Alamar Blue Method Research Articles

New Bis 4-Bromobenzene Sulphonate Compound: Synthesis, Characterizaton, DFT study, Antibacterial and Leishmanicidal activity

The new bis 4-bromobenzene sulphonate compound called as ((1E,1'E)-(1,4-phenylenebis(azanylylidene))bis(methanylylidene))bis(4,1-phenylene) bis(4-bromobenzenesulfonate) (I) was synthesized. Fourier transform infrared (FTIR), proton and carbon-13 nuclear magnetic resonance (1H- and 13C-NMR) methods were used to confirm the molecular structure. All calculations were performed at DFT/B3LYP/6-311G(d,p) level. IR and NMR spectral data were compared with experimental ones and the details of molecular structure were investigated. The antimicrobial activity of the synthesized compound (I) was determined by Alamar blue microdilution method against seven different established standard bacteria and one parasite isolate. The compound I exhibited antibacterial activity at different concentrations. Compound I was found to have antimicrobial activity on all bacterial species and L. infantum parasite, albeit at different concentrations. Compound I has the most effective antibacterial activity on S. flexneri and the least effect on S. aureus and E. cloacea bacteria. In order for the synthesized compound to be used as a drug candidate, in vivo control studies in a series of experimental animal models and whether it has toxic effects on human cells should be tested. In silico analysis was conducted on four different proteins using the synthesized compound I to investigate the essential interactions responsible for its antibacterial activity. The docking results of compound I supported its antibacterial activity, revealing high inhibition constants.

Evaluation of Selected Plant Phenolics via Beta-Secretase-1 Inhibition, Molecular Docking, and Gene Expression Related to Alzheimer's Disease.

Background: The goal of the current study was to investigate the inhibitory activity of six phenolic compounds, i.e., rosmarinic acid, gallic acid, oleuropein, epigallocatechin gallate (EGCG), 3-hydroxytyrosol, and quercetin, against β-site amyloid precursor protein cleaving enzyme-1 (BACE1), also known as β-secretase or memapsin 2, which is implicated in the pathogenesis of Alzheimer's disease (AD). Methods and Results: The inhibitory potential against BACE1, molecular docking simulations, as well as neurotoxicity and the effect on the AD-related gene expression of the selected phenolics were tested. BACE1 inhibitory activity was carried out using the ELISA microplate assay via fluorescence resonance energy transfer (FRET) technology. Molecular docking experiments were performed in the human BACE1 active site (PDB code: 2WJO). Neurotoxicity of the compounds was carried out in SH-SY5Y, a human neuroblastoma cell line, by the Alamar Blue method. A gene expression analysis of the compounds on fourteen genes linked to AD was conducted using the real-time polymerase chain reaction (RT-PCR) method. Rosmarinic acid, EGCG, oleuropein, and quercetin (also used as the reference) were able to inhibit BACE1 with their respective IC50 values 4.06 ± 0.68, 1.62 ± 0.12, 9.87 ± 1.01, and 3.16 ± 0.30 mM. The inhibitory compounds were observed to occupy the non-catalytic site of the BACE1. However, hydrogen bonds were found to be present between rosmarinic acid and EGCG and aspartic amino acid D228 in the catalytic site. Oleuropein and quercetin effectively suppressed the expression of PSEN, APOE, and CLU, which are recognized to be linked to the pathogenesis of AD. Conclusions: The outcomes of the work bring quercetin, EGCG, and rosmarinic acid to the forefront as promising BACE1 inhibitors.

The Potential of Paku Gajah (Angiopteris evecta) as Antitumor Through In Vitro and In Silico Studies

ABSTRACT. Paku gajah (Angiopteris evecta) is one of the largest ferns which has been used empirically by the Dayak tribe of Kalimantan, Indonesia as a traditional medicine to treat various diseases, one of which is tumors. This research aims to determine the potential utilization of A. evecta stem extract as an antitumor by secondary metabolites analysis, toxicity and antitumor assay. The methods used in this study were gradual maceration using three solvents (n-hexane, ethyl acetate, and methanol), phytochemical screening, toxicity test, antitumor activity assay with the Alamarblue method, Liquid Chromatography-Mass Spectrometry (LC-MS) analysis, and molecular docking analysis. This study indicated that A. evecta stem extract contained secondary metabolites such as flavonoids, saponins, tannins, and steroids. The ethyl acetate and methanol were found as toxic extracts with LC50 130.67 and 314.31 µg/mL, respectively. In line with the toxicity, the antitumor activity of the ethyl acetate extract was the highest with an IC50 of 240.94 µg/mL and phytochemical analysis revealed the presence of violanthin and angiopteroside in the extract. Molecular docking showed that the binding energy and inhibition constants of violanthin and angiopteroside against receptors were higher than standard ligand F82. The interaction between violanthin and the receptor results five hydrogen-bond (H-bond) with Lys920, Cys919, Asp1046, and Leu840, while the angiopteroside produces four H-bonds with Leu836, Leu834, and Arg831. Keywords: Antitumor, molecular docking, paku gajah (Angiopteris evecta).

Cu-doped 70S bioactive glass fibrous membranes produced using solution blow spinning (SBS)

3D fibrous membranes of undoped and Cu-doped 70S glass were developed, by solution blow spinning. The concentration of copper ions ranged from 1 to 4 % in mol. As-spun fibers were calcined/stabilized at 700 °C. The produced membranes' morphological and biological properties and the release of Ca and Cu ions were evaluated. Amorphous materials were obtained in pure and doped with 1 % Cu2+ 70S glass, while metallic copper was observed in 4 % Cu2+ doped glass. All samples have fibers with average diameter at the submicron scale, ranging from 700 to 850 nm. The amount of Cu ions influenced the degradation of the samples and the release of Cu and Ca ions in the Tris solution, the greater the amount of copper the more these ions were degraded and released in the solution. In vitro biological results, showed that adding copper favored the biomineralization process. No toxicity was observed in the MMT test for all samples. Alamar Blue method showed cell viability for fibroblasts greater than 90 % for all samples and for osteoblasts greater than 70 % for the BG-70S.2Cu sample. The developed fibrous membranes presented antimicrobial characteristics against Staphylococcus aureus and Candida albicans.

Development and validation of a colorimetric antifungal susceptibility testing method for the dimorphic fungus Talaromyces marneffei.

Antifungal drug resistance is an emerging cause of treatment failure in invasive fungal infections, and antifungal susceptibility testing (AFST) may inform treatment decisions. Currently, there are no established AFST guidelines for Talaromyces marneffei (Tm) or other dimorphic fungi. We developed a colorimetric AFST method using a fluorescent redox indicator alamarBlue, which changes from blue to pink in proportion to cellular metabolic activity. We determined the optimal time for alamarBlue addition to be 24h post-inoculation and for MIC reading to be 72h post-inoculation. Our method allows three ways to determine minimum inhibitory concentration (MIC): visual inspection of color change, optical density, and fluorescence intensity. We validated the assay by determining the MICs for seven antifungals against 32 Tm clinical isolates and assessed the essential agreement (EA) and inter-rater reliability between our alamarBlue and the Clinical Laboratory Standard Institute (CLSI) broth microdilution methods. The MIC ranges (from low to high) were: 0.008-0.025 μg/ml for itraconazole, 0.004-0.13 μg/ml for voriconazole, 0.03-0.13μg/ml for posaconazole, 0.06-0.5µg/ml for flucytosine, 0.5-1µg/ml for amphotericin B, 0.5-4µg/ml for caspofungin, and 0.5-16µg/ml for fluconazole. The EAs were 100% between all three MIC readouts of the alamarBlue method, and 94%-100% between the alamarBlue and CLSI methods. Our alamarBlue method had substantially higher inter-rater agreement and offers a more reliable method that can be standardized across laboratories in both high- and low-resource settings compared to the established CLSI methodology.

Comparison of cytotoxic, apoptotic and oxidative properties of Akacid plus and chlorhexidine in corneal epithelial cell culture.

This study aims to compare the cytotoxic, apoptotic, and oxidative effects of a new cationic disinfectant, Akacid Plus, with chlorhexidine, on the human corneal epithelial cell line. Time-dependent cytotoxicity studies were performed with the Alamar Blue method. Apoptotic activity was investigated by flow cytometric methods. Reactive oxygen species levels were measured with the ROS cellular test kit. BAX, BCL2 and caspase 3, 9, 12 mRNA expressions were evaluated by PCR, as well as BAX and BCL2 protein expressions by Western-Blot. At the fifth minute of the treatment, the viability was 68.15% with Akacid Plus and 43.95% with chlorhexidine. At the 15th minute, no significant difference was observed with both solutions. In the apoptotic evaluation, Akacid Plus significantly increased the early and late apoptotic activity in the cell line (p < 0.0001), while a significant increase was observed in late apoptosis and necrosis levels with chlorhexidine (p < 0.001). Chlorhexidine also induced gene expression of BAX, BCL2, caspase 3, 9 and BAX proteins (p < 0.05), while reducing protein expression of BCL2 (p < 0.001). Akacid Plus induced the gene expressions of BCL2, CASP3 and caspase 9, reduced gene expressions of BAX and caspase 12 and protein expression of BCL2 (p < 0.05). No significant difference was observed in the ROS level with both solutions (p > 0.05). Due to the widespread use of cationic polymers in ophthalmology, this new molecule with high antimicrobial activity and relatively low cytotoxicity may be of interest for clinical use. Further investigations are necessary to fully understand the ophthalmologic potential of this solution.

Biocompatibility of graphene oxide nanosheets functionalized with various amino acids towards mesenchymal stem cells

Graphene and its derivatives have gained popularity due to their numerous applications in various fields, such as biomedicine. Recent reports have revealed the severe toxic effects of these nanomaterials on cells and organs. In general, the chemical composition and surface chemistry of nanomaterials affect their biocompatibility. Therefore, the purpose of the present study was to evaluate the cytotoxicity and genotoxicity of graphene oxide (GO) synthesized by Hummer's method and functionalized by different amino acids such as lysine, methionine, aspartate, and tyrosine. The obtained nanosheets were identified by FT-IR, EDX, RAMAN, FE-SEM, and DLS techniques. In addition, trypan blue and Alamar blue methods were used to assess the cytotoxicity of mesenchymal stem cells extracted from human embryonic umbilical cord Wharton jelly (WJ-MSCs). The annexin V staining procedure was used to determine apoptotic and necrotic death. In addition, COMET and karyotyping techniques were used to assess the extent of DNA and chromosome damage. The results of the cytotoxicity assay showed that amino acid modifications significantly reduced the concentration-dependent cytotoxicity of GO to varying degrees. The GO modified with aspartic acid had the lowest cytotoxicity. There was no evidence of chromosomal damage in the karyotyping method, but in the comet assay, the samples modified with tyrosine and lysine showed the greatest DNA damage and rate of apoptosis. Overall, the aspartic acid-modified GO caused the least cellular and genetic damage to WJ-MSCs, implying its superior biomedical applications such as cell therapy and tissue engineering over GO.

Metabolomic profiling and bactericidal effect of Polyalthia longifolia (Sonn.) Twaites. stem bark against methicillin-resistant Staphylococcus aureus.

Objective. The present study was carried out to establish the chemical profile of the methanolic extract of Polyalthia longifolia stem bark and investigate its antibacterial property against some human pathogenic bacteria. Methods. The extract was analysed using liquid and gas chromatography coupled to mass spectrometry. Antibacterial activity of P. longifolia extract against some human pathogenic bacteria was screened using the AlamarBlue method, and MIC and MBC were determined. Results and Conclusion. Liquid chromatography-mass spectrometry (LC-MS) revealed the presence of 21 compounds among which 12 were identified. Gas chromatography-mass spectrometry (GC-MS) allowed identification of 26 compounds, the three major ones being the following: cis vaccenic acid (17.79 %), 3-ethyl-3-hydroxyandrostan-17-one (13.80 %) and copaiferic acid B (12.82 %). P. longifolia extract was active against Gram-positive bacteria with MIC ranging from 1 to 2 mg ml-1 and MBC from 2 to 6 mg ml-1. This study demonstrated the bactericidal effect of the methanolic extract of Polyalthia longifolia stem bark against some human pathogenic bacteria, including methicillin-resistant S. aureus . This effect could be related to the presence in the extract of a broad diversity of well-known compounds with established pharmacological properties. These results support the ethnomedicinal use of P. longifolia stem bark in Cameroon for the management of methicillin-resistant S. aureus (MRSA)-related infections.

Combination Therapy of Curcumin and Disulfiram Synergistically Inhibits the Growth of B16-F10 Melanoma Cells by Inducing Oxidative Stress

Oxidative stress plays a central role in the pathophysiology of melanoma. Curcumin (CUR) is a polyphenolic phytochemical that stimulates reactive oxygen species (ROS) production, while disulfiram (DSS) is a US FDA-approved drug for the treatment of alcoholism that can act by inhibiting the intracellular antioxidant system. Therefore, we hypothesized that they act synergistically against melanoma cells. Herein, we aimed to study the antitumor potential of the combination of CUR with DSS in B16-F10 melanoma cells using in vitro and in vivo models. The cytotoxic effects of different combination ratios of CUR and DSS were evaluated using the Alamar Blue method, allowing the production of isobolograms. Apoptosis detection, DNA fragmentation, cell cycle distribution, and mitochondrial superoxide levels were quantified by flow cytometry. Tumor development in vivo was evaluated using C57BL/6 mice bearing B16-F10 cells. The combinations ratios of 1:2, 1:3, and 2:3 showed synergic effects. B16-F10 cells treated with these combinations showed improved apoptotic cell death and DNA fragmentation. Enhanced mitochondrial superoxide levels were observed at combination ratios of 1:2 and 1:3, indicating increased oxidative stress. In vivo tumor growth inhibition for CUR (20 mg/kg), DSS (60 mg/kg), and their combination were 17.0%, 19.8%, and 28.8%, respectively. This study provided data on the potential cytotoxic activity of the combination of CUR with DSS and may provide a useful tool for the development of a therapeutic combination against melanoma.

Synthesis, characterization and in vitro anti mycobacterial activity of some novel benzothiazole derivatives

World Health Organization considers Mycobacterium tuberculosis is the most dangerous chronic transmittable disease. Development of resistance by Mycobacterium tuberculosis is a common problem world Wide. To combat these problems there could be a solution that we should search newer molecules with different mechanism of action to fight against such diseases. A series of some novel benzothiazole derivatives (I-X) have been synthesized and evaluated for their in vitro antimycobacterial activity against Mycobacterium tuberculosis H37Rv using Microplate Alamar Blue Assay Method (MABA). Structures of the synthesized compounds were supported by means of IR, NMR and MASS spectral studies. Among the synthesized compounds, compound [IX], 2-[5-(4-Bromophenyl)-2-furylcarboxamido]-6-methoxy-l,3-benzothiazole exhibited a significant activity when compared with the standard drug Isoniazid. This could be the remarkable starting point to develop new lead molecules with potential antitubercular activity.

Poly(lactic-co-glycolic acid) (PLGA) nanoparticles loaded with tetramethylpyrazine and conjugated with TED-1 polypeptide reduce formation of β-amyloid and alleviate oxidative stress in alzheimer’s disease

Controlling β-amyloid and oxidative stress is indicated to improve Alzheimer’s disease (AD). In this study, we loaded Poly Lactic-co-Glycolic Acid (PLGA) nanoparticles with ligustrazine and coupled them with TED-1 polypeptide, to explore an effective drug for the condition. Tet-1 peptide was coupled to Tetramethylpyrazine (TMP)-PLGA nanoparticles. Glioma cells were then treated with PLGA nanoparticles, TMP, TMP-PLGA nanoparticles, and TMP-PLGA-Tet-1 nanoparticles. Microscope and flow cytometry were then conducted to assess the impact of nanoparticles on cell morphology and toxicity of the nanoparticles. Oxidative stress of the nanoparticles was detected by 2,2-Diphenyl-1-picrylhydrazyl (DPPH) method. TMP-PLGA-Tet-1 nanoparticles were found to have significant ability to scavenge free radicals, also showing significantly increased anti-β-amyloid activity. Finally, Alamar Blue method and 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) method proved that, the TMP-PLGA-Tet-1 nanoparticles enhanced the efficiency of drug delivery to neurons without toxicity. Moreover, Tet-1-conjugated TMP-PLGA nanoparticles reduced amyloid and alleviated oxidative stress, and may hence be a potential option for treatment of AD. Labeling Tet-1 to TMP-PLGA could therefore improve drug delivery.

Cytotoxic, Apoptotic, and Oxidative Effects of Preserved and Preservative-Free Brimonidine in a Corneal Epithelial Cell Line.

Purpose: This study aims to compare the cytotoxic, apoptotic, and oxidative effects of preserved and preservative-free forms of brimonidine 0.15% on the human corneal epithelial cell (HCEC) line. Methods: Time-dependent cytotoxicity studies were performed with the Alamar Blue method. For apoptotic studies, PE Annexin V and 7-amino-actinomycin (7-AAD) staining and flow cytometry were performed. Messenger RNA (mRNA) expressions of Bax, Bcl-2, and caspase-3, -9, -12, and protein expressions of Bax and Bcl-2 were evaluated by quantitative real-time polymerase chain reaction and Western blot method, respectively. Results: Cell viability was 76.4% with the preserved solution and 36.05% with the preservative-free solution at the fifth minute. No significant difference was observed with either solution at the 15-min mark, whereas cell viability did not change significantly after 1 h. In the apoptosis evaluation, it was observed that the preservative-free solution increased the early apoptotic activity to a greater degree (P < 0.05). Preservative-free solution also induced gene expression of proapoptotic Bax, caspase-9 and -12, and protein expression of Bax while reducing the protein expression of anti-apoptotic Bcl-2 (P < 0.0001). Preserved solution induced only the gene expression of caspase-12, and reduced the protein expression of Bcl-2 (P < 0.0001). No significant difference was observed in the reactive oxygen species (ROS) levels of either solution compared with the control group (P > 0.05). Conclusion: It was demonstrated that the preserved solution is less cytotoxic to the HCEC line in the early period, has less early apoptotic activity, and does not significantly increase ROS levels.

Effects of four efflux pump inhibitors on the activities of clarithromycin against Mycobacterium abscessus

Objective: To detect the effects of four efflux pump inhibitors on the minimum inhibitory concentration of clarithromycin (CLA) against Mycobacterium abscessus (M. abscessus) in vitro, and to explore the role of efflux pump in CLA resistance of M. abscessus. Methods: Four frequently-used efflux pump inhibitors (Carbonyl Cyanide 3-chlorophenylhydrazone, CCCP, N, N'-dicyclohexylcarbodiimide, DCC, Verapamil, VP, Reserpine, RSP) were evaluated in this study. The minimum inhibitory concentration (MIC) values of clarithromycin against M. abscessus reference strain and 60 clinical strains with or without efflux pump inhibitors were detected by Alamar Blue method. Sequence analysis of erm(41) and rrl genes known to be associated with CLA resistance in M. abscessus was performed to analyze the correlation between the effect of efflux pump inhibitors on MIC and mutation of resistance-related genes. Results: CCCP, DCC, VP and RSP could reduce the MIC of M. abscessus to CLA, and the effect of RSP was weaker than the other three efflux pump inhibitors. Among the sixty M. abscessus clinical strains, ten strains were resistant to clarithromycin, seven of which had rrl gene mutation. The CLA resistance rate of smooth phenotype isolates was higher than that of rough phenotype isolates. At 3 day of clarithromycin incubation, the MICs of resistant strains were all reduced by efflux pump inhibitors. Compared with the strains with rrl gene mutation, efflux pump inhibitors had a greater effect on the strains without rrl gene mutation. At 14 day of clarithromycin incubation, 83% of M. abscessus subsp. abscessus, were induced to be resistant, and all of them were T28 sequence type of erm(41). With the occurrence of induced drug resistance, the effect of efflux pump inhibitor on CLA MIC decreased. Efflux pump inhibitors had no statistically significant diffence in the effect of effcux pump inhibitors on CLA MIC levels in different phenotypes of isolates. Conclusions: Efflux pump is involved in the resistance process of M. abscessus to CLA. Efflux pump inhibitors reduce the drug resistance to clarithromycin against M. abscessus in different degrees. The use of efflux pump inhibitors may provide a new way to alleviate the drug resistance of M. abscessus.

Structural Design, Synthesis and Antioxidant, Antileishmania, Anti-Inflammatory and Anticancer Activities of a Novel Quercetin Acetylated Derivative.

Quercetin (Q) is a bioflavonoid with biological potential; however, poor solubility in water, extensive enzymatic metabolism and a reduced bioavailability limit its biopharmacological use. The aim of this study was to perform structural modification in Q by acetylation, thus, obtaining the quercetin pentaacetate (Q5) analogue, in order to investigate the biological potentials (antioxidant, antileishmania, anti-inflammatory and cytotoxicity activities) in cell cultures. Q5 was characterized by FTIR, 1H and 13C NMR spectra. The antioxidant potential was evaluated against the radical ABTS•+. The anti-inflammatory potential was evaluated by measuring the pro-inflammatory cytokine tumor necrosis factor (TNF) and the production of nitric oxide (NO) in peritoneal macrophages from BALB/c mice. Cytotoxicity tests were performed using the AlamarBlue method in cancer cells HepG2 (human hepatocarcinoma), HL-60 (promyelocytic leukemia) and MCR-5 (healthy human lung fibroblasts) as well as the MTT method for C6 cell cultures (rat glioma). Q and Q5 showed antioxidant activity of 29% and 18%, respectively, which is justified by the replacement of hydroxyls by acetyl groups. Q and Q5 showed concentration-dependent reductions in NO and TNF production (p < 0.05); Q and Q5 showed higher activity at concentrations > 40µM when compared to dexamethasone (20 µM). For the HL-60 lineage, Q5 demonstrated selectivity, inducing death in cancer cells, when compared to the healthy cell line MRC-5 (IC50 > 80 µM). Finally, the cytotoxic superiority of Q5 was verified (IC50 = 11 µM), which, at 50 µM for 24 h, induced changes in the morphology of C6 glioma cells characterized by a round body shape (not yet reported in the literature). The analogue Q5 had potential biological effects and may be promising for further investigations against other cell cultures, particularly neural ones.

Antioxidant and Cytotoxicity Activities of the Fermentation Extract of the Endophytic Fungi from the Marine Biota of Colt Coral

Indonesia has the largest and most biodiverse coral reef in the world. Colt coral has not been studied and explored especially endophytic fungi associated with the coral. Endophytic fungi are highly potential for the production of antioxidant and anticancer compounds. This research aimed to study the antioxidant and cytotoxic activities of fermentation extract from endophytic fungi from colt coral. Filtrate and mycelium extracts were obtained from static and shake fermentations of isolate SKF 15. Antioxidant and cytotoxic assays were conducted by free radical scavenger 1,1-diphenyl-2-picrylhydrazyl (DPPH) and AlamarBlue methods, respectively. The result showed that FD extract provided the highest antioxidant activity with inhibition of 49.36% at 200 ppm of DPPH. Variation of fermentation time (3-21 days) demonstrated the highest activity with inhibition of 66.97% for antioxidant assay (7 days) and 81.13% for cytotoxic assay (3 days). FTIR analysis presented the existence of hydroxyl groups O-H (3452.58 cm-1), C=C groups (1668.43 cm-1); C-O hydroxyl group (1230.58 cm -1), and C-H sp3 groups (2941.44 cm-1). Based on LC-MS analysis, FD extract has a mass of m/z 305.63, [M+H]+, predicted as dihydroquercetin (C15H24O7). Keywords: Antioxidant assay, cytotoxic assay, endophytic fungi, colt coral, DPPH method, AlamarBlue method

Antimicrobial and cytotoxic activity of fungal mycelial extracts from aquatic environments in the Amazon

Fungi are a prolific source of biologically active metabolites, including a wide range of clinically important drugs. Therefore, this study aims to evaluate the antimicrobial and cytotoxic activity of secondary metabolites extracted from fungal mycelia isolated from freshwater samples in the state of Amazonas. Mycelial extracts from 12 fungal were used, extracted with MeOH/AcOEt (1:1) according to the criteria established by Souza et al. (2004). For antimicrobial activity, the extracts were tested against the pathogens Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Enterococcus feacalis, Candida albicans and C. tropicalis. To identify the minimum inhibitory dosage (MID) the microdilution method was used. To perform the cytotoxicity assay, the VERO strain (ATCC® CCL-81TM) was used. The assays were determined by the Alamar Blue method according to Ahmed et al. (1994). The tested extracts did not show antibacterial activity. Five extracts (41.7%), obtained from the fungi Aspergillus - 1283, Chrysoporther - 1169, Diaporther – 1203, Fusarium – 1085, and Trichoderma, showed antifungal activity against C. albicans. Diaporther extract (8.3%) - 1203 was active against C. tropicalis. In the cytotoxicity assay, 58.3% of the evaluated extracts showed no significant toxic effect. Five extracts, Cladosporium - 1135, Chrysoporther - 1169, Cytospora - 1098, Fusarium - 1085, and Talaromyces - 1244, showed cytotoxic potential, exhibiting viability lower than 70%. The results obtained suggest that mycelial extracts of fungi isolated from water samples from the Amazon region have potential against yeasts of medical interest. Only two of the active extracts were revealed potentially cytotoxic.

1,2,3-triazole derivative: Synthesis, characterization, DFT, molecular docking study and antibacterial-antileishmanial activities

In this study, 4-((1-(4-chlorobenzyl)-1H-1,2,3-triazole-4-yl)methoxy)-3-methoxybenzaldehyde (I) was synthesized and molecular structure of compound I was confirmed by FTIR and NMR (1H and 13C NMR) spectroscopic methods. The geometric structure of compound I was optimized by DFT/B3LYP method using 6–311++G(d, p) basis set. The molecular docking study was carried out against six different proteins. The antibacterial and antileishmanial activities of compound I were tested by microdilution broth with Alamar blue method and minimum inhibitor concentrations (MIC) were determined. According to the test results, it was found to be effective against eleven types of bacteria at different concentrations (MIC: 312–5000 ​μg/mL). In addition, compound I was not effective against leishmania species at the concentrations that were examined.

A bifunctional and multi-responsive fluorescent sensor for toxic analytes in the aqueous medium: Easy synthesis, NIR-visible effect, imaging in living cells

This work aims to develop a new cheap-facile and bifunctional and multi-responsive fluorescent sensor, which based-on isophthaloyl dichloride/nitro-benzofurazan (isophthalo-NBD), for toxic anions and cations. This paper is based on fluorometric&colorimetric methods for the determination of Fe3+&Cu2+ and F−&CN− ions through an easy-to-make sensor having double binding modes in aqueous media. The binding mode of the sensor, isophthalo-NBD, with Fe3+&Cu2+ and F−&CN− ions was confirmed by classical spectroscopic data such as absorption, fluorescence intensity, Job plot, response time, the limit of detection, COSY-NMR and 1HNMR titrations, etc. The sensor showed a remarkable color change from neon yellow to bright-red along with both an increase in fluorescence intensity, and a red-shift of emission wavelength in the presence of F− and CN− anions, respectively. Otherwise, the sensor exhibited an observable quenching effect without changing wavelength in the presence of Fe3+ and Cu2+, respectively, via the Chelation Enhanced Fluorescence effect (CHEF). The Alamar Blue method was used to determine cell viability by treating HeLa and Hep-2 cells with isophthalo-NBD. The fluorescence images of isophthalo-NBD in living cells to detect the presence of F− and CN− anions and Fe3+ and Cu2+, respectively, were performed by using a confocal laser scanning microscope.

Mineralized collagen scaffold bone graft accelerate the osteogenic process of HASCs in proper concentration

PurposeTo investigate the feasibility and the optimum condition of human adipose-derived stem cells cultured on the mineralized collagen material; and to further explore the mechanism of osteogenic differentiation of the human Adipose-derived stem cells stimulated by the mineralized collagen material.MethodsPrimary human adipose-derived stem cells (HADSCs) were isolated from human adipose tissue using centrifugal stratification, which had been passed repeatedly to later generations and purified. Human adipose-derived stem cells were cultured on the bone graft material and the optimum concentration was explored by Alamar blue colorimetric method. The rest experiment was conducted according to the result. The experimental groups are shown below: group A (HADSCs + bone graft material); group B (HADSCs). Morphological observation was taken by scanning electronic microscope (SEM). Alkaline phosphatase activities were tested by histochemical method. Calcium deposition was investigated by alizarin red staining. The quantity access of osteogenic-related mRNA: ALP (alkaline phosphatase), BMP2 (bone morphogenetic protein 2) and RUNX2 (runt-related transcription factor 2) were detected using RT-PCR.ResultsThe cultured cells grew stably and proliferated rapidly. The optimum condition was 0.5 mg/cm2 bone graft material coated on the bottom of medium. After culturing on the material 14 days, the alizarin red staining showed that more calcium deposition was detected in group A and alkaline phosphatase activities of group A was higher than group B (p ˃ 0.05). Similarly, after culturing for 14 days, the ALP, BMP2 and RUNX2 transcription activity of group A was higher than group B (p ˃ 0.05).ConclusionHuman adipose-derived stem cells cultured on bone graft material were dominantly differentiated into osteoblast in vitro. Thus it provided a new choice for bone tissue engineering.

Unveiling the Anti-tubercular Properties of Biscoumarins, through Biological Evaluation and Docking Studies

Background: Biscoumarin scaffolds are known for their promising pharmacological properties. These compounds have not been studied for their activity against tuberculosis strains. Objective: Unveil the antitubercular properties of biscoumarin scaffolds. Methods: Biscoumarin derivatives (3a-3l) were synthesized using lemon juice as a catalyst and were investigated for their in-vitro anti-tubercular activity against the H37Rv strain of Mycobacterium tuberculosis using Microplate Alamar Blue Assay Method (MABA). Their binding interaction was investigated by Molecular Docking Studies using InhA with PDB-ID: 2NSD as target receptors in the H37Rv strain of Mycobacterium tuberculosis. These derivatives (3a-3l) were subjected to the neutrophil function test. Results: The results revealed that compounds 3b, 3c, 3d, 3f, 3i, 3j showed excellent activity with MIC 1.6μg/mL. Molecular docking interactions for their antitubercular activity proved that the derivatives (3a-3l) can easily bind into the pockets of the enzyme. Neutrophil function test signified that they exhibit moderate neutrophil functions assuring that they do not harm the functioning of Neutrophils. Conclusion: These studies have awakened the property of Biscoumarins as promising antitubercular scaffolds.