Antioxidant and anticholinesterase activities of five wild mushroom species with total bioactive contents

Abstract

Context: Recently, mushrooms are interesting natural products to be investigated due to exhibiting various bioactivities.

Objective: This study determines the antioxidant and anticholinesterase activities of various extracts of five wild mushroom species. In addition, the total bioactive contents, namely, ascorbic acid, β-carotene, and lycopene along with phenolic and flavonoid contents were also determined spectrophotometrically.

Materials and methods: Antioxidant activity was tested by using five complementary tests; namely, β-carotene-linoleic acid, DPPH^• scavenging, ABTS^•+ scavenging, cupric-reducing antioxidant capacity (CUPRAC), and metal chelating assays. The in vitro anticholinesterase activity was tested against acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes using the Ellman method. The spectrophotometric methods were used to determine the total phenolic, flavonoid, ascorbic acid, β-carotene, and lycopene contents.

Results: The current study has shown that ethyl acetate extracts of Ganoderma lucidum (Curtis) P. Karst (IC₅₀: 1.55 ± 0.05 µg/mL) and Funalia trogii (Berk.) Bondartsev & Singer (IC₅₀: 4.31 ± 0.18 µg/mL) exhibited good lipid peroxidation inhibitory activity. The DPPH, ABTS, and CUPRAC assays supported this activity. The ethyl acetate and methanol extracts of Funalia trogii and Ganoderma lucidum indicated good anticholinesterase activity. Ganoderma lucidum had rich phenolic and flavonoid contents, indicating 98.67 ± 0.32 mg PEs/g extract and 160.38 ± 1.25 mg QEs/g extract, respectively.

Discussion and conclusion: The results demonstrate that some of the mushroom species tested herein could be used in food and pharmaceutical industries as natural antioxidants.

• Funalia trogii
• Ganoderma lucidum
• Gyromitra esculenta
• Lyophyllum decastes
• Pleurotus ostreotus

Introduction

All over the world, mushrooms are used as valuable healthy foods since they are low in calories, fat, and essential fatty acids whereas they are rich in proteins, vitamins, and minerals (Pereira et al., 2012; Reis et al., 2012). Evaluation of studies in the literature on mushroom species indicate that polysaccharides, β-glucans, polysaccharide-protein complexes, lanosten-type triterpenoids, cyathane-type diterpenoids, steroids, alkaloids, and phenolic compounds were isolated from mushroom species previously. These isolated compounds exhibited anticancer, antioxidant, antitumor, antiviral, antibacterial, antifungal, anti-inflammatory, and immunomodulator activities (Moradali et al., 2007; Tong et al., 2009; Türkoğlu et al., 2007).

Funalia trogii (Berk.) Bondartsev & Singer is a non-poisonous inedible mushroom species that grows on trees. So far a few studies on F. trogii have been reported. The antioxidant activity of methanol and water extracts has been studied using DPPH radical scavenging, metal chelating, and lipid peroxidation inhibitory assays (Yegenoglu et al., 2011). Ganoderma lucidum (Curtis) P. Karst has been used as a medicinal mushroom in traditional Chinese medicine for more than 2000 years. There are many reports on G. lucidum. In previous studies, several biological activities such as antioxidant (Yegenoglu et al., 2011), anticancer (Trajkovic et al., 2009), antitumor (Joseph et al., 2011; Liu et al., 2012), immunomodulatory (Wang et al., 2013), and anti-inflammatory properties (Joseph et al., 2011) of G. lucidum were investigated. Although, potentially fatal if eaten raw, Gyromitra esculenta (Pers. ex Pers.) Fr. is a popular delicacy in Scandinavia, Eastern Europe, and some regions of North America. There are some studies performed on G. esculenta mushroom (Arshadi et al., 2006; Leal et al., 2013). Lyophyllum decastes (Fries: Fries) Singer, commonly known as the fried chicken mushroom, is an edible species in the Lyophyllaceae family. In previous studies, several biological activities of L. decastes such as antitumor (Ukawa et al., 2000), hypocholesterolemic (Ukawa et al., 2001), and antidiabetic (Miura et al., 2002) properties were investigated. Pleurotus ostreotus (Jacq. ex Fr.) P.Kumm., the oyster mushroom, is another common edible mushroom. Antioxidant (Yim et al., 2010) and antimicrobial activities (Vamanu, 2013) of P. ostreotus were studied.

The antioxidant activity by using ABTS radical scavenging and cupric-reducing antioxidant capacity (CUPRAC) assays of F. trogii and antioxidant activity of G. esculenta were studied for the first time in this study. In addition, the anticholinesterase activity of all studied mushroom species was investigated for the first time.

Some diseases, such as atherosclerosis, diabetes, cancer, and cirrhosis have been related with oxidative damage caused by oxygen-centered free radicals. In fact, oxidation is necessary for many living organisms (Halliwell & Gutteridge, 1984). Nowadays, especially in food and pharmaceutical industries, synthetic antioxidants have been used against free radicals due to their easy accessibility (Ding et al., 2010). To the best of our knowledge, since synthetic antioxidants are suspected of being responsible for liver damage and carcinogenesis, some of them were restricted to be used in the food industry by some countries (Grice, 1988). After that, some of the antioxidants occurring naturally have received much attention as safe antioxidants (Scalbert et al., 2005). In contrast, it is also reported that oxygen-centered free radicals contribute to cellular ageing and neuronal damage (Sastre et al., 2000). According to Soholm (1998), an excess amount of free radical species causing oxidative stress is also associated with the pathology of many diseases including Alzheimer’s disease. Supportively, the lack of vitamin E, a natural antioxidant, enhanced Alzheimer’s disease on a mouse model (Nishida et al., 2006). Thus, the development and utilization of more effective antioxidants of natural origin as well as anticholinesterase compounds are desired.

Regarding the consumption of G. lucidum, G. esculenta, L. decastes, and P. ostreotus, and the usage of F. trogii, collected from Uşak-Turkey, we aimed to investigate the antioxidant and anticholinesterase activities, and the total bioactive compounds of these species. The main goal of the study is to compare the results with each other and those in the literature.

Materials and methods

Mushroom materials

The species names, collection localities and dates, family, and edibility of five mushroom species were listed in Table 1. All species were identified by Dr. Aziz Türkoğlu. Voucher specimen have been deposited in the Fungarium of Department of Biology, Muğla Sıtkı Koçman University.

Table 1. Collection localities and dates, family and edibility of the mushroom species studied.

No.	Mushroom	Collection localities and dates	Family	Edibility
1	Funalia trogii (Berk.) Bondartsev & Singer	Uşak-Eşme, September 2009	Polyporaceae	Not edible
2	Ganoderma lucidum (Curtis) P. Karst	Uşak-Eşme, September 2009	Ganodermataceae	Not edible/consumed as tea
3	Gyromitra esculenta (Pers. ex Pers.) Fr.	Uşak-Banaz, May 2009	Discinaceae	Edible after cooking
4	Lyophyllum decastes (Fries: Fries) Singer	Uşak-Banaz, November 2008	Lyophyllaceae	Edible
5	Pleurotus ostreotus (Jacq. ex Fr.) P.Kumm.	Uşak-Banaz, May 2008	Pleurotaceae	Edible

Spectral measurements and chemicals

Bioactivity measurements were carried out on a 96-well microplate reader, SpectraMax 340PC³⁸⁴ (Molecular Devices, Silicon Valley, CA). The measurements and calculations of the activity results were evaluated by using Softmax PRO v5.2 software (Molecular Devices, Silicon Valley, CA).

Methanol, n-hexane, pyrocatechol, quercetin ethanol, ferrous chloride, metaphosphoric acid, copper (II) chloride, ascorbic acid, 2,6-dichlorophenolindophenol, and ethylenediaminetetraacetic acid (EDTA) were purchased from E. Merck (Darmstadt, Germany). Polyoxyethylene sorbitan monopalmitate (Tween-40), Folin–Ciocalteu’s reagent (FCR), butylatedhydroxyl anisole (BHA), neocuproine, β-carotene, linoleic acid, α-tocopherol, 1,1-diphenyl-2-picryl-hydrazyl (DPPH), 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) salt 3-(2-pyridyl)-5,6-di(2-furyl)-1,2,4-triazine-5′,5′′-disulfonic acid disodium salt (Ferene), acetylcholinesterase (AChE) from electric eel (Type-VI-S, EC 3.1.1.7, 425.84 U/mg, Sigma, St. Louis, MO), butyrylcholinesterase (BChE) from horse serum (EC 3.1.1.8, 11.4 U/mg, Sigma, St. Louis, MO), 5,5′-dithiobis (2-nitrobenzoic) acid (DTNB), galantamine, acetylthiocholine iodide, and butyrylthiocholine chloride were purchased from Sigma Chemical Co. (Sigma-Aldrich GmbH, Sternheim, Germany). All other chemicals and solvents were of analytical grade.

Extraction

Each mushroom species was extracted separately with 2.5 L hexane, ethyl acetate, and methanol (4 × 24 h), successively, at room temperature (25 °C), filtered, and evaporated to dryness in vacuo. The crude extracts were stored in a refrigerator until required.

Determination of total bioactive compounds

The phenolic content in methanolic extracts of five wild mushroom species was determined with FCR according to the method of Slinkard and Singleton (1977) as described in the literature and expressed as micrograms of pyrocatechol equivalents per g extract (PEs). The following standard curve which was obtained by pyrocatechol standard was used to calculate total phenolic contents: The aluminum nitrate method was used to determine the flavonoid content in methanolic extracts (Öztürk et al., 2011) and the results were expressed as quercetin equivalents per g extract (QEs). The following standard curve obtained by quercetin standard was used to calculate total flavonoid contents: Ascorbic acid content in methanolic extracts was determined as ascorbic acid equivalents per g extract (AAEs). The method was based on the extraction of the sample with metaphosphoric acid and then the reaction with 2,6-dichlorophenolindophenol (Barros et al., 2008). The following standard curve, which was obtained by authentic ascorbic acid, was used to calculate the ascorbic acid content: β-Carotene and lycopene contents in methanol extracts were determined according to the method of Barros et al. (2008). The method was based on the measurement of the absorbance of methanol extract, solved in acetone–hexane mixture (6:4, v/v), at 453 (A₄₅₃), 505 (A₅₀₅), and 663 (A₆₆₃) nm wavelengths.

Contents of β-carotene and lycopene were calculated according to the following equations:

Determination of antioxidant activity

β-Carotene/linoleic acid assay

The lipid peroxidation inhibitory activity was evaluated using β-carotene/linoleic acid assay as previously described (Marco, 1968; Öztürk et al., 2011). Ethanol was used as a control while BHA and α-tocopherol were used as antioxidant standards. The results were given as 50% inhibition concentration, µg/mL (IC₅₀).

DPPH free radical scavenging assay

The free radical scavenging activity was determined spectrophotometrically by the DPPH assay described by Blois (1958) with a slight modification (Öztürk et al., 2011). Ethanol was used as a control while BHA and α-tocopherol were used as antioxidant standards. The results were given as IC₅₀ µg/mL.

ABTS cation radical decolorization assay

The ABTS^•+ scavenging activity was determined according to the method of Re et al. (1999) with slight modifications (Öztürk et al., 2011). Ethanol was used as a control while BHA and α-tocopherol were used as antioxidant standards. The results were given as IC₅₀ µg/mL.

Cupric reducing antioxidant capacity

The cupric reducing antioxidant capacity (CUPRAC) was determined according to the method of Apak et al. (2004) with slight modifications (Öztürk et al., 2011). BHA and α-tocopherol were used as antioxidant standards. Results were given as A_0.50, (µg/mL) that corresponds to the concentration providing 0.500 absorbance.

Metal-chelating activity

The metal-chelating activity of the extracts on Fe²⁺ was measured spectrophotometrically (Decker & Welch, 1990). Ethanol was used as a control, and EDTA was used as a chelating standard for comparison of the activity. The results were given as IC₅₀ µg/mL.

Determination of anticholinesterase activity

AChE and BChE inhibitory activities were measured by slightly modifying the spectrophotometric method developed by Ellman et al. (1961). AChE from electric eel (5.32 × 10⁻³ U in 20 µL phosphate buffer for each well) and BChE from horse serum (6.85 × 10⁻³ U in 20 µL phosphate buffer for each well) were used as enzymes, while acetylthiocholine iodide and butyrylthiocholine chloride were employed as substrates of the reaction. DTNB [5,5′-dithio-bis(2-nitrobenzoic) acid] was used for the measurement of the cholinesterase activity. All conditions were described in our earlier publication (Öztürk et al., 2011). Galantamine which is a standard drug for the treatment of mild Alzheimer's disease was used as a reference compound. The results were given as percentage inhibition against 25, 50, 100, and 200 µg/mL concentrations, the IC₅₀ was also given.

Statistical analysis

All data on antioxidant and anticholinesterase activities as well as bioactive contents were the averages of triplicate analyses. Data were recorded as mean ± S.E.M. Significant differences between means were determined by Student’s t-test, p values < 0.05 were regarded as significant.

Results and discussions

Total bioactive compounds

Table 2 presents total phenols and flavonoids, ascorbic acid, and carotenoids concentrations in the extracts obtained from the mushroom species. Total phenolic and flavonoid contents of the extracts of mushroom species determined as pyrocatechol equivalents (PEs) and quercetin equivalents (QEs), respectively. The G. lucidum afforded rich phenolic content, exhibiting 98.67 ± 0.32 mg PEs/g extract. The most flavonoid rich mushroom was found to be G. lucidum (160.38 ± 1.25 mg QEs/g extract), while G. esculanta (7.64 ± 0.14 mg QEs/g extract) was the poorest (Table 2).

Table 2. Total bioactive contents of methanol extract of five wild mushroom species^a.

Mushroom species	Total phenolic content (mg/g)	Total flavonoid content (mg/g)	Ascorbic acid (mg/g)	β-Carotene (mg/g)	Lycopene (mg/g)
Funalia trogii	78.03 ± 1.02	33.14 ± 0.05	4.33 ± 0.01	0.43 ± 0.02	0.23 ± 0.04
Ganoderma lucidum	98.67 ± 0.32	160.38 ± 1.25	5.60 ± 0.40	nd	0.67 ± 0.02
Gyromitra esculenta	28.03 ± 0.25	7.64 ± 0.14	2.86 ± 0.38	0.15 ± 0.01	0.07 ± 0.01
Lyophyllum decastes	7.76 ± 0.05	36.95 ± 0.74	4.77 ± 0.41	0.07 ± 0.03	0.03 ± 0.00
Pleurotus ostreotus	7.16 ± 0.08	14.18 ± 0.48	2.11 ± 0.48	0.11 ± 0.00	0.05 ± 0.00

^aValues expressed are means ± S.D. of three parallel measurements (p < 0.05). nd: not detected.

Ascorbic acid was found to be between 2.11 and 5.60 mg/g extract in the all mushroom species. β-Carotene and lycopene were found to be between 0.07–0.43 and 0.03–0.67 mg/g extract, respectively (Table 2).

Antioxidant activity

Table 3 shows the antioxidant activity of the extracts of mushroom species. β-Carotene-linoleic acid, DPPH^• scavenging, ABTS^•+ scavenging, CUPRAC, and metal-chelating assays were used to determine the antioxidant activity. BHA and α-tocopherol, and EDTA were the positive standards for comparison of the activity. The results were found to be statistically significant (p < 0.05) when compared with those of controls in each test. In general, the ethyl acetate extracts and the methanol extracts of the mushroom species exhibited good lipid peroxidation inhibitory activity. The highest activity was observed in ethyl acetate extract of G. lucidum (GLEA, IC₅₀: 1.55 ± 0.05 µg/mL), followed by ethyl acetate extract of F. trogii (FTEA, IC₅₀: 4.31 ± 0.18 µg/mL), the methanol extract of P. ostreotus (POM, IC₅₀: 32.5 ± 1.01 µg/mL) and the methanol extract of G. esculenta (IC₅₀: 40.2 ± 0.10 µg/mL) in β-carotene linoleic acid assay. The hexane extracts, however, showed weak activity. As shown in Table 3, the lipid peroxidation inhibition effect of the mushroom extracts and standards decreased in the following order: BHA > GLEA > α-tocopherol > FTEA > FTM > POM > GEM > GEEA > LDM > LDEA > GLM > POEA > LDH > FTH. To the best of our knowledge, G. lucidum is known as a good antioxidant source (Yegenoglu et al., 2011). When the activity of all mushroom species compared with those of antioxidant standards, none of the mushroom species except G. lucidum showed better activity than the standards (Table 3).

Table 3. Antioxidant activity of the extracts of five wild mushroom species by the β-carotene-linoleic acid, DPPH^•, ABTS^•+, CUPRAC, and metal chelating assays^a.

		Antioxidant activity
		β-Carotene-linoleic acid assay	DPPH^• assay	ABTS^•+ assay	CUPRAC assay	Metal chelating assay
Mushroom species	Extracts	IC50 (µg mL^−1)	IC50 (µg mL^−1)	IC50 (µg mL^−1)	A0.50 (µg mL^−1)	IC50 (µg mL^−1)
Funalia trogii	Hexane (FTH)	482.6 ± 1.25	>200	>200	285.8 ± 2.15	>200
	Ethyl acetate (FTEA)	4.31 ± 0.18	132.9 ± 1.37	43.4 ± 0.48	59.3 ± 1.87	47.7 ± 0.65
	Methanol (FTM)	32.2 ± 0.24	206.2 ± 1.23	98.8 ± 0.65	227.8 ± 2.01	>200
Ganoderma lucidum	Hexane (GLH)	>200	>200	>200	309.1 ± 2.76	>200
	Ethyl acetate (GLEA)	1.55 ± 0.05	49.1 ± 1.06	1.06 ± 0.05	19.2 ± 1.11	>200
	Methanol (GLM)	123.7 ± 1.69	47.6 ± 1.02	1.27 ± 0.07	20.7 ± 1.21	101.7 ± 0.78
Gyromitra esculenta	Hexane (GEH)	>200	>200	>200	247.6 ± 2.18	120.9 ± 0.89
	Ethyl acetate (GEEA)	52.7 ± 0.12	>200	197.3 ± 1.16	76.1 ± 1.98	>200
	Methanol (GEM)	40.2 ± 0.10	202.6 ± 1.18	88.7 ± 1.09	256.8 ± 2.14	262.9 ± 1.18
Lyophyllum decastes	Hexane (LDH)	316.2 ± 1.78	>200	>200	405.6 ± 2.87	376.5 ± 1.25
	Ethyl acetate (LDEA)	80.9 ± 1.24	>200	>200	114.3 ± 1.16	408.1 ± 1.36
	Methanol (LDM)	60.26 ± 1.45	>200	53.1 ± 0.63	440.1 ± 2.41	>200
Pleurotus ostreotus	Hexane (POH)	>200	>200	>200	236.4 ± 2.56	250.8 ± 1.17
	Ethyl acetate (POEA)	127.3±1.86	>200	142.1 ± 1.26	102.3 ± 1.05	>200
	Methanol (POM)	32.5 ± 1.01	>200	192.1 ± 1.63	433.3 ± 2.56	>200
Standards	BHA^b	1.34 ± 0.04	45.37 ± 0.47	4.10 ± 0.06	24.4 ± 0.68	NT
	α-Tocopherol^b	2.10 ± 0.08	7.31 ± 0.17	4.31 ± 0.10	89.4 ± 0.79	NT
	EDTA^b	NT	NT	NT	NT	3.47 ± 0.35

^aIC₅₀ values represent the means ± S.E.M. of three parallel measurements (p < 0.05).

^bReference compounds; NT, not tested; BHA, butylatedhydroxyl anisole; EDTA, ethylenediaminetetraacetic acid.

The results of DPPH^• and ABTS^•+ assays were also given in Table 3. In DPPH^• assay, the methanol (GLM) and the ethyl acetate extracts of G. lucidum (GLEA) exhibited close activity to those of antioxidant standards. The methanol extract of G. lucidum exhibited the highest activity (GLM, IC₅₀: 47.6 ± 1.02 µg/mL), followed by ethyl acetate extract of G. lucidum (GLEA, IC₅₀: 49.1 ± 1.06 µg/mL), and the ethyl acetate extract of F. trogii (FTEA, IC₅₀: 132.9 ± 1.37 µg/mL). In ABTS^• +assay, however, the ethyl acetate extract (GLEA, IC₅₀: 1.06 ± 0.05 µg/mL) and the methanol extract (GLM, IC₅₀: 1.27 ± 0.07 µg/mL) of G. lucidum also showed higher radical scavenging activity. For these extracts, the ABTS^•+ assay supported the DPPH^• assay almost in all extracts of mushroom species. The difference between the tested extracts and control was statistically significant (p < 0.05) in both antiradical assays. The scavenging effects of the mushroom extracts and standards on the DPPH radical decreased in the following order: α-tocopherol > BHA > GLM > GLEA > FTEA > GEM > FTM. As shown in Table 3, the scavenging effect on the ABTS cation radical, however, decreased in the following order: GLEA > GLM > BHA > α-tocopherol > FTEA > LDM > GEM > FTM > POEA > POM > GEEA.

The CUPRAC of the extracts of mushroom species was given in Table 3. The difference between the tested extracts and the control was statistically significant (p < 0.05) in the assay. The ethyl acetate extract of G. lucidum (GLEA, A_0.50: 19.2 ± 1.11 µg/mL) was found to be the best reductant followed by its methanol extract (GLM, A_0.50: 20.7 ± 1.21 µg/mL) and the ethyl acetate extract of F. trogii (FTEA, A_0.50: 59.3 ± 1.87 µg/mL). Moreover, the methanol and ethyl acetate extract of G. lucidum exhibited higher activity than those of antioxidant standards in CUPRAC and ABTS^•+ assays, which supported each other. The cupric reducing antioxidant capacity of the mushroom extracts and standards decreased in the following order: BHA >GLEA > GLM > FTEA > GEEA > α-tocopherol > POEA > LDEA > FTM > POH > GEH > GEM > FTH > GLH > LDH> POM > LDM.

Ferrous ions are considered as one of the effective pro-oxidants because the ferrous state of iron accelerates lipid oxidation by breaking down hydrogen and lipid peroxides to reactive free radicals through the Fenton reaction (Halliwell & Gutteridge, 1984). In fact, the reaction is very slow, and peroxidation accelerates when catalyzed by ferrous state iron. Table 3 shows the chelating effects of the extracts of mushroom species compared with EDTA on ferrous ions. The ethyl acetate extract of F. trogii (FTEA, IC₅₀: 47.7 ± 0.65 µg/mL) showed the highest metal-chelating activity among the other extracts studied. However, none of the extracts have comparable results with that of EDTA. As seen in Table 3, the metal-chelating effect of the extracts and EDTA decreased in the following order: EDTA > FTEA > GLM > GEH > POH > GEM > LDH > LDEA.

Anticholinesterase activity

Table 4 shows the AChE and BChE inhibitory activities of the extracts compared with that of galantamine. Against AChE enzyme, the ethyl acetate extract of F. trogii (FTEA, IC₅₀: 94.6 ± 1.03 µg/mL) exhibited significant activity, followed by ethyl acetate extract of G. lucidum (GLEA, IC₅₀: 174.3 ± 1.39 µg/mL). Against BChE enzyme, however, the most active extract was the methanol extract of G. lucidum (GLM, IC₅₀: 121.8 ± 1.54 µg/mL). At the same conditions, the IC₅₀ values of galantamine were 5.0 ± 0.1 and 11.6 ± 0.9 against AChE and BChE, respectively. In general, the studied mushroom species exhibited mild inhibitory activity against both enzymes. However, F. trogii possessed interesting AChE inhibitory activity. Therefore, in order to obtain AChE inhibitory compounds, isolation studies by bioactivity-guided fractionation are needed.

Table 4. Acetyl- and butyryl-cholinesterase inhibitory activities of the extracts of five wild mushroom species^a.

Anticholinesterase activity
	Acetylcholinesterase activity						Butyrylcholinesterase activity
Mushroom species	Extract	25 µg	50 µg	100 µg	200 µg	IC50 (µg mL^−1)	25 µg	50 µg	100 µg	20 µg	IC50 (µg mL^−1)
Funalia trogii	Hexane	NA	NA	NA	NA	NA	NA	5.93 ± 1.05	6.13 ± 1.41	12.92 ± 1.04	>200
	Ethyl acetate	25.14 ± 1.25	26.41 ± 2.45	52.84 ± 2.49	61.78 ± 0.42	94.6 ± 1.03	NA	NA	NA	9.67 ± 1.49	>200
	Methanol	3.79 ± 0.32	16.72 ± 1.36	28.34 ± 2.70	52.87 ± 2.98	188.5 ± 1.65	NA	NA	NA	NA	NA
Ganoderma lucidum	Hexane	NA	NA	NA	NA	NA	2.77 ± 0.94	4.33 ± 0.90	9.91 ± 1.49	13.59 ± 1.51	>200
	Ethyl acetate	23.77 ± 1.22	34.20 ± 2.14	40.24 ± 1.65	53.37 ± 2.58	174.3 ± 1.39	NA	NA	3.97 ± 0.25	12.29 ± 0.66	>200
	Methanol	1.77 ± 0.51	3.97 ± 0.21	7.95 ± 1.67	8.44 ± 2.32	>200	22.64 ± 2.02	34.46 ± 2.68	48.55 ± 1.91	55.18 ± 1.09	121.8 ± 1.54
Gyromitra esculenta	Hexane	NA	NA	NA	NA	NA	4.72 ± 0.96	6.87 ± 2.25	11.36 ± 0.56	21.15 ± 0.03	>200
	Ethyl acetate	15.66 ± 3.17	19.85 ± 3.04	20.98 ± 3.01	21.42 ± 1.83	>200	14.29 ± 1.73	20.09 ± 0.17	24.59 ± 0.94	38.20 ± 1.60	286.7 ± 1.73
	Methanol	10.18 ± 2.27	16.36 ± 2.09	19.96 ± 2.71	24.52 ± 2.41	>200	NA	1.63 ± 0.42	11.82 ± 0.10	23.39 ± 0.79	>200
Lyophyllum decastes	Hexane	NA	NA	NA	NA	NA	3.28 ± 0.64	5.60 ± 1.13	7.82 ± 0.51	15.63 ± 0.39	>200
	Ethyl acetate	20.42 ± 1.04	22.74 ± 0.45	22.98 ± 1.77	24.46 ± 0.74	>200	8.50 ± 1.03	9.84 ± 1.13	11.39 ± 0.97	17.90 ± 0.61	>200
	Methanol	17.81 ± 2.11	18.29 ± 0.18	26.00 ± 2.55	28.06 ± 0.73	>200	NA	9.86 ± 1.90	18.55 ± 0.61	30.77 ± 0.32	>200
Pleurotus ostreotus	Hexane	NA	NA	NA	4.79 ± 0.02	>200	NA	6.47 ± 1.75	11.08 ± 1.61	19.04 ± 0.77	>200
	Ethyl acetate	NA	0.89 ± 0.01	2.60 ± 0.12	3.91 ± 0.15	>200	5.03 ± 1.19	6.85 ± 0.69	10.17 ± 0.35	17.55 ± 1.31	>200
	Methanol	NA	0.50 ± 0.03	4.71 ± 1.71	13.09 ± 1.47	>200	NA	NA	NA	5.60 ± 0.93	>200
Standard	Galantamin^b	75.99 ± 0.45	78.08 ± 1.08	78.76 ± 0.52	80.41 ± 0.98	5.0 ± 0.1	62.48 ± 0.02	66.47 ± 0.46	71.43 ± 0.06	76.90 ± 0.20	11.6 ± 0.9

NA, not active.

^aValues expressed are means ± S.E.M. of three parallel measurements (p < 0.05).

^bReference compounds.

Conclusion

The results showed the antioxidant and anticholinesterase importance of the mushroom species. Four of them are commonly consumed as edible mushrooms in Anatolia as well as in some parts of the world due to their delicious taste. Thus, consuming these species may help people to protect them against lipid peroxidation and free radical damage. In addition, their extracts will probably be used for the development of safe food products and additives. Moreover, the ethyl acetate extracts of F. trogii and G. lucidum, against acetylcholinesterase, and methanol extract of G. lucidum, and ethyl acetate extract of G. esculenta against butyrylcholinesterase, demonstrated mild cholinesterase inhibitory activity. In order to treat mild Alzheimer’s disease, these species may be useful as mild cholinesterase inhibitory agents, particularly against AChE. However, further studies, especially in vivo activity tests on extracts and isolated constituents, are needed.

Declaration of interest

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.