The green leaves were dried in a stove with hot air circulation a

The green leaves were dried in a stove with hot air circulation and thermostatized

at 40 °C during 4 days. After grinding them, the resulting material was subjected to extraction with ethyl acetate and ethanol (3:1), using 9 L of this mixture for each extraction. This process was repeated four times with an interval of 7 days between extractions. The total weight of the extract obtained corresponded to 5.5% of fresh plant mass ( Cotinguiba et al., 2009). The essential oils from L. sidoides CH5424802 research buy and M. piperita were obtained at the Embrapa Western Amazon Research Station from plants cultivated in Manaus, Amazonas state, Brazil. The leaves of L. sidoides and M. piperita were cut at ground level and placed in a freezer until extraction. After separation of the leaves, two samples of 20 g were used to determine moisture

by drying an oven at 65 °C for 3 days. Two other samples of 100 g each were used to extract the essential oil by hydrodistillation in a Clevenger type apparatus for 3 h. H. crepitans latex was collected in the trees located in the city of Porto Velho, Rondônia state by employees of Embrapa Rondônia. The seed oil of C. guianensis was produced and acquired in the local market of Porto Velho. The active substances from P. tuberculatum used in the present trial were previously described by Cotinguiba et al. (2009). Chemical analyses of the L. sidoides and M. piperita essential oils, C. guianensis oil and H. crepitans latex were performed by the Embrapa Food Agribusiness Research Unit (CTAA). The identification of the essential oil components BMN 673 mouse was carried out by gas chromatography coupled to mass spectrometry (GC-MS) in an Agilent 5973N system (Agilent Technologies, Delaware, USA) equipped with an HP-5MS capillary column (5% diphenyl, 95% dimethylsilicone, 30 m × 0.25 mm; film thickness 0.25 μm). Helium was used as the carrier gas (1.0 mL min−1), with injection of 1.0 mL of a 1% solution of the essential

oil in dichloromethane in an injector heated to 250 °C, operating in split mode (split ratio 1:100). either Oven temperature was varied from 60 to 240 °C at a rate of 3 °C min−1. The mass detector was operated in electron ionization (70 eV) with the mass analyzer maintained at 150 °C, the ionization source at 220 °C and transfer line at 260 °C. To obtain the quantification, the essential oils were also analyzed in an Agilent 7890A chromatograph (Agilent Technologies, Delaware, USA) equipped with a flame ionization detector (FID) kept at 280 °C and fitted with an HP-5 capillary column (5% diphenyl–95%-dimethyl silicone; 30 m × 0.32 mm; film thickness 0.25 μm). The same injection and chromatographic conditions above were applied, but hydrogen was used as the carrier gas, at 1.5 mL min−1. The results were indicated through relative area (% area). Linear retention indices were calculated by injection of a series of n-alkanes (C7–C26) in the same column and conditions stated for GC-FID analyses.

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