carbons.

Resumo Este artigo de pesquisa descreve os biomarcadores saturados identificados (n־alcanos, hopanos y esteranos) em extratos obtidos a partir de carvão betuminoso, tratado com ácido e um solvente não convencional. Extratos de carvão betuminoso venezuelano foram obtidos utilizando־se clorofórmio (solvente convencional) e N,N־ dimetilformamida ־DMF־ como solvente extrativo elevado, com e sem tratamento ácido prévio, com o objectivo de avaliar o rendimento e distribuição de biomarcadores característicos. O rendimento de extração com DMF atingiu 44% p/p; com CHCl3 o valor não excedeu 2% p/p. A desmineralização prévia com HCl aumentou o rendimento da extração sem afetar a distribuição dos biomarcadores, exceto na proporção de hidrocarbonetos leves e pesados. DMF é proposto como o solvente de eleição para obter informações geoquímicas sobre os carvões intemperizados.

Research on the inflammatory injury induced by N,N-dimethylformamide in H9c2 cardiomyocytes / 中国职业医学

OBJECTIVE: To explore the effect of N,N-dimethylformamide( DMF)-induced inflammatory injury in H9c2 cardiomyocytes and its mechanism. METHODS: H9c2 cardiomyocytes were cultured in vitro and randomly divided into 4different groups: control group,50 mmol / L-group,100 mmol / L-group,200 mmol / L-group. These 4 groups of cells were treated with different DMF concentrations( 0,50,100,200 mmol / L) for 12 hours. The cells were also divided into 6groups and treated with 200 mmol / L DMF at different time points( 0,2,4,6,8,12 h) : control group,2 h-group,4 hgroup,6 h-group,8 h-group and 12 h-group. The level of lactate dehydrogenase( LDH) was detected by colorimetry. The levels of creatine kinase( CK) and isoenzyme of creatine kinase( CK-MB) were detected by ultraviolet spectrometry. The levels of tumor necrosis factor-α( TNF-α),interleukin( IL)-1β,IL-6,and IL-8 were detected by enzyme linked immunosorbent assay. The level of reactive oxygen species( ROS) was detected by fluorescence probe. The location of nuclear factor-kappa B( NF-κB) p65 protein was detected by immunofluorescence cytochemistry( IFC) staining. RESULTS: The levels of LDH,CK and CK-MB in the 50 mmol / L-group,100 mmol / L-group and 200 mmol / L-group were higher than that of the control group( P < 0. 05) and showed a significant dose-effect( P < 0. 05). The levels of LDH,CK and CK-MB in the 6 h-group,8 h-group and 12 h-group were higher than that of the control group( P < 0. 01) and showed a significant time-effect( P < 0. 01). The levels of TNF-α,IL-1β,IL-6 and IL-8 of the 200 mmol / L-group were higher than the control group( P < 0. 05). Compared with the control group,the levels of TNF-α of the 4 h-group,12 h-group were higher( P < 0. 05),the levels of IL-1β of the 2 h-group,4 h-group,6 h-group,8 h-group and 12 h-group were higher( P < 0. 05),the levels of IL-6 of the 2 h-group and 4 h-group were higher( P < 0. 05),the level of IL-8 of the 2 h-group was higher( P < 0. 05). In addition,the levels of TNF-α,IL-1β and IL-6 reached a peak at 4 h-group and the level of IL-8 reached a peak at 2 h-group. The ROS levels of the 2 h-group,4 h-group and 6 h-group were higher than the control group( P < 0. 01),and the level of ROS reached a peak at 2 h-group. Furthermore,IFC staining showed that the fluorescence intensity of NF-κB p65 protein in nucleus of the 2h-group and 4 h-group increased after treatment with DMF,comparing with the control group. CONCLUSION: DMF leads to inflammatory injury in H9c2 cardiomyocytes. ROS and NF-κB might be involved in the process.

Rapid Determination of N-Methylcarbamoyl Adduct in Hemoglobin of Workers Exposed to N,N-Dimethylformamide by Ultra High Performance Liquid Chromatography-Mass Spectrometry / 分析化学

A rapid analytical method was developed for the determination of N-methylcarbamoyl adduct in Hemoglobin of workers exposed to N,N-dimethylformamide by ultra high performance liquid chromatography-mass spectrometry ( UPLC/MS/MS). About 0. 1 g of hemoglobin sample, 40 μmol/L of 3-methyl-5-isopropylhydantoin (MIH) as the internal standard and 4. 75 mL of HCl-acetic acid (2∶1, V/V) were added in the centrifuge tube, and mixed for 3 min. Then the tube was heated in boiling water bath for 1h. After cooling down, 200 μL of the mixture and 600 μL of formic acid-acetonitrile (1%) were added into 96-well extract plate. The vacuum pump pressure was controlled to make the sample collection elute within 2-4 min.The purified collection was transferred into the sample vial, and 3-methyl-5-isopropylhydantoin ( MVH ) as degration product of N-methylcarbamoyl adduct was quantified by UPLC/MS/MS in multiple reaction monitoring ( MRM ) by internal standard method. A good linear relationship was obtained in the MVH concentration range of 0 . 01-1 . 0 μmol/L with the correlation coefficient of 0 . 999 . The recovery of added MVH in the blank sample was 97 . 3% and the relative standard deviation was 1 . 7%. The limit of detection (LOD) was 0. 01 μmol/g. This method was proved to be fast and efficient.

Transmission FTIR derivative spectroscopy for estimation of furosemide in raw material and tablet dosage form

A Fourier transform infrared derivative spectroscopy (FTIR-DS) method has been developed for determining furosemide (FUR) in pharmaceutical solid dosage form. The method involves the extraction of FUR from tablets with N,N-dimethylformamide by sonication and direct measurement in liquid phase mode using a reduced path length cell. In general, the spectra were measured in transmission mode and the equipment was configured to collect a spectrum at 4 cm(-1) resolution and a 13 s collection time (10 scans co-added). The spectra were collected between 1400 cm(-1) and 450 cm(-1). Derivative spectroscopy was used for data processing and quantitative measurement using the peak area of the second order spectrum of the major spectral band found at 1165 cm(-1) (SO2 stretching of FUR) with baseline correction. The method fulfilled most validation requirements in the 2 mg/mL and 20 mg/mL range, with a 0.9998 coefficient of determination obtained by simple calibration model, and a general coefficient of variation <2%. The mean recovery for the proposed assay method resulted within the (100±3)% over the 80%-120% range of the target concentration. The results agree with a pharmacopoeial method and, therefore, could be considered interchangeable.

Effect of acute exposure of N, N-Dimethylformamide, an industrial solvent on lipid peroxidation and antioxidants in liver and kidney of rats.

N, N-Dimethylformamide (DMF), an industrial solvent widely used throughout the world is a known toxic compound. Here, we studied the effects of acute exposure of DMF on liver and kidney in rats. Rats were treated intraperitoneally with a single dose of DMF (1.5 g/kg) for 24 and 48 h. Hepatic and nephrotoxicity was confirmed based on the significant increase in the serum levels of aspartate amino transferase, alanine amino transferase, alkaline phosphatase, g-glutamyl transferase, urea, creatinine and electrolytes. Oxidative stress was assessed by determining the levels of lipid peroxidation (LPO) and antioxidants in liver and kidney. The LPO levels were elevated in both the tissues upon DMF exposure, whereas the activities of enzymatic antioxidants SOD, CAT and Gpx and non-enzymatic antioxidants (glutathione and vitamin C) were declined. The hepatic- and nephrotoxicity was further confirmed by the increasing incidence of inflammation in the histopathological studies. The findings indicate that acute exposure of DMF results in oxidative stress, antioxidant deficiency, attenuating liver and kidney marker enzymes, resulting in tissue inflammation and damage.

Comparação de métodos diretos de extração e quantificação dos teores de clorofilas em folhas do capim-Tifton 85 / Comparison of direct methods of extraction and chlorophyll contents quantification in leaves of Tifton 85 bermudagrass

O objetivo deste estudo foi verificar qual é o melhor protocolo para extração das clorofilas a, b e total (a + b) a ser aplicado em estudos com a gramínea forrageira Tifton 85 (Cynodon spp.). Os resultados revelaram que os melhores extratores, tomando-se como base os teores de clorofila total (a + b), foram o N,N-dimetilformamida, o dimetilsulfóxido e a acetona 80 por cento (com as equações propostas por ARNON, 1949). A variação temporal dos teores das clorofilas a, b e total ajustaram-se a um modelo hiperbólico comum a todos os solventes, sendo o período de 48 horas considerado suficiente para uma adequada extração.

This study aimed to assess which is the most appropriate protocol for extraction of chlorophylls a, b and total (a + b) to be applied in studies with Tifton 85 (Cynodon spp.) forage grass. Results revealed that, based on total (a + b) chlorophyll contents, the best extractors were N,N-dimethylformamide, dimethylsulfoxide and acetone 80 percent (using the equations proposed by ARNON, 1949). Temporal changes in the concentrations of chlorophyll a, b and total were well described by a hyperbolic model common to all solvents, being the period of 48 hours considered sufficient for an appropriate extraction.

Determination of N,N-dimethylformamide in Air by Capillary Gas Chromatography / 环境与健康杂志

0.999) . The limit of detection was 4.0 ?10-4 ?g. The relative standard deviation was 0.9%-7.5% and the desorption efficiency of DMF was 86.5%-94.0%. Conclusion The method is applicable to the determination of DMF in the air.

Phenotypic and Functional Differentiation of Promyelocytic Cell Line HL-60 by N-N-dimethylformamide

PURPOSE: HL-60 is a promyelocytic cell line. Fc receptors and complement receptor 3 (CR3) play important role in the protective response of granulocytes and monocytes against microbial infection. We quantified the expression of Fc I, Fc II, Fc III, and CD11b/CD18 during differentiation using HL-60 cells by N-N-dimethylformamide (DMF). Functional studies, such as phagocytic activity, respiratory burst and ADCC, were also performed. METHODS: HL-60 cells were induced to differentiate by adding 0.8% DMF. On the 4th and 7th day after stimulation as well as before stimulation, the cells were analyzed for phenotypic and functional differentiaton. Phenotypic analysis was performed by flow cytometry after staining the cells with PE-conjugated anti-human CD64, CD32, CD16, CD11b, CD18, and isotype controls. And the measured fluorescent intensity was transformed into Molecules of Equivalent Soluble Fluorochromes (MESF). Phagocytic activity was also measured by flow cytometry after incubation with fluorochrome-conjugated beads. Respiratory burst was measured by chemiluminescence assay of cells incubated with luminol after stimulation with PMA. ADCC was measured by hemoglobin release assay. RESULTS: The expression of CD11b, CD18 and CD64 on HL-60 cells markedly increased on the 4th day and slightly decreased on the 7th day. Expression of CD32 was already induced before differentiation induction and slightly increased by DMF. CD16 was not expressed during differentiation. In phagocytic assay, the phagocytic cell fraction increased by stimulation on 4th and 7th day. Chemiluminescence showed the DMF increased the respiratory burst of HL-60 cells on the 4th and 7th day. In ADCC, DMF increased the target cell lysis continuously. CONCLUSION: HL-60 cells which were differentiated with DMF for are good models for studying opsonophagocytic assay of immunized sera.

The Metabolism and Liver Toxicity of N, N-dimethylformamide in the Isolated Perfused Liver / 대한산업의학회지

N, N-dimethylformamide (DMF) is a solvent which is widely used in the industrial workplace. It causes the liver damages to the chronically exposed workers and is also well known as the harzadous material to generate occupational malignancies. DMF is mainly metabolized into N-hydroxymethyl-N-methylformamide (HMMF) by the microsomal cytochrome p-450. HMMF breaks down to NMF. However, the detailed mechanism of its toxicity are unknown. In this experiment, the metabolism and the toxicity of DMF was investigated using an isolated perfumed liver model. DMF (0, 10, 25mM) were added into recirculating perfusate of the isolated perfused rat liver. Samples were collected at 0, 30, 45, 60, 75, 90 minutes from inferior vena cava. The gas-chromatography was used to analyze the metabolite of DMF, The changes in the oxygen consumption rate by DMF were monitored during perfusion. The enzyme activity (AST, ALT, LDH) in the perfusate were treasured to find out whether DMF causers hepatotoxicity. As perfusion continued, DMF concentration in the perfusate decreased, and NMF 1.16mM was detected. The oxygen consumption rate increased both at 10mM and 25mM DMF concentration. However, when SKF 525A, a known inhibitor of cytochrome p-450, had been pretreated (300uM before DMF addition, the oxygen consumption rate was significantly inhibited, indicating that cytochrome p-450 system is responsible for the conversion to NMF. With DMF addition, the activity of AST, ALT, and LDH significantly increased time dependently and dose dependently. However, the pretreatment of perfused liver with SKF 525A shoved that the release of AST, ALT and LDH was inhibited. In summary, it is found that DMF is metabolized to NMF in liver, and that cytochrome p-450 mono-oxygenase is suggested to play a role in the biotransformation of NMF. The time course of BMF toxicity in relation to NMF formation is compatible with hypothesis that the hepatotoxicity of DMF is mediated via NMF. Further study combined with in vivo experiment through the toxicological approaches is expected.

Reproductive Toxicity of N, N-dimethylformamide / 环境与健康杂志

N, N-dimethylformamide (DMF) is a kind of organic compound which is widely used in the modern industry. With the increasing utilization of DMF year by year, the interest in the research of DMF toxicity has increased dramatically. The data of animal experiments and epidemiological investigations show that DMF has harmful effect on reproductive function of human and animals. The effect of DMF on the reproductive function of animal and occupational crowd were reviewed in this paper, the research perspective was also presented.

Determination of N,N-Dimethylformamide in oligosaccharide of medicine by Gas Chromatography / 中成药

AIM: The quantitative determination of N,N-dimethylf in olgosaccharide was carried out. METHODS: Gas Chromatographic was employed. RESULTS: Experimental results showed that a good separation of DMF with other contents of sample was achieved on FFAP(50 m?0.32 mm) column,and the separation degree was more than 1.5.The linear range was 3.58～0.000358 mg/mL with r=0.999 9 and the relative standard deviation was 1.52%(n=5).and the limit of detection was 1.0 mg/mL.The recovery of the method was in the range of 95.7%～98.4%. CONCLUSION: The method is simple,rapid,accurate and precise.