Fábio Lazzarotto de Oliveira

Sesquiterpene lactones from Vernonia scorpioides and their in vitro cytotoxicity

Fresh leaves of Vernonia scorpioides are widely used in Brazil to treat a variety of skin disorders. Previous in vivo studies with extracts of this species had also demonstrated a high antitumor potential. This paper reports isolation of four sesquiterpene lactones (hirsutinolides and glaucolides), together with diacetylpiptocarphol, 8-acetyl-13-etoxypiptocarphol, luteolin, apigenin, and ethyl caffeate from fresh leaves and flowers of Vernonia scorpioides. The hypothesis that hirsutinolide 3 is formed during extraction was verified theoretically using Density Functional Theory. The effects of isolated compounds on in vitro tumor cells were investigated, as well as their genotoxicity by means of an in vitro comet assay. The results indicate that glaucolide 2 and hirsutinolide 4 are toxic to HeLa cells. These compounds were genotoxic in vitro, a property that appears to be related to the presence of their epoxy groups, which has been a more reliable indication of toxicity than substitution on C-13 or the presence of alpha,beta-unsaturated keto-groups. These results need to be replicated in vivo in order to ascertain their toxicity.

Sesquiterpene lactones from the leaves of Hedyosmum brasiliense (Chloranthaceae).

Hedyosmum brasiliense Miq. is an endemic aromatic arborescent shrub that is the only representative of the Chloranthaceae in Brazil. There have been few studies seeking to determine its chemical constituents and/or pharmacological effects. This work describes the isolation and identification of sesquiterpene lactones from the leaves, including guaianolides, elemanolides and a lindenanolide. These were tested against Mycobacterium tuberculosis, together with podoandin, onoseriolide and some other common phenolics. The structures of the isolated compounds were determined based on extensive analysis of 1D and 2D NMR spectroscopic and MS data, as well as comparison with published data. The compounds found were the guaianolides, 1,2-epoxy-10α-hydroxy-podoandin and 1-hydroxy-10,15-methylenepodoandin, the elemenolide 15-acetoxy-isogermafurenolide and the lindenanolide 8α/β,9α-hydroxy-onoseriolide, along with the previously isolated guaianolide podoandin, the lindenanolide onoseriolide and the elemenolide 15-hydroxy-isogermafurenolide. The phenolic compounds isolated were scopoletin, vanillin, vanillic acid, protocatechuic aldehyde and ethyl caffeate. The isolated sesquiterpene lactones did not show anti-mycobacterial activity against isoniazid-sensitive M. tuberculosis cultures at concentrations of 1-30 μM.

Crystal structures of the apo form and a complex of human LMW-PTP with a phosphonic acid provide new evidence of a secondary site potentially related to the anchorage of natural substrates

Low molecular weight protein tyrosine phosphatases (LMW-PTP, EC 3.1.3.48) are a family of single-domain enzymes with molecular weight up to 18 kDa, expressed in different tissues and considered attractive pharmacological targets for cancer chemotherapy. Despite this, few LMW-PTP inhibitors have been described to date, and the structural information on LMW-PTP druggable binding sites is scarce. In this study, a small series of phosphonic acids were designed based on a new crystallographic structure of LMW-PTP complexed with benzylsulfonic acid, determined at 2.1Å. In silico docking was used as a tool to interpret the structural and enzyme kinetics data, as well as to design new analogs. From the synthesized series, two compounds were found to act as competitive inhibitors, with inhibition constants of 0.124 and 0.047 mM. We also report the 2.4Å structure of another complex in which LMW-PTP is bound to benzylphosphonic acid, and a structure of apo LMW-PTP determined at 2.3Å resolution. Although no appreciable conformation changes were observed, in the latter structures, amino acid residues from an expression tag were found bound to a hydrophobic region at the protein surface. This regions is neighbored by positively charged residues, adjacent to the active site pocket, suggesting that this region might be not a mere artefact of crystal contacts but an indication of a possible anchoring region for the natural substrate-which is a phosphorylated protein.

(1S,2R,6R,7aS)-1,2,6-Trihy­droxy­hexa­hydro-1H-pyrrolizin-3-one

In the title compound, C7H11NO4, prepared via a Morita–Baylis–Hillman adduct, the five-membered ring bearing three O atoms approximates to a twisted conformation, whereas the other ring is close to an envelope, with a C atom in the flap position. The dihedral angle between their mean planes (all atoms) is 23.11 (9)°. The new stereocenters are created in a trans-diaxial configuration. In the crystal, O—H⋯O and O—H⋯(O,O) hydrogen bonds link the molecules, generating a three-dimensional network. A weak C—H⋯O interaction also occurs.

(1S,2S,6R,7aR)-2-Benzyl-1,6-dihy­droxy­hexa­hydro­pyrrolizin-3-one

In the title compound, C14H17NO3, the dihedral angles show that the H atoms at two stereocenters are in a trans-diaxial configuration. In the crystal, the molecules are linked by O—H⋯O hydrogen bonds. The absolute configuration of the molecule has been established on the basis of refinement of the Hooft and Flack parameters.

(1S,2E,6R,7aR)-1,6-Dihy-droxy-2-(4-nitro-benzyl-idene)-2,3,5,6,7,7a-hexa-hydro-1H-pyrrolizin-3-one.

The crystal structure of the title compound, C14H14N2O5, contains two distinct conformers in the asymmetric unit. The compound has three defined stereocenters, two of them contiguous, and a C=C double bond with an E conformation. The stereocenters exhibit the same chirality in both conformers, with significant differences in the conformation of the five-membered rings of the pyrrolizine unit (both either in a twist or in an envelope form) and in the dihedral angles between the corresponding mean planes and the benzene rings. A prominent feature is a change from almost coplanar rings in one conformer to a new conformation in the second conformer, in which the mean plane of a five-membered ring is almost perpendicular to the benzene ring, with a dihedral angle 87.19 (8)°; the corresponding angle in the first conformer is 14.02 (10)°. In the crystal, molecules are linked by O—H⋯O and C—H⋯O hydrogen bonds. Crystallographic data were essential to confirm the configuration of the double bond, which was unclear from the available two-dimensional NMR data. In addition, reliable Flack and Hooft parameters were obtained, allowing for the correct absolute structure to be determined.

(1S,2E,6R,7aR)-2-Benzyl-idene-1,6-dihy-droxy-2,3,5,6,7,7a-hexa-hydro-1H-pyrrolizin-3-one.

In the title compound, C14H15NO3, the conformation of the double bond was determined to be E, confirming the result obtained from two-dimensional NMR data. The five-membered rings of the pyrrolizine unit exhibit C-envelope conformations, with C atoms displaced from the mean planes formed by the remaining rings atoms by 0.1468 (15) and 0.5405 (17) Å. The mean planes of these rings (through all ring atoms) have a dihedral angle of 49.03 (10)°. In the crystal, molecules are linked by O—H⋯O hydrogen bonds. The absolute configuration of the molecule was established, as judged by the, as judged by the obtained values for the Hooft and Flack parameters.