The essential oils of Thymus caespititius, T. camphoratus, T. capitellatus, T. carnosus, T. pulegioides, T. zygis subsp. zygis and T. zygis subsp. sylvestris collected in diff erent regions of Portugal, including the Azores islands, were evaluated for preventing lipid peroxidation and scavenging free radicals, and hydroxyl and superoxide anions. T. zygis subsp. zygis oil possessed the best capacity for preventing lipid peroxidation (IC 50 = 0.030 ± 0.003 mg/ml), immediately followed by the oils isolated from T. zygis subsp. sylvestris collected in diff erent regions of Portugal. IC 50 values ranged from 0.066 ± 0.003 to 0.087 ± 0.001 mg/ml in oils isolated from samples from Alcanena and Duas Igrejas, respectively. The oils isolated from T. zygis subsp. sylvestris, independent of the harvesting region, were shown to be the most eff ective for scavenging free radicals (0.358-0.453 mg/ml). The best capacity for scavenging hydroxyl radicals was found in the oils of T. camphoratus collected in Espartal and of T. caespititius of Lordelo, with IC 50 < 0.5 mg/ml. The low oil amount did not allow IC 50 values for the superoxide anion scavenging determination, therefore considering a unique oil concentration (5 mg/ml), the highest inhibition percentages (>50%) were registered with the essential oils of T. zygis subsp. sylvestris from Duas Igrejas and Covão do Coelho, and of T. capitellatus from Sines.

The identification, isolation and functional characterization of two genes encoding two monoterpene synthases-γ-terpinene synthase (Tctps2) and α-terpineol synthase (Tctps5)-from three chemically distinct Thymus caespititius (Lamiaceae) genotypes were performed. Genomic exon-intron structure was also determined for both terpene synthase genes, revealing an organization with seven exons and six introns. The cDNA of Tctps2 was 2,308 bp long and had an open reading frame of 1,794 bp encoding for a protein with 598 amino acids. Tctps5 was longer, mainly due to intron sequences, and presented high intraspecific variability on the plants analyzed. It encoded for a protein of 602 amino acids from an open reading frame of 1,806 bp comprising a total of 2,507 bp genomic sequence. The amino acid sequence of these two active Tctps genes shared 74 % pairwise identity, ranging between 42 and 94 % similarity with about 50 known terpene synthases of other Lamiaceae species. Gene expression revealed a multi-product Tctps2 and Tctps5 enzymes, producing γ-terpinene and α-terpineol as major components, respectively. These enzymatic results were consistent with the monoterpene profile present in T. caespititius field plants, suggesting a transcriptional regulation in leaves. Herewith reported for the first time for this species, these two newly characterized Tctps genes improve the understanding of the molecular mechanisms of reaction responsible for terpene biosynthesis and chemical diversity found in T. caespititius.

Pinewood nematode (PWN), Bursaphelenchus xylophilus, is the causal agent of pine wilt disease, a serious threat to global forest populations of conifers, especially Pinus spp. A time-course study of the essential oils (EOs) of 2-year-old Pinus halepensis, Pinus pinaster, Pinus pinea and Pinus sylvestris following inoculation with the PWN was performed. The constitutive and nematode inoculation induced EOs components were analyzed at both the wounding or inoculation areas and at the whole plant level. The enantiomeric ratio of optically active main EOs components was also evaluated. External symptoms of infection were observed only in P. pinaster and P. sylvestris 21 and 15 days after inoculation, respectively. The EO composition analysis of uninoculated and unwounded plants revealed the occurrence of chemotypes for P. pinaster, P. halepensis and P. sylvestris, whereas P. pinea showed a homogenous EO composition. When whole plants were evaluated for EO and monoterpene hydrocarbon enantiomeric chemical composition, no relevant qualitative and quantitative differences were found. Instead, EO analysis of inoculated and uninoculated wounded areas revealed an increase of sesquiterpenes and diterpenic compounds, especially in P. pinea and P. halepensis, comparatively to healthy whole plants EOs.

The antioxidant and antiproliferative activities of the essential oils from Thymbra capitata and Thymus species grown in Portugal were evaluated. Thymbra and Thymus essential oils were grouped into two clusters: Cluster I in which carvacrol, thymol, p-cymene, α-terpineol, and γ-terpinene dominated and Cluster II in which thymol and carvacrol were absent and the main constituent was linalool. The ability for scavenging ABTS•+ and peroxyl free radicals as well as for preventing the growth of THP-1 leukemia cells was better in essential oils with the highest contents of thymol and carvacrol. These results show the importance of these two terpene-phenolic compounds as antioxidants and cytotoxic agents against THP-1 cells.

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Aerial parts of Pittosporum undulatum were collected fortnightly for 1 year at the Parque de Saúde de Lisboa. Collective samples of two populations were investigated, i.e. from pruned and non-pruned trees, and in addition samples from one pruned and one non-pruned individual. An SEM study of the aerial parts showed the presence of nonglandular T-shaped trichomes, and capitate trichomes with an elongated stalk and two to four round-shaped cells on the top. Numerous canals were seen at the level of the capsule wall as well as in the leaves and stems. Essential oil samples were isolated from the (deep-frozen) aerial parts by hydrodistillation to estimate the oil yields, and by distillation-extraction to determine their percentage composition, and analysed by GC and GC-MS. Monoterpenes (59-97%) were dominant in all oils, sabinene (5-58%), limonene (2-37%) and terpinen-4-ol (4 -27%) being the major components, although in different relative amounts during the year of collection. Benzyl benzoate (8%) and benzyl salicylate (10%) attained high relative amounts in the oils from fully developed flowers, whereas in the oils from the other plant parts their relative amounts did not exceed 1%. The enantiomeric ratio of sabinene, limonene and terpinen-4-ol was investigated in the leaf, flower and capsule oils, showing dominance of the (+ + + + +)-forms and homogeneity between all the oil samples.

The essential oils isolated from the vegetative aerial parts (mature branches with leaves) of 16 Eucalyptus species, grown in Mata Experimental do Escaroupim (Salvaterra de Magos, Portugal) were studied. The essential oils from E.