Study of the anti-cancer activity of a mesoporous silica nanoparticle surface coated with polydopamine loaded with umbelliprenin.

A mesoporous silica nanoparticle (MSN) coated with polydopamine (PDA) and loaded with umbelliprenin (UMB) was prepared and evaluated for its anti-cancer properties in this study. Then UMB-MSN-PDA was characterized by dynamic light scattering (DLS), Field emission scanning electron microscopy (FESEM), Transmission electron microscopy (TEM) and FTIR methods. UV-visible spectrometry was employed to study the percentage of encapsulation efficiency (EE%). UMB-MSN-PDA mediated cell cytotoxicity and their ability to induce programmed cell death were evaluated by MTT, real-time qPCR, flow cytometry, and AO/PI double staining methods. The size of UMB-MSN-PDA was 196.7 with a size distribution of 0.21 and a surface charge of -41.07 mV. The EE% was 91.92%. FESEM and TEM showed the spherical morphology of the UMB-MSN-PDA. FTIR also indicated the successful interaction of the UMB and MSN and PDA coating. The release study showed an initial 20% release during the first 24 h of the study and less than 40% during 168 h. The lower cytotoxicity of the UMB-MSN-PDA against HFF normal cells compared to MCF-7 carcinoma cells suggested the safety of formulation on normal cells and tissues. The induction of apoptosis in MCF-7 cells was indicated by the upregulation of P53, caspase 8, and caspase 9 genes, enhanced Sub-G1 phase cells, and the AO/PI fluorescent staining. As a result of these studies, it may be feasible to conduct preclinical studies shortly to evaluate the formulation for its potential use in cancer treatment.

Discovery of a Novel Inhibitor Structure of Mycobacterium tuberculosis Isocitrate Lyase.

Tuberculosis remains a global threat to public health, and dormant Mycobacterium tuberculosis leads to long-term medication that is harmful to the human body. M. tuberculosis isocitrate lyase (MtICL), which is absent in host cells, is a key rate-limiting enzyme of the glyoxylic acid cycle and is essential for the survival of dormant M. tuberculosis. The aim of this study was to evaluate natural compounds as potential MtICL inhibitors through docking and experimental verification. Screening of the TCMSP database library was done using Discovery Studio 2019 for molecular docking and interaction analysis, with the putative inhibitors of MtICL, 3-BP, and IA as reference ligands. Daphnetin (MOL005118), with a docking score of 94.8 and -CDOCKER interaction energy of 56 kcal/mol, was selected and verified on MtICL in vitro and M. smegmatis; daphnetin gave an IC50 of 4.34 µg/mL for the MtICL enzyme and an MIC value of 128 µg/mL against M. smegmatis, showing enhanced potential in comparison with 3-BP and IA. The interactions and essential amino acid residues of the protein were analyzed. In summary, natural daphnetin may be a promising new skeleton for the design of inhibitors of MtICL to combat dormant M. tuberculosis.

Genetic Incorporation of Fluorescent Amino Acid into Fatty Acid Binding Protein for Fatty Acid Detection.

Fatty acids play critical roles in biological processes, such as energy storage, metabolism, signal transduction, and immune regulation. Therefore, it is necessary to develop in-vitro fluorescent sensors to detect free fatty acids. By genetically incorporating a synthetic fluorescent amino acid (L-(7-hydroxycoumarin-4-yl) ethylglycine, Cou) into fatty acid-binding protein (FABP), we obtained a fluorescent sensor that has a turn-on signal in the presence of the fatty acids. Its response to medium-chain and long-chain fatty acids can be increased by 5.8-fold within several minutes, highlighting its potential applications in fatty acids-related biological processes. Our newly developed fatty acid detection system based on genetic expansion technology has extended the molecular toolboxes available for important biological molecular analysis.

Structural Basis for Blocked Excited State Proton Transfer in a Fluorescent, Photoacidic Non-Canonical Amino Acid-Containing Antibody Fragment.

The fluorescent non-canonical amino acid (fNCAA) L-(7-hydroxycoumarin-4-yl)ethylglycine (7-HCAA) contains a photoacidic 7-hydroxycoumarin (7-HC) side chain whose fluorescence properties can be tuned by its environment. In proteins, many alterations to 7-HCAA's fluorescence spectra have been reported including increases and decreases in intensity and red- and blue-shifted emission maxima. The ability to rationally design protein environments that alter 7-HCAA's fluorescence properties in predictable ways could lead to novel protein-based sensors of biological function. However, these efforts are likely limited by a lack of structural characterization of 7-HCAA-containing proteins. Here, we report the steady-state spectroscopic and x-ray crystallographic characterization of a 7-HCAA-containing antibody fragment (in the apo and antigen-bound forms) in which a substantially blue-shifted 7-HCAA emission maximum (∼70 nm) is observed relative to the free amino acid. Our structural characterization of these proteins provides evidence that the blue shift is a consequence of the fact that excited state proton transfer (ESPT) from the 7-HC phenol has been almost completely blocked by interactions with the protein backbone. Furthermore, a direct interaction between a residue in the antigen and the fluorophore served to further block proton transfer relative to the apoprotein. The structural basis of the unprecedented blue shift in 7-HCAA emission reported here provides a framework for the development of new fluorescent protein-based sensors.

LC-MS/MS for determination of aesculetin in rat plasma and its application to a pharmacokinetic study.

Aesculetin, a coumarin compound present in the sancho tree and chicory, exhibits excellent antioxidant and anti-inflammatory activities in the vascular and immune system. In this study, a rapid and sensitive ultra-high performance liquid chromatography electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) method was established and validated for the determination of aesculetin in rat plasma. Plasma samples were prepared by protein precipitation with acetonitrile. Chromatographic separation was performed on an Acquity UPLC HSS T3 C18 column (2.1 × 100 mm, 1.8 µm) with gradient elution at a flow rate of 0.3 ml/min, using mobile phase consisting of 0.1% formic acid (A) and acetonitrile (B). Aesculetin and puerarin (internal standard) were detected by multiple reaction monitoring in negative ion mode. The method was fully validated according to the US Food and Drug Administration guidelines. The calibration curve was linear over the range of 2-1,000 ng/ml with correlation coefficient >0.9980. The carry-over, matrix effect, extraction recovery, dilution effect, intra- and inter-day precision and the accuracy were within acceptable limits. The method was then applied to a pharmacokinetic study of aesculetin in rats. After oral administration at doses of 5, 10 and 20 mg/kg, the plasma concentration reached peaks of 95.7, 219.9, 388.6 ng/ml at times of 1.22-1.78 h. The oral bioavailability was calculated as 15.6-20.3% in rat plasma. The result provided pre-clinical information for further application of aesculetin.

Aesculetin Exhibits Strong Fluorescent Photoacid Character.

Coumarins are bioactive molecules that often serve as defenses in plant and animal systems, and understanding their fundamental behavior is essential for understanding their bioactivity. Aesculetin (6,7-dihydroxycoumarin) has recently attracted attention due to its ability to act as an antioxidant, but little is known about its photophysical properties. The fluorescence lifetimes of its neutral and anion form in water are 19 ± 2 ps and 2.3 ± 0.1 ns, respectively. Assuming the short lifetime of the neutral is determined by ESPT, we estimate kPT ~ 5 × 1010 s-1. Using steady-state and time-resolved fluorescence spectroscopy, we determine its ground and excited-state [Formula: see text] to be 7.3 and -1, respectively, making it one of the strongest photoacids of the natural coumarins. Aesculetin exhibits a strong pH dependence of the relative fluorescence quantum yield becoming much more fluorescent above [Formula: see text]. The aesculetin anion [Formula: see text] slightly photobasic character. We also report that aesculetin forms a fluorescent catechol-like complex with boric acid, and this complex has a [Formula: see text] of 5.6.

Twice as Nice: The Duff Formylation of Umbelliferone Revised.

More efficient and preferably more convenient and greener synthetic solutions in coumarin scaffold functionalization are in steady demand. The Duff ortho-formylation of unsubstituted umbelliferone was revised in this study. The reaction conditions were optimized based upon data from the literature analysis and resulted in unexpectedly rapid ortho-formylation of umbelliferone, yielding a mixture of ortho-formyl position isomers. Thorough studies on the separation of ortho-formylated umbelliferones using chromatographic and recrystallization methods as well as the evaluation of their solubility in common organic solvents led to complete resolution of 8-formyl- and 6-formylumbelliferones. The precise protocol for simultaneous preparation, extraction, and purification of 8-formyl- and 6-formylumbelliferones is provided, and the prospective studies of biological and pharmacological activities of these compounds are synopsized.

A Natural Alternative Treatment for Urinary Tract Infections: Itxasol©, the Importance of the Formulation.

Genito-urinary tract infections have a high incidence in the general population, being more prevalent among women than men. These diseases are usually treated with antibiotics, but very frequently, they are recurrent and lead to the creation of resistance and are associated with increased morbidity and mortality. For this reason, it is necessary to develop new compounds for their treatment. In this work, our objective is to review the characteristics of the compounds of a new formulation called Itxasol© that is prescribed as an adjuvant for the treatment of UTIs and composed of ß-arbutin, umbelliferon and n-acetyl cysteine. This formulation, based on biomimetic principles, makes Itxasol© a broad-spectrum antibiotic with bactericidal, bacteriostatic and antifungal properties that is capable of destroying the biofilm and stopping its formation. It also acts as an anti-inflammatory agent, without the adverse effects associated with the recurrent use of antibiotics that leads to renal nephrotoxicity and other side effects. All these characteristics make Itxasol© an ideal candidate for the treatment of UTIs since it behaves like an antibiotic and with better characteristics than other adjuvants, such as D-mannose and cranberry extracts.

Computational Design of a Molecularly Imprinted Polymer for the Biomonitoring of the Organophosphorous Metabolite Chlorferron.

Coumaphos is an organophosphorus compound used as insecticide and frequently used by beekeepers for the management of parasitic mites. The most important metabolite, chlorferron (CFN), has been identified in biological samples and foodstuff. The need to quickly identify the presence of typical metabolites, as an indication of interaction with coumaphos has driven the need to produce a highly sensitive electrochemical method for chlorferron analysis, based on molecularly imprinting polymers (MIP) technology. It showed irreversible behaviour with mixed diffusion/adsorption-controlled reactions at the electrode surface. A monoelectronic mechanism of reaction for oxidation has also been suggested. The linear range observed was from 0.158 to 75 µM. Median precision in terms of %RSD around 3% was also observed. For DPV, the limit of detection (LOD) and the limit of quantitation (LOQ) for the CFN-MIP were 0.158 µM and 0.48 µM, respectively. The obtained median % recovery was around 98%. The results were also validated to reference values obtained using GC-MS. Urine and human synthetic plasma spiked with CFN were used to demonstrate the usability of the method in biological samples, showing the potential for biomonitoring. The developed imprinted sensor showed maximum signal change less than 16.8% when related metabolites or pesticide were added to the mix, suggesting high selectivity of the MIP sensor toward CFN molecules. The results from in vitro metabolism of CMP analysed also demonstrates the potential for detection and quantification of CFN in environmental samples. The newly developed CFN-MIP sensor offers similar LoDs than chromatographic methods with shorter analysis time.

Exploring cardioprotective potential of esculetin against isoproterenol induced myocardial toxicity in rats: in vivo and in vitro evidence.

BACKGROUND: Esculetin is a natural coumarin derivative from various plants with multiple pharmacological effects. Hence, the present study was undertaken to explore the cardio protective potential of esculetin against isoproterenol induced myocardial toxicity in rats. METHODS: The treatment schedule was fixed for 28 days and the rats were divided into five groups of six each. Rats of group I received the normal saline and served as normal control, group II was received ISO (100 mg/kg body weight) for last two consecutive days of the study and served as disease control. Groups III and IV received esculetin 10 and 20 mg/kg body weight respectively once a day per oral for 28 days along with ISO for last two consecutive days of the study. Cardiac biomarkers such as CK-MB and LDH, membrane bound Na+ /K+ ATPases activity, myocardial lysosomal enzymes activity and tissue antioxidants status were estimated in the heart tissue samples. The histopathological changes in the myocardium were also assessed. Further, DPPH assay was done to evaluate the free radicals scavenging potential of esculetin. Cytoxicity assay, intracellular ROS levels by DCFDA assay and m-RNA expression of TNF-α, IL-6 and NF-κB by quantitative RT-PCR in H9c2 cell lines. RESULTS: The increased levels of CK-MB, LDH, LPO, myocardial lysosomal enzymes and membrane bound Na+ /K+ ATPase levels by ISO administration was significantly increased with concomitant decrease in tissue antioxidant enzymes such as GSH, Catalase, and SOD. Pre-treatment with esculetin for 28 days has significantly decreased the levels of cardiac bio-markers, lysosomal enzymes, membrane bound Na+ /K+ ATPase levels as well as Lipid peroxides which is in contrary to the ISO group. Amelioration of the antioxidant levels were also found in esculetin treated groups. Histopathological examination of heart reveals that myocardial degeneration, mononuclear cell infiltration was noticed in ISO treated rats, whereas the same was restored with esculetin treatment. In H9C2 cell lines esculetin could effectively reduced intracellular ROS inhibition and m-RNA expression of pro-inflammatory cytokines including TNF-α, IL-6 and NF-κB to prevent apoptosis or cell necrosis. CONCLUSION: The study provides the evidence of cardioprotective potentials of esculetin against isoproterenol induced myocardial infarction by antioxidant and myocardial membrane stabilization along with in vitro protection from arsenic induced ROS cell necrosis or apoptosis in H9C2 cells.

Evaluation of ThT augmentation and RLS inner filter effect caused by highly fluorescent coumarin derivative and establishing it as true inhibitor of amyloid fibrillation.

Screening of inhibitors that slow down or suppress amyloid fibrils formation relies on some simple but sensitive spectroscopy techniques. Thioflavin T (ThT) fluorescence assay is one of the most common, amyloid specific and sensitive method. However, if an inhibitor is itself fluorescent in the ThT fluorescence range, its screening becomes complicated and require complementary assays. One of such molecules, 6, 7-dihydroxycoumarin (6, 7-DHC, also known as aesculetin, esculetin, and cichorigenin) is fluorescent in the ThT emission range and absorbs in the ThT excitation range. Therefore, it can produce a subtractive effect attributed to primary inner filter effect and/or additive effect due to its self-fluorescence in ThT assay. Our study shows that 6, 7-DHC produces an additive effect in ThT fluorescence, which is minimized at high concentration of ThT and decrease in ThT fluorescence is solely due to its inhibitory effect against HSA fibrillation. These ThT fluorescence-based results are verified through other complementary assays, such as Rayleigh and dynamic light scattering and amyloid-specific Congo red binding assay. Furthermore, hydrophobicity reduction is studied through Nile red (NR) and kinetics through far-UV circular dichroism (far-UV CD) in place of the most commonly employed ThT assay owing to extremely high fluorescence of 6, 7-DHC during initial incubation period.

Modulators of immunoregulatory exonucleases PLD3 and PLD4 identified by high-throughput screen.

PLD3 and PLD4 have recently been revealed to be endosomal exonucleases that regulate the innate immune response by digesting the ligands of nucleic acid sensors. These enzymes can suppress RNA and DNA innate immune sensors like toll-like receptor 9, and PLD4-deficent mice exhibit inflammatory disease. Targeting these immunoregulatory enzymes presents an opportunity to indirectly regulate innate immune nucleic acid sensors that could yield immunotherapies, adjuvants, and nucleic acid drug stabilizers. To aid in delineating the therapeutic potential of these targets, we have developed a high-throughput fluorescence enzymatic assay to identify modulators of PLD3 and PLD4. Screening of a diversity library (N = 17952) yielded preferential inhibitors of PLD3 and PLD4 in addition to a PLD3 selective activator. The modulation models of these compounds were delineated by kinetic analysis. This work presents an inexpensive and simple method to identify modulators of these immunoregulatory exonucleases.

The First Step of Biodegradation of 7-Hydroxycoumarin in Pseudomonas mandelii 7HK4 Depends on an Alcohol Dehydrogenase-Type Enzyme.

Coumarins are well known secondary metabolites widely found in various plants. However, the degradation of these compounds in the environment has not been studied in detail, and, especially, the initial stages of the catabolic pathways of coumarins are not fully understood. A soil isolate Pseudomonas mandelii 7HK4 is able to degrade 7-hydroxycoumarin (umbelliferone) via the formation of 3-(2,4-dihydroxyphenyl)propionic acid, but the enzymes catalyzing the α-pyrone ring transformations have not been characterized. To elucidate an upper pathway of the catabolism of 7-hydroxycoumarin, 7-hydroxycoumarin-inducible genes hcdD, hcdE, hcdF, and hcdG were identified by RT-qPCR analysis. The DNA fragment encoding a putative alcohol dehydrogenase HcdE was cloned, and the recombinant protein catalyzed the NADPH-dependent reduction of 7-hydroxycoumarin both in vivo and in vitro. The reaction product was isolated and characterized as a 7-hydroxy-3,4-dihydrocoumarin based on HPLC-MS and NMR analyses. In addition, the HcdE was active towards 6,7-dihydroxycoumarin, 6-hydroxycoumarin, 6-methylcoumarin and coumarin. Thus, in contrast to the well-known fact that the ene-reductases usually participate in the reduction of the double bond, an alcohol dehydrogenase catalyzing such reaction has been identified, and, for P. mandelii 7HK4, 7-hydroxycoumarin degradation via a 7-hydroxy-3,4-dihydrocoumarin pathway has been proposed.

Novel Fluorinated 7-Hydroxycoumarin Derivatives Containing an Oxime Ether Moiety: Design, Synthesis, Crystal Structure and Biological Evaluation.

A series of fluorinated 7-hydroxycoumarin derivatives containing an oxime ether moiety have been designed, synthesized and evaluated for their antifungal activity. All the target compounds were determined by 1H-NMR, 13C-NMR, FTIR and HR-MS spectra. The single-crystal structures of compounds 4e, 4h, 5h and 6c were further confirmed using X-ray diffraction. The antifungal activities against Botrytis cinerea (B. cinerea), Alternariasolani (A. solani), Gibberella zeae (G. zeae), Rhizoctorzia solani (R. solani), Colletotrichum orbiculare (C. orbiculare) and Alternaria alternata (A. alternata) were evaluated in vitro. The preliminary bioassays showed that some of the designed compounds displayed the promising antifungal activities against the above tested fungi. Strikingly, the target compounds 5f and 6h exhibited outstanding antifungal activity against B. cinerea at 100 µg/mL, with the corresponding inhibition rates reached 90.1 and 85.0%, which were better than the positive control Osthole (83.6%) and Azoxystrobin (46.5%). The compound 5f was identified as the promising fungicide candidate against B. cinerea with the EC50 values of 5.75 µg/mL, which was obviously better than Osthole (33.20 µg/mL) and Azoxystrobin (64.95 µg/mL). Meanwhile, the compound 5f showed remarkable antifungal activities against R. solani with the EC50 values of 28.96 µg/mL, which was better than Osthole (67.18 µg/mL) and equivalent to Azoxystrobin (21.34 µg/mL). The results provide a significant foundation for the search of novel fluorinated 7-hydroxycoumarin derivatives with good antifungal activity.

Chemical composition, antioxidant and antimicrobial activities of Himalayan Fraxinus micrantha Lingelsh leaf extract.

The present investigation determines the phytochemical analysis, antioxidant and antibacterial potential of methanol extract of Himalayan Fraxinus micrantha (Lingelsh) leaves. Chemical analysis of extract revealed the presence of fifty five compounds. The extract was strongly characterised by tyrosol, esculetin, ß-sitosterin, l-(+)-ascorbic acid 2,6-dihexadecanoate, (Z,Z)-6,9-cis-3,4-epoxy nonadecadiene as main components (62.6%). The F. micrantha extract presents IC50, 43.93 µg/ml and 68.90 µg/ml using DPPH and H2O2 scavenging assay, respectively. The antibacterial activity of F. micrantha was evaluated for five microorganisms showed that the extract had a remarkable inhibitory potential with a mean zone diameter of inhibition ranging from 11 to 22 mm. The MIC of the extracts ranged from 7.8 to 250 µg/ml and MBC value range 31.25 to 125 µg/ml. F. micrantha showed significant antibacterial activity against B. subtilis (22 mm, ZOI) with MIC and MBC values 7.8 and 31.25 µg/ml respectively.

Syntheses and evaluation of daphnetin derivatives as novel G protein-coupled receptor inhibitors and activators.

A series of daphnetin (7,8-dihydroxycoumarin) derivatives 1-22 were synthesized including sixteen new compounds (1-5, 7-14, 18, 21 and 22) and six known compounds (6, 15-17, 19 and 20). Their pharmacological activities on G protein-coupled receptors (GPCRs) were evaluated by double antibody sandwich ELISA (DAS-ELISA) in vitro. Daphnetin derivatives with various substitution patterns/groups were obtained from inhibitors to activators on GPCRs. Derivatives 2-5, 8, 15, 16 and 18-20 possessed moderate activation potency on GPCRs. Among them, derivatives 3-5, 16 and 19 presented significant activation potency on GPCRs with EC50 values in the range of 1.18-1.91 nM. Derivatives 6, 11, 14 and 18 showed significant inhibitory potency on GPCRs with IC50 values in the range of 1.26-1.38 nM. Moreover, the structure-activity relationships (SARs) of daphnetin derivatives were discussed in detail. The new daphnetic-based GPCRs activators and inhibitors have potentials as future drug candidates for the treatment of metabolic diseases.

7-Hydroxycoumarins Are Affinity-Based Fluorescent Probes for Competitive Binding Studies of Macrophage Migration Inhibitory Factor.

Macrophage migration inhibitory factor (MIF) is a cytokine with key roles in inflammation and cancer, which qualifies it as a potential drug target. Apart from its cytokine activity, MIF also harbors enzyme activity for keto-enol tautomerization. MIF enzymatic activity has been used for identification of MIF binding molecules that also interfere with its biological activity. However, MIF tautomerase activity assays are troubled by irregularities, thus creating a need for alternative methods. In this study, we identified a 7-hydroxycoumarin fluorophore with high affinity for the MIF tautomerase active site (Ki = 18 ± 1 nM) that binds with concomitant quenching of its fluorescence. This property enabled development of a novel competition-based assay format to quantify MIF binding. We also demonstrated that the 7-hydroxycoumarin fluorophore interfered with the MIF-CD74 interaction and inhibited proliferation of A549 cells. Thus, we provide a high-affinity MIF binder as a novel tool to advance MIF-oriented research.

Structural Characterization of Mono and Dihydroxylated Umbelliferone Derivatives.

Coumarin derivatives are a class of compounds with a pronounced wide range of applications, especially in biological activities, in the medicine, pharmacology, cosmetics, coatings and food industry. Their potential applications are highly dependent on the nature of the substituents attached to their nucleus. These substituents modulate their photochemical and photophysical properties, as well as their interactions in their crystalline form, which largely determines the final field of application. Therefore, in this work a series of mono and dihydroxylated coumarin derivatives with different chemical substituents were synthesized and characterized by UV-Visible spectroscopy, thermal analysis (differential scanning calorimetry (DSC) and TGA), 1H NMR and X-Ray Diffraction to identify limitations and possibilities as a function of the molecular structure for expanding their applications in polymer science.

Evaluation of cobalt(III) complexes as potential hypoxia-responsive carriers of esculetin.

Differentiation between hypoxic and normoxic tissues have been exploited for the development of selective chemotherapeutic agents. In this context, cobalt(III)-based coordination compounds have been designed and investigated as prospective hypoxia-responsive drug delivery systems. Three cobalt(III) complexes, namely [CoIII(esc)(py2en)]ClO4·(CH3OH)2 (1) [CoIII(esc)(TPA)]ClO4·3H2O (2) and [CoIII(bipy)2(esc)]ClO4·2.5H2O (3) (py2en = N,N'-bis(pyridin-2-ylmethyl)ethylenediamine, TPA = tris(2-pyridylmethyl)amine, bipy = 2,2'-bipyridine and esc = 6,7-dihydroxycoumarin or esculetin), were prepared and investigated as potential carriers of esculetin. The spectroscopic and electrochemical properties of 1-3 were investigated and compared. Reactions of the complexes with biologically relevant reducing agents, viz. ascorbic acid, cysteine and glutathione, were monitored spectroscopically for 24 h, in pH 6.2 and 7.4 PBS phosphate buffer saline (PBS) solutions at 37 °C, under air, argon and dioxygen atmospheres. Dissociation of esculetin was observed upon Co3+/Co2+ reduction preferably under hypoxic conditions, with more effective conversion rates for 3 > 2 > 1. These results illustrate the importance to modulate the Co3+/Co2+ redox potential through the donor-acceptor properties of the ancillary ligands. Complex 3 is cytotoxic against HCT-116 but not against HT-29 and HEK-293 cells. In addition, DNA-binding studies indicate that interactions of 1 and 3 with the biomolecule are electrostatic.

Synthesis, computational studies and assessment of in vitro inhibitory activity of umbelliferon-based compounds against tumour-associated carbonic anhydrase isoforms IX and XII.

Coumarins are widely diffused secondary metabolites possessing a plethora of biological activities. It has been established that coumarins represent a peculiar class of human carbonic anhydrase (hCA) inhibitors having a distinct mechanism of action involving a non-classical binding with amino acid residues paving the entrance of hCA catalytic site. Herein, we report the synthesis of a small series of new coumarin derivatives 7-11, 15, 17 prepared via classical Pechmann condensation starting from resorcinol derivatives and suitable ß-ketoesters. The evaluation of inhibitory activity revealed that these compounds possessed nanomolar affinity and high selectivity towards tumour-associated hCA IX and XII over cytosolic hCA I and hCA II isoforms. To investigate the binding mode of these new coumarin-inspired inhibitors, the most active compounds 10 and 17 were docked within hCA XII catalytic cleft.