Search

Ornithine decarboxylase (ODC) catalyzes the decarboxylation of ornithine to produce putrescine, the first step in the metabolism of polyamines (putrescine, spermidine, and spermine), which are essential growth factors in eukaryotic cells. ODC is active as a homodimer and depends on pyridoxal 5′-phosphate (PLP) as a cofactor. An increase in the concentration of polyamines has been associated with carcinogenesis. Therefore, there is much interest in identifying inhibitors of this pathway as potential chemotherapeutic and chemopreventive agents. The best-known inhibitor of mammalian ODC is α-difluoromethylornithine (DFMO), a highly selective compound that alkylates Cys-360 (a residue of the ODC active site). Although DFMO was initially developed for the treatment of cancer, the World Health Organization recommends its use in combination with nifurtimox for the treatment of human African trypanosomiasis. Considering the importance of ODC as a promising target for the treatment of various types of cancer and other infectious diseases, choosing the right method for screening potential inhibitors can help to accelerate the discovery of new drugs. Several methods for the determination of ODC activity are found in the literature. Among these, we can mention analysis with radioactive markers, colorimetric assays using auxiliary enzymes to detect CO2 or H2O2 release, chromatographic separations with putrescine derivatization, mass spectrometry, and spectroscopic techniques. In this review, the main analysis methods used will be described, highlighting their advantages and disadvantages, as well as identifying the most promising methods for high-throughput screening.

The aim of this study was to perform the isolation and characterization of vasodilatory flavonoids from Tapirira guianensis Aubl. (Annacardiaceae) leaves. In this context, ethyl acetate fraction (E...

Nucleoside hydrolases are a strategic target for the development of drugs to treat leishmaniasis, a neglected disease that affects 700 thousand to one million people annually. The present study aimed to identify Leishmania donovani nucleoside hydrolase (LdNH) inhibitors from the leaves of Ormosia arborea, a tree endemic to Brazilian ecosystems, through a strategy based on 1H NMR analyses and chemometrics. The aqueous EtOH extract of O. arborea leaves inhibited LdNH activity by 95%. The extract was fractionated in triplicate (13 in each step, making a total of 39 fractions). Partial least squares discriminant analysis (PLS-DA) was used to correlate the 1H NMR spectra of the fractions with their LdNH inhibitory activity and thus to identify the spectral regions associated with the bioactivity. The strategy aimed at isolating the probable bioactive substances and led to two new A-type proanthocyanidins, linked to a p-coumaroyl unit (1 and 2), which appeared as noncompetitive inhibitors of LdNH (IC50: 28.2 ± 3.0 μM and 25.6 ± 4.1 μM, respectively). This study confirms the usefulness of the NMR-based chemometric methods to accelerate the discovery of drugs from natural products.

Conformational stabilization mediated by hydrogen bonds can play an important role in the drug-target recognition process. Non-classical hydrogen bonding is receiving great attention, where C–H⋯Y hydrogen bonds are found in some biological systems and are important for molecular recognition, leading to the possibility of exploration of this interaction in drug design projects. Studies that identify and characterize new non-classical interactions in different systems can assist in the rational application of these interactions. In this paper, we describe the serendipitous identification and characterization of a new non-classical hydrogen bond donor moiety found in 3-acyl-substituted furan. We applied a theoretical and experimental approach in the study of several N-acylhydrazones containing 3-acyl-substituted furan, where we describe the presence of a hydrogen bond between the C(2)–H bond of 3-acyl-substituted furan with the imine nitrogen in the syn conformer of the N-acylhydrazone moiety. In addition, we evaluated the strength of this interaction with intermolecular models. Thus, we believe that the 3-acyl-substituted furan moiety can be rationally explored as a hydrogen bond donor moiety in drug design projects.

Ocotea notata (Lauraceae) is popularly known as white-cinnamon. Ocotea species have several medicinal uses, especially for treating chest pain, rheumatism and wounds. The present study aimed to analyze the chemical composition of O. notata n-hexane fraction, in addition to its anti-mycobacterial and immunomodulatory activities. The n-hexane fraction was analyzed by GC-MS and was chromatographed to afford 15 subfractions (SF1-15), where SF5 was identified, by GC-MS and NMR, as the sesquiterpene spathulenol. The n-hexane fraction was the most potent in inhibiting nitric oxide (NO) and tumor necrosis factor-alpha (TNF-α) production on LPS-stimulated macrophages (IC50 8.3 ± 0.9 and 5.9 ±1.0 μg/mL, respectively). SF4, a major subfraction, that presents a spathulenol analogous as a constituent, also inhibited NO and TNF-α production. Spathulenol only modulated NO production (IC50 45.6 ± 1.4 μg/mL). The n-hexane fraction, SF4, and spathulenol revealed antimycobacterial activity against Mycobacterium bovis BCG, M. tuberculosis H37Rv, and M299 strains. Spathulenol inhibited the growth of Mtb H37Rv with MIC50 36.9 ± 1.5 μg/mL (167.5 ± 6.8 μM), and Mtb M299 with MIC5042.1 ± 0.5 μg/mL (191.0 ± 2.2 μM). This is the first report describing the isolation of spathulenol from O. notata leaves and its anti-mycobacterial activity. Resumo Ocotea notata (Lauraceae) é popularmente conhecida como canela-branca. As espécies do gênero Ocotea têm vários usos medicinais, especialmente no tratamento de dores no peito, reumatismo e feridas. O presente estudo teve como objetivo analisar a composição química da fração n-hexano de O. notata, além de suas atividades antimicobacteriana e imunomoduladora. A fraçao n-hexano foi analisada por GC-MS e fracionada fornecendo 15 subfrações (SF1-15), onde SF5 foi identificado, por GC-MS e RMN, como o sesquiterpeno espatulenol. A fração n-hexano foi a mais potente na inibição da produção de óxido nítrico (NO) e do fator de necrose tumoral-alfa (TNF-α) em macrófagos estimulados por LPS (IC50 8,3 ± 0,9 e 5,9 ±1,0 μg/mL, respectivamente). SF4, urna subfração majoritária, que apresenta um análogo do espatulenol como constituinte, também inibiu a produção de NO e TNF-α. O espatulenol modulou apenas a produção de NO (IC50 45,6 ±1,4 μg/mL). A fração n-hexano, SF4 e espatulenol apresentaram atividade antimicobacteriana contra as cepas de Mycobacterium bovis BCG, M. tuberculosis (Mtb) H37Rv e M299. O espatulenol inibiu o crescimento de Mtb H37Rv com MIC50 36,9 ± 1,5 μg/mL (167,5 ± 6,8 μM) e Mtb M299 com MIC50 42,1 ± 0,5 μg/mL (191,0 ± 2,2 μM). Este é o primeiro relato que descreve o isolamento do espatulenol das folhas de O. notata e sua atividade antimicobacteriana.

Among the compounds of natural origin, diterpenes have proved useful as drugs for the treatment of cancer. Marine organisms, such as soft corals and algae, are a promising source of diterpenes, being a rich and unexplored source of cytotoxic agents. This study evaluated a library of 32 natural and semisynthetic marine diterpenes, including briarane, cembrane, and dolabellane nuclei, with the aim of determining their cytotoxicity against three human cancer cell lines (A549, MCF7, and PC3). The three most active compounds were submitted to a flow cytometry analysis in order to determine induction of apoptosis against the A549 cell line. An NMR analysis was conducted to determine and evaluate the interactions between active diterpenes and tubulin. These interactions were characterized by a computational study using molecular docking and MD simulations. With these results, two cembrane and one chlorinated briarane diterpenes were active against the three human cancer cell lines, induced apoptosis in the A549 cell line, and showed interactions with tubulin preferably at the taxane-binding site. This study is a starting point for the identification and optimization of the marine diterpenes selected for better antitumor activities. It also highlights the power of integrating NMR studies, computational predictions, and in vitro assays in the search for compounds with antitumor activity.

The flavonoid rutin presents several pharmacological effects, despite this, its application in the pharmaceutical industry can be significantly limited by its low bioavailability. The development of lipophilic derivatives by esterification of hydroxyl groups with fatty acid chains may be an effective strategy to change their physicochemical properties. This work aims to use high-speed countercurrent chromatography (HSCCC) to isolate rutin esters produced by esterification reactions catalyzed by the immobilized lipase from Candida antarctica (Novozyme 435®). The lipase-catalyzed synthesis of rutin esters (R3-R18:2) in 2-methyl-2-butanol exhibited conversions that ranged from 16 to 40% and highest conversion for short chain fatty acids (R4-R12). After initial partitioning tests of the reaction mixture in different solvent proportions followed by thin layer chromatography (TLC) analysis, the biphasic solvent systems consisting in HEMWat (hexane/ethyl acetate/methanol/water) in different proportions were chosen to separate rutin esters. These esters were separated for the first time to the reaction mixture via HSCCC. This technique proved to be more advantageous than the traditionally one since it allowed quick isolation and high purity (≥ 90%) of the products. It also permitted a substrate recovery for reuse in other enzymatic reactions. Furthermore, the results add valuable information, specially concerning the structures elucidation of different rutin esters not previously described.

The treatment of diseases with animal resources or their derivatives is a traditional practice worldwide, representing a wide field of research for the elaboration of strategies of management and conservation of the fauna, and contributing to the search for sustainable therapy alternatives. This study presents the therapeutic applications of animal fats to the treatment of several diseases in Pimenteiras do Oeste, state of Rondonia, Brazil. Twelve animals including: mammals, fishes, reptiles and birds are reported as a source of medicinal lard for the treatment of respiratory illnesses (asthma, flu, bronchitis, cough), rheumatism, and earache and as a healing agent (dislocation and wounds). The ethnopharmacological focus of the study was on Green-anaconda fat (Eunectes murinus), which stands out for its frequent local use and the lack of previous chemical studies. The chemical composition of E. murinus fat was analyzed by gas and liquid chromatography, both coupled to mass spectrometry. The main fatty acids identified were oleic, linoleic and palmitic acids, which were also predominant in the composition of the triglycerides. Pharmacological analysis of Green-anaconda fat showed a significant anti-inflammatory effect, which is related to its use by traditional communities. Having confirmed the pharmacological potential of Green-anaconda fat, its fatty acid composition was used as a parameter in the search for vegetable oils from the Amazon Region with a similar composition. This comparative analysis can be of help by proposing therapeutic alternatives for the Amazonian population. The use of plant sources can contribute to the conservation of the aforementioned species.

The aim of this study was to perform the isolation and characterization of vasodilatory flavonoids from

This study presents new inhibitors of the nucleoside hydrolase from Leishmania donovani (LdNH) with in vitro leishmanicidal activity. Biological screening of 214 Brazilian plant extracts was performed to select plants with enzyme inhibitory activity. Two plants were selected for their results, and for their lack of prior phytochemical description: Leandra amplexicaulis DC. (Melastomataceae) and Urvillea rufescens Cambess (Sapindaceae). Three flavonoids were isolated by bioguided fractionation of the hydroethanolic extracts: kaempferol 3-O-α-L-rhamnopyranoside (1) and kaempferol 3-O-β-D-xylopyranosyl-(1→2)-α-L-rhamnopyranoside (2) from L. amplexicaulis, as well as tricetin-4′-O-methyl flavone (3) from U. rufescens. These flavonoids showed inhibitory activities (IC50) of 197.4 μM (1), 74.7 μM (2) and 1.1 μM (3) on the LdNH. Their binding mode was proposed based on molecular docking with LdNH and by NMR Saturation Transfer Difference studies. Kinetic studies demonstrate that the most potent inhibitor (3) acts by uncompetitive inhibition. This study reports for the first time the inhibition of LdNH by naturally sourced flavonoids.

Sigma-hole (σ-hole) bonds are interactions that are gaining special attention in medicinal chemistry. This type of interaction, initially assigned to the halogens (group 17 of the periodic table), has been extended to atoms of groups 14, 15 and 16. Sulfur atoms have been outstanding for describing these interactions at the intramolecular level (to induce conformational stability) and the intermolecular level (participating in molecular recognition of bioactive compounds by their respective targets). Thus, this work describes the theoretical and experimental characterization of a 1,4-N⋯S σ-hole intramolecular interaction in the N-acylhydrazone cardioactive prototype LASSBio-294 (1), which leads to conformational stabilization and has a direct influence on the molecular properties of this inotropic prototype compared to a negative control for the interaction, LASSBio-897 (2), which is the regioisomer at the thiophene ring. Our theoretical results were reached using the B3LYP/6-311G(d) level of theory, including analysis of conformational, orbital and electrostatic properties. We performed experimental studies using IR, Raman, UV and NMR spectroscopies, which corroborated our theoretical data, showing significant differences between LASSBio-294 (1) and LASSBio-897 (2) in relation to the bond strength of the groups involved in the N⋯S interaction (S–C and NC bonds), the energies of the orbitals associated with the S lone pair (Lp(S)) and the antibonding NC π bond (π*(NC)), as well as the 15N chemical shifts in both systems. Together, our results show how this unusual interaction can influence the molecular properties of some organic compounds.

Earthworm metabolism is recognized as a useful tool for monitoring environmental insults and measuring ecotoxicity, yet extensive earthworm metabolic profiling using 1H nuclear magnetic resonance (NMR) spectroscopy has been limited in scope. This study aims to expand the embedded metabolic material in earthworm coelomic fluid, coelomocytes, and tissue to aid systems toxicology research. Fifty-nine metabolites within Eisenia fetida were identified, with 47 detected in coelomic fluid, 41 in coelomocytes, and 54 in whole-worm samples and tissue extracts. The newly detected but known metabolites 2-aminobutyrate, nicotinurate, Nδ,Nδ,Nδ-trimethylornithine, and trigonelline are reported along with a novel compound, malylglutamate, elucidated using 2D NMR and high-resolution MS/MS. We postulate that malylglutamate acts as a glutamate/malate store, chelator, and anionic osmolyte and helps to provide electrolyte balance.

The aim of this study was to perform the isolation and characterization of vasodilatory flavonoids from Tapirira guianensis Aubl. (Annacardiaceae) leaves. In this context, ethyl acetate fraction (EA fraction) was obtained and subjected to fractionation batches by HSCCC affording: myricetin 3-O-α-L-rhamnopyranoside (myricitrin, 1); quercetin 3-O-(6”-O-galloyl)-β-D-galactopyranoside (2); quercetin 3-O-α-L-arabinofuranoside (avicularin, 3); and quercetin 3-O-α-L-rhamnopyranoside (quercitrin, 4). Myricitrin (1) induced a relaxation of 56.07 ± 13.04% at 300 μM (P 2) (IC50 1.0 ± 0.3 µM) was the strongest peroxidase inhibitor. In conclusion, it was possible to verify that myricitrin together with quercetin are mainly responsible for vasodilatory potential, besides flavonoid 2 for myeloperoxidase inhibition. Together these flavonoids seem to be responsible for Tapirira guianensis cardiovascular effects.

Ocimum basilicum and Ocimum gratissimum (Lamiaceae) are used to treat diabetes mellitus in Africa. In a previous work, we identified chicoric acid as a hypoglycemic substance in O. gratissimum. This study aims to compare the chemical metabolite profile and the hypoglycemic activity of unfractionated aqueous extracts from leaves of both Lamiaceae species. The metabolite composition of OB and OG decoctions (10% w/v) was analyzed using HPLC-DAD and NMR tools. Chicoric acid showed to be the major phenolic in both extracts, besides caftaric, caffeic, and rosmarinic acids; nevertheless, there is approximately three times more of this substance in OG. From 1D- and 2D-NMR analyses, 19 substances were identified in OB, while 12 in OG. The in vivo acute hypoglycemic activity of the extracts was assessed intraperitoneally in streptozotocin (STZ)-induced diabetic mice. The doses of 100 and 200 mg/kg of both extracts significantly reduced their glycemia, compared to controls (P < 0.05). OB was a little more effective than OG, despite the lower content of chicoric acid in OB. This result strongly suggests that components other than chicoric acid contribute to the hypoglycemic activity of the two extracts. Despite the abundance of caffeic and rosmarinic acids in OB, their hypoglycemic activity observed at 8.3 μmol/kg was low. This is the first chemical profile of crude extracts from Ocimum species by NMR. Our findings confirmed the potential of both species in DM treatment in spite of marked differences in their chemical composition. However, long-term studies are necessary in order to identify the most promising of the two species for the development of an herbal medicine.

Due to the lack of effective pharmacotherapy options to treats Alzheimer’s disease, new strategies have been approached in the search for multi-target molecules as therapeutic options. In this work, four indole alkaloids, geissoschizoline, geissoschizone, geissospermine, and 3′,4′,5′,6′-tetradehydrogeissospermine were isolated from Geissospermum vellosii (Pao pereira) and evaluated for their anticholinesterase activities. While geissospermine inhibited only butyrylcholinesterase (BChE), the other alkaloids behaved as non-selective inhibitors of acetylcholinesterase (AChE) and BChE. In cell viability tests, only geissoschizoline was not cytotoxic. Therefore, geissoschizoline actions were also evaluated in human cholinesterases, where it was twice as potent inhibitor of hBChE (IC50 = 10.21 ± 0.01 µM) than hAChE (IC50 = 20.40 ± 0.93 µM). On enzyme kinetic studies, geissoschizoline presented a mixed-type inhibition mechanism for both enzymes. Molecular docking studies pointed interactions of geissoschizoline with active site and peripheral anionic site of hAChE and hBChE, indicating a dual site inhibitor profile. Moreover, geissoschizoline also played a promising anti-inflammatory role, reducing microglial release of NO and TNF-α at a concentration (1 μM) ten and twenty times lower than the IC50 values of hBChE and hAChE inhibition, respectively. These actions give geissoschizoline a strong neuroprotective character. In addition, the ability to inhibit hAChE and hBChE, with approximate inhibitory potencies, accredits this alkaloid for therapeutic use in the moderate to severe phase of AD. Thus, geissoschizoline emerges as a possible multi-target prototype that can be very useful in preventing neurodegeneration and restore neurotransmission.

As part of the ongoing efforts in discovering potentially bioactive natural products from medicinal plants, the present study was conducted to isolate a new complex triterpenoid saponin from the barks of Albizia lebbeck. It was isolated by using chromatographic methods and its structural elucidation was performed using detailed analyses of 1H and 13C NMR spectra including 2D-NMR (COSY, TOCSY, HSQC and HMBC) spectroscopic techniques, high-resolution electrospray ionization mass spectrometry (HRESIMS) analysis and chemical conversions. Its structure was established as 21-[[(2E,6S)-6-[6-deoxy-4-O-[(2E,6S)-6-hydroxy-2-(hydroxymethyl)-6-methyl-1-oxo-2,7-octadienyl]-[(β- d -glucopyranosyl)oxy]-2-(hydroxymethyl)-6-methyl-1-oxo-2,7-octadienyl]-[(β- d -glucopyranosyl)oxy]-2,6-dimethyl-1-oxo-2,7-octadienyl]oxy]-16-hydroxy-3-[[O-β- d -xylopyranosyl-(1 → 2)-O-α- l -arabinopyranosyl-(1 → 6)-2-(acetylamino)-2-deoxy-β- d -glucopyranosyl]oxy]-(3β,16α,21β)-olean-12-en-28-oic acid O-α- l -arabinofuranosyl-(1 → 4)-O-[β- d -glucopyranosyl-(1 → 3)]-O-6-deoxy-α- l -mannopyranosyl-(1 → 2)-β- d -glucopyranosyl ester (1). Additionally, this study aimed to investigate the permeability property of 1, its activity on membrane integrity and supramolecular interactions with cellular constituents using in vitro experimental models.

Nucleoside hydrolase is an important target for the development of new leishmanicidal agents due to its role in parasite proliferation. Using the principles of fragment-based drug discovery, a library of 111 fragments was assembled for screening by nuclear magnetic resonance using the saturation transfer difference. Five fragments were selected as ligands of Leishmania donovani nucleoside hydrolase (NHLd) and kinetics studies revealed that fragment 3 acts as a new non-competitive inhibitor. Its binding mode was proposed on the basis of molecular docking, using a structural model of NHLd constructed by homology. An intermolecular interaction between fragments 3 and 5 was found by NOESY and docking studies. These data could be used to design more potent NHLd inhibitors and to aid the development of leishmanicidal drugs.

In this paper, the in vitro microsomal hepatic metabolism of the antiasthmatic prototype LASSBio-448 and the structural identification of its major phase I metabolites were described. Incubation with pooled rat liver microsomes converted LASSBio-448 to the following major metabolites: O-demethyl-LASSBio-448 (M1) and 3,4-dihydroxyphenyl- LASSBio-448 (M2). These metabolites were formed by the dealkylation step of 3,4-dimethoxyphenyl and 1,3- benzodioxole subunits, respectively, in agreement with the in silico prediction using MetaSite Program. The development of a reproducible analytical methodology for the major metabolites by using HPLC-MS showed that both reactions require NADPH generating system and appeared to be catalyzed by cytochrome P450 (CYP). The identification of which isoenzyme was involved in the oxidative metabolism of LASSBio-448 was carried out by pre-incubations with the selective inhibitors sulfaphenazole (CYP2C9), quinidine (CYP2D6), furafylline (CYP1A2), p-nitrophenol (CYP2E1), ticlopidine (CYP2C19) and ketoconazole (CYP3A4). CYP1A2, CYP2C19 and CYP3A4 were demonstrated to be involved in the oxidative biotransformation of LASSBio-448.

Ocimum gratissimum L. is popularly used to treat diabetes mellitus. The hypoglycemic activity of this medicinal species has been confirmed by in vivo studies. The present study conducted a chemical investigation of a leaf decoction (10% p/v) of O. gratissimum monitored by in vivo hypoglycemic activity assays. Four phenolic substances were identified: l-caftaric acid (1), l-chicoric acid (2), eugenyl-β-d-glucopyranoside (3) and vicenin-2 (4). The acute hypoglycemic activity of the O. gratissimum decoction fractions Og1-S (300mg/kg), Og1-A (240mg/kg) and Og1-B (80mg/kg) was evaluated intraperitoneally in normal and streptozotocin-induced diabetic mice. They reduced glycemia by 63%, 76% and 60% (in 120min), respectively, in the diabetic mice. Subfractions of Og1-A were also evaluated under the same conditions: Og1-AS (200mg/kg) and Og1-AP (40mg/kg) produced a decrease of only 37% and 39%, respectively. Among the major phenolic substances, only chicoric acid (2; 3mg/kg) reduced significantly the glycemic levels of diabetic mice by 53%, 120min after treatment. This is the first study describing the hypoglycemic activity of chicoric acid in an animal model of diabetes mellitus. In addition, we suggest that there may be other substances contributing to this activity. Thus, for the first time, a correlation is established between the hypoglycemic activity of O. gratissimum and its chemical composition.

The aim of this research was to perform a phytochemical study of the methanol leaves extract of T. guianensis (MET) guided by vasodilatory and antioxidant activities. The chemical profile of MET and the ethyl acetate fraction (EA fraction) was determined by HPLC-UV-MS and EA fraction guided fractionation by reverse-phase chromatography. The vasorelaxant effects of MET, fractions, sub-fractions and constituents were assessed on rat aorta pre-contracted with phenylephrine. Antioxidant activity was evaluated by using a DPPH assay. The results show that MET-induced vasodilation was dependent on NO/cGMP; and that the PI3K/Akt pathway seems to be the main route involved in eNOS activation. The EA fraction showed greater vasodilatory and antioxidant potency and was submitted to further fractionation. This allowed the isolation and characterization of quercetin, quercetin 3-O-(6″-O-galloyl)-β-d-galactopyranoside and 1,4,6-tri-O-galloyl-β-d-glucose. Also, galloyl-HHDP-hexoside and myricetin deoxyhexoside were identified by HPLC-UV-MS. These compounds are being described for the first time for T. guianensis. 1,4,6-tri-O-galloyl-β-d-glucose and quercetin 3-O-(6″-O-galloyl)-β-d-galactopyranoside showed no vasodilatory activity. Quercetin and myricetin glycoside seems to contribute to the MET activity, since they have been reported as vasodilatory flavonoids. MET-induced vasodilation could contribute to the hypotensive effect of T. guianensis previously reported.

Hybrid nanoparticles (NPs) are emerging as an important family of multifunctional nanoscale materials. Due to its unique properties cyclodextrins (CDs) present the possibility to be employed in nanostructured devices for multiple applications. Therefore the study of hybrids NPs containing CDs, metal nanoparticles and an organic molecular guest is promising for applications in separation science, sensing, optics, catalysis and medical therapy. To investigate the hybrid nanoparticles properties in light of technological and medical science interfaces we chose tramadol hydrochloride as the model molecular guest. Using 2D 1H NMR and FTIR spectroscopies and ESI-MS we studied the topology and host-guest stoichiometry of inclusion compound between 2-hydroxypropyl-α-cyclodextrin and tramadol. These techniques confirmed that the inclusion compounds present 1:1 stoichiometry with the tramadol aromatic moiety inserted in the CD cavity and the amine part interacting with the CD superficial OHs. Performing capillary electrophoresis experiments, the stability constants of inclusion compounds were obtained and showed that the increase of temperature during the synthesis of these NPs can decrease their stabilities. Precipitated palladium nanoparticles were determined to have 80-200 nm of size distribution and the incorporation of tramadol was evidenced by additional 1H NMR experiments.

In this study the recombinant enzyme nucleoside hydrolase of Leishmania donovani (r Ld NH) was expressed in Escherichia coli in connection with maltose binding protein (MBP). The r Ld NH–MBP showed efficient a significant in vitro activity with inosine as substrate. From the coupled reaction with xanthine oxidase (XO) it was possible to determine the kinetic constants of r Ld NH–MBP as K M (434 ± 109 μM) and V max (0.20 ± 0.02 μM). In addition, two nucleoside analogs (compounds 1 and 2 ) were tested as prototypes of r Ld NH inhibitors. These compounds presented high affinity for the enzyme with K i values of 1.6 ± 0.2 and 17.0 ± 2.1 μM, respectively, as well as 271 and 26 folds higher than the affinity constant found for inosine. We also determined the type of enzyme inhibition, using double-reciprocal plot for these two compounds and the results confirmed a competitive inhibition. Additional docking studies showed the binding manner of compounds 1 and 2 inside the active site of Ld NH revealing the essential residues for an effective inhibition. These results confirm that compounds 1 and 2 are strong r Ld NH–MBP inhibitors.

Phytochemical investigation of Mimosa xanthocentra Mart. (Leguminosae: Mimosoideae), an herb comprising the diet of marsh deer and pampas deer in the Brazilian Pantanal, led to the isolation of flavones isovitexin-2″- O -α- l -rhamnopyranoside 1 and vitexin-2″- O -α- l -rhamnopyranoside 2 , and a mixture of flavonols avicularin 3a and reynoutrin 3b . At room temperature, compound 1 was observed in the 1 H and 13 C NMR spectra as a mixture of two rotamers in equilibrium in a DMSO- d 6 solution. Molecular modeling using MM2 calculations led to the first proposal of three-dimensional solution structure of both observed conformers. The energy barrier between both rotamers was evaluated theoretically by molecular modeling and experimentally by variable-temperature NMR studies and calculation of Δ G ‡ of rotation using Eyring equation. This is the first report on flavonoids from M. xanthocentra .

Context: Triosephosphate isomerase (TIM) is a ubiquitous enzyme that has been targeted for the discovery of small molecular weight compounds with potential use against Trypanosoma cruzi, the causative agent of Chagas disease. We have identified a new selective inhibitor chemotype of TIM from T. cruzi (TcTIM), 1,2,4-thiadiazol-5(4H)-one.Objective: Study the mechanism of TcTIM inhibition by a 1,2,4-thiadiazol derivative.Methods: We performed the biochemical characterization of the interaction of the 1,2,4-thiadiazol derivative with the wild-type and mutant TcTIMs, using DOSY-NMR and MS experiments. Studies of T. cruzi growth inhibition were additionally carried out.Results and conclusion: At low micromolar concentrations, the compound induces highly selective irreversible inactivation of TcTIM through non-covalent binding. Our studies indicate that it interferes with the association of the two monomers of the dimeric enzyme. We also show that it inhibits T. cruzi growth in culture.

(±)-1,2- O -Isopropylidene-3,6-di- O -benzyl- myo -inositol is a relevant starting material in the synthesis of inositol phosphates and their analogs. In this study, we disclose our efforts toward an efficient methodology for the kinetic resolution of this compound by lipase B of Candida antarctica (Novozym 435). This reaction selectively affords L-(−)-1,2- O -isopropylidene-5- O -acetyl-3,6-di- O -benzyl- myo -inositol. From a conversion of 34% with EtOAc as an acylating agent, the use of vinyl acetate increased the yield to over 49%, while maintaining a very high ee (>99%). The combination of the latter reagent with TBME as a solvent accelerates the reaction.

This work reports the nutritional potential and the antiradical activity of the cladodes of the Brazilian Opuntia monacantha Haw. (Cactaceae), as well the isolation of kaempferol and isorhamnetin through activity-guided fractionation from its active MeOH extract. TLC, HPLC-DAD and NMR techniques were used to detect and identify the compounds. The water (91.1%), ash, protein, fibre and lipid contents (15.0, 5.4, 18.5 and 1.4 g/100 g, respectively) were shown to be quite similar to the mean values of other Opuntia spp., some widely used as food and forage. The antioxidant activity of their MeOH extract and of its n-hexane, EtOAc and n-BuOH fractions was assessed by measuring the ability of the fractions to scavenge DPPH radical, showing an activity enhancement for the fractions in comparison to the crude MeOH extract. The well-known free-radical scavenging activity of the isolated flavonoids reinforces the contribution of these compounds to the presented activity of the O. monacantha cladodes. (C) 2010 Elsevier Ltd. All rights reserved.

NMR is one of the most powerful techniques for ligand-biomolecule interaction studies and drug screening and design. There are several methods that are strongly used, including chemical shift perturbation (CSP), saturation transfer difference (STD) and diffusion coefficients. However, one of the most useful and easy to apply NMR parameters in medicinal chemistry studies is the spin-lattice relaxation data, which can be employed to investigate the strength and topology of intermolecular interactions, such as drug-drug, drug-protein, drug-DNA, drug-micelle (or vesicle) and biomolecule-biomolecule interactions. This review deals with the newest applications of T(1) in different studies of interest for drug design and evaluation.

This work investigates the chiral recognition of d - and l -tryptophan enantiomers using 2-hydroxypropyl-α- or β-cyclodextrins (HPαCD and HPβCD) as selectors. Capillary electrophoresis experiments showed that only the α-form was effective for enantiomeric separation. NMR measurements (T1 and ROESY) and circular dichroism experiments were carried out to provide information about the topologies of interaction between the tryptophan isomers and cyclodextrins. For HPβCD, the inclusion of both isomers was verified, while for HPαCD only d -tryptophan was inserted in the cavity. These results agree with the stability constant values obtained from NMR diffusion experiments. We were able to determine that, together with charged groups of tryptophan ( NH 3 + and COO−), the indole moiety is the third guest interaction point with the cyclodextrin.

As part of our search of bioactive saponins from Brazilian plants, phytochemical study of the seeds of Inga laurina led to the isolation of a new complex triterpenoid saponin, named ingasaponin. It is the first saponin isolated from a species of Inga genus. It was isolated by using chromatographic methods and its structural elucidation was performed using detailed analyses of (1)H and (13)C NMR spectra including 2D-NMR spectroscopic techniques and chemical conversions. Its structure was established as 21-[[(2E,6S)-6-[[6-deoxy-4-O-[(2E,6S)-6-[(β-D-glucopyranosyl)oxy]-2,6-dimethyl-1-oxo-2,7-octadienyl]-[(β-D-glucopyranosyl)oxy]-2,6-dimethyl-1-oxo-2,7-octadienyl]-[(β-D-glucopyranosyl)oxy]-2,6-dimethyl-1-oxo-2,7-octadienyl]-[(β-D-glucopyranosyl)oxy]-2-(hydroxymethyl)-6-methyl-1-oxo-2,7-octadienyl]oxy]-16-hydroxy-3-[[O-β-D-xylopyranosyl-(1 → 2)-O-α-L-arabinopyranosyl-(1 → 6)-2-(acetylamino)-2-deoxy-β-D-glucopyranosyl]oxy]-(3β,16α,21β)-olean-12-en-28-oic acid O-α-L-arabinofuranosyl-(1 → 4)-O-[β-D-glucopyranosyl-(1 → 3)]-O-6-deoxy-α-L-mannopyranosyl-(1 → 2)-β-D-glucopyranosyl ester (1). The hemolytic potential of 1 was evaluated using in vitro assays, and its adjuvant activity on the cellular immune response against ovalbumin antigen was investigated using in vivo models.

The importance of flavonoids for the antileishmanial activity of Kalanchoe pinnata was previously demonstrated by the isolation of quercitrin, a potent antileishmanial flavonoid. In the present study, the aqueous leaf extract from the medicinal plant K. pinnata (Crassulaceae) afforded a kaempferol di-glycoside, named kapinnatoside, identified as kaempferol 3-O-alpha-L-arabinopyranosyl (1-->2) alpha-L-rhamnopyranoside (1). In addition, two unusual flavonol and flavone glycosides already reported, quercetin 3-O-alpha-L-arabinopyranosyl (1-->2) alpha-L-rhamnopyranoside (2) and 4',5-dihydroxy-3',8-dimethoxyflavone 7-O-beta-D-glucopyranoside (3), have been isolated. Their structures were determined via analyses of mono and bi-dimensional (1)H and (13)C NMR spectroscopic experiments and HR-MALDI mass spectra. Because of its restricted occurrence and its abundance in K. pinnata, flavonoid (2) may be a chemical marker for this plant species of high therapeutic potential. The three flavonoids were tested separately against Leishmania amazonenis amastigotes in comparison with quercitrin, quercetin and afzelin. The quercetin aglycone - type structure, as well as a rhamnosyl unit linked at C-3, seem to be important for antileishmanial activity.

Coupling constant values ( J NH and J NC ) obtained from 1 H and 13 C NMR spectra of acetylcholine chloride (ACh) in several solvents with a wide range of permittivity constants ( e ) are remarkably invariant, indicating the absence of solvent effects in the conformational equilibrium of this compound. The values show that the OCH 2 CH 2 N + system occurs in a gauche arrangement. J NH and J NC are observable in most solvents, but not in chlorine-containing solvents, and are not dependent on solvent viscosity. This behavior was explained using data from T 1 measurements. The measurement of NMR diffusion coefficients shows that ACh has a greater tendency to aggregate when dissolved in chlorinated solvents, a fact that could explain the observed differences in 14 N T 1 .

A mechanistic 1H NMR kinetic study on the solvolysis of complexes [Ru2(μ-O)(μ-O2CCH3)2(4-picoline)6](PF6)2 (1), [Ru2(μ-O)(μ-O2CCH3)2(pyridine)6](PF6)2 (2), and [Ru2(μ-O)(μ-O2CCH3)2(4-picoline)4(pyridine)2](PF6)2 (3) in DMSO-d6, CD3OD and CD3CN was carried out. All rate constants were calculated from the kinetic data, which fit in a series of reversible pseudo first order and second order reactions. The comparative kinetic study and the large ΔH‡ and positive ΔS‡ values for all solvolysis support the dissociative nature of the ligand exchange mechanism. The leaving ligand effect, determined by comparison of the forward reactions for complexes 1 and 3, showed that the rate constants for 4-picoline substitution are smaller than for pyridine. An apparent contradiction with the proposed mechanism for the reverse reaction in DMSO-d6 where the less basic ligand coordinates faster, was explained by a ligand–solvent interaction, which was demonstrated by our kinetics results, molecular modeling and through the determination of diffusion coefficients in DMSO-d6 and CD3OD using DOSY.

A study of the nature of the anthelmintic p-cresol:piperazine complex in chloroform solution has been conducted using different NMR techniques: self-diffusion coefficients using DOSY; NOE, NULL, and double-selective T1 measurements to determine inter-molecular distances; and selective and non-selective T1 measurements to determine correlation times. The experimental results in solution and CP-MAS were compared to literature X-ray diffraction data using molecular modeling. It was shown that the p-cresol:piperazine complex exists in solution in a very similar manner as it does in the solid state, with one p-cresol molecule hydrogen bonded through the hydroxyl hydrogen to each nitrogen atom of piperazine. The close correspondence between the X-ray diffraction data and the inter-proton distances obtained by NULL and double selective excitation techniques indicate that those methodologies can be used to determine inter-molecular distances in solution.

Two new alkaloids containing Nb-methyl-triptamine units linked to bis-quinoline moieties, N-8”-formyl-calycosidine and N-8”-methyl-N-1’-demethyl-iso-calycosidine, were isolated from the aerial part of Margaritopsis carrascoana, along with known calycosidine and hodgkinsine. The cytotoxicity of the isolated alkaloids has been evaluated against cancer cell lines. The first two compounds showed only weak activity against HL-60 cell line, while hodgkinsine was the most active compound against HCT-116, OVCAR-8, HL-60 and SF-295 cancer cell lines. The structures of the isolated compounds were established by spectroscopic and molecular mechanical methods.

The species Piper ottonoides Yuncker (Piperaceae), known as “joao-brandim”, is a shrub that occurs in the Brazilian Amazon rainforest. Its roots and leaves are used in traditional medicine as local anesthetic to treat toothache and sore throat. In this study, the structural characterization in mixture of isobutyl amides present in semi-purified fractions from fruits, leaves, stems and roots of P. ottonoides was achieved by employing gas chromatography-mass spectrometry (GC-MS) and a combination of nuclear magnetic resonance (NMR) techniques (1H, 1H-1H COSY, 1H-13C HSQC, 1H-13C HMBC, TOCSY and 1H-1H J-RES). The MS fragmentation patterns and the NMR chemical shifts, multiplicities, coupling constants and the signal correlations in the two-dimensional spectra were carefully analyzed also taking into account some key elements of differentiation among the compounds. The data set allowed identifying unequivocally the new amide N-isobutyl-7-(4’- methoxyphenyl)-2E,4E-heptadienamide, which we named ottonoidenamide as well as piperovatine and chingchengenamide A in all parts of the plant, the additional presence of pipercallosine and pipercallosidine in roots and of dihydropiperlonguminine in fruits and leaves. The presence of piperovatine and pipercallosine in P. ottonoides should be associated to its traditional use.

PW2 (HPLKQYWWRPSI) was selected from phage display libraries through an alternative panning method using living sporozoites of Eimeria acervulina as target. Synthetic PW2 shows anticoccidial activity against E. acervulina and Eimeria tenella with very low hemolytic activity. It also displays antifungal activity but no activity against bacteria. We present the solution structure of the PW2 bound to SDS micelles. In the absence of an interface, PW2 is in random coil conformation. In micelles, structural calculation shows that Trp-7 forms the hydrophobic core that is important for the peptide folding. Lys-4, Tyr-6, Trp-8, and Arg-9 are in the same surface, possibly facing the micelle interface. This possibility was supported by the fact that chemical shift differences for these residues were more pronounced when compared with PW2 in water and in SDS. PW2 gains structure upon binding to SDS micelles. Lys-4, Tyr-6, Trp-8, and Arg-9 were found to bind to the micelle. Trp-7, Trp-8, and Arg-9 composed the WW+ consensus found in the sequence of the peptides selected with the phage display technique against E. acervulina sporozoites. This suggested that Trp-7, Trp-8, and Arg-9 are probably key residues not only for the peptide interaction with SDS micelles but also for the interaction with E. acervulina sporozoites surface.

Resveratrol is a polyphenol found in black grapes and red wine and has many biological activities. In this study, we evaluated the effect of resveratrol alone and in association with amphotericin B (AMB) against Leishmania amazonensis . Our results demonstrate that resveratrol possesses both antipromastigote and antiamastigote effects, with 50% inhibitory concentrations (IC 50 s) of 27 and 42 μM, respectively. The association of resveratrol with AMB showed synergy for L. amazonensis amastigotes, as demonstrated by the mean sums of fractional inhibitory index concentration (mean ΣFIC) of 0.483, although for promastigotes, this association was indifferent. Treatment with resveratrol increased the percentage of promastigotes in the sub-G 0 /G 1 phase of the cell cycle, reduced the mitochondrial potential, and showed an elevated choline peak and CH 2 -to-CH 3 ratio in the nuclear magnetic resonance (NMR) spectroscopy analysis; all these features indicate parasite death. Resveratrol also decreased the activity of the enzyme arginase in uninfected and infected macrophages with and without stimulation with interleukin-4 (IL-4), also implicating arginase inhibition in parasite death. The anti- Leishmania effect of resveratrol and its potential synergistic association with AMB indicate that these compounds should be subjected to further studies of drug association therapy in vivo .

A new polysaccharide with an estimated weight-average molar mass of 2.6 × 10 3 was isolated from Allium ampeloprasum var. porrum by hot water extraction, and purified by Sephacryl S-300 HR high-resolution chromatography. It was composed of d -fructose and d -glucose in 10:6 molar ratio, respectively. The structure of the glucofructan was investigated by chemical and spectroscopic methods, including methylation analysis, nuclear magnetic resonance, and electrospray mass spectrometry (ES-MS). The results permitted the structure of the glucofructan to be written as α- d -Glc p -(1→1)-β- d -Fru f -(2→1)-{[α- d -Glc p -(1→6)-β- d -Fru f -(2→6)]-β- d -Fru f -(2→1)} 4 -β- d -Fru f -(2↔1)-α- d -Glc p . Results of the present study indicated that this new glucofructan exhibited significant gastroprotective property, using in vivo experimental models.

The genusOcotea(Lauraceae) is distributed mainly in tropical and subtropical regions. Some species of this genus asO. puberulaandO. quixoshave been described in the literature, showing antibacterial activity. AndOcotea macrophyllashowed anti-inflammatory activity with inhibition of COX-1, COX-2, and LOX-5. The purpose of this study was the phytochemical investigation of the plant speciesOcotea notatafrom Restinga Jurubatiba National Park, Macaé, RJ, Brazil, and the search for antimycobacterial fractions and compounds. The crude extract was evaluated for antimycobacterial activity and presented95.75±2.53% of growth inhibition at 100 µg/mL. Then, it was subjected to a liquid-liquid partition and subsequently was chemically investigated by HPLC, revealing the major presence of flavonoids. In this process the partition fractions hexane, ethyl acetate, and butanol are shown to be promising in the antimycobacterial assay. In addition, ethyl acetate fraction was chromatographed and afforded two flavonoids identified by MS and NMR as afzelin and isoquercitrin. The isolated flavonoids afzelin and isoquercitrin were evaluated for their antimycobacterial activity and for their ability to inhibit NO production by macrophages stimulated by LPS; both flavonoids isoquercitrin (Acet22) and afzelin (Acet32) were able to inhibit the production of NO by macrophages. The calculated IC50of Acet22 and Acet32 was 1.03 and 0.85 µg/mL, respectively.

A polysaccharide with an estimated weight-average molar mass of 5.35 × 10 5 was obtained from an aqueous extract of pseudobulbs of Cyrtopodium andersonii R. Br. It was composed of d -glucose and d -mannose in 1:3 molar ratio. Chemical and spectroscopic analyses revealed a linear structure of the polymer with a backbone composed of (1 → 4)-linked β- d -glucopyranosyl and mannopyranosyl units slightly branched at C-2, C-3, and C-6 by side chains, as terminal non reducing residues of d -mannopyranose and d -glucopyranose. It was found to contain 14.6% of acetyl groups substituted at C-2 of (1 → 4)-linked β- d -mannopyranosyl units. The acetylated glucomannan demonstrated antiinflammatory and antiulcerogenic activities.

Equilibrium dialysis and NMR spin-lattice relaxation time (T1) studies of the interaction of the trypanocidal drug 3-nitrobenzaldehyde guanyl hydrazone (3NBGH) with bovine serum albumin (BSA) were used to show that this interaction is weak and nonspecific, making 3NBGH a potential agent for the prophylaxis of Trypanosoma cruzi infected blood.

The effects of the changes in sample concentration on the NMR chemical shifts and on the selective and non-selective spin-lattice relaxation rates (R1S and R1NS) of the three isomers of nitrobenzaldeyde guanyl hydrazone (NBGH) pure and with bovine serum albumin (BSA) were measured in solution. The results were used to determine the correlation times (tc), showing that the degree of intermolecular drug-drug association varies with the nitro group position on the ring and that this degree of association interferes with the interaction of these drugs with BSA. The results suggest that the degree of drug-drug and drug-BSA association are related to the in vitro anti-Trypanosoma cruzi activity of these compounds.

MARCKS (Myristoylated Alanine-Rich C Kinase substrate) is a natively unfolded protein that interacts with actin, Ca(2+)-Calmodulin, and some plasma membrane lipids. Such interactions occur at a highly conserved region that is specifically phosphorylated by PKC: the Effector Domain. There are two other conserved domains, MH1 (including a myristoylation site) and MH2, also located in the amino terminal region and whose structure and putative protein binding capabilities are currently unknown. MH2 sequence contains a serine that we described as being phosphorylated only in differentiating neurons (S25 in chick). Here, Circular Dichroism (CD) and Nuclear Magnetic Resonance (NMR) spectroscopy were used to characterize the phosphorylated and unphosphorylated forms of a peptide with the MARCKS sequence surrounding S25. The peptide phosphorylated at this residue is recognized by monoclonal antibody 3C3 (mAb 3C3). CD and NMR data indicated that S25 phosphorylation does not cause extensive modifications in the peptide structure. However, the sharper lines, the absence of multiple spin systems and relaxation dispersion data observed for the phosphorylated peptide suggested a more ordered structure. Surface Plasmon Resonance was employed to compare the binding properties of mAb 3C3 to MARCKS protein and peptide. SPR showed that mAb 3C3 binds to the whole protein and the peptide with a similar affinity, albeit different kinetics. The slightly ordered structure of the phosphorylated peptide might be at the origin of its ability to interact with mAb 3C3 antibody, but this binding did not noticeably modify the peptide structure.

Piper claussenianum inflorescences crude methanol extract was tested for hypoglycemic effect in streptozotocin-induced diabetic rats. The blood glucose levels of rats treated with methanol extract were reduced from 318.4±28.1 mg/dl before treatment to 174.2±38.3 mg/dl after 12 days of treatment (P

A series of 22 aromatic guanyl hydrazones, prepared by condensation of several aldehydes with aminoguanidine hydrochloride, were fully characterized by NMR techniques and tested in vitro against the trypomastigote form of Trypanosoma cruzi , the causative agent of Chagas disease. Most of the compounds, especially those without hydrogen bonding groups and possessing ortho -substitution, were significantly more active than crystal violet (ID 50 536 μM). The most active compound has an ID 50 value of 17 μM (25 times more potent than gentian violet).