José L. Guil-Guerrero

Occurrence and characterization of oils rich in γ-linolenic acid. Part I : Echium seeds from Macaronesia

Nineteen species of the genus Echium (Fam. Boraginaceae) collected in Macaronesia were surveyed in a search for new sources of gamma-linolenic acid (GLA, 18:3omega6). High amounts of this acid were found in all of them, ranging from 9.15% (E. plantagineum) to 26.31% (E. callithyrsum) of total seed fatty acids. The amounts of GLA related to total seed weight were also significant, ranging from 1.77% (E. sventenii) to 5.02% (E. nervosum). In addition, considerable amounts of stearidonic acid (SA, 18:4omega3) were detected, ranging from 3.03% (E. auberianum) to 12.94% (E. plantagineum) of total fatty acids. These data allow us to consider tile members of the genus Echium from Macaronesia as one of the richest sources of gamma-linolenic acid found so far in nature. The results obtained from multivariable data analysis and the taxonomic relationships among the species is discussed.

Occurrence and characterization of oils rich in γ-linolenic acid (III) : the taxonomical value of the fatty acids in Echium (Boraginaceae)

Fourteen species of the genus Echium (Fam. Boraginaceae) collected in the Macaronesia were surveyed in a search for high levels of gamma-linolenic acid (GLA, 18:3omega6) in the seed oil. High amounts of this fatty acid were found in all of them, ranging from 18.85% (E. pitardii var. pitardii) to 27.42% (E. gentianoides) on total seed fatty acids. The GLA content related to total seed weight was also significant, ranging from 1.26% (E. handiense) to 8.22% (E. gentianoides). In addition, considerable amounts of stearidonic acid (SA, 18:4omega3) were detected, ranging from 3.78% (E. bonnetii var. bonnetii) to 8.81% (E. pininana) on total fatty acids. Besides all the perennial species, the four herbaceous Echium taxa endemic to the Macaronesia also showed high GLA percentages. This is in contrast to the low GLA level found in continental Echium species, all of them bearing an herbaceous habit. These results are in good agreement with the available genetic data and show the ability of GLA to discriminate between Macaronesian and continental Echium species. The analysis of five other Macaronesian species belonging to plant families rich in GLA are also reported.

Δ6-Desaturase sequence evidence for explosive Pliocene radiations within the adaptive radiation of Macaronesian Echium (Boraginaceae).

The oceanic islands of Macaronesia provide an ideal temporal and spatial context to test hypotheses of plant evolution using a novel set of phylogenetic markers, Delta(6)-desaturase sequences. In contrast to the limited resolution of standard molecular markers (nrDNA and plastid sequences), the Delta(6)-desaturase sequence phylogeny of Echium unequivocally reconstructs its active colonization across islands and archipelagos (Madeira, the Canary Islands, and Cape Verde), as well as its subsequent geographical and ecological speciation. Molecular-clock estimates using penalized likelihood and Bayesian inference reveal two radiation processes coincident with two dramatic climatic changes recorded in the region: the advent of the cold Canarian sea current (ca. 4 Ma) and the establishment of a strong seasonality in the Pleistocene (1.8 Ma). Though Echium had available all the diversity of present-day Macaronesian environments (xeric and mesic scrub, laurisilva, pine forest, and subalpine habitats) in the Miocene, evolutionary divergence appears to have been triggered by an extension of fluctuating xeric and mesic habitats with the advent of Pliocene conditions. These Echium radiations not only fulfill traditional predictions of adaptive radiation (i.e., common ancestry, rapid speciation, and phenotype-environment correlation), but also, uniquely among Macaronesian species, trait utility of woodiness. A Pliocene transition from annuality to a bush or tree-like condition occurred in early Echium lineages. Maintenance of woodiness in major lineages, and reversal to an herbaceous condition by three independent events, is reported for the first time in plants of oceanic islands.

Occurrence and characterization of oils rich in γ-linolenic acid. Part II: fatty acids and squalene from Macaronesian Echium leaves.

Leaves from 25 Macaronesian Echium (Boraginaceae) species have been surveyed for hydrocarbon compounds. These plants were previously reported as the major source of gamma-linolenic acid so far found in nature. In addition, six European Echium species and the common Borago officinalis have been analysed for comparative purposes. High squalene amounts were found in all Echium plants from the Macaronesia, ranging from 3.73%, in E. simplex to 20.1% in E. fastousum. Squalene was almost absent from all European Echium species, and the same is true for B. officinalis. The relatively high oil content (2.27%) in leaves of E. fastuosum raises the total squalene amount to about 0.46% within this tissue. The main fatty acid component in the leaf was alpha-linolenic acid (ALA, 18:3omega3), ranging in the Macaronesian Echium from 9.32% in E. acanthocarpum to 54.45% in E. simplex. Possible utilisation of these plants as a commercial source of squalene and hypotheses about its physiological role in the plant are discussed.

The fat from frozen mammals reveals sources of essential fatty acids suitable for Palaeolithic and Neolithic humans.

The elucidation of the sources of n-3 fatty acids available for the humans in the Upper Palaeolithic and Neolithic is highly relevant in order to ascertain the availability of such nutrients in that time frame as well as to draw useful conclusions about healthy dietary habits for present-day humans. To this end, we have analysed fat from several frozen mammals found in the permafrost of Siberia (Russia). A total of 6 specimens were included in this study: 2 mammoths, i.e. baby female calf called “Lyuba” and a juvenile female called “Yuka”, both specimens approximately from the same time, i.e. Karginian Interstadial (41,000 and 34,000 years BP); two adult horses from the middle Holocene (4,600 and 4,400 years BP); and two bison very close to the Early Holocene (8,200 and 9,300 years BP). All samples were analysed by gas-liquid chromatography-mass spectrometry (GLC-MS) and GLC-flame ionization detector (GLC-FID). As demonstrated in this work, the fat of single-stomached mammals often consumed by Palaeolithic/Neolithic hunters contained suitable amounts of n-3 and n-6 fatty acids, possibly in quantities sufficient to meet the todays recommended daily intake for good health. Moreover, the results also suggest that mammoths and horses at that time were hibernators.

Docosahexaenoic acid at the sn-2 position of structured triacylglycerols improved n-3 polyunsaturated fatty acid assimilation in tissues of hamsters.

In this study, we hypothesized that the incorporation of docosahexaenoic acid (DHA) in tissues will be higher when it is ingested as triacylglycerols (TAG) structured at the sn-2 position, which enhances efficacy and health benefits of dietary DHA n-3 supplementation. Ten-week-old Golden Syrian male hamsters were randomly allocated into 4 dietary groups with 10 animals in each: linseed oil (LSO; control group), fish oil (FO), fish oil ethyl esters (FO-EE), and structured DHA at the sn-2 position of TAG (DHA-SL). After 12 weeks, there were no variations in the hamsters body composition parameters across dietary groups. The DHA-SL diet had the lowest values of total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, total lipids, and aspartate aminotransferase activity, whereas the inverse was observed for the FO diet. Glucose was increased in the LSO diet without affecting insulin and insulin resistance markers. Whereas n-3 polyunsaturated fatty acid was increased in the brain of hamsters fed the DHA-SL diet, higher levels of n-6 polyunsaturated fatty acid were observed in the liver and erythrocytes of the LSO. The highest omega-3 index was obtained with the DHA-SL diet. The principal component analyses discriminated DHA from other metabolites and set apart 4 clusters matching the 4 diets. Similarly, liver, erythrocytes, and brain were separated from each other, pointing toward an individual signature on fatty acid deposition. The structured sn-2 position DHA-containing TAG ameliorated blood lipids and fatty acid incorporation, in particular eicosapentaenoic acid and DHA in liver, erythrocytes, and brain, relative to commercially FOs, thus improving the health benefits of DHA due to its higher bioavailability.

Phytochemical composition and in vitro anti-tumour activities of selected tomato varieties

BACKGROUNDnPrevious studies indicated that tomato is a rich source of phytochemicals that act on different tumours. In this research, the phytochemical composition of selected tomato varieties was assessed by GLC and UHPLC/HPLC-MS, as well as their anti-tumour activities on HT-29 colorectal cancer cells.nnnRESULTSnSignificant differences were found among tomato varieties; lycopene was high in Racimo, phenolics in Pera, sterols in Cherry, and linoleic acid predominated in all varieties. The MTT and LDH assays showed significant time- and concentration-dependent inhibitory/cytotoxic effects of all tomato varieties on HT-29 cells. Furthermore, the joint addition of tomato carotenoids and olive oil to HT-29 cell cultures induced inhibitory effects significantly higher than those obtained from each of them acting separately, while no actions were exercised in CCD-18 normal cells.nnnCONCLUSIONnTomato fruits constitute a healthy source of phytochemicals, although differences exist among varieties. In vitro, all of them inhibit colorectal cancer cell proliferation with Racimo variety at the top, and exercising a selective action on cancer cells by considering the lack of effects on CCD-18 cells. Furthermore, synergy was observed between olive oil and tomato carotenoids in inhibiting HT-29 cancer cell proliferation; conversely, phenolics showed no significant effects and hindered carotenoids actions.

Phytochemical Composition and Antitumor Activities of New Salad Greens: Rucola (Diplotaxis tenuifolia) and Corn Salad (Valerianella locusta)

D. tenuifolia and V. locusta, two greens, were analyzed for active compounds and antitumor actions on colorectal cancer cells. Phenolics were determined by UHPLC-Orbitrap-MS; carotenoids and glucosinolates by HPLC-MS; and sterols and fatty acids by gas–liquid chromatography (GLC). For antitumor effects, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) tests were run on HT-29 colorectal cancer cells, and in CCD-18 untransformed enterocyte cells. Six main carotenoids were identified in both vegetables, while total carotenoids accounted for 3520 and 2970xa0μgu2009·u2009g−1 dry weight in D. tenuifolia and V. locusta, respectively. Six phenolics were detected in D. tenuifolia (68,600xa0μgu2009·u2009g−1 dry weight) and five in V. locusta (139,000xa0μgu2009·u2009g−1 dry weight). Three glucosinolates (GSL) were found in D. tenuifolia (1960xa0μgu2009·u2009g−1 dry wt. total). Low-polarity extracts from V. locusta and D. tenuifolia showed IC50u2009~u2009150 and ~200xa0μgu2009·u2009mL−1 on HT-29 cells, while both plants lacked actions on CCD-18 cells. V. locusta inhibited HT-29 cancer cells viability more efficiently than D. tenuiofolia, but induced less cytotoxicity. This work highlights the importance of functional foods for colorectal cancer prevention.

Various Acylglycerols from Common Oils Exert Different Antitumor Activities on Colorectal Cancer Cells

ABSTRACT Colorectal cancer is one of the leading causes of death in Western countries; therefore, the implementation of healthy dietary habits in order to prevent its occurrence is a desirable action. We show here that both free fatty acids (FFAs) and some acylglycerols induce antitumoral actions in the colorectal cancer cell line HT-29. We tested several C18 polyunsaturated fatty acid-enriched oils (e.g., sunflower and Echium) as well as other oils, such as arachidonic acid-enriched (Arasco®) and docosahexaenoic acid-enriched (Marinol® and cod liver oil), in addition to coconut and olive oils. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide test indicated inhibitory effects on HT-29 cells viability for FFAs, and monoacylglycerol and diacylglycerol (DAG) species, while the lactate dehydrogenase test proved that FFAs were the more effective species to induce membrane injury. Conversely, all species did not exhibit actions on CCD-18 normal human colon cells viability. Furthermore, transmission electron microscopy showed the presence of necrosis and apoptosis, while the monoacylglycerol lipase (MAGL) inhibition test demonstrated high activity for 2-monoacylglycerols derived from Arasco and sunflower oils. However, different monoacylglycerols and DAGs have also the potential for MAGL inhibition. Therefore, checking for activity on colon cancer cells of specifically designed acylglycerol-derivative species would be a suitable way to design functional foods destined to avoid colorectal cancer initiation.

The PUFA-Enriched Fatty Acid Profiles of some Frozen Bison from the Early Holocene found in the Siberian Permafrost

Knowledge concerning the availability of n-3 fatty acids for humans in prehistoric times is highly relevant in order to draw useful conclusions on the healthy dietary habits for present-day humans. To this end, we have analysed fat from several frozen bison found in the permafrost of Siberia (Russia). A total of 3 bison were included in this study, all them very close to the early Holocene (8,000; 8,200; and 9,300 years BP). All samples were analysed by gas-liquid chromatography-mass spectrometry (GLC-MS) and GLC flame-ionization detection (GLC-FID). Fat samples from two bison showed two well-differenced areas, i.e. brown and white, the latter being saturated fatty acid enriched, corresponding to an intermediate stage of adipocere formation, while the brown ones yielded α-linolenic acid in higher percentages than found in present-day bison. As demonstrated in this work, the subcutaneous fat of bison consumed by Mesolithic hunters contained amounts of n-3 fatty acids in higher quantities than those found in current bison; thus, the subcutaneous fat of bison could have contributed to meet todays recommended daily intake of essential fatty acids for good health in the Mesolithic to a greater extent than previously thought.