Second generation of pyrimidin-quinolone hybrids obtained from virtual screening acting as sphingosine kinase 1 inhibitors and potential anticancer agents.

We report here the virtual screening design, synthesis and activity of eight new inhibitors of SphK1. For this study we used a pre-trained Graph Convolutional Network (GCN) combined with docking calculations. This exploratory analysis proposed nine compounds from which eight displayed significant inhibitory effect against sphingosine kinase 1 (SphK1) demonstrating a high level of efficacy for this approach. Four of these compounds also displayed anticancer activity against different tumor cell lines, and three of them (5), (6) and (7) have shown a wide inhibitory action against many of the cancer cell line tested, with GI50 below 5 µM, being (5) the most promising with TGI below 10 µM for the half of cell lines. Our results suggest that the three most promising compounds reported here are the pyrimidine-quinolone hybrids (1) and (6) linked by p-aminophenylsulfanyl and o-aminophenol fragments respectively, and (8) without such aryl linker. We also performed an exhaustive study about the molecular interactions that stabilize the different ligands at the binding site of SphK1. This molecular modeling analysis was carried out by using combined techniques: docking calculations, MD simulations and QTAIM analysis. In this study we also included PF543, as reference compound, in order to better understand the molecular behavior of these ligands at the binding site of SphK1.These results provide useful information for the design of new inhibitors of SphK1 possessing these structural scaffolds.

Inhibitory effect of galantamine and donepezil combination against cholinesterase: An in silico and in vitro study.

This study aimed to evaluate the in silico and in vitro inhibitory effect of the combined use of galantamine (GAL) and donepezil (DON) against acetylcholinesterase and butyrylcholinesterase (BuChE) enzymes. In silico and in vitro cholinesterase analysis were carried out for GAL and DON alone and combined. Molecular modeling studies were carried out (docking analysis, molecular dynamics simulation, and quantum theory of atoms in molecules). Cholinesterase's inhibitory activities by modified Ellman's method and the drug combination effect using the Chou-Talalay method were assayed. GAL/DON combination showed the co-occupancy of the ligands in both enzymes through in silico studies. Regarding in vitro BuChE inhibition analyses, three of five combinations showed an interaction between GAL and DON at the threshold of additive affect (0.9 < CI < 1.1), with a tendency toward a synergistic effect for higher concentrations. This is the first report showing the efficacy of the GAL/DON combinations inhibiting BuChE, showing the importance of analyzing the behavior of different ligands when co-occupancy into the active site is possible. These combinations might be a possible therapy to improved efficacy, reduced doses, minor side effects, and high levels of the neurotransmitter in the synaptic space for Alzheimer's disease.

The Mechanism of Antimicrobial Small-Cationic Peptides from Coarse-Grained Simulations.

Antimicrobial cationic peptides (AMPs) are excellent candidates for use as therapeutic antimicrobial agents. Among them, short peptides possessing sequences of 9-11 amino acids have some advantages over long-sequence peptides. However, one of the main limitations of short peptides is that their mechanism of action at the molecular level is not well-known. In this article, we report a model based on multiscale molecular dynamics simulations of short peptides interacting with vesicles containing palmitoyl-oleoyl-phosphatidylglycerol (POPG)/palmitoyl-oleoyl-phosphatidylethanolamine (POPE). Simulations using this approach have allowed us to understand the different behaviors of peptides with antimicrobial activity with respect to those that do not produce this effect. We found remarkable agreement with a series of experimental results directly supporting our model. Moreover, these results allow us to understand the mechanism of action at the molecular level of these short peptides. Our simulations suggest that mechanical inhomogeneities appear in the membrane, promoting membrane rupture when a threshold concentration of peptides adsorbed on the membrane is achieved. These results explain the high structural demand for these peptides to maintain a delicate balance between the affinity for the bilayer surface, a low peptide-peptide repulsion (in order to reach the threshold concentration), and an acceptable tendency to penetrate into the bilayer. This mechanism is different from those proposed for peptides with long amino acid sequences. Such information is very useful from the medicinal chemistry point of view for the design of new small antimicrobial peptides.

Evaluating the conformational space of the active site of D2 dopamine receptor. Scope and limitations of the standard docking methods.

We report here for the first time the potential energy surfaces (PES) of phenyletilamine (PEA) and meta-tyramine (m-OH-PEA) at the D2 dopamine receptor (D2DR) binding site. PESs not only allow us to observe all the critical points of the surface (minimums, maximums, and transition states), but also to note the ease or difficulty that each local minima have for their conformational inter-conversions and therefore know the conformational flexibility that these ligands have in their active sites. Taking advantage of possessing this valuable information, we analyze how accurate a standard docking study is in these cases. Our results indicate that although we have to be careful in how to carry out this type of study and to consider performing some extra-simulations, docking calculations can be satisfactory. In order to analyze in detail the different molecular interactions that are stabilizing the different ligand-receptor (L-R) complexes, we carried out quantum theory of atoms in molecules (QTAIM) computations and NMR shielding calculations. Although some of these techniques are a bit tedious and require more computational time, our results demonstrate the importance of performing computational simulations using different types of combined techniques (docking/MD/hybrid QM-MM/QTAIM and NMR shielding calculations) in order to obtain more accurate results. Our results allow us to understand in details the molecular interactions stabilizing and destabilizing the different L-R complexes reported here. Thus, the different activities observed for dopamine (DA), m-OH-PEA, and PEA can be clearly explained at molecular level.

Arylated analogues of cypronazole: fungicidal effect and activity on human fibroblasts. Docking analysis and molecular dynamics simulations.

Triadimefon (TDM) and cyproconazole (CPZ) are two triazoles widely used as fungicides. Several azoles were synthesised starting from commercial TDM and CPZ. The compounds were evaluated against phytopathogenic filamentous fungi, including Aspergillus fumigatus (AF), A. niger (AN), A. ustus (AU), A. japonicus (AJ), A. terreus (AT), Fusarium oxysporum and Botrytis cinerea isolated from grapevine in the province of San Juan, Argentina. Three of the synthesised compounds (1-(Biphenyl-4-yloxy)-3,3-dimethyl-1-(1H-1,2,4-triazol-1-yl)butan-2-one, 1; 2-(Biphenyl-4-yl)-3-cyclopropyl-1-(1H-1,2,4-triazol-1-yl)butan-2-ol, 3; 3-Cyclopropyl-2-(4'-fluorobiphenyl-4-yl)-1-(1H-1,2,4-triazol1-yl)butan-2-ol, 4) presented remarkable in vitro fungicidal properties, with better effects than TDM and CPZ on some of the target fungi. Cytotoxicity was assessed using human lung fibroblasts MRC5. Derivative 1, with IC50 values of 389.4 µM, was less toxic towards MRC-5 human lung fibroblasts than commercial TDM (248.5 µM) and CPZ (267.4 µM). Docking analysis and molecular dynamics simulations suggest that the compounds present the same interaction in the binding pocket of the CYP51B enzyme and with the same amino acids as CPZ. The derivatives investigated could be considered broad-spectrum but with some selectivity towards imperfect fungi.

Pseudorotaxane formation affected by stereo-electronic effects. A theoretical and experimental study.

We report a theoretical and experimental study on different complexes of pseudorotaxanes possessing pyridine axles. In order to evaluate the stereo-electronic effects of the methyl substituents in the pyridine ring, complexes with different substitution patterns were synthesized. In this way, it was possible to analyze the different behaviors of these complexes according to the positions of their methyl substituents. Combined techniques of molecular dynamics and quantum mechanical calculations with the help of molecular electrostatic potentials for a simpler visualization of the electronic effects were employed. We have sought experimental support of NMR spectroscopy analysis to corroborate the conclusions obtained from the molecular simulations. Our results not only clearly demonstrate that both electronic and steric effects play key roles in the feasibility of the formation of such complexes, but also the simulations reported here might predict the degree of difficulty of their formation. The combination of computational techniques employed here seems to be an excellent approach to be able to predict whether or not a complex can be formed and with what degree of difficulty. In addition, our experimental and theoretical results have allowed us to visualize the formation of external complexes in the rotaxanes reported here. In this case, the use of bolaforms with trimethylammonium groups at both ends was very useful to evaluate in detail the formation of the so-called external complexes in these systems.

Covalence and π-electron delocalization influence on hydrogen bonds in proton transfer process of o-hydroxy aryl Schiff bases: A combined NMR and QTAIM analysis.

Within the framework of the density functional theory approach, we studied the relationship between the chemical nature of intramolecular hydrogen bonds (HBs) and nuclear magnetic resonance (NMR) parameters, J-couplings and 1H-chemical shifts [δ(1H)], of the atoms involved in such bonds in o-hydroxyaryl Schiff bases during the proton transfer process. For the first time, the shape of the dependence of the degree of covalence in HBs on 1J(N-H), 1J(O-H), 2hJ(O-N), and δ(1H) during the proton transfer process in o-hydroxyaryl Schiff bases was analyzed. Parameters obtained from Bader's theory of atoms in molecules were used to assess the dependence of covalent character in HBs with both the NMR properties. The influence of π-electronic delocalization on 2hJ(N-O) under the proton transfer process was investigated. 2hJ(O-N) in a Mannich base was also studied in order to compare the results with an unsaturated system. In addition, substituent effects on the phenolic ring were investigated. Our results indicate that the covalent character of HBs on both sides of the transition state undergoes a smooth exponential increase as the δ(1H) moves downfield. The degree of covalence of the N⋯H (O⋯H) bond increases linearly as 1J(N-H) (1J(O-H)) becomes more negative, even after reaching the transition state. Non-vanishing values of spin dipolar (SD) and paramagnetic spin orbital terms of 2hJ(O-N) show that π-electronic delocalization has a non-negligible effect on tautomeric equilibrium and gives evidence of the presence of the resonance assisted HB.Variation of the SD term of 2hJ(O-N) follows a similar pattern as the change in the para-delocalization aromaticity index of the chelate ring.

New short cationic antibacterial peptides. Synthesis, biological activity and mechanism of action.

We report a theoretical and experimental study on a new series of small-sized antibacterial peptides. Synthesis and bioassays for these peptides are reported here. In addition, we evaluated different physicochemical parameters that modulate antimicrobial activity (charge, secondary structure, amphipathicity, hydrophobicity and polarity). We also performed molecular dynamic simulations to assess the interaction between these peptides and their molecular target (the membrane). Biophysical characterization of the peptides was carried out with different techniques, such as circular dichroism (CD), linear dichroism (LD), infrared spectroscopy (IR), dynamic light scattering (DLS), fluorescence spectroscopy and TEM studies using model systems (liposomes) for mammalian and bacterial membranes. The results of this study allow us to draw important conclusions on three different aspects. Theoretical and experimental results indicate that small-sized peptides have a particular mechanism of action that is different to that of large peptides. These results provide additional support for a previously proposed four-step mechanism of action. The possible pharmacophoric requirement for these small-sized peptides is discussed. Furthermore, our results indicate that a net +4 charge is the adequate for 9 amino acid long peptides to produce antibacterial activity. The information reported here is very important for designing new antibacterial peptides with these structural characteristics.

A Potent N-(piperidin-4-yl)-1H-pyrrole-2-carboxamide Inhibitor of Adenylyl Cyclase of G. lamblia: Biological Evaluation and Molecular Modelling Studies.

In this work, we report a derivative of N-(piperidin-4-yl)-1H-pyrrole-2-carboxamide as a new inhibitor for adenylyl cyclase of Giardia lamblia which was obtained from a study using structural data of the nucleotidyl cyclase 1 (gNC1) of this parasite. For such a study, we developed a model for this specific enzyme by using homology techniques, which is the first model reported for gNC1 of G. lamblia. Our studies show that the new inhibitor has a competitive mechanism of action against this enzyme. 2-Hydroxyestradiol was used as the reference compound for comparative studies. Results in this work are important from two points of view. on the one hand, an experimentally corroborated model for gNC1 of G. lamblia obtained by molecular modelling is presented; on the other hand, the new inhibitor obtained is an undoubtedly excellent starting structure for the development of new metabolic inhibitors for G. lamblia.

Hydroxynaphthalenecarboxamides and substituted piperazinylpropandiols, two new series of BRAF inhibitors. A theoretical and experimental study.

The oncogenic mutated kinase BRAFV600E is an attractive molecular target because it is expressed in several human cancers, including melanoma. To present, only three BRAF small inhibitors are approved by the FDA for the treatment of patients with metastatic melanoma: Vemurafenib, Dabrafenib and Encorafenib. Although many protocol treatments have been probed in clinical trials, BRAF inhibition has a limited effectiveness because patients invariably develop resistance and secondary toxic effects associated with the therapy. These limitations highlight the importance of designing new and better inhibitors with different structures that could establish different interactions in the active site of the enzyme and therefore decrease resistance progress. Considering the data from our previous report, here we studied two series of derivatives of structural scaffolds as potential BRAF inhibitors: hydroxynaphthalenecarboxamides and substituted piperazinylpropandiols. Our results indicate that structural analogues of substituted piperazinylpropandiols do not show significantly better activities to that previously reported. In contrast, the hydroxynaphthalenecarboxamides derivatives significantly inhibited cell viability and ERK phosphorylation, a measure of BRAF activity, in Lu1205 BRAFV600E melanoma cells. In order to better understand these experimental results, we carried out a molecular modeling study using different combined techniques: docking, MD simulations and quantum theory of atoms in molecules (QTAIM) calculations. Thus, by using this approach we determined that the molecular interactions that stabilize the different molecular complexes are closely related to Vemurafenib, a well-documented BRAF inhibitor. Furthermore, we found that bi-substituted compounds may interact more strongly respect to the mono-substituted analogues, by establishing additional interactions with the DFG-loop at the BRAF-active site. On the bases of these results we synthesized and tested a new series of hydroxynaphthalenecarboxamides bi-substituted. Remarkably, all these compounds displayed significant inhibitory effects on the bioassays performed. Thus, the structural information reported here is important for the design of new BRAFV600E inhibitors possessing this type of structural scaffold.

Conformational and electronic study of dopamine interacting with the D2 dopamine receptor.

We report an exhaustive conformational and electronic study on dopamine (DA) interacting with the D2 dopamine receptor (D2 DR). For the first time, the complete surface of the conformational potential energy of the complex DA/D2 DR is reported. Such a surface was obtained through the use of QM/MM calculations. A detailed study of the molecular interactions that stabilize and destabilize the different molecular complexes was carried out using two techniques: Quantum Theory of Atoms in Molecules computations and nuclear magnetic shielding constants calculations. A comparative study of the behavior of DA in the gas phase, aqueous solution, and in the active site of D2 DR has allowed us to evaluate the degree of deformation suffered by the ligand and, therefore, analyze how rustic are the lock-key model and the induced fit theory in this case. Our results allow us to propose one of the conformations obtained as the "biologically relevant" conformation of DA when it is interacting with the D2 DR.

Design of new quinolin-2-one-pyrimidine hybrids as sphingosine kinases inhibitors.

Sphingosine-1-phosphate is now emerging as an important player in cancer, inflammation, autoimmune, neurological and cardiovascular disorders. Abundance evidence in animal and humans cancer models has shown that SphK1 is linked to cancer. Thus, there is a great interest in the development new SphK1 inhibitors as a potential new treatment for cancer. In a search for new SphK1 inhibitors we selected the well-known SKI-II inhibitor as the starting structure and we synthesized a new inhibitor structurally related to SKI-II with a significant but moderate inhibitory effect. In a second approach, based on our molecular modeling results, we designed new structures based on the structure of PF-543, the most potent known SphK1 inhibitor. Using this approach, we report the design, synthesis and biological evaluation of a new series of compounds with inhibitory activity against both SphK1 and SphK2. These new inhibitors were obtained incorporating new connecting chains between their polar heads and hydrophobic tails. On the other hand, the combined techniques of molecular dynamics simulations and QTAIM calculations provided complete and detailed information about the molecular interactions that stabilize the different complexes of these new inhibitors with the active sites of the SphK1. This information will be useful in the design of new SphK inhibitors.

Searching new structural scaffolds for BRAF inhibitors. An integrative study using theoretical and experimental techniques.

The identification of the V600E activating mutation in the protein kinase BRAF in around 50% of melanoma patients has driven the development of highly potent small inhibitors (BRAFi) of the mutated protein. To date, Dabrafenib and Vemurafenib, two specific BRAFi, have been clinically approved for the treatment of metastatic melanoma. Unfortunately, after the initial response, tumors become resistant and patients develop a progressive and lethal disease, making imperative the development of new therapeutic options. The main objective of this work was to find new BRAF inhibitors with different structural scaffolds than those of the known inhibitors. Our study was carried out in different stages; in the first step we performed a virtual screening that allowed us to identify potential new inhibitors. In the second step, we synthesized and tested the inhibitory activity of the novel compounds founded. Finally, we conducted a molecular modelling study that allowed us to understand interactions at the molecular level that stabilize the formation of the different molecular complexes. Our theoretical and experimental study allowed the identification of four new structural scaffolds, which could be used as starting structures for the design and development of new inhibitors of BRAF. Our experimental data indicate that the most active compounds reduced significantly ERK½ phosphorylation, a measure of BRAF inhibition, and cell viability. Thus, from our theoretical and experimental results, we propose new substituted hydroxynaphthalenecarboxamides, N-(hetero)aryl-piperazinylhydroxyalkylphenylcarbamates, substituted piperazinylethanols and substituted piperazinylpropandiols as initial structures for the development of new inhibitors for BRAF. Moreover, by performing QTAIM analysis, we are able to describe in detail the molecular interactions that stabilize the different Ligand-Receptor complexes. Such analysis indicates which portion of the different molecules must be changed in order to obtain an increase in the binding affinity of these new ligands.

Synthesis and biological evaluation of sphingosine kinase 2 inhibitors with anti-inflammatory activity.

The synthesis of inhibitors of SphK2 with novel structural scaffolds is reported. These compounds were designed from a molecular modeling study, in which the molecular interactions stabilizing the different complexes were taken into account. Particularly interesting is that 7-bromo-2-(2-phenylethyl)-2,3,4,5-tetrahydro-1,4-epoxynaphtho[1,2-b]azepine, which is a selective inhibitor of SphK2, does not exert any cytotoxic effects and has a potent anti-inflammatory effect. It was found to inhibit mononuclear cell adhesion to the dysfunctional endothelium with minimal impact on neutrophil-endothelial cell interactions. The information obtained from our theoretical and experimental study can be useful in the search for inhibitors of SphK2 that play a prominent role in different diseases, especially in inflammatory and cardiovascular disorders.

The nitrone spin trap 5,5­dimethyl­1­pyrroline N­oxide binds to toll-like receptor-2-TIR-BB-loop domain and dampens downstream inflammatory signaling.

The nitrone spin trap 5,5­dimethyl­1­pyrroline N­oxide (DMPO) dampens endotoxin-induced and TLR4-driven priming of macrophages, but the mechanism remains unknown. The available information suggests a direct binding of DMPO to the TIR domain, which is shared between TLRs. However, TLR2-TIR domain is the only TLR that have been crystallized. Our in silico data show that DMPO binds to four specific residues in the BB-loop within the TLR2-TIR domain. Our functional analysis using hTLR2.6-expressing HEKs cells showed that DMPO can block zymosan-triggered-TLR2-mediated NF-κB activation. However, DMPO did not affect the overall TLR2-MyD88 protein-protein interaction. DMPO binds to the BB-loop in the TIR-domain and dampens downstream signaling without affecting the overall TIR-MyD88 interaction. These data encourage the use of DMPO-derivatives as potential mechanism-based inhibitors of TLR-triggered inflammation.

New substituted aminopyrimidine derivatives as BACE1 inhibitors: in silico design, synthesis and biological assays.

We report in this work new substituted aminopyrimidine derivatives acting as inhibitors of the catalytic site of BACE1. These compounds were obtained from a molecular modeling study. The theoretical and experimental study reported here was carried out in several steps: docking analysis, Molecular Dynamics (MD) simulations, Quantum Theory Atom in Molecules (QTAIM) calculations, synthesis and bioassays and has allowed us to propose some compounds of this series as new inhibitors of the catalytic site of BACE1. The QTAIM study has allowed us to obtain an excellent correlation between the electronic densities and the experimental data of IC50. Also, using combined techniques (MD simulations and QTAIM calculations) enabled us to describe in detail the molecular interactions that stabilize the different L-R complexes. In addition, our results allowed us to determine what portion of these compounds should be changed in order to increase their affinity with the BACE1. Another interesting result is that a sort of synergism was observed when the effects of these new catalytic site inhibitors were combined with Ac-Tyr5-Pro6-Tyr7-Asp8-Ile9-Pro10-Leu11-NH2, which we have recently reported as a modulator of BACE1 acting on its exosite.

Antinociceptive effect of neo-clerodane diterpenes obtained from Baccharis flabellata.

We report here for the first time antinociceptive effects of extracts from Baccharis flabellata. Two extracts in this analysis, one obtained in summer and the other during winter time. Our results indicate that both extract show strong antinociceptive effects, being the extracts obtained during the summer significantly more active. Our results suggest that this activity is mainly due to the presence of the diene-acid clerodane ent-15,16-epoxy-19-hydroxy-1,3,13(16),14-clerodatetraen-18-oic acid (DAC) and its dimer called DACD. Employing naloxone as an antagonist of opioid receptors, we demonstrated that both compounds act on opioid receptors, being the antinociceptive effect of DACD stronger than DAC. Thus, the antinociceptive activity of DACD was almost two times stronger than DAC (44.8 over 24.6 s in the hot-plate test) after one hour of treatments. In order to better understand the mechanism of action at molecular level of these compounds, we conducted a molecular modeling study analyzing the molecular interactions of DAC and DACD complexes with the κ-ORs. Our results suggest interactions for both DAC and DACD with Gln115, Val118, Tyr119, Asn122 and Tyr313 stabilizing their complexes; however, these interactions are significantly stronger for DACD with respect to DAC. This finding could explain why DACD have a higher affinity for the κ-ORs. These results are in agreement with the obtained antinociceptive effect. In addition, our results indicate that these neoclerodanes would have a mechanism of action similar to that of salvinorin A; such information can be very useful for the design of new inhibitors of κ-ORs.

Theoretical models to predict the inhibitory effect of ligands of sphingosine kinase 1 using QTAIM calculations and hydrogen bond dynamic propensity analysis.

We report here the results of two theoretical models to predict the inhibitory effect of inhibitors of sphingosine kinase 1 that stand on different computational basis. The active site of SphK1 is a complex system and the ligands under the study possess a significant conformational flexibility; therefore for our study we performed extended simulations and proper clusterization process. The two theoretical approaches used here, hydrogen bond dynamics propensity analysis and Quantum Theory of Atoms in Molecules (QTAIM) calculations, exhibit excellent correlations with the experimental data. In the case of the hydrogen bond dynamics propensity analysis, it is remarkable that a rather simple methodology with low computational requirements yields results in excellent accord with experimental data. In turn QTAIM calculations are much more computational demanding and are also more complex and tedious for data analysis than the hydrogen bond dynamic propensity analysis. However, this greater computational effort is justified because the QTAIM study, in addition to giving an excellent correlation with the experimental data, also gives us valuable information about which parts or functional groups of the different ligands are those that should be replaced in order to improve the interactions and thereby to increase the affinity for SphK1. Our results indicate that both approaches can be very useful in order to predict the inhibiting effect of new compounds before they are synthesized.

The Antimicrobial Activity of Annona emarginata (Schltdl.) H. Rainer and Most Active Isolated Compounds against Clinically Important Bacteria.

Annona emarginata (Schltdl.) H. Rainer, commonly known as "arachichú", "araticú", "aratigú", and "yerba mora", is a plant that grows in Argentina. Infusions and decoctions are used in folk medicine as a gargle against throat pain and for calming toothache; another way to use the plant for these purposes is chewing its leaves. Extracts from bark, flowers, leaves, and fruits from A. emarginata were subjected to antibacterial assays against a panel of Gram (+) and Gram (-) pathogenic bacteria according to Clinical and Laboratory Standards Institute protocols. Extracts from the stem bark and leaves showed moderate activity against the bacteria tested with values between 250⁻1000 µg/mL. Regarding flower extracts, less polar extracts (hexane, dichloromethane) showed very strong antibacterial activity against methicillin-sensitive Staphylococcus aureus ATCC 25923 and methicillin-resistant S. aureus ATCC 43300 with values between 16⁻125 µg/mL. Additionally, hexane extract showed activity against Klebsiella pneumoniae (MIC = 250 µg/mL). The global methanolic extract of the fruits (MeOHGEF) was also active against the three strains mentioned above, with MICs values 250⁻500 µg/mL. Bioassay-guided fractionation of MeOHGEF led to the isolation of a new main compound-(R)-2-(4-methylcyclohex-3-en-1-yl)propan-2-yl (E)-3-(4-hydroxyphenyl)acrylate (1). The structure and relative configurations have been determined by means of 1D and 2D NMR techniques, including COSY, HMQC, HMBC, and NOESY correlations. Compound 1 showed strong antimicrobial activity against all Gram (+) species tested (MICs = 3.12⁻6.25 µg/mL). In addition, the synthesis and antibacterial activity of some compounds structurally related to compound 1 (including four new compounds) are reported. A SAR study for these compounds was performed based on the results obtained by using molecular calculations.

Cholinesterase-inhibitory effect and in silico analysis of alkaloids from bulbs of Hieronymiella species.

BACKGROUND: In Argentina, the Amaryllidaceae family (59 species) comprises a wide variety of genera, only a few species have been investigated as a potential source of cholinesterases inhibitors to treat Alzheimer disease (AD). PURPOSE: To study the acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities of the basic dichloromethane extracts (E) from Hieronymiella aurea, H. caletensis, H. clidanthoides, H. marginata, and H. speciosa species, as well as the isolated compounds from these plant extracts. STUDY DESIGN AND METHODS: AChE and BChE inhibitory activities were evaluated with the Ellman's spectrophotometric method. The alkaloids composition from the E was obtained by gas chromatography-mass spectrometry (GC-MS). The E were successively chromatographed on a silica gel column and permeated on Sephadex LH-20 column to afford the main alkaloids identified by means of spectroscopic data. Additionally, an in silico study was carried out. RESULTS: Nine known alkaloids were isolated from the E of five Hieronymiella species. Galanthamine was identified in all the species by GC-MS standing out H. caletensis with a relative abundance of 9.79% of the total ion current. Strong AChE (IC50 = 1.84 - 15.40 µg/ml) and moderate BChE (IC50 = 23.74 - 136.40 µg/ml) inhibitory activities were displayed by the extracts. Among the isolated alkaloids, only sanguinine and chlidanthine (galanthamine-type alkaloids) demonstrated inhibitory activity toward both enzymes. The QTAIM study suggests that sanguinine has the strongest affinity towards AChE, attributed to an additional interaction with Ser200 as well as stronger molecular interactions Glu199 and His440.These results allowed us to differentiate the molecular behavior in the active site among alkaloids possessing different in vitro inhibitory activities. CONCLUSION: Hieronymiella species growing in Argentina represent a rich and widespread source of galanthamine and others AChE and BChE inhibitors alkaloids. Additionally, the new trend towards the use of natural extracts as pharmaceuticals rather than pure drugs opens a pathway for the development of a phytomedicine derived from extracts of Hieronymiella spp.