Acetylcholinesterase inhibitory activity of Ocotea pomaderroides extracts: HPLC-MS/MS characterization and molecular modeling studies.

The present work describes the acetylcholinesterase inhibitory activity of Ocotea pomaderroides extracts besides the chemical composition of chromatographic fractions. The hexane, dichloromethane and ethyl acetate extract soluble fractions showed high Electrophorus electricus acetylcholinesterase (EelAChE) inhibition (92.18, 71.86 and 74.25%, respectively) while the butanolic and aqueous extracts showed moderate to low activities (44.48 and 20.74%, respectively). The high-performance liquid chromatography coupled with electrospray ionization multiple-stage mass spectrometry (HPLC-ESI-MSn) analysis led to the identification of the alkaloids and flavonol glycoside derivatives present in these extracts. The binding profile of the alkaloids and their atomic effect on 3D structure of Electrophorus electricus AchE (EelAChE) were assessed with molecular modeling.

Hydroxycinnamic acid-spermidine amides from Tetragonisca angustula honey as anti-Neospora caninum: In vitro and in silico studies.

Tetragonisca angustula honey was fractioned in a SiO2 column to furnish three fractions (A-C) in which four hydroxycinnamic acid-Spermidine amides (HCAAs), known as N', N″, N‴-tris-p-coumaroyl spermidine, N', N″-dicaffeoyl, N‴-coumaroyl spermidine, N', N″, N‴-tris-caffeoyl spermidine and N', N″-dicaffeoyl and N‴-feruloyl spermidine were identified in the fractions B and C by electrospray ionization tandem mass spectrometry. A primary culture model previously infected with Neospora caninum (72 h) was used to evaluate the honey fractions (A-C) for two-time intervals: 24 and 72 h. Parasitic reduction ranged from 38% on fraction C (12.5 µg/ml), after 24 h, to 54% and 41% with fractions B and C (25 µg/ml) after 72 h of treatment, respectively. Additionally, HCAAs did not show any cell toxicity for 24 and 72 h. For infected cultures (72 h), the active fractions B (12.5 µg/ml) and C (25 µg/ml) decreased their NO content. In silico studies suggest that HCAAs may affect the parasite's redox pathway and improve the oxidative effect of NO released from infected cells. Here, we presented for the first time, that HCAAs from T. angustula honey have the potential to inhibit the growth of N. caninum protozoa.

γ-Lactones from Persea americana and Persea fulva - in Vitro and in Silico Evaluation of Trypanosoma cruzi Activity.

In the present study, five known γ-lactones (majoranolide B - 1, majorenolide - 2, majorynolide - 3, lincomolide D - 4, and isolinderanolide E - 5), as well as a new one (perseanolide - 6), were isolated from Persea fulva and P. americana. All isolated compounds exhibited potential activity against trypomastigote forms of Trypanosoma cruzi, whereas compounds 2 (EC50 of 4.8 µM) and 6 (EC50 of 3.6 µM) displayed superior activity than the positive control benznidazole (EC50 of 16.4 µM), with selectivity index (SI) values of 17.8 and >55.6, respectively (benznidazole, SI>12.2). Molecular docking studies were performed for 1-6 against six T. cruzi molecular targets. Using this approach, we observed that, even though perseanolide (6) showed favorable docking to several studied targets, the results were especially promising for hypoxanthine phosphoribosyl transferase (PDB 1TC1). As PDB 1TC1 is associated to the transference of a monophosphorylated ribose from phosphoribosylpyrophosphate (PRPP) in the ribonucleotide synthesis pathway, this interaction may affect the survival of T. cruzi in mammalian cells. The data herein also indicate that possible intermolecular interactions between 6 and PDB 1TC1 derive from (i) hydrogen bonds in the α,ß-unsaturated-γ-lactone unity and (ii) hydrophobic interactions in the long-chain alkyl group. Based on our results, perseanolide (6), reported for the first time in this work, can auspiciously contribute to future works regarding new trypanocidal agents.

Chemical profiling and separation of bioactive secondary metabolites in Maca (Lepidium peruvianum) by normal and reverse phase thin layer chromatography coupled to desorption electrospray ionization-mass spectrometry.

Maca is a Peruvian tuberous root of the Brassicaceae family grown in the central Andes between altitudes of 4000 and 4500 m. The medicinal plant is a nutraceutical with important biological activities and health effects. In this study, we report a rapid high-performance thin layer chromatography (HPTLC)-(-)desorption electrospray ionization (DESI)-mass spectrometry (MS) method to profile and separate intact glucosinolates without prior biochemical modifications from the hydromethanolic extracts of two phenotypes, red and black Maca (Lepidium peruvianum) seeds. In the first stage of the plant's life cycle, aromatic glucosinolates were the main chemical constituents whereby six aromatic, three indole, and one aliphatic glucosinolate were tentatively identified. At the seedling stage, glucolepigramin/Glucosinalbin was the most predominant precursor, rather than Glucotropaeolin, which is mainly found in hypocotyls and roots. These findings lead us to suggest that glucolepigramin/glucosinalbin play a major role as active precursors in the biosynthetic pathways of other secondary metabolites in the early stages of plant development. Between red and black Maca seeds, only minor differences in the relative abundances of glucosinolates were observed rather than different plant metabolites. For the first time, we report six potential plant antibiotics, phytoanticipins: glycosylated ascorbigens and dihydroascorbigens from Maca seeds. We also investigated a targeted reverse phase C18 functionalized TLC-DESI-MS method with high sensitivity and specificity for Brassicaceae fatty acids in Maca seeds and health supplements such as black Maca root lyophilized powder and tinctures. The investigation of secondary metabolites by normal and reverse phase TLC-DESI-MS methods, described in this study, can aid in their identification as they begin to emerge in later stages of development in plant tissues such as leaves, hypocotyls, and roots.

Identification of Sassafras albidum alkaloids by high-performance thin-layer chromatography tandem mass spectrometry and mapping by desorption electrospray ionization mass spectrometry imaging.

Sassafras albidum is an important tree species that occurs across North America. The presence of benzylisoquinoline and aporphine alkaloids has been previously described; however, the spatial distribution of these compounds within S. albidum and other plants of Lauraceae family is still unclear. Mass spectrometry imaging has become an important tool in analysis of plants metabolites, uncovering important contributions about the functional role, biosynthetic pathway, and accumulation of these compounds in the plant. This work aimed to identify further alkaloids present in S. albidum roots, twigs, and leaves by high-performance thin-layer chromatography coupled to desorption electrospray ionization multistage mass spectrometry (HPTLC DESI-MSn ) and to map the spatial distribution of these compounds by DESI-MS imaging. A total of 12 alkaloids were indentified in the roots and twigs, and six of them were detected for the first time in S. albidum. A high number of alkaloids was found in S. albidum roots; however, alkaloids were not detected in the leaves. Cross sections of roots and twigs were blotted onto TLC plates assisted by heating and solvent extraction, and these imprints were analyzed by DESI-MS imaging. The profile of alkaloid spatial distribution in DESI-MS images showed different accumulation patterns across and within different plant parts. Most alkaloids displayed higher intensities in the outer-most layer of the roots and twigs. The detailed spatial localization pattern of these alkaloids analyzed by DESI-MS imaging in different plant parts could contribute to a better understanding of the profile of distribution, accumulation, and biosynthesis of benzylisoquinoline and aporphine alkaloids.

Rapid structural characterisation of benzylisoquinoline and aporphine alkaloids from Ocotea spixiana acaricide extract by HPTLC-DESI-MSn.

INTRODUCTION: Lauraceae alkaloids are a structurally diverse class of plant specialised secondary metabolites that play an important role in modern pharmacotherapy, being useful as well as model compounds for the development of synthetic analogues. However, alkaloids characterisation is challenging due to low concentrations, the complexity of plant extracts, and long processes for accurate structural determinations. OBJECTIVE: The use of high-performance thin layer chromatography coupled with desorption electrospray ionisation multistage mass spectrometry (HPTLC DESI-MSn ) as a fast tool to identify alkaloids present in Ocotea spixiana extract and evaluate the extract's acaricide activity. METHODS: Ocotea spixiana twigs were extracted by conventional liquid-liquid partitioning. HPTLC analysis of the ethyl acetate extract was performed to separate isobaric alkaloids prior to DESI-MSn analysis, performed from MS3 up to MS7 . The extract's acaricide activity against Rhipicephalus microplus was evaluated by in vitro (larval immersion test) and in silico tests. RESULTS: HPTLC-DESI-MSn analysis was performed to identify a total of 13 aporphine and four benzylisoquinoline-type alkaloids reported for the first time in O. spixiana. In vitro evaluation of the extract and the alkaloid boldine showed significant activity against R. microplus larvae. It was established in silico that boldine had important intermolecular interactions with R. microplus acetylcholinesterase enzyme. CONCLUSION: The present study demonstrated that HPTLC-DESI-MSn is a useful analytical tool to identify isoquinoline alkaloids in plant extracts. The acaricide activity of the O. spixiana ethyl acetate extract can be correlated to the presence of alkaloids.