1 Introduction

Wild mushrooms are valuable non-timber forest resources and their use has been documented in many countries around the world (Jones and Whalley 1994; Garibay-Orijel et al. 2006). They are sold in traditional markets or commercially exploited as food or medicines. Mushrooms grown in the wild have been found to be nutritious and medicinally important (Barros et al. 2008; Kalac 2009). Mushrooms are also demanding because of their toughness, meaty taste, desirable flavor and medicinal values. The consumption of wild edible mushrooms is increasing in the developed countries, for being a rich source of proteins as well as other nutrients and minerals such as amino acids, sugar, vitamins with low fat and cholesterol content (Barros et al. 2008; Kalac 2009). Mushrooms are one of the natural sources of vitamin D (Mattila et al. 1994), which is essential for healthy bones and teeth. They are also a good source of the B vitamins; riboflavin (B2), niacin (B3) and pantothenic acid (B5) (Kalac 2009; Caglarlrmak et al. 2002). These vitamins help break down proteins, fats and carbohydrates so they can be used for energy. Mushrooms are not only sources of nutrients but also have been reported as therapeutic foods, useful in preventing diseases such as hypertension, hypercholesterolemia and cancer (Bobek and Galbavy 1999; Kidd 2000). These functional characteristics are mainly due to the presence of dietary fiber such as chitin and beta glucans (Kidd 2000; Lindequist et al. 2005). Recently, a number of bioactive molecules have been identified from various mushrooms which are known for their antibacterial, antioxidant, antitumor, antiviral, and immunomodulating properties (Lindequist et al. 2005; Rai et al. 2005). The bioactive compounds of mushrooms include phenolics, flavonoids, glycosides, alkaloids, carotenoids, ascorbic acid, tocopherols, folates, proteins, polysaccharides, volatile oils, organic aids etc. These molecules were quantified in many different species mainly from Finland, India, Korea, Poland, Portugal, Taiwan and Turkey (Ferreira et al. 2010). Other important phytochemicals have been isolated from medicinal mushrooms and developed as cytostatic polysaccharide drugs in Japan. These are ‘‘Krestin” (PSK) from the cultured mycelium of Kawaratake (Trametes versicolor), ‘‘Lentinan” from the fruiting bodies of Shiitake (Lentinus edodes) and ‘‘Schizophyllan” (Sonifilan) from the culture fluid of Suehirotake (Schizophyllum commune) (Mizuno 1993). Lentinan and Schizophyllan are pure β-glucans, whereas PSK is a protein bound polysaccharide (Larone 2002). The biological activity of these three products is related to their immunomodulating properties, which enhance the body’s defences against various forms of infectious disease. However, wild mushrooms are popular and favorite all over the world for their characteristics flavor as well as for nutritional and functional properties. For instance, many people collect wild edible mushrooms from forest for substantially contributing to food intake and also for ethno-medicinal purposes.

Similipal Biosphere Reserve (SBR) in Odisha, India is one of the tropical forest ecosystems rich in diversity of flora and fauna including mushrooms. Wild mushrooms are a popular food source among the tribes of SBR. This region is a high rainfall area with dense forest. The high humidity level during the monsoon season (June–October) provides ideal atmospheric conditions for the growth of many saprophytes, including the mushrooms. Several wild mushrooms growing in the forests of SBR are frequently consumed by the tribals/local people. Mushrooms are picked up from the forest and they form an integral part of diet during the monsoon season when these are abundantly available. In spite of immense popularity of this food in the region, data are not available till date regarding nutritional and medicinal value of these wild mushroom varieties available in SBR. Thus the objective of present study was to determine the nutrients composition and nutraceuticals potential (antioxidant and antibacterial activities) of three selected wild edible mushrooms viz., Russula vesca Fr., Russula delica Fr., Termitomyces eurrhizus (Berk.) Heim. from SBR along with their mineral compositions.

2 Results

Three collected wild edible mushrooms (Russula vesca, Russula delica and Termitomyces eurrhizus) from the forest soil of Similipal Biosphere Reserve were found non toxic. The results of macronutrient composition and energetic value of three wild edible mushrooms are given in Table 1. The moisture content of all mushrooms ranged from 7 g/100 g dw in Termitomyces eurrhizus to 15 g/100 g fw in Russula delica. Crude protein contents were found to be high levels and varied between 22.83 g/100 g dw in Termitomyces eurrhizus and 35 g/100 g dw in Russula vesca. The amount of free amino acids in the mushrooms ranged from 2.81 g/100 g dw in Russula delica to 6.26 g/100 g dw in Termitomyces eurrhizus. Crude fat values ranged from 2.03 g/100 g dw in Russula vesca to 4.62 g/100 g dw in Termitomyces eurrhizus, making ideal to be included in low caloric diets. Total carbohydrate contents of mushrooms was calculated by discounting protein and fat levels, were the most abundant macronutrient which ranged from 46 g/100 g dw in Russula delica to 63 g/100 g dw in Termitomyces eurrhizus. Starch is an important polysaccharide of carbohydrate in wild mushrooms which ranged between 8.09 g/100 g dw in Termitomyces eurrhizus to 12.35 g/100 g dw in Russula vesca. Reducing sugars were only a small part of carbohydrate, found between 1.17 g/100 g dw in Russula delica and 2.37 g/100 g dw in Russula vesca. Ash content of the mushrooms varied from 14.31 g/100 g dw in Russula vesca to 36.04 g/100 g dw in Termitomyces eurrhizus. On the basis of proximate analysis, it was observed that 100 g dw of these wild mushrooms contained highest value of energy in Russula vesca (394 kcal) and lowest energy in Russula delica (322 kcal). Micronutrients such as vitamins and carotenoids content were also determined and the results are presented in Table 2. The thiamine contents in mushrooms varied between 0.17 mg/g dw in Russula delica to 0.96 mg/g dw in Russula vesca. Riboflavin was not found in Termitomyces eurrhizus. Among other two mushrooms, Russula delica contained highest riboflavin (0.13 mg/g dw) than that of Russula vesca (0.11 mg/g dw). Ascorbic acid was found most abundantly in all three wild mushrooms. The highest level of ascorbic acid was found in Termitomyces eurrhizus (24.66 mg/g dw) and the lowest value was found in Russula delica (6.97 mg/g dw). Carotenoids were found in lower amounts; the highest levels of β-carotene and lycopene were observed in Termitomyces eurrhizus (2.24 μg/g dw) and Russula vesca (1.2 μg/g dw), respectively than that of other wild mushrooms.

Table 1 Macronutrient composition of three wild edible mushrooms
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Table 2 Micronutrient composition of three wild edible mushrooms
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The results of mineral contents such as phosphorus, potassium, manganese, cobalt, nickel, cadmium and iron in wild edible mushrooms are presented in Table 3. The phosphorus content in wild edible mushrooms ranged from 0.47 g/100 gm dw in Russula delica to 1.65 g/100 gm in Termitomyces eurrhizus. The highest potassium content was present in Russula vesca (1.8 g/100 g dw) and Termitomyces eurrhizus showed lower quantity of potassium (0.81 g/100 gm). The mean heavy metal concentrations across all the mushrooms species were in the order Fe > Mn > Ni > Co > Cd. The iron concentration ranged from 2.78 mg/kg dw in Russula delica to 15.6 mg/kg dw in Termitomyces eurrhizus. The highest level of manganese and cobalt was detected in Termitomyces eurrhizus (0.406 and 0.008 mg/kg dw, respectively) compared to the other mushrooms whereas cadmium was not detected in all the three wild mushrooms tested. The relationship between various minerals were correlated statistically and presented in Table 4. The results showed that there is a strong linear and positive correlation between the phosphorus with manganese and iron content (r = 0.918 and 0.975, respectively) and manganese with iron content (r = 0.965) in all the three wild mushrooms (Table 4). There is also strong negative correlation between phosphorous and potassium (r = −0.884) and potassium with manganese (r = −0.991) and iron content (r = −0.954) (Table 4).

Table 3 Mineral contents of three wild edible mushrooms
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Table 4 Correlations between the mineral contents
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The percentage yields of three different extracts (ethanol, methanol and aqueous) of the studied mushrooms ranged from 11.7 to 17.3 % of the dry weight (data not shown). The percentage yield of extracts from Russula vesca were found to be 14.3, 13.5 and 16.4 %, yield of extracts of Russula delica were 15.5, 11.3 and 17 % and yield of extracts of Termitomyces eurrhizus were 14, 11.7 and 17.3 % in ethanol, methanol and aqueous extracts, respectively. The composition of non-nutrients and in vitro antioxidant activity (scavenging potential) in the studied wild mushrooms are presented in Table 5 and Figs. 1, 2, 3, 4. The phenolic content in all the three studied wild mushrooms varied from 21.92 to 41.99 mg/g dw in three different extracts (ethanol, methanol and aqueous). Among the three extracts of mushrooms, the methanol extract of Termitomyces eurrhizus showed highest phenolic content (41.99 mg/g dw) whereas ethanol extract of Russula vesca showed lowest quantity (21.92 mg/g dw) (Table 5). The flavonoid content of studied wild mushrooms was found to be very less as compared to phenolic content. The flavonoid content of wild mushrooms ranged from 2.53 to 7.52 mg/g dw in three different extracts. Among the three wild mushrooms, Russula delica showed the highest amount of flavonoid (7.52 mg/g dw) in ethanol extracts whereas Russula vesca showed the lowest value (2.53 mg/g dw) in aqueous extract. Further, total antioxidant capacities of studied wild mushrooms were found in higher amount as compared to phenol and flavonoid. Total antioxidant capacity of Russula vesca ranged from 44.48 mg/g dw in ethanol extract to 48.18 mg/g dw in methanol extract. Total antioxidant capacity of Russula delica varied between 52.45 mg/g dw in ethanol extract to 56.44 mg/g dw in aqueous extract. In case of Termitomyces eurrhizus, the total antioxidant capacity was found to be highest in aqueous extract (47.21 mg/g dw) and lowest value was found in ethanol extract (40.30 mg/g dw).

Table 5 Non-nutrient composition and in vitro antioxidant properties of three wild edible mushrooms
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The results of antioxidant properties of all these three wild mushrooms were presented in Figs. 1, 2, 3, 4. The ABTS free radical scavenging activity of the ethanol and aqueous extract from Termitomyces eurrhizus showed significantly high activity (27.29 and 63.37 %) among the studied mushrooms at a concentration 100 μg/ml whereas methanol extract of Russula delica showed significantly high scavenging activity (39.10 %) at a concentration 100 μg/ml which is comparable with the standard BHT (Fig. 1).

Fig. 1
figure 1

ABTS free radical scavenging activities of ethanol (a), methanol (b) and aqueous extract (c) from three different wild mushrooms

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Fig. 2
figure 2

DPPH free radical scavenging activities of ethanol (a), methanol (b) and aqueous extract (c) from three different wild mushrooms

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Fig. 3
figure 3

H2O2 radical scavenging activities of ethanol (a), methanol (b) and aqueous extract (c) from three different wild mushrooms

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Fig. 4
figure 4

Metal chelating activities of ethanol (a), methanol (b) and aqueous extract (c) from three different wild mushrooms

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The DPPH radical scavenging activities of all the three extracts (ethanol, methanol and aqueous) from studied mushroom species with standard BHT are given in Fig. 2. In the DPPH radical scavenging assay, all these three extracts (ethanol, methanol and aqueous) from Termitomyces eurrhizus showed significantly high activity (12.89, 29.24 and 16.75 %, respectively) than the other two studied mushrooms (Russula vesca and Russula delica) at a concentration 100 μg/ml which is less comparable with standard BHT (Fig. 2).

The results of H2O2 free radical scavenging activity of wild mushrooms compared with that of BHT are presented in Fig. 3. In the H2O2 free radical scavenging assay, ethanol and aqueous extract of Termitomyces eurrhizus showed high H2O2 free radical scavenging activity (13.99 and 12.27 %, respectively) than the other studied mushrooms at concentration 100 μg/ml whereas methanol extract of Russula delica showed significantly high scavenging activity (12.07, 21.22 and 28.48 %) at three different concentration 50, 75 and 100 μg/ml, respectively which is comparable with standard BHT (Fig. 3).

The metal chelating activity of three different extracts (ethanol, methanol and aqueous) from wild mushrooms with BHT as standard are presented in Fig. 4. In comparison with BHT, the ethanol extract of Russula delica revealed the highest metal chelating ability (3.26 %) at concentration 100 μg/ml whereas methanol and aqueous extract of Termitomyces eurrhizus showed high chelating ability (4.16 and 4.78 %, respectively) among the studied mushrooms at concentration 100 μg/ml which is comparable with standard BHT (Fig. 4). The results of antibacterial activity in different crude extracts of wild mushrooms against six human pathogenic bacterial strains are presented in Table 6. All the three extracts (ethanol, methanol and aqueous) of Russula vesca showed highest antibacterial activity (inhibition zone in mm) against Escherichia coli (26–29.01 mm) and Staphylococcus aureus (23.07–27.16 mm) than the other two wild mushrooms (Russula delica and Termitomyces eurrhizus) (Table 6). The results of antibacterial activity depended largely upon the mushroom species, solvent used for extraction and the organisms tested.

Table 6 Antibacterial activity (inhibition zone in millimeter) of mushroom extracts
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3 Discussion

Mushrooms are widely appreciated food due to their unique taste and flavor, but also for their chemical and nutritional properties (Kalac 2009). Dry matter/moisture content is an important factor during the nutritional evaluation of mushrooms, which directly affects the nutrient content of mushrooms (Mattila et al. 2002). Present moisture contents in mushrooms of Similipal Biosphere Reserve (SBR) are in agreement with earlier published data, which state that fresh mushroom contained 70–96 % moisture (Agrahar-Murugkar and Subbuakshmi 2005; Grangeia et al. 2011; Pereira et al. 2012). However, variability of moisture content (70–96 %) is dependent on the mushroom species and other parameters such as environmental temperature, relative humidity during growth and relative amount of metabolic water that may be produced or utilized during storage (Crisan and Sands 1978). The studied species of mushrooms from SBR have high proteins and carbohydrate contents, in contrast to low fat levels, which make them suitable to incorporate into low calorie diets. Energetic contributions in wild mushrooms of SBR differ from species to species due to their nutrition contents. Despite the above nutritional values, Termitomyces eurrhizus also contained higher amount of ash as compared to other two mushrooms (Russula vesca and Russula delica). The findings of the present study are in agreement with different studies reported by several authors (Agrahar-Murugkar and Subbuakshmi 2005; Grangeia et al. 2011; Pereira et al. 2012). It is well known that free amino acids, especially highly basic amino acid and glutamic acid contribute to the flavor properties of mushrooms (Sugahara et al. 1975; Maga 1981) thus high levels of free amino acids in Termitomyces eurrhizus and Russula vesca probably contribute most to their characteristics flavor which was evaluated in our study. However, the mineral contents of wild mushrooms were in relatively lower quantity compared to earlier published report (Sugahara et al. 1975; Maga 1981; Ouzouni et al. 2009). This may be due to geographic location and environmental factors of SBR.

Besides macronutrients and minerals, studied wild mushrooms have also important micronutrients such as vitamins, carotenoids and non-nutrients (e.g. phenol and flavonoids) with bioactive properties such as antioxidant and antibacterial potential, which are in agreement with other reports concerning vitamins (thiamine, riboflavin and ascorbic acid), carotenoids (β-carotene and lycopene) and non nutrients (phenol and flavonoids) quantification in different mushrooms (Singdevsachan et al. 2013; Barros et al. 2007). However, some authors have already reported a direct correlation between mushrooms antioxidant activity and total phenolic content, although the antioxidant action is raised by other substances such as vitamins and carotenes (Cheung et al. 2003; Barros et al. 2007). In present study we tested whole extracts of total mushrooms, taking advantage of the idea that complex mixture of phytochemicals may have potential additive or synergistic effects (Cheung et al. 2003; Barros et al. 2007). Three different extracting solvents were used in order to obtain low molecular weight compounds such as phenolic antioxidants (ethanol and methanol) and high molecular weight compounds such as polysaccharides (water). The percentage yields of mushroom extracts in studied wild mushrooms are found in moderate quantities whereas mushrooms are reported to contain more polar constituents (Vaskovsky et al. 1998). The group such as phenolic acids, lignans, flavonoids with structures containing –OH and –COOH functional groups are easily extracted by the polar solvent in samples. The discrepancy in the yield from fruiting bodies of the mushrooms might be due to the differences in strains and harvest times (Mau et al. 2005). In fact, the bioactivity of phenolics may be related to their ability to chelate metals, inhibit lipoxygenase and scavenge free radicals (Decker 1997). Flavonoid can act as free radical scavengers and terminate the radical chain reactions that occur during the oxidation of triglycerides in food systems (Roedig-Penman and Gordon 1998). These molecules can also play protective role in diseases related to oxidative stress, such as cancer and cardiovascular diseases (Ferreira et al. 2010). The extracts of the studied mushroom species were demonstrated with a capacity to scavenge free radicals such as ABTS and DPPH, high inhibition power of HO− and Fe2+ chelating abilities which may be due to the flavonoids.

Free radical scavenging effect has been known as an established phenomenon in inhibiting lipid oxidation, which otherwise can be deleterious to the cellular components and cellular function. Natural antioxidant can be used to replace the synthetic antioxidant in the food industry such as BHT, BHA and TBHQ, which may possess mutagenic activity (Namiki 1990). With the presence of radical scavenging activity, consumption of wild mushrooms might be beneficial to protect human body against oxidative damage, which can be further developed into health related degenerative illnesses. The ABTS free radical scavenging assay is based on the inhibition of the absorbance of the radical cation ABTS+ which has a characteristic long wave length absorbance spectrum (Rice-Evans and Miller 1997). The results imply that all the three extracts (ethanol, methanol and aqueous) of the studied species inhibit or scavenge the ABTS+ radicals in increasing orders with the increasing concentration of extracts (50–100 μg/ml), which supports the earlier findings of several workers on both inhibition and scavenging properties of antioxidant towards ABTS+ radicals (Rice-Evans and Miller 1997; Baskar et al. 2008). The free radical DPPH· possesses a characteristic absorption at 517 nm (purple in color), which decreases significantly on exposure to radical scavengers (by providing hydrogen atoms or by electron donation). A lower absorbance at 517 nm indicates a higher radical scavenging activity of the extract. Free radical scavenging is one of the well known mechanisms by which antioxidants inhibit lipid oxidation. This antioxidant assay offers a rapid technique for screening the radical scavenging activity of specific compound or extracts (Barros et al. 2007). The DPPH radical scavenging activity of studied wild mushrooms with three different extracts (ethanol, methanol and aqueous) were found to be higher and are comparable with the standard BHT which were also reported by several authors in different species of mushrooms (Barros et al. 2007; Pereira et al. 2012). Hydrogen peroxide itself is not very reactive, but it can give highly reactive species HO− through Fenton reaction (Halliwell 1978). It was suggested that H2O2 could induce DNA break in the intact cell and purified DNA (Imlay et al. 1988). Thus, removal of H2O2 is crucial for medicinal importance. The H2O2 reducing capacity of compound or extract may serve as indicator of its potential antioxidant capacity (Meir et al. 1995). The H2O2 reducing capacities of extracts in the studied wild mushrooms were found high as compared to standard BHT. It is an agreement which will be beneficial for human health through consumption of wild mushrooms. In the metal chelating assay, extracts of the mushroom species interfered with the formation of ferrous and ferrozine complex, suggesting that they have chelating activity and capture ferrous ions before ferrozine. Iron stimulates lipid peroxidation by the Fenton reaction and accelerates peroxidation by decomposing lipid hydroperoxides into peroxyl and alkoxyl radicals that can abstract hydrogen and perpetuate the chain reaction of lipid peroxidation (Halliwell 1991). The extracts of wild edible mushrooms in the present study demonstrated a marked capacity for iron binding, suggesting that their action as peroxidation protector may be related to its iron binding capacity. The ethanol, methanol and aqueous extract of Russula vesca, Russula delica and Termitomyces eurrhizus, respectively were found to be better chelators for ferrous ion, since ferrous ions are the most effective pro-antioxidants in food system where the high ferrous ion chelating abilities of extracts from wild edible mushrooms would be beneficial.

Further, antibacterial activity (inhibition zone) in three extracts of studied wild mushroom was tested against six pathogenic bacterial strains in comparison with the standard antibiotic Amphoxyllin. All three extracts of Russula vesca showed higher antibacterial activity against studied pathogenic strains than other two mushrooms (Russula delica and Termitomyces eurrhizus) which is comparable with standard antibiotic Amphoxyllin. The results of antibacterial activity are in agreement with previous report (Yaltirak et al. 2009; Nwachukwu and Uzoeto 2010). Thus, the study could provide valuable information to support wild mushrooms as an excellent source of antioxidants and antibacterial agents in human diet especially to the low-income community or tribal peoples of SBR those are regularly consuming such kind of mushrooms.

4 Conclusion

The studied wild edible mushrooms of Similipal Biosphere Reserve are non-toxic. They contain all the nutrients (high carbohydrate and protein levels with low fat content and toxic metals) which make them ideal food for consumption and no doubt is a good source of nutrients for tribal peoples. Based on their antioxidant potential and bioactive compounds (vitamins and phenolics), they might have been used in ethno-medicine for curing of free radical related diseases. Besides, these mushrooms also showed antibacterial properties. The present study indicates that the daily intake of these mushrooms could provide a natural dose of antibiotic to fight against common pathogens. Further, as these are a source of nutrient, important antioxidants and antibacterial agents, the wild mushrooms of SBR can be used in the diet as nutraceuticals and/or functional foods maintaining and promoting health, longevity and life quality without any health risk.