ABSTRACT
Two new heptapeptides, [1-7-NαC]-crocaorbs A1 (1) and A2 (2), were isolated from the latex of Croton campanulatus. Their structures were determined using NMR spectroscopic techniques, ESI-HRMS data, Marfey's method, and further refined using molecular dynamics with simulated annealing (MD/SA). Molecular dynamics calculations of peptides 1 and 2 demonstrated greater stability in simulations using a biological solvent compared to those using DMSO. Compound 1, the most abundant peptide in latex, was assessed for NO production, antiplasmodial and cytotoxicity activities. The peptide significantly increased nitric oxide (NO) production at concentrations of 40, 20 or 10 µM (17.932 ± 1.1, 18.270 ± 0.9, 18.499 ± 0.7, respectively). Its antiplasmodial activity exhibited limited efficacy, with only 5% inhibition of Plasmodium falciparum 3D7 growth at a concentration of 50 µM. Also, it exhibited no cytotoxic effects in the J774A.1 murine macrophages cell line. This study represents the first report of a phytochemical investigation of the species C. campanulatus, which showed orbitides with distinctive sequences in contrast to other peptides described for the genus Croton and contributes to the study of structural diversity within this particular class of compounds.
ABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Croton floribundus Spreng. (Euphorbiaceae), popularly known as Capixinguí, stands out due to its widespread use in traditional medicine to treat wounds, syphilis, hemorrhoids, eye diseases and as a purgative. AIM OF THE STUDY: To characterize clerodanes diterpenes from C. floribundus and to evaluate the effects of the fraction and diterpenes (1-5) on inhibition of nitrite production. MATERIALS AND METHODS: The hydroethanolic root extract of C. floribundus was fractionated on a solid phase extraction column to obtain the fraction named Fr80%. From this, five compounds were obtained and characterized. The absolute configuration of compound 1 was determined by a combination of electronic and vibrational circular dichroism spectroscopies. Additionally, compounds 1-5 were evaluated for their inhibitory effects on nitrite production induced by lipopolysaccharide (LPS) in RAW 264 macrophage cell. RESULTS: Five clerodane diterpenoids were characterized, and the absolute stereochemistry of 1 was established as 3R,4R,5R,8R,9R,10S,12S. The IC50 values obtained through inhibition of nitrite production were 28.52⯱â¯2.21⯵M (1), 40.26⯱â¯2.79⯵M (2), 25.47⯱â¯2.16⯵M (3), 35.78⯱â¯2.93⯵M (4) and 40.58⯱â¯4.78⯵M (5). In the tested concentrations, the samples presented low toxicity in macrophages. CONCLUSIONS: Four new diterpenes were characterized from C. floribundus, these being croflorins A-D (1-4) and a known halimane (5). These compounds exhibited inhibitory effect on nitrite production.
Subject(s)
Croton/chemistry , Diterpenes/chemistry , Diterpenes/pharmacology , Nitrites/metabolism , Animals , Cell Line , Circular Dichroism/methods , Diterpenes, Clerodane/chemistry , Diterpenes, Clerodane/pharmacology , Euphorbiaceae/chemistry , Macrophages/drug effects , Macrophages/metabolism , Medicine, Traditional/methods , Mice , RAW 264.7 CellsABSTRACT
Two new bracelet cyclotides from roots of Pombalia calceolaria with potential anticancer activity have been characterized in this work. The cyclotides Poca A and B (1 and 2) and the previously known CyO4 (3) were de novo sequenced by MALDI-TOF/TOF mass spectrometry (MS). The MS2 spectra were examined and the amino acid sequences were determined. The purified peptides were tested for their cytotoxicity and effects on cell migration of MDA-MB-231, a triple-negative breast cancer cell line. The isolated cyclotides reduced the number of cancer cells by more than 80% at 20 µM, and the concentration-related cytotoxic responses were observed with IC50 values of 1.8, 2.7, and 9.8 µM for Poca A (1), Poca B (2), and CyO4 (3), respectively. Additionally, the inhibition of cell migration (wound-healing assay) exhibited that CyO4 (3) presents an interesting activity profile, in being able to inhibit cell migration (50%) at a subtoxic concentration (2 µM). The distribution of these cyclotides in the roots was analyzed by MALDI imaging, demonstrating that all three compounds are present in the phloem and cortical parenchyma regions.