Genotyping Analysis by RAD-Seq Reads Is Useful to Assess the Genetic Identity and Relationships of Breeding Lines in Lavender Species Aimed at Managing Plant Variety Protection.

Lavender species are widely distributed in their wild forms around the Mediterranean Basin and they are also cultivated worldwide as improved and registered clonal varieties. The economic interest of the species belonging to the Lavandula genus is determined by their use as ornamental plants and important source of essential oils that are destinated to the production of cosmetics, pharmaceuticals and foodstuffs. Because of the increasing number of cases of illegal commercialization of selected varieties, the protection of plant breeders' rights has become of main relevance for the recognition of breeding companies' royalties. With this aim, genomic tools based on molecular markers have been demonstrated to be very reliable and transferable among laboratories, and also much more informative than morphological descriptors. With the rising of the next-generation sequencing (NGS) technologies, several genotyping-by-sequencing approaches are now available. This study deals with a deep characterization of 15 varietal clones, belonging to two distinct Lavandula species, by means of restriction-site associated DNA sequencing (RAD-Seq). We demonstrated that this technology screens single nucleotide variants that enable to assess the genetic identity of individual accessions, to reconstruct genetic relationships among related breeding lines, to group them into genetically distinguishable main subclusters, and to assign their molecular lineages to distinct ancestors. Moreover, a number of polymorphic sites were identified within genes putatively involved in biosynthetic pathways related to both tissue pigmentation and terpene production, useful for breeding and/or protecting newly registered varieties. Overall, the results highlighted the presence of pure ancestries and interspecific hybrids for the analyzed Lavandula species, and demonstrated that RAD-Seq analysis is very informative and highly reliable for characterizing Lavandula clones and managing plant variety protection.

Phenolic Composition and Bioactivity of Lavandula pedunculata (Mill.) Cav. Samples from Different Geographical Origin.

The aim of this study was to characterize the phenolic composition and evaluate the bioactivity of several samples of Lavandula pedunculata (Mill.) Cav, and to compare aqueous and hydroethanolic extracts. Plant materials were obtained by growing some accessions (seed samples) of various wild populations from different regions of Portugal conserved at the Portuguese Genebank in Braga. Phenolic compounds were analised by HPLC-DAD-ESI/MSn, antioxidant potential through in vitro assays (DPPH radical scavenging activity, reducing power and inhibition of lipid peroxidation), cytotoxicity on tumor cells (MCF-7, NCI-H460, HeLa and HepG2) and non-tumor (PLP2) cells, anti-inflammatory activity in rat RAW 264.7 macrophages, by the ability to inhibit NO production and antimicrobial potential by the microdilution method with INT dye (iodonitrotetrazolium chloride). Thirteen compounds were identified, being salvianolic acid B, rosmarinic acid and luteolin-7-O-glucuronide, the main compounds present, with values ranging between 44.3⁻582, 50.9⁻550, and 24.36⁻101.5 mg/g extract, respectively. L. pedunculata aqueous extract revealed a higher antioxidant potential (EC50 values between 14 to 530 µg/mL), which could be related to its higher concentration in phenolic compounds; however, the hydroethanolic extract showed a higher anti-inflammatory (lower EC50 values than 124 µg/mL) potential and antiproliferative capacity (lower GI50 values than 34 µg/mL). Thus, this study highlights the bioactive effects of this species and opens up possibilities of uses in food and pharmaceutical formulations. However, there are potential differences in such properties according to geographical origin of plant material, as in general, the samples from Alentejo presented higher results in all the bioactivities, compared with Trás-os-Montes samples.

Anti-inflammatory activity of hydroalcoholic extracts of Lavandula dentata L. and Lavandula stoechas L.

ETHNOPHARMACOLOGICAL RELEVANCE: Plants from genus Lavandula have been used as anti-inflammatory drugs in Mediterranean traditional medicine. Nowadays, there is a growing interest for complementary medicine, including herbal remedies, to treat inflammatory bowel disease (IBD). AIM OF THE STUDY: To test the anti-inflammatory properties of Lavandula dentata and Lavandula stoechas extracts in two inflammatory experimental models: TNBS model of rat colitis and the carrageenan-induced paw edema in mice, in order to mimic the intestinal conditions and the extra-intestinal manifestations of human IBD, respectively. MATERIAL AND METHODS: The extracts were characterized through the qualitative HPLC analysis. Then, they were assayed in vitro and in vivo. In vitro studies were performed in BMDMs and CMT-93 epithelial cells with different concentrations of the extracts (ranging from 0.1 to 100µg/ml). The extracts were tested in vivo in the TNBS model of rat colitis (10 and 25mg/kg) and in the carrageenan-induced paw edema in mice (10, 25 and 100mg/kg). RESULTS: L. dentata and L. stoechas extracts displayed immunomodulatory properties in vitro down-regulating different mediators of inflammation like cytokines and nitric oxide. They also showed anti-inflammatory effects in the TNBS model of colitis as evidenced by reduced myeloperoxidase activity and increased total glutathione content, indicating a decrease of neutrophil infiltration and an improvement of the oxidative state. Besides, both extracts modulated the expression of pro-inflammatory cytokines and chemokines, and ameliorated the altered epithelial barrier function. They also displayed anti-inflammatory effects in the carrageenan-induced paw edema in mice, since a significant reduction of the paw thickness was observed. This was associated with a down-regulation of the expression of different inducible enzymes like MMP-9, iNOS and COX-2 and pro-inflammatory cytokines, all involved in the maintenance of the inflammatory condition. CONCLUSION: L. dentata and L. stoechas extracts showed intestinal anti-inflammatory effect, confirming their potential use as herbal remedies in gastrointestinal disorders. In addition, their anti-inflammatory effect was also observed in other locations, thus suggesting a possible use for the treatment of the extra-intestinal symptoms of IBD.

Composition and Variability of the Essential Oil of the Flowers of Lavandula stoechas from Various Geographical Sources.

Samples of flowers of wild Lavandula stoechas L. spp. stoechas populations were collected in three areas of Sicily (Italy) and were characterized in agronomic and chemical terms. Essential oil (EO) was extracted by hydrodistillation and analyzed by GC-FID and GC-MS. GC-FID and GC-MS analyses permitted identification of 89 compounds from the EO. The samples were separated into 3 groups using PCA (Principal Component Analysis) statistical method, with reference to the chemical composition of the EO. All three Sicilian populations of lavender were identified as the fenchone chemotype with percentage content ranged between 52.8-71.1%. The population of Partinico showed the highest dry weight of flowers per plant (221.3 g), but the lowest EO yield (0.37%). The essential oils of the three wild Sicilian populations of L. stoechas L. spp. stoechas showed a greater chemical differentiation than those obtained from other Mediterranean areas.

Lavandula L. - aplicação da cultura in vitro à produção de óleos essenciais e seu potencial económico em Portugal / Lavandula L. - application of in vitro cultivation for the essential oils production and their economic potential in Portugal

RESUMO Os óleos essenciais são metabolitos secundários que possuem diversas propriedades com elevado interesse, nomeadamente as biológicas. Estas propriedades englobam todas as atividades que esta mistura de compostos voláteis (principalmente monoterpenos, sesquiterpenos e fenilpropanóides) exerce sobre os seres humanos, animais e outras plantas. Os óleos essenciais apresentam grande valor económico, sendo os do género Lavandula dos mais comercializados e estudados devido à sua aplicabilidade industrial e propriedades terapêuticas As lavandulas são colhidas na natureza ou propagadas por técnicas convencionais, nomeadamente por estacaria. Mais recentemente, protocolos de micropropagação foram desenvolvidos para algumas espécies, permitindo a produção de plantas em larga escala disponíveis em qualquer período do ano e sem comprometer a biodiversidade das espécies. O desenvolvimento de plantas tetraplóides capazes de aumentar a produção de óleo essencial nas suas flores é outro meio eficaz para aumentar potencialmente o valor das espécies de Lavandula. Em Portugal existem 5 espécies nativas do género Lavandula, amplamente distribuídas pelo país. Contudo, o seu potencial industrial permanece praticamente inexplorado e em termos de mercado o seu reconhecimento é muito reduzido. Trabalhos recentes, baseados na avaliação das propriedades biológicas dos óleos essenciais e a forma como eles podem exercer os seus efeitos contribuíram para a valorização do potencial económico das lavandulas em Portugal. Esta revisão tenta dar uma visão geral de que forma a aplicação das culturas in vitro pode levar a uma maior produção de óleos essenciais em Lavandula spp., dando especial ênfase às lavandulas nativas de Portugal.

ABSTRACT Essential oils are secondary metabolites plants and, among other features, they own several biological properties. The term “biological” includes all activities that these mixtures of volatile compounds (mainly monoterpenes, sesquiterpenes, and phenylpropanoids) maintain on humans, animals and other plants. Essential oils have great economic value, being the genus Lavandula one of the most commercialized and studied type due to its dits industrial applicability and therapeutic properties. Lavenders are harvested in the nature or propagated using traditional methodologies. More recently micropropagation protocols have been developed for several Lavandula species allowing a production of a high amount of plants available at any time of the year without compromising the biodiversity of the species. Another efficient way to potentially increase the value of Lavandula species is through the development of tetraploid plants, capable of raising the production of essential oils in flowers. In Portugal, there are 5 native species of the genus Lavandula, widely distributed throughout the country. However, native Portuguese lavenders remain mainly unexplored and have received poor recognition in markets. A recent study was carried out in order to evaluate the biological properties of the oils and also to understand how they may exercise their effects, contributing to enhance the economic potential of lavandulas in Portugal. This review attempts to provide an overview about the application of in vitrocultivations for the production of essential oils in Lavandulaspp., with special emphasis on native lavandulas of Portugal.

[Histo-anatomical researches of two subspecies of Lavandula angustifolia Mill]. / Cercetari histo-anatomice privind doua subspecii de Lavandula angustifolia Mill.

AIM: The structure of the inflorescences of two subspecies of Lavandula angustifolia sL. angustifolia ssp. angustifolia and. L. angustifolia ssp. pyrenaica (D.C.) Guineat was studied to determine the range of variation in certain histo-anatomical characters. METHODS: The flower and leaf structure has been analysed on cross and superficial section using the usual techniques and methods applied in plant histo-anatomical research. RESULTS: In cross-section, inflorescence axis has a squared-shape contour, with four proeminent ribs. On the epiderme of inflorescence axis there are present stomata, trichoms and secretory hairs. The trichoms are pluricelullar and T-branched. The secretory hairs have a short pedicel and a uni-or bicellular head. CONCLUSIONS: The study revealed that there are only quantitative differences, referring to the degree of sclerification and lignification of mechanical elements and the size of the vascular bundles.

Essential oil variation among natural populations of Lavandula multifida L. (Lamiaceae).

Volatiles from twelve wild Tunisian populations of Lavandula multifida L. growing in different bioclimatic zones were assessed by GC (RI) and GC/MS. Thirty-six constituents, representing 83.48% of the total oil were identified. The major components at the species level were carvacrol (31.81%), beta-bisabolene (14.89%), and acrylic acid dodecyl ester (11.43%). These volatiles, together with alpha-pinene, were also the main compounds discriminating the populations. According to these dominant compounds, one chemotype was revealed, a carvacrol/beta-bisabolene/acrylic acid dodecyl ester chemotype. However, a significant variation among the populations was observed for the majority of the constituents. A high chemical-population structure, estimated both by principal component analysis (PCA) and unweighted pair group method with averaging (UPGMA) cluster analysis based on Euclidean distances, was observed. Both methods allowed separation of the populations in three groups defined rather by minor than by major compounds. The population groups were not strictly concordant with their bioclimatic or geographic location. Conservation strategies should concern all populations, because of their low size and their high level of destruction. Populations exhibiting particular compounds other than the major ones should be protected first.

Enantiomeric composition studies in Lavandula species using supercritical fluids.

Characteristic aroma compounds in plants and essential oils of Lavandula from different varieties were examined. The study of the qualitative and quantitative composition of the major volatile components was faced by developing a method based on the use of supercritical fluid extraction-GC-MS (SFE-GC-MS). The optimization of a variety of parameters affecting SFE extraction enabled RSDs from three replicates lower than 2% to be achieved. Equally, recoveries of up to 59% were obtained by applying the proposed method. The use of multidimensional GC was necessary to enantiomerically resolve the target compounds. The obtained results showed enantiomeric purities >90% for all studied compounds in all varieties considered, proving the natural invariability of the enantiomeric composition of the compounds of interest. This information can be useful in authenticity studies as well as in selecting natural sources of enantiomerically pure compounds.