Jair Mafezoli

Structure of Trypanosoma cruzi glycosomal glyceraldehyde-3-phosphate dehydrogenase complexed with chalepin, a natural product inhibitor, at 1.95 Å resolution

The structure of the glycosomal glyceraldehyde‐3‐phosphate dehydrogenase (gGAPDH) from Trypanosoma cruzi complexed with chalepin, a natural product from Pilocarpus spicatus, has been determined by X‐ray crystallography to 1.95 Å resolution. The structure is in the apo form without cofactors in the subunits of the tetrameric gGAPDH in the asymmetric unit. Unequivocal density corresponding to the inhibitor was clearly identified in one monomer. The final refined model of the complex shows extensive conformational changes when compared with the native structure. The mode of binding of chalepin to gGAPDH and its implications for inhibitor design are discussed.

In vitro activity of Rutaceae species against the trypomastigote form of Trypanosoma cruzi.

The activity of crude plant extracts of nine species of Rutaceae against the trypomastigote form of Trypanosoma cruzi was evaluated at 4 mg/ml. Thirty-two crude extracts were tested and eight of them showed significant activity (>80%). The most active extract was obtained from the stems of Pilocarpus spicatus (97.3%). Fractionation of the active crude extracts provided 25 fractions which were tested against the trypomastigote form of T. cruzi at 2 mg/ml. Of these six showed significant activity (>80%). The most active fractions (100%) were obtained from the leaves of Almeidea coerulea (butanol fraction) and Conchocarpus inopinatus (dichloromethane fraction).

Strategies for the isolation and identification of trypanocidal compounds from the Rutales

Crude extracts of Rutales species were tested in vitro against the trypomastigote form of Trypanosoma cruzi at 4 mg/mL, and 20% of them showed significant activity (80%). Their inhibitory activity against the glycolytic enzyme GAPDH from T. cruzi has also been evaluated at the concentrations of 100 and 200 mg/mL. Additionally, the inhibitory activity of 13 purified coumarins were also assayed against T. cruzi-GAPDH. Chalepin was the most active substance with IC50 = 64 mM. The 3D structure of the complex chalepin-enzyme was elucidated by X-ray crystallography, revealing the architecture of the interactions between the inhibitor and the enzyme active site.

Estudo químico e biológico de Tephrosia toxicaria Pers.

The ethanol extracts from leaves, stems, pods and roots were assayed against the 3rd instar Aedes aegypti larvae and the highest activity was observed in the roots extracts (LC50 47.86 ppm). This extract was submitted to partition with hexane, chloroform, ethyl acetate and methanol. The respective fractions were bioassayed and the best larvicidal activities were identified in the hexane (LC50 23.99 ppm) and chloroform (LC50 13.80 ppm) fractions. Antioxidant activity (DDPH method) was observed in the ethanol extract (IC50 276 µg/mL) from roots of T. toxicaria. Fractions from this extract were also tested and the highest antioxidant activity (IC50 89 µg/mL) was found in the methanol fraction. The flavonoids iso-obovatin (1), obovatin (2), 6a,12a-dehydro-β-toxicarol (3), 6a,12a-dehydro-α-toxicarol (4) and α-toxicarol (5) were isolated and bioassayed against A. aegypti. The flavonoid 5 showed the best larvicidal activity (LC50 24.55 ppm). The antioxidant activity of 2 was investigated and showed IC50 3.370 µg/mL. The antioxidant and larvicidal activities of Tephrosia toxicaria are reported for the first time.

Synthesis and Antifungal Activity In Vitro of Isoniazid Derivatives against Histoplasma capsulatum var. capsulatum

ABSTRACT Histoplasmosis is a severe infection that affects millions of patients worldwide and is endemic in the Americas. Amphotericin B (AMB) and itraconazole are highly effective for the treatment of severe and milder forms of the disease, but AMB is toxic, and the bioavailability of itraconazole is erratic. Therefore, it is important to investigate new classes of drugs for histoplasmosis treatment. In this study, a series of nine isoniazid hydrazone derivatives were synthesized and evaluated for their antifungal activities in vitro against the dimorphic fungus Histoplasma capsulatum var. capsulatum. The drugs were tested by microdilution in accordance with CLSI guidelines. The compound N′-(1-phenylethylidene)isonicotinohydrazide had the lowest MIC range of all the compounds for the yeast and filamentous forms of H. capsulatum. The in vitro synergy of this compound with AMB against the planktonic and biofilm forms of H. capsulatum cells was assessed by the checkerboard method. The effects of this hydrazone on cellular ergosterol content and membrane integrity were also investigated. The study showed that the compound alone is able to reduce the ergosterol content of planktonic cells and can alter the membrane permeability of the fungus. Furthermore, the compound alone or in combination with AMB showed inhibitory effects against mature biofilms of H. capsulatum. N′-(1-Phenylethylidene)isonicotinohydrazide alone or combined with AMB might be of interest in the management of histoplasmosis.

New pyrone and quinoline alkaloid from Almeidea rubra and their trypanocidal activity

O estudo da fracao acetato de etila do extrato metanolico das folhas de Almeidea rubra A. St.- Hil. (Rutaceae) permitiu o isolamento de duas substâncias ineditas: 4-metoxi-6-(2-(metilamino)fenil)- 2H-piran-2-ona e acetato de rel-(7R,8R)-8-((E)-3-hidroxi-3-metil-1-butenil)-4,8-dimetoxi-5,6,7,8- tetraidrofuro(2,3-b)quinolin-7-ila; e dos alcaloides arborinina, N-metil-1-hidroxi-3-metoxiacridona, esquimianina, cocusagina, isodutaduprina, isoesquimianina e isococusagina. Atraves da analise dos dados espectroscopicos foram estabelecidas as estruturas quimicas das substâncias isoladas sendo que para os alcaloides ineditos tais dados sao descritos pela primeira vez. Alem disso, os ensaios bilogicos sobre as formas tripomastigotas do Trypanosoma cruzi das substâncias isoladas mostraram que elas possuem atividade tripanocida moderada. The investigation of the ethyl acetate fraction of methanol extract from leaves of Almeidea rubra A. St.-Hil. (Rutaceae) afforded two new compounds 4-methoxy-6-(2-(methylamino)phenyl)-2H- pyran-2-one and rel-(7R,8R)-8-((E)-3-hydroxy-3-methyl-1-butenyl)-4,8-dimethoxy-5,6,7,8- tetrahydrofuro(2,3-b)quinoline-7-yl acetate, along with the known compounds arborinine, N-methyl- 1-hydroxy-3-methoxyacridone, skimmianine, kokusagine, isodutaduprine, isoskimmianine, and isokokusagine. Their structures were established based on their spectral data, and for the new compounds these data are described herein. Additionally, these compounds were assayed on the tripomastigote forms of Trypanosoma cruzi showing moderate trypanocidal activity.

Rotenoids from Tephrosia toxicaria with larvicidal activity against Aedes aegypti, the main vector of dengue fever

In the search for new larvicides from plants, we have investigated the potential activity of the rotenoids deguelin (1), 12a-hydroxy-α-toxicarol (2) and tephrosin (3), isolated from the bioactive ethanol extract of roots of Tephrosia toxicaria Pers., against Aedes aegypti, the main vector of dengue. The absolute configuration of these compounds was determined by circular dichroism (CD) spectra. The LC50 values of the compounds evaluated justify the potential of T. toxicaria as a new natural larvicide.

Volatile Constituents of Different Populations of Pilocarpus spicatus Saint Hill. (Rutaceae) from the Northeast of Brazil

Abstract The chemical composition of essential oils from leaves, stem bark, stem wood, root bark and root wood of five populations of Pilocarpus spicatus, collected in the northeast of Brazil, were compared by GC/MS. The aliphatic ketones 2-undecanone, 2-tridecanone and 2-pentadecanone were present in samples of all populations. 2-Tridecanone (1.7–84.7 %) was detected in 30 out of 34 samples analyzed. It was the main component in all samples of root barks, except one where 2-pentadecanone (24.7%) was the major component. 2-Undecanone, β-eudesmol and sabinene were the major components of leaf oils.

Cytotoxic compounds from the marine-derived fungus Aspergillus sp. recovered from the sediments of the Brazilian coast.

A fungal strain of Aspergillus sp. (BRF 030) was isolated from the sediments collected in the northeast coast of Brazil, and the cytotoxic activity of its secondary metabolites was investigated against HCT-116 tumour cell line. The cytotoxicity-guided fractionation of the extracts from this fungus cultured in potato-dextrose-sea water for 14 days at room temperature yielded the hetero-spirocyclic γ-lactams pseurotin A (1), pseurotin D (2) and pseurotin FD-838 (7), the alkaloids fumitremorgin C (5), 12,13-dihydroxy fumitremorgin C (6), methylsulochrin (4) and bis(dethio)bis(methylthio)gliotoxin (3). Among them, fumitremorgin C (5) and 12,13-dihydroxy fumitremorgin C (6) were the most active. The cytotoxic activities of the extracts from Aspergillus sp. grown from 7 to 28 days were investigated, and they were associated with the kinetic production of the compounds. The most active extracts (14 and 21 days) were those with the highest relative concentrations of the compounds fumitremorgin C (5) and 12,13-dihydroxy fumitremorgin C (6).

Bioprospection of cytotoxic compounds in fungal strains recovered from sediments of the Brazilian coast.

The cytotoxic activities of extracts (50 μg/ml) from 48 fungal strains, recovered from sediments of Pecéms offshore port terminal (Northeast coast of Brazil), against HCT‐116 colon cancer cell lines were investigated. The most promising extract was obtained from strain BRF082, identified as Dichotomomyces cejpii by phylogenetic analyses of partial RPB2 gene sequence. Thus, it was selected for bioassay‐guided isolation of the cytotoxic compounds. Large‐scale fermentation of BRF082 in potato dextrose broth, followed by chromatographic purification of the bioactive fractions from the liquid medium, yielded gliotoxin (4) and its derivatives acetylgliotoxin G (3), bis(dethio)bis(methylsulfanyl)gliotoxin (1), acetylgliotoxin (5), 6‐acetylbis(dethio)bis(methylsulfanyl)gliotoxin (2), besides the quinazolinone alkaloid fiscalin B. All isolated compounds were tested for their cytotoxicities against the tumor cell lines HCT‐116, revealing 4 and 3 as the most cytotoxic ones (IC50 0.41 and 1.06 μg/ml, resp.).