Daedaleopsis Confragosa
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Cent. Eur. J. Biol.• 6(4) • 2011 • 575-582 DOI: 10.2478/s11535-011-0029-5 Central European Journal of Biology The antioxidant properties of polypore mushroom Daedaleopsis confragosa Research Article Senka Vidović1,*, Zoran Zeković1, Ibrahim Mujić2, Žika Lepojević1, Marija Radojković1, Jelena Živković3 1Faculty of Technology, Department of Biotechnology and Pharmaceutical Engineering, The University of Novi Sad, 21000 Novi Sad, Serbia 2Collegium Fluminense Polytechnic of Rijeka, 52441 Poreč, Croatia 3Faculty of Medicine, The University of Niš, The Department of Pharmacy, 18000 Niš, Serbia Received 17 November 2010; Accepted 15 February 2011 Abstract: Daedaleopsis confragosa belong to a large and remarkable group of mushrooms called polypores. This type of mushroom could be easily said to be quite unexplored and unused when it comes to its antioxidant properties. Thus, in order to evaluate its antioxidant activity, the investigation had to include the total phenolics and flavonoide content, the content of Selenium, the content of Zinc, the scavenging capacity on DPPH˙and OH˙ radicals, reducing power and capacity to remove lipid peroxidation. The investigated mushroom extract contained 54.17 mg GAE/g of total phenols and 48.46 mg CE/g of total flavonoides. Zinc and Selenium were detected and quantified by using inductively coupled plasma mass spectroscopy. The scavenging activity of the radicals was found to exhibit 50% of the inhibition value (IC50 value) at the extract concentration of 0.015±0.007 mg/ml for the investigated D. confragosa extract. By using electron paramagnetic resonant spectroscopy it was found that the investigated extract does not have a significant role in the prevention of lipid peroxidation. It was effective in scavenging on ˙OH radical, RI was 56±2%. Keywords: Mushrooms • Daedaleopsis confragosa • Antioxidant activity • Antioxidant compounds • Zinc • Selenium © Versita Sp. z o.o. 1. Introduction is believed that one species was used for its antibacterial properties and the other one for starting fire [3]. There Daedaleopsis confragosa belong to the order of the are two famous medicinal polypore mushrooms that Polyporales, the family of Polyporaceae and genus are in use today: Ganoderma lucidum, well known as Daedaleopsis. It is also known by its common name reishi, and Trametes versicolor. They have been used “the blushing bracket”. The polyporales are among in traditional medicine and in many scientific medicinal the most common and easily identifiable group of wild researches related to immune system recovery and mushrooms. They are usually inedible, because of its cancer [4]. As D. confragosa belong to this large and hardness and woody structure, except for a few special significant group of mushrooms, the further research of edible species [1,2]. Almost all of these mushrooms the latter could be of tremendous importance. grow on wooden bases such as trees, logs and stumps. Considering the growing interest for the natural Some of the polypore mushrooms have been used in sources of antioxidants on one hand, and the unexplored rituals many centuries ago. Famous Otzi the Iceman, the and unused field of polypore mushrooms on the other, the mummy of a man from about 53 centuries ago, who was main objective of this research was the investigation of the found in 1991 in a glacier on the border between Austria antioxidant properties and some antioxidant compounds and Italy, was carrying two different polypore species. It of the woody D. confragosa mushroom, collected from * E-mail: [email protected] 575 The antioxidant properties of polypore mushroom Daedaleopsis confragosa the Istrian peninsula in Croatia during late summer 2008. total phenolics and flavonoide content, the content of The powerful antioxidant properties of these mushrooms Selenium, the content of Zinc, the scavenging capacity refer to different antioxidant compounds. Mushrooms are on DPPH˙ and OH˙ radicals, the reducing power and generally considered to be a good source of protein and capacity to remove or prevent lipid peroxidation. phenolic antioxidants, which is the case with variegatic acid and diboviquinone [5]. Bearing in mind the presence of the conjugated ring structures and the number and 2. Experimental Procedures arrangement of the polar hydroxyl groups [6], many phenolic compounds can fuction as antioxidants by 2.1 Chemicals and Reagents scavenging superoxide anion [7], singlet oxygen [8], and 1,1-Diphenyl-2-picryl-hydrazyl-hydrate (DPPH), lipid peroxyl radicals [9], and by stabilizing free radicals that Folin-Ciocalteu reagent, L-α-phosphatidylcholine, are involved in oxidative processes through hydrogenation (±)-catechin, hypoxanthine and xanthine oxidase or complexing with the oxidizing species [10]. ˙OH radical was purchased from Sigma (Sigma-Aldrich GmbH, is one of the most reactive radicals in biological systems Sternheim, Germany). The gallic acid was purchased and is marked as a highly damaging species in free radical from Sigma (Sigma, St. Luis, MO, USA). Potassium pathology, capable of damaging bio-molecules of living ferricyanide, FeSO4 was purchased from Merck cells [11]. Thus, the inhibition of this destructive activity can (Darmstadt, Germany). 7-DS (2-(5-carboxypentyl)-2- be seen as crucial. Now, most active phenolic antioxidants, undecyl-4,4-dimethyloxazolidine-3-oxylstearate) was in view of their chemical structure, are flavonoides. purchased from Molecular Probes (Junction City, OR, Selenium and Zinc act as integral parts of the USA). DEPMPO (5-diethoxyphosphoryl-5-methyl- antioxidant systems. Alone, these micronutrients cannot 1-pyrroline-N-oxide) was purchased from Alexis be considered as antioxidant compounds. Selenium is Biochemical (Laursen, Switzerland). All other chemicals important as constituents of protective enzymes via special and reagents were of the analytical reagent grade. amino acids (selenocysteine, selenomethionine), or structures like selenium-proteins. Selenium-proteins have 2.2 Sample Preparation a key role in a variety of biological processes, including Voucher specimens (Daedaleopsis confragosa (Bolton) antioxidant defence [12]. For example, we may consider J. Schröt., Croatia, Istria, Ćićarija Mune, 08.2008. (det.: four glutathione peroxidases, among them cytosolic Maja Karaman) were confirmed and deposited at the glutathione peroxidase, the first Selenium-protein that Herbarium of the Department of Biology and Ecology was identified, which protects cells against peroxidative (BUNS Herbarium), Faculty of Natural Sciences, damage by reducing hydrogen peroxide, free fatty acid University of Novi Sad [18]. The mushrooms fruiting hydroperoxides, and phospholipids hydroperoxides [13]. bodies were collected in their natural environment, after This trace element is established as an essential element they were gently cleansed of any residual compost. of fundamental importance to human health. It is known The fruiting bodies of the fresh mushrooms were air- primarily for its antioxidant activity and its therapeutic dried and then stored in an air-tight plastic bag at room aspects, for its chemo-preventive, anti-inflammatory and temperature. The dried mushroom samples were milled antiviral properties [14]. Zinc antioxidant properties belong in a blender before the extraction. The mushroom to some other structures, for example Zn-metallothionein samples (10.0 g) were extracted by using 50% ethanol or antioxidant enzyme Cu-Zn superoxide dismutase. Zinc (50.0 ml). The extraction process was carried out using deficiency leads to oxidative damage of lipids, proteins, the ultrasonic bath (B-220, Branson and SmithKline DNA oxidation [15], or diseases such as cancer and Company, USA) at 45°C for 40 minutes. After the atherosclerosis. This second essential mineral could well filtration, the extraction solvent was removed by a perform its antioxidant actions using different mechanisms rotary evaporator (Devarot, Elektromedicina, Ljubljana, including sulfhydryl protection against oxidation, the Slovenia) under a vacuum. The obtained extract was induction of metallothionein, a cystein rich protein with the then dried at 60°C to the constant mass. In the end, capacity to scavenge oxidants and bind redox active metals, the dry extract was placed in glass bottles and stored and as a constituent of the intracellular and extracellular at -4°C to prevent oxidative damage until the analysis. antioxidant enzyme Cu-Zn superoxide dismutase [16]. It can replace the redox active metals, Copper and Iron, in 2.3 The Determination of Antioxidant membrane binding sites and thus inhibit the oxidation of Compounds and trace elements Selenium liposomes [17]. and Zinc For evaluating D. confragosa antioxidant activity, The content of total phenolic compounds in a dry the following elements have been investigated: the mushroom extract was determined by Folin-Ciocalteu 576 S. Vidović et al. procedure [19] using gallic acid as a standard. The for 20 min at 50°C. After the incubation, 2.5 ml of absorbance was measured at 765 nm. The content of 10% trichloroacetic acid solution was added to the total phenolic compounds was expressed as mg of gallic mixture, and the mixture was centrifuged for 10 minutes acid equivalent (GAE) per g of dry mushroom extract. (1509 x g). The obtained supernatant (2.5 ml) was mixed The total flavonoids content was determined by with bidestilated water (2.5 ml) and 0.1% FeCl3 solution aluminium chloride colorimetric assay [20] using catehin (0.5 ml). The absorbance was measured at 700 nm. The as