In vitro antiproliferative and apoptotic effects of thiosemicarbazones based on (-)-camphene and R-(+)-limonene in human melanoma cells.

A series of 38 thiosemicarbazone derivatives based on camphene and limonene were evaluated for their antiproliferative activity. Among them, 19 were synthesized and characterized using proton and carbon-13 nuclear magnetic resonance (1H and 13C NMR). For initial compound selection, human melanoma cells (SK-MEL-37) were exposed to a single concentration of a compound (100 µM) for 24, 48, and 72 hours, and cell detachment was visually observed. Cell viability was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. Nineteen compounds (4, 6, 8, 11, 13, 14, 15, 16, 17, 18, 20, 22, 25, 26, 31, 3', 4', 6', and 9') yielded cell viability below 20%. Subsequently, IC50 values for these compounds were determined, ranging from 11.56 to 55.38 µM, after 72 hours of treatment. Compound 17 (o-hydroxybenzaldehyde (-)-camphene-based thiosemicarbazone) demonstrated the lowest IC50 value, followed by compound 4 (benzaldehyde (-) camphene-based thiosemicarbazone) at 12.84 µM. Regarding compound 4, we observed the induction of a characteristic ladder pattern of DNA fragmentation through gel electrophoresis. Furthermore, fluorescence, flow cytometry and scanning microscopy assays revealed morphological changes consistent with apoptosis induction. Additionally, the measurement of caspase 6 and 8 activity in cellular extracts after treatment for 2, 4, 6, and 24 hours suggested the potential involvement of the extrinsic apoptosis pathway in the mechanism of action of compound 4. Further investigations, including molecular docking studies, are required to fully explore the potential of compound 4 and the other selected compounds, highlighting their promising role in future melanoma therapy research.

Kopsanone and N-oxide isolated from Aspidosperma macrocarpon Mart. (Apocynaceae) leaves and their MAO-A inhibitory activity.

Aspidosperma macrocarpon Mart., popularly known as 'guatambu' or 'peroba', is found from North American (Mexico) to South American (Argentina) continents and in Brazil. Two indole alkaloids were isolated from leaves of A. macrocarpon, kopsanone (1) and unreported N(4)-oxide-kopsanone (2).

4-Fluorobenzaldehyde limonene-based thiosemicarbazone induces apoptosis in PC-3 human prostate cancer cells.

AIMS: This study evaluated parameters of toxicity and antiproliferative effects of (+)-N(1)-4-fluorobenzaldehyde-N(4)-{1-methyl-1-[(1R)-4-methylcyclohexene-3-il]-ethyl}-thiossemicarbazone (4-FTSC) in PC-3 adenocarcinoma prostate cells. MAIN METHODS: Cytotoxicity of 4-FTSC in PC-3 cells was evaluated using MTT assay. Morphology examination of PC-3 cells treated with 4-FTSC was also performed as well as the cell death mechanisms induced were investigated using flow cytometry. Parameters of toxicity of 4-FTSC was conducted by the investigation of its potential myelotoxicity and lymphotoxicity, hemolytic activity and acute oral toxicity profile. KEY FINDINGS: 4-FTSC showed promising cytotoxic effects against PC-3 cells (IC50 = 18.46 µM). It also triggered apoptotic morphological changes, phosphatidylserine externalization and a significant increase of DNA fragmentation in PC-3 cells. Moreover, 4-FTSC did not show changes in the PC-3 cell cycle with levels of p21, p27, NFĸB and cyclin D1 similar to those found in both control and treated cells. 4-FTSC also promoted an increase of p53 levels associated with mitochondrial impairment through loss of ∆Ψm and ROS overproduction. 4-FTSC-induced cell death mechanism in PC-3 cells involved activation of caspase-3/-7 through apoptosis intrinsic pathway via caspase-9. Regarding toxicological profile, 4-FTSC showed in vitro lymphotoxicity, although with low cytotoxicity for bone marrow progenitors and no hemolytic potential. Moreover, it was classified as GHS category 5 (LD50 > 2000-5000 mg/Kg), suggesting it has low acute oral systemic toxicity. SIGNIFICANCE: 4-FTSC seems to be a promising candidate to be used as a clinical tool in prostate cancer treatment. Further studies are required to better clarify its toxicopharmacological effects found in this compound.

Generating nanoparticles containing a new 4-nitrobenzaldehyde thiosemicarbazone compound with antileishmanial activity.

Thiosemicarbazones are an important class of compounds that have been extensively studied in recent years, mainly because of their broad profile of pharmacological activity. A new 4-nitrobenzaldehyde thiosemicarbazone compound (BZTS) that was derived from S-limonene has been demonstrated to have significant antiprotozoan activity. However, the hydrophobic characteristic of BZTS limits its administration and results in low oral bioavailability. In the present study, we proposed the synthesis of nanoparticle-based block copolymers that can encapsulate BZTS, with morphological evaluation of the nanoparticle suspensions being performed by transmission and cryo-transmission electronic microscopy. The mean particle sizes of the nanoparticle suspensions were determined by static light and dynamic light scattering (SLS/DLS), and the hydrodynamic radius (Rh) was determined using the Stokes-Einstein equation. The zeta potential (ζ) and polydispersity index (PDI) were also determined. The entrapment encapsulation efficiency of the BZTS nanoparticles was measured by ultraviolet spectrophotometry. In vitro activity of BZTS nanoparticle suspensions against intracellular amastigotes of Leishmania amazonensis and cytotoxic activity were also evaluated. The results showed the production of spherical nanoparticles with varied sizes depending on the hydrophobic portion of the amphiphilic diblock copolymers used. Significant concentration-dependent inhibitory activity against intracellular amastigotes was observed, and low cytotoxic activity was demonstrated against macrophages.

4-Nitrobenzaldehyde thiosemicarbazone: a new compound derived from S-(-)-limonene that induces mitochondrial alterations in epimastigotes and trypomastigotes of Trypanosoma cruzi.

Trypanosoma cruzi is the causative agent of Chagas' disease, a parasitic disease that remains a serious health concern with unsatisfactory treatment. Drugs that are currently used to treat Chagas' disease are partially effective in the acute phase but ineffective in the chronic phase of the disease. The aim of the present study was to evaluate the antitrypanosomal activity and morphological, ultrastructural and biochemical alterations induced by a new molecule, 4-nitrobenzaldehyde thiosemicarbazone (BZTS), derived from S-(-)-limonene against epimastigote, trypomastigote and intracellular amastigote forms of T. cruzi. BZTS inhibited the growth of epimastigotes (IC50 = 9·2 µ m), intracellular amastigotes (IC50 = 3·23 µ m) and inhibited the viability of trypomastigotes (EC50 = 1·43 µ m). BZTS had a CC50 of 37·45 µ m in LLCMK2 cells. BZTS induced rounding and distortion of the cell body and severely damaged parasite mitochondria, reflected by extensive swelling and disorganization in the inner mitochondrial membrane and the presence of concentric membrane structures inside the organelle. Cytoplasmic vacuolization, endoplasmic reticulum that surrounded organelles, the loss of mitochondrial membrane potential, and increased mitochondrial O2 •- production were also observed. Our results suggest that BZTS alters the ultrastructure and physiology of mitochondria, which could be closely related to parasite death.

Cell death and ultrastructural alterations in Leishmania amazonensis caused by new compound 4-Nitrobenzaldehyde thiosemicarbazone derived from S-limonene.

BACKGROUND: The treatment of leishmaniasis with pentavalent antimonials is problematic because of their toxicity. Investigations of potentially active molecules are important to discover less toxic drugs that are viable economic alternatives for the treatment of leishmaniasis. Thiosemicarbazones are a group of molecules that are known for their wide versatility and biological activity. In the present study, we examined the antileishmania activity, mechanism of action, and biochemical alterations produced by a novel molecule, 4-nitrobenzaldehyde thiosemicarbazone (BZTS), derived from S-limonene against Leishmania amazonensis. RESULTS: BZTS inhibited the growth of the promastigote and axenic amastigote forms, with an IC50 of 3.8 and 8.0 µM, respectively. Intracellular amastigotes were inhibited by the compound with an IC50 of 7.7 µM. BZTS also had a CC50 of 88.8 µM for the macrophage strain J774A1. BZTS altered the shape, size, and ultrastructure of the parasites, including damage to mitochondria, reflected by extensive swelling and disorganization of the inner mitochondrial membrane, intense cytoplasmic vacuolization, and the presence of concentric membrane structures inside the organelle. Cytoplasmic lipid bodies, vesicles inside vacuoles in the flagellar pocket, and enlargement were also observed. BZTS did not induce alterations in the plasma membrane or increase annexin-V fluorescence intensity, indicating no phosphatidylserine exposure. However, it induced the production of mitochondrial superoxide anion radicals. CONCLUSIONS: The present results indicate that BZTS induced dramatic effects on the ultrastructure of L. amazonensis, which might be associated with mitochondrial dysfunction and oxidative damage, leading to parasite death.

Novel R-(+)-limonene-based thiosemicarbazones and their antitumor activity against human tumor cell lines.

In an attempt to develop potent and selective antitumor agents, a series of novel thiosemicarbazones derived from a natural monoterpene R-(+)-limonene was synthesized and their antitumor activity was evaluated. Overall, the majority of tested compounds exhibited considerable inhibitory effects on the growth of a wide range of cancer cell lines. Almost all of tested thiosemicarbazones were especially sensitive to prostate cells (PC-3). Derivatives 5, 6, 8, 9, 10, 11 and 13 presented the most potent antitumor activity against PC-3 cells. These compounds showed lower value of GI50 (0.04-0.05 µM) than the reference drug paclitaxel, besides a high selectivity for the same cell line. The 4-fluorobenzaldehyde derivative 10 was the most selective compound for prostate cells, while 2-hydroxybenzaldehyde derivative 8 was the most active compound, with potent antitumor activity against all tested cell lines.

Benzaldehyde thiosemicarbazone derived from limonene complexed with copper induced mitochondrial dysfunction in Leishmania amazonensis.

BACKGROUND: Leishmaniasis is a major health problem that affects more than 12 million people. Treatment presents several problems, including high toxicity and many adverse effects, leading to the discontinuation of treatment and emergence of resistant strains. METHODOLOGY/PRINCIPAL FINDINGS: We evaluated the in vitro antileishmanial activity of benzaldehyde thiosemicarbazone derived from limonene complexed with copper, termed BenzCo, against Leishmania amazonensis. BenzCo inhibited the growth of the promastigote and axenic amastigote forms, with IC(50) concentrations of 3.8 and 9.5 µM, respectively, with 72 h of incubation. Intracellular amastigotes were inhibited by the compound, with an IC(50) of 10.7 µM. BenzCo altered the shape, size, and ultrastructure of the parasites. Mitochondrial membrane depolarization was observed in protozoa treated with BenzCo but caused no alterations in the plasma membrane. Additionally, BenzCo induced lipoperoxidation and the production of mitochondrial superoxide anion radicals in promastigotes and axenic amastigotes of Leishmania amazonensis. CONCLUSION/SIGNIFICANCE: Our studies indicated that the antileishmania activity of BenzCo might be associated with mitochondrial dysfunction and oxidative damage, leading to parasite death.

N-trans-feruloyltyramine and flavonol glycosides from the leaves of Solanum sordidum

Chemical investigation of the leaves of Solanum sordidum Sendtn., Solanaceae, resulted in the isolation and identification of sitosterol, stigmasterol, 3β-O-β-glycopyranosyl stigmasterol, 3β-O-β-glycopyranosyl sitosterol, kaempferol-3-O-α-rhamnopyranosyl-(1-6)-α-glycopyranoside, rutin, and N-trans-feruloyltyramine. The structures of these compounds were established by analysis of 1D and 2D NMR spectrometric data and comparison with data in the literature. The evaluation of antioxidant activity showed an IC50 of 159.5 ppm for the chloroformic fraction and IC50 of 77.5 ppm for the hydromethanolic fraction.

Anti-leishmanial activity of alkaloidal extracts obtained from different organs of Aspidosperma ramiflorum.

The present study was designated to evaluate semi-quantitative antileishmanial activity of alkaloidal extracts that were obtained from 1g of different parts of Aspidosperma ramiflorum (leaves, roots, seeds, and stem barks). Alkaloidal extracts of barks and leaves presented a good activity against the extracellular form (promastigotes) of Leishmania (L.) amazonensis. It is known that compounds responsible for the antileishmanial activity in the alkaloidal extracts from A. ramiflorum are the monoterpenoid indole alkaloids ramiflorine A and ramiflorine B, therefore extracts obtained from different plant parts were analyzed by electrospray ionization mass spectrometry (ESI-MS) in order to evidence the presence of these bioactive alkaloids. Based on these findings, alkaloidal extract from leaves was fractionated on preparative thin-layer chromatography in a bioassay-guided fractionation affording individual purified ramiflorines A and B. Both ramiflorines A and B showed significant activity against Leishmania (L.) amazonensis (LD(50) values of 18.5±6.5µg/ml and 12.63±5.52µg/ml, respectively). Our results are showing that alkaloidal extract from leaves is a promising alternative to the use of stem barks from A. ramiflorum.

Constituintes químicos e avaliação das atividades antibacteriana e antiedematogênica de Aloysia gratissima (Gillies & Hook.) Tronc. e Aloysia virgata (Ruiz & Pav.) Pers., Verbenaceae / Chemical composition and evaluation of antibacterial and antiedematogenic Activities of Aloysia gratissima (Gillies & Hook.) Tronc. e Aloysia virgata (Ruiz & Pav.) Pers., Verbenaceae

O estudo químico de Aloysia gratissima (Gillies & Hook.) Tronc. e Aloysia virgata (Ruiz & Pav.) Pers., Verbenaceae, resultou no isolamento de quatorze substâncias. Suas estruturas foram determinadas com base nos dados de IV, RMN de ¹H e de 13C uni e bidimensionais e comparação com dados da literatura. A atividade antibacteriana do extrato bruto das folhas e caules e das frações de A. virgata, bem como as substâncias hoffmaniacetona e seu monoacetato, verbascosídeo e arenariosídeo, foram avaliadas através do método de bioautografia e a atividade antiedematogênica foi avaliada pelo modelo de edema de orelha induzido para os extratos brutos e frações metanólicas de A. gratissima e A. virgata..

Phytochemical study of Aloysia gratissima (Gillies & Hook.) Tronc. e Aloysia virgata (Ruiz & Pav.) Pers., Verbenaceae, afforded fourteen compounds. The structures were established using IR, 1D and 2D NMR and by comparison of its spectroscopic data to those of literature. The antibacterial activity of the crude extract from the leaves and branches and the fractions from the crude extract of A. virgata, besides the isolated compounds hoffmaniacetone, hoffmaniacetone monoacetate, verbascoside and arenarioside were evaluated by means of bioautography and the antiedematogenic activity was evaluated using induced ear oedema model to the crude extracts and methanol fractions for A. gratissima and A. virgata.

Antimicrobial activity of Hymenaea martiana towards dermatophytes and Cryptococcus neoformans.

The biological activity of crude extract and fractions of Hymenaea martiana was evaluated against a panel of human pathogenic fungi. The crude extracts and hydroalcoholic fractions (E) showed a high activity against Cryptococcus neoformans species complex isolates with MICs between 2 and 64 µg ml(-1). The methanolic (C) and butanolic (D) fractions were the most active against Trichopyton rubrum, Trichopyton mentagrophytes and Microsporum canis with MICs between 8 and 256 µg ml(-1). None of the extracts was active against the yeast Malassezia furfur, Malassezia obtusa and Malassezia sympodialis.