In this project we will proceed to the study of Pieris brassicae, an insect whose larvae constitutes a frequent pest of some Brassica species, which are an important way of subsistence in the Trás-os-Montes region (Portugal). With this study we intend to take advantage of the damage caused, using P. brassicae as a source of compounds with interest for the health. Simultaneously, several Brassica, of high consumption and from which the insect feeds, will be valorised: tronchuda or Portuguese cabbage (Brassica oleracea var. costata), kale (Brassica oleracea var. acephala) and turnip (Brassica rapa var. rapa).
The chemical composition of tronchuda cabbage is already partially defined and its antioxidant activity is also described by our research group (POCI/AGR/57399/2004). The two other Brassica are little known, at both chemical and biological levels. Thus, samples of these Brassica, obtained from different conditions of cultivation, will be studied.
In what concerns the insect, the culture conditions will be optimized, with respect to moisture, temperature, photoperiod and food. Under the selected conditions the development of P. brassicae in its different stages (larvae, pupae and butterfly) will be accompanied. Individuals and its excrements will be analysed, in order to esteem taxes and products of metabolization.
With respect to the chemical composition of all matrices, phenolic compounds, amino acids, organic acids and carotenoids will be determined by HPLC-PAD-ESI/MS/MS, HPLC/UV-vis, HPLC/UV and HPLC/DAD, respectively.
To evaluate the antioxidant capacity it will be studied the ability of to act as a scavenger of reactive species of oxygen (superoxide radical, hydroxyl radical and hypochlorous acid) and of nitrogen (nitric oxide and peroxynitrite). Superoxide radical will be generated by the xanthine/xanthine oxidase and phenazine methosulphate/NADH systems and the prevention of nitroblue tetrazolium reduction induced by superoxide radical will be used as the measured endpoint. Hydroxyl radical will be generated by the Fe 3+-EDTA/ascorbate Fenton system, and assayed by the evaluation of deoxyribose degradation using the thiobarbituric acid method. Hypochlorous acid scavenging activity will be tested by measuring the inhibition of hypochlorous acid-induced 5-thio-2-nitrobenzoic acid oxidation to 5,5`-dithiobis (2-nitrobenzoic) acid. Nitric oxide will be generated by the sodium nitroprusside and the nitrite resulting of its autoxidation will be measured appealing to the Greiss reagent. Peroxynitrite will be synthesised from potassium nitrite and its effect in the 3-nitrotyrosine formation will be determined by HPLC/UV.
It will also be evaluated the antimicrobial activity of P. brassicae and of the three Brassica species of which it feeds, relatively to Gram positive (Bacillus cereus, B. subtilis and Staphylococcus aureus) and Gram negative (Pseudomonas aeruginosa, Escherichia coli and Klebsiella penumoniae) bacteria and to fungi (Candida albicans and Cryptococcus neoformans).
At the end of the project its conclusions will be transmitted to the adequate entities, namely to “Ministry of Agriculture, Rural Development and Fishing” and to cooperatives of farmers producers of the involved spec