Gouregine, an α-Gem-Dimethyltetradehydrocularine Alkaloid, and Other Aporphinoid Alkaloids from the Bark of Guatteria olivacea (Annonaceae) and Their In Vitro Cytotoxic Activities.

Guatteria olivacea R.E. Fries is an Amazonian species known as 'envira-bobó' and 'envira-fofa' and is common in the states of Amazonas, Acre, and Pará. Recently, the essential oil from the leaves of this species has shown promising antitumor activity both in vitro and in vivo. The presence of isoquinoline-derived alkaloids, including aporphinoids and tetrahydroprotoberberine alkaloids, has also been previously reported. In our ongoing search for bioactive compounds from Annonaceae Amazonian plants, the bark of G. olivacea was investigated via classical chromatography techniques, which revealed nine compounds, eight isoquinoline-derived alkaloids, a rare alkaloid with a α-gem-dimethyltetradehydrocularine structure known as gouregine, seven known aporphinoid alkaloids: isopiline, O-methylisopiline, melosmine, 9-hydroxyiguattescine, dihydromelosmine, lysicamine, and guattouregidine, and one known pimaradiene diterpene: acanthoic acid. All the isolated compounds were described for the first time in the bark of G. olivacea, and their structures were elucidated by extensive analyses of their 1D and 2D NMR spectra in combination with MS data. The NMR data of the alkaloids isopiline, O-methylisopiline, melosmine, dihydromelosmine, and guattouregidine were revised due to incomplete data in the literature and some ambiguities. The in vitro cytotoxic activities of the isolated compounds were evaluated against human cancer (HepG2, KG-1a, and HCT116) and noncancerous (MRC-5) cell lines via the Alamar blue assay after 72 h of incubation. Among the compounds evaluated against human cancer cell lines, the most active was the oxoaporphine alkaloid lysicamine, which has strong activity against HCT116 cells, with an IC50 value of 6.64 µg/mL (22.79 µmol/L). Melosmine had a moderate effect on HCT116 cells, with an IC50 value of 16.77 µg/mL (49.70 µmol/L), whereas acanthoic acid had moderate effects on HepG2 and HCT116 cells, with IC50 values of 14.63 µg/mL (48.37 µmol/L) and 21.25 µg/mL (70.25 µmol/L), respectively.

Antitumor Effect of Guatteria olivacea R. E. Fr. (Annonaceae) Leaf Essential Oil in Liver Cancer.

Guatteria olivacea R. E. Fries (synonym Guatteria punctata (Aubl.) R.A. Howard) is a tree of 10-27 m tall popularly known as "envira-bobó", "envira-fofa", "envireira", "embira", "embira-branca", "embira-preta", envira-branca", and "envira-preta", which can be found in the Brazilian Amazon biome. In this study, we evaluated the cytotoxic and antitumor effects of the essential oil (EO) obtained from the leaves of G. olivacea against liver cancer using HepG2 cells as a model. EO was obtained using a hydrodistillation Clevenger-type apparatus and was qualitatively and quantitatively characterized using GC-MS and GC-FID, respectively. The alamar blue assay was used to assess the cytotoxic potential of EO in a panel of human cancer cell lines and human non-cancerous cells. In HepG2 cells treated with EO, YO-PRO-1/propidium iodide staining, cell cycle distribution, and reactive oxygen species (ROS) were examined. In C.B-17 SCID mice with HepG2 cell xenografts, the efficacy of the EO (20 and 40 mg/kg) was tested in vivo. GC-MS and GC-FID analyses showed germacrene D (17.65%), 1-epi-cubenol (13.21%), caryophyllene oxide (12.03%), spathulenol (11.26%), (E)-caryophyllene (7.26%), bicyclogermacrene (5.87%), and δ-elemene (4.95%) as the major constituents of G. olivacea leaf EO. In vitro cytotoxicity of EO was observed, including anti-liver cancer action with an IC50 value of 30.82 µg/mL for HepG2 cells. In HepG2 cells, EO treatment increased apoptotic cells and DNA fragmentation, without changes in ROS levels. Furthermore, the EO inhibited tumor mass in vivo by 32.8-57.9%. These findings suggest that G. olivacea leaf EO has anti-liver cancer potential.

In vitro and in vivo growth inhibition of human acute promyelocytic leukemia HL-60 cells by Guatteria megalophylla Diels (Annonaceae) leaf essential oil.

Guatteria megalophylla Diels (Annonaceae) is an 8-10 m tall tree that grows near streams and is widely spread throughout Colombian, Ecuadorian, Peruvian, Brazilian and Guianese Amazon rainforest. Herein, we investigated for the first time the chemical composition and in vitro and in vivo anti-leukemia potential of G. megalophylla leaf essential oil (EO) using human promyelocytic leukemia HL-60 cells as model. EO was obtained by a hydrodistillation clevenger-type apparatus and characterized quali- and quantitatively by GC-MS and GC-FID, respectively. In vitro cytotoxic potential of EO was evaluated in human cancer cell lines (HL-60, MCF-7 CAL27, HSC-3, HepG2 and HCT116) and in human non-cancer cell line (MRC-5) by Alamar blue method. Annexin V/propidium iodide staining, cell cycle distribution and reactive oxygen species (ROS) were assessed by flow cytometry for HL-60 cells treated with EO. In vivo efficacy of EO (50 and 100 mg/kg) was evaluated in C.B-17 SCID mice with HL-60 cell xenografts. Chemical composition analyses showed spathulenol, γ-muurolene, bicyclogermacrene, ß-elemene and δ-elemene as main constituents of assayed sample. EO displayed in vitro cytotoxicity, including anti-leukemia effect with IC50 value of 12.51 µg/mL for HL-60 cells. EO treatment caused augment of phosphatidylserine externalization and DNA fragmentation without increasing of ROS in HL-60 cells. In vivo tumor mass inhibition rates of EO was 16.6-48.8 %. These data indicate anti-leukemia potential of G. megalophylla leaf EO.