Gamma radiation preserves chemical and bioactive properties of Boletus edulis wild mushrooms

Boletus edulis Bull: Fr. is an edible mushroom quite appreciated for its organoleptic and nutritional properties. However, the seasonality and perishability cause some difficulties in its distribution and marketing in fresh form; losses associated with this type of food during marketing can reach 40% [1]. Irradiation is recognized as a safe and effective method for food preservation, being used worldwide to increase shelf life of fresh and dehydrated products (e.g. fruits, vegetables and spices) [2]. In particular, gamma irradiation has already been applied to cultivated mushrooms (especially Agaricus, Lentinula and Pleurotus Genus) and proved to be an interesting conservation technology [3]. However, the studies with added-value wild species are scarce. In this work, the effects of gamma irradiation on chemical and antioxidant properties of wild B. edulis, were evaluated. Fruiting bodies were obtained in Trás-os-Montes, in the Northeast of Portugal, in November 2012. The irradiation was performed in experimental equipment with 60Co sources at 1 and 2 kGy. All the results were compared with nonirradiated samples (control). Macronutrients and energy value were determined following official procedures of food analysis; fatty acids were analyzed by gas-chromatography coupled to flame ionization detection (GC-FID), while sugars and tocopherols were determined by high performance liquid chromatography (HPLC) coupled to refraction index (RI) and fluorescence detectors, respectively. Antioxidant activity was evaluated in the methanolic extracts by in vitro assays measuring DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging activity, reducing power, inhibition of β- carotene bleaching and inhibition of lipid peroxidation using thiobarbituric acid reactive substances (TBARS) assay. Total phenolics were also determined by the Folin-Ciocalteu assay. The nutritional profiles were not affected in high extension. Fatty acids and sugars were slightly affected, decreasing with the increasing doses. The performed assays for antioxidant activity, indicate that irradiated samples tended to have lower scavenging activity and reducing power, but higher lipid peroxidation inhibition. Despite the detected differences in individual compounds, the results of nutritional parameters, the most relevant in terms of mushroom acceptability by consumers, were less affected, indicating an interesting potential of gamma-irradiation to be used as an effective conservation technology for the studied mushrooms.

Study of antioxidant, antiproliferative and apoptosis-inducing properties of wild mushrooms from the Northeast of Portugal

Mushrooms are known as a powerful source of bioactive compounds including antioxidants, inhibitors of human tumour cell lines growth, inducers of apoptosis and enhancers of immunity. Indeed, many pre-clinical studies have been conducted in human tumour cell lines and in some cases a number of compounds isolated from mushrooms have followed to clinical trials. The Northeast of Portugal is one of the European regions with higher wild mushrooms diversity. However, to our knowledge, no studies had been conducted so far to verify their bioactivities. The main aim of this work was the evaluation of the bioactive properties (antioxidant properties and growth inhibitory potential on human tumour cell lines) of wild edible mushrooms collected in the Northeast of Portugal. Once properly identified, methanolic, ethanolic and boiling water extracts were prepared from thirty eight wild mushroom species collected in that region. Chemical characterization was obtained by high performance liquid chromatography (HPLC) coupled to a photodiode array detector (DAD) or to a refraction index detector (RI). Antioxidant activity assays were carried out in those extracts, including evaluation of 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals scavenging capacity, reducing power and inhibition of β-carotene bleaching. Extract-induced cell growth inhibition was assessed with the sulforhodamine B assay in four human tumour cell lines (NCI-H460 - lung cancer, MCF-7 -breast cancer, HCT-15 -colon cancer and AGS - gastric cancer). The effects on cell cycle profile and apoptosis were evaluated by flow cytometry and the effect on the expression levels of proteins related to cell cycle and apoptosis was further investigated by Western blotting. Three wild edible mushroom species revealed growth inhibitory activity in the studied human tumour cell lines: Clitocybe alexandri ethanolic extract, Lepista inversa methanolic extract and Suillus collinitus methanolic extract. C. alexandri ethanolic extract induced an S-phase cell cycle arrest and increased the percentage of apoptotic cells, in the NCI-H460 cell line. The analysed mushroom species also provided interesting antioxidant potential, mainly the boiling water extract of L. inversa which showed the highest DPPH radical scavenging activity, reducing power and β-carotene bleaching inhibition. S. collinitus methanolic extract induced a slight increase in the number of cells in G1, with a concomitant decrease in the percentage of cells in the S phase of the cell cycle and an increase in the percentage of apoptotic cells, in the MCF-7 cell line. The combined use of the S. collinitus methanolic extract and etoposide caused a greater decrease in the percentage of cell growth, when compared to either of them used individually, indicating the potential benefit of this combination. The tested extracts were chemically characterized and protocatechuic, p-hydroxybenzoic, p-coumaric and cinnamic acids were the main compounds identified on the phenolic (methanolic and ethanolic) extracts, while mannitol, trehalose and arabinose were the main sugars found in the polysaccharidic (boiling water) extracts after hydrolysis. The individual compounds identified in the extracts were submitted to a screening of tumour cells growth inhibitory activity, but only the phenolic acids and a related compound, cinnamic acid, presented activity. This compound was found to be the most potent one regarding cell growth inhibition in the NCI-H460 cell line. The effect of the individual and combined treatment with the identified compounds was also evaluated. Cinnamic and protochatequic acids caused a statistically significantly reduction in the number of viable cells. In addition, p-hydroxybenzoic acid did not show any significantly reduction in the viable cell number. Nevertheless, it was verified that the concomitant use of the three compounds provided the strongest decrease in the viable cell number, suggesting a possible concomitant effect of those compounds. Overall, the present work has contributed to further understand the bioactive potential of wild edible mushrooms from the Northeast of Portugal. This study allowed to identify some species with antioxidant or tumour cell growth inhibitory potential.

From the field to the table: ionizing radiation as a feasible postharvest treatment for fresh and dried plant foods

Food irradiation is a treatment that involves subjecting in-bulk or packaged food to a controlled dose of ionizing radiation, with a clearly defined goal. It has been used for disinfestation and sanitization of food commodities and to retard postharvest ripening and senescence processes, being a sustainable alternative to chemical agents 1 . Doses up to 10 kGy are approved by several international authorities for not offering negative effects to food from a nutrition and toxicology point of view 2 . However, the adoption of this technology for food applications has been a slow process due to some misunderstandings by the consumer who often chooses non-irradiated foods. In this study, the effects of the ionizing radiation treatment on physical, chemical and bioactive properties of dried herbs and its suitability for preserving quality attributes of fresh vegetables during cold storage were evaluated. The studied herbs, perennial spotted rockrose (Tuberaria lignosa (Sweet) Samp.) and common mallow (Malva neglecta Wallr.) were freeze-dried and then irradiated up to 10 kGy in a Cobalt-60 chamber. The selected vegetables, watercress (Nasturtium officinale R. Br.) and buckler sorrel (Rumex induratus Boiss. Reut.) were rinsed in tap water, packaged in polyethylene bags, submitted to irradiation doses up to 6 kGy and then were stored at 4 C for a period of up to 12 days. Physical, chemical and bioactive parameters of irradiated and non-irradiated samples were evaluated using different methodologies the colour was measured with a colorimeter, individual chemical compounds were analyzed by chromatographic techniques, antioxidant properties were evaluated using in vitro assays based on different reaction mechanisms, and other quality analyses were performed following official methods of analysis. The irradiation treatment did not significantly affect the colour of the perennial spotted rockrose samples, or its phenolic composition and antioxidant activity 3 . Medium doses preserved the colour of common mallow and a low dose did not induce any adverse effect in the organic acids profile. The green colour of the irradiated vegetables was maintained during cold storage but the treatment had pros and cons in other quality attributes. The 2 kGy dose preserved free sugars and favoured polyunsaturated fatty acids (PUFA) while the 5 kGy dose favoured tocopherols and preserved the antioxidant properties in watercress samples. The 6 kGy dose was a suitable option for preserving PUFA and the ω-6 ω-3 fatty acids ratio in buckler sorrel samples. This comprehensive experimental work allowed selecting appropriate processing doses for the studied plant foods in order to preserve its quality attributes and edibility.

Nutritional and biochemical profiling of Leucopaxillus candidus (Bres.) singer wild mushroom

The wild mushroom Leucopaxillus candidus (Bres.) Singer was studied for the first time to obtain information about its chemical composition, nutritional value and bioactivity. Free sugars, fatty acids, tocopherols, organic and phenolic acids were analysed by chromatographic techniques coupled to different detectors. L. candidus methanolic extract was tested regarding antioxidant potential (reducing power, radical scavenging activity and lipid peroxidation inhibition). L. candidus was shown to be an interesting species in terms of nutritional value, with high content in proteins and carbohydrates, but low fat levels, with the prevalence of polyunsaturated fatty acids. Mannitol was the most abundant free sugar and β-tocopherol was the main tocopherol isoform. Other compounds detected were oxalic and fumaric acids, p-hydroxybenzoic and cinnamic acids. The methanolic extract revealed antioxidant activity and did not show hepatoxicity in porcine liver primary cells. The present study provides new information about L. candidus.