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Research Article

Distribution of Antioxidant Activities and Total Phenolic

Contents in Acetone, Ethanol, Water and Hot Water
Extracts from 20 Edible Mushrooms via Sequential
Extraction
Yuxi Wang and Baojun Xu*
Abstract
Department of Food Science and Technology Program,
Distribution of antioxidative properties and total phenolic content (TPC) of
Beijing Normal University-Hong Kong Baptist University,
20 commonly consumed edible mushrooms in four extraction solvents were
China
investigated. The results showed that aqueous extracts (water extracts and
*Corresponding author: Baojun Xu, Food Science and hot water extracts) possessed much higher antioxidant properties than organic
Technology Program, Beijing Normal University-Hong solvent extracts (acetone extracts and ethanol extracts). Among all extracts, the
Kong Baptist University United International College, water extract of Agaricus subrufescens presented the best antioxidant activity.
China, Tel.: +86 756 3620636; Fax: +86 756 3620882; Similar as antioxidants, the TPC values of the samples extracted with water and
E-mail: [email protected] hot water were much higher than those extracted with acetone and ethanol. In
general, water extract of Ophiocordyceps sinensis exhibited the highest TPC
Received: January 10, 2014; Accepted: February 10, 2014;
value among all extracts.
Published: February 17, 2014
Keywords: Antioxidant properties; Edible mushrooms; Extraction solvent;
Phenolics.

Introduction in 20 commonly consumed edible mushrooms in China. The study

compared the antioxidant activities of different mushroom species
Mushrooms have been a part of human diet in many regions of
and how different solvents affected the effectiveness of the extraction.
the world for centuries due to their organoleptic characteristics as
well as the nutritional values [1,2]. The consumption of mushrooms Materials and Methods
has even increased remarkably over the past few decades [3]. Mushroom samples
Generally, mushrooms provide low energy while usually contain large
One species (Ophiocordyceps sinensis) (fruiting bodies) was
amounts of dietary fibers, proteins, vitamins and minerals [4]. Some
produced in Shanghai Academy of Agricultural Sciences. Other
mushrooms are even consumed for medicinal purpose as they contain
mushroom samples (fruiting bodies) were purchased from several
valuable bioactive components, for example, Ganoderma lucidum [5].
supermarkets in Zhuhai and Harbin, China. Among the samples
Polysaccharides and polysaccharide-protein complexes extracted
collected, Boletus pinophilus was produced in Inner Mongolia, China;
from some edible mushrooms were found to have stimulation effects
Agrocybe aegerita and Grifola frondosa were produced in Zhejiang
on non-specific immune system and have anti-tumor properties by
Province, China; Pleurotus citrinopileatus and Pleurotus eryngii were
stimulating the defense system of the host in animal experiments [6].
produced in Fujian Province, China; Boletus edulis, Boletus luridus and
It has been proven that the polysaccharides contained in cell Boletus aereus were produced in Wutian, Yunnan Province, China;
wall of mushroom possess free radical scavenging properties [7]. In Auricularia auricular, Tremella fuciformis, Hypsizygus tessellates,
general, wild mushrooms contain significant amount of antioxidants Agaricus subrufescens, Hericium erinaceus, Coprinus comatus, Phallus
and phenols [8]. It has been widely accepted that mushrooms are a indusiatus, Pholiota nameko, Armillaria mellea, Hohenbuehelia
good source of nutrients including high level of antioxidants [9]. The reniformis and Lentinus edodes were all produced in Heilongjiang
intake of dietary antioxidants may help the prevention of free radical Province, China. All samples were dry mushroom and stored in a cool
damage in human body [10]. Antioxidants can scavenge free radicals place before treatment. The information of mushrooms was listed in
through the inhibition of the initiation process or interruption of Table 1.
the propagation process of lipid oxidation and provide preventive Chemicals and reagents
function by several actions [10].
Absolute acetone and absolute ethanol were offered by Guangzhou
The objective of this study was to determinate the antioxidative Chemical Reagent Factory (Guangzhou, China). 2-Diohenyl-1-
properties and the antioxidant-related composition of phenolics picryhydrazyl (DPPH) was supplied by Shanghai Yuanye Biological

Austin J Nutri Food Sci - Volume 2 Issue 1 - 2014 Citation: Wang Y, Xu B. Distribution of Antioxidant Activities and Total Phenolic Contents in Acetone, Ethanol,
Submit your Manuscript | www.austinpublishinggroup.org Water and Hot Water Extracts from 20 Edible Mushrooms via Sequential Extraction. Austin J Nutri Food Sci.
Xu et al. © All rights are reserved 2014;2(1): 5.
Xu Austin Publishing Group

Table 1: Scientific name, common name and sources of mushroom samples.

No. Common name Scientific name Mode of production Place of origin

1 Jew’s ear Auricularia auricula Cultivated Suifenhe, Heilongjiang Province

2 Tremella Tremella fuciformis Cultivated Suifenhe, Heilongjiang Province

3 Shimeji Hypsizygus tessellatus Cultivated Suifenhe, Heilongjiang Province

4 Princess matsutake Agaricus subrufescens Cultivated Suifenhe, Heilongjiang Province

5 Lion's mane mushroom Hericium erinaceus Cultivated Suifenhe, Heilongjiang Province

6 Shaggy ink cap Coprinus comatus Cultivated Suifenhe, Heilongjiang Province

7 Bamboo fungus Phallus indusiatus Cultivated Suifenhe, Heilongjiang Province

8 Golden oyster mushroom Pleurotus citrinopileatus Cultivated Gutian, Fujian Province

9 Apricot oyster mushroom Pleurotus eryngii Cultivated Gutian, Fujian Province

10 Porcino Boletus edulis Cultivated Wuding, Yunnan Province

11 Lurid bolete Boletus luridus Cultivated Wuding, Yunnan Province

12 Porcino nero Boletus aereus Cultivated Wuding, Yunnan Province

13 Caterpillar fungus Ophiocordyceps sinensis Cultivated Shanghai

14 Nameko Pholiota nameko Cultivated Shangzhi, Heilongjiang Province

15 Stump mushroom Armillaria mellea Cultivated Shangzhi, Heilongjiang Province

16 Winter mushroom Hohenbuehelia reniformis Cultivated Shangzhi, Heilongjiang Province

17 Shiitake Lentinus edodes Cultivated Shangzhi, Heilongjiang Province

18 Pine bolete Boletus pinophilus Wild Chifeng, Inner Mongolia

19 Poplar mushroom Agrocybe aegerita Cultivated Lishui, Zhejiang Province

20 Maitake Grifola frondosa Cultivated Lishui, Zhejiang Province

Technology Co., Ltd (Shanghai, China). Folin-Ciocalteu reagent determination of aqueous extracts, 0.2 mL of each water extract, hot
was provided by Shanghai Sanjie Biotechnology Co., Ltd (Shanghai, water extract, blank and standard solutions was added into a pre-
China). Trolox was purchased from Aldrich Co. (St. Louis, MO, labeled centrifuge tube and mixed with 3.8 mL of DPPH solution. The
mixtures were mixed evenly by vortexing and stood in the dark at
U.S.A.). Sodium carbonate was supplied by Tianjin Nuoke Technology
room temperature for 20 min. Then the mixtures were centrifuged
Development Co., Ltd (Tianjin, China). Gallic acid was purchased by a centrifuge (Weierkang Xiangying Centrifuge Co., Ltd, Changsha,
from Tianjin Damao Chemical Reagent Co., Ltd (Tianjin, China). All Hunan, China) at 3000 rpm for 10 min. The absorbance of each
the chemicals were of analytical grade. mixture was measured by an UV-Visible spectrophotometer (TI-
1901, Beijing Purkinje General Instrument Co., Ltd, Beijing, China)
Extraction of mushroom samples
at 517 nm against solvent blank. The results were expressed as Trolox
Each of the twenty dry mushroom samples was pulverized; 5.0 equivalents (µmole of TE/g sample) in accordance to the standard
g of each mushroom powder was accurately weighed into a set of curve of Trolox.
centrifuge tubes (each of the sample prepared in triplicate); 40 mL
of absolute acetone was added into each tube for the preparation of Determination of total phenolic content (TPC)
acetone extracts. The samples were extracted for 24 hours and then For determination of water extracts, 50 µL of each water extract,
centrifuged by a centrifuge (Weierkang Xiangying Centrifuge Co., Ltd, hot water extract, blank and standard solution was mixed with 3 mL
Changsha, Hunan, China) at 8000 rpm for 10 min. The supernatants of distilled water, 250 µL of Folin-Ciocalteu reagent and 750 µL of 7%
were transferred into new centrifuge tubes and stored at -20 degree Na2CO3 solution and the mixtures were mixed evenly by vortexing.
C. The residues were dried by vaporizing and collected; 40 mL of After standing at room temperature for 8 min, 950 µL of distilled water
absolute ethanol was added into the dry residues for the preparation was added into each mixture and the mixtures were allowed to stand
of ethanol extracts. The same extraction procedures were repeated to at room temperature for 1 hour. For organic solvent extraction, 500 µL
prepare water extracts as acetone and ethanol extraction and finally of each acetone extract, ethanol extract, blank and standard solution
hot water extraction was done at 99°C in water bath for 2 hours. All was mixed with 2.55 mL of distilled water, 250 µL of Folin-Ciocalteu
the extracts were stored at -20°C in the dark for further usage. reagent and 750 µL of 7% Na2CO3 solution. The absorbance of each
mixture was measured by the UV-Visible spectrophotometer (TI-
Determination of antioxidant properties 1901, Beijing Purkinje General Instrument Co., Ltd, Beijing, China)
For determination of organic extracts, 1 mL of each acetone at 765 nm by using distilled water as the blank. The total phenolic
extract, ethanol extract, blank and standard solutions was added into content (TPC) was expressed as gallic acid equivalents (mg of GAE/g
a pre-labeled test tube and mixed with 3 mL of DPPH solution. For sample) in accordance to the standard curve of gallic acid.

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Statistical analysis Results and Discussion

The data obtained were analyzed by ANOVA with the use of SPSS
Antioxidant properties of various extracts from
Statistics (Version 17.0, SPSS Inc., Chicago, IL, U.S.A.). Duncan’s mushroom
multiple range test was used to test whether there was any significant
difference in antioxidant activity or total phenolic content among Free radical scavenging is one of the mechanisms involved
in inhibiting lipid oxidation; therefore it is normally used for
different mushroom species.

Table 2: Antioxidant activities of mushroom samples extracted by different solvents.

DPPH free radical scavenging capacities (µmole TE/g)

No. Scientific name
Acetone extracts Ethanol extracts Water extracts Hot water extracts
1 Auricularia auricula 0.57 ± 0.02g 0.35±0.02i 2.42±0.09h 2.18±0.08j
2 Tremella fuciformis 0.28 ± 0.02m 0.17±0.02k 3.62±0.07d 3.66±0.15f
3 Hypsizygus tessellatus 0.37 ± 0.03kl 0.24±0.01j 1.92±0.06j 3.72±0.12f
4 Agaricus subrufescens 1.04 ± 0.01e 1.18±0.02c 4.94±0.09a 3.76±0.08ef
5 Hericium erinaceus 0.39 ± 0.01jk 0.39±0.03h 2.85±0.06f 2.28±0.14ij
6 Coprinus comatus 0.53 ± 0.04gh 0.71±0.04e 4.88±0.11a 4.07±0.08d
7 Phallus indusiatus 1.21 ± 0.06d 1.53±0.04b 2.35±0.12h 2.17±0.02j
8 Pleurotus citrinopileatus 0.37 ± 0.01k 0.47±0.02fg 4.11±0.10c 3.76±0.20ef
9 Pleurotus eryngii 0.39 ± 0.04jk 0.48±0.05f 3.72±0.07d 4.45±0.13ab
10 Boletus edulis 1.75 ± 0.06b 2.18±0.04a 4.58±0.04b 4.19±0.12cd
11 Boletus luridus 1.79 ± 0.03b 2.16±0.02a 4.82±0.23a 2.84±0.21h
12 Boletus aereus 1.93 ± 0.09a 2.20±0.02a 4.87±0.12a 4.33±0.10bc
13 Ophiocordyceps sinensis 1.29 ± 0.01c 0.95±0.03d 2.68±0.09g 3.40±0.15g
14 Pholiota nameko 0.50 ± 0.03hi 0.43±0.03gh 2.13±0.08i 3.99±0.18de
15 Armillaria mellea 0.31 ± 0.02lm 0.43±0.03fgh 2.50±0.05h 3.78±0.19ef
16 Hohenbuehelia reniformis 0.30 ± 0.03m 0.05±0.01l 1.32±0.06l 1.74±0.04k
17 Lentinus edodes 0.40 ± 0.04jk 0.28±0.02j 1.40±0.09l 2.93±0.05h
18 Boletus pinophilus 0.27 ± 0.01m 1.52±0.01b 3.75±0.08d 3.29±0.08g
19 Agrocybe aegerita 0.66 ± 0.03f 1.51±0.03b 3.32±0.06e 4.64±0.20a

20 Grifola frondosa 0.44 ± 0.03ij 0.41±0.02h 1.75±0.04k 2.47±0.10i

Note: Data were expressed as mean ± standard deviation (n = 3), small letters indicate significantly different within the species at 95% confident interval (p < 0.05).

Table 3: Total phenolic content of mushroom samples extracted by different solvents

Total phenolic content (mg GAE/g)
No. Scientific name
Acetone extracts Ethanol extracts Water extracts Hot water extracts
1 Auricularia auricula 0.12 ± 0.01h 0.09 ± 0.01i 0.82 ± 0.04o 0.35 ± 0.07m
2 Tremella fuciformis 0.11 ± 0.01hi 0.06 ± 0.01j 0.81 ± 0.06o 0.66 ± 0.03l
3 Hypsizygus tessellatus 0.09 ± 0.01i 0.07 ± 0.01j 2.16 ± 0.03n 2.37 ± 0.06g
4 Agaricus subrufescens 0.25 ± 0.01d 0.30 ± 0.01d 2.82 ± 0.01i 1.01 ± 0.03k
5 Hericium erinaceus 0.16 ± 0.01g 0.13 ± 0.01h 3.08 ± 0.09k 1.75 ± 0.08i
6 Coprinus comatus 0.19 ± 0.01f 0.18 ± 0.01g 2.41 ± 0.06m 4.12 ± 0.05b
7 Phallus indusiatus 0.28 ± 0.01c 0.27 ± 0.01e 3.21 ± 0.16k 2.58 ± 0.02f
8 Pleurotus citrinopileatus 0.10 ± 0.01i 0.10 ± 0.01i 9.42 ± 0.16d 2.96 ± 0.10d
9 Pleurotus eryngii 0.09 ± 0.01j 0.07 ± 0.01j 3.65 ± 0.11j 1.52 ± 0.06j
10 Boletus edulis 0.28 ± 0.01c 0.30 ± 0.01d 5.17 ± 0.09g 3.98 ± 0.15bc
11 Boletus luridus 0.36 ± 0.01b 0.38 ± 0.01b 10.43 ± 0.01b 2.72 ± 0.11ef
12 Boletus aereus 0.29 ± 0.01c 0.32 ± 0.01c 10.18 ± 0.13c 3.95 ± 0.07c
13 Ophiocordyceps sinensis 0.22 ± 0.01e 0.24 ± 0.01f 10.82 ± 0.19a 9.44 ± 0.24a
14 Pholiota nameko 0.06 ± 0.01j 0.06 ± 0.01j 7.31 ± 0.09e 2.15 ± 0.10h
15 Armillaria mellea 0.10 ± 0.01hi 0.11 ± 0.01i 4.81 ± 0.09n 2.13 ± 0.10h
16 Hohenbuehelia reniformis 0.06 ± 0.01j 0.03 ± 0.01k 2.35 ± 0.04m 1.37 ± 0.07j
17 Lentinus edodes 0.06 ± 0.01j 0.06 ± 0.01j 4.26 ± 0.01i 1.82 ± 0.01i
18 Boletus pinophilus 0.91 ± 0.06a 0.62 ± 0.01a 4.26 ± 0.07i 2.86 ± 0.10de
19 Agrocybe aegerita 0.13 ± 0.01h 0.27 ± 0.01e 5.71 ± 0.07f 2.99 ± 0.09d
20 Grifola frondosa 0.11 ± 0.01hi 0.11 ± 0.01i 3.78 ± 0.12j 2.34 ± 0.09g

Note: Data were expressed as mean ± standard deviation (n = 3) with small letters indicate significantly different within the species at 95% confident interval (p < 0.05).

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determination of antioxidant activity [11]. The DPPH free radical

scavenging activity of extracts from 20 different mushrooms was
tested. DPPH provides a violet color when dissolves in ethanol and
the discoloration occurs when antioxidants donate protons to DPPH
[11,12].
Antioxidant capacities of the 80 extracts from 20 different
mushrooms with four different extraction solvents were presented
in Table 2. Three mushroom species (Boletus edulis, B. luridus and
B. aereus) provided a relative higher antioxidant activity for all four
extraction solvents as compared to the other 17 species. Figure 1
presented antioxidant activities of various extracts from mushrooms
B. edulis, B. luridus and B. aereus together with their total phenolic
contents. In this study, the results showed that the extracts of
genus Boletus provided higher antioxidant activities comparing to
mushrooms of other genuses.
For all mushrooms, the water extracts and hot water extracts
exhibited much higher antioxidant properties than that of acetone
extracts and ethanol extracts. However, the differences between water
extracts and hot water extracts and the differences between acetone
extracts and ethanol extracts were not significant (p > 0.05). Among
all the extracts, the water extracts of Agaricus subrufescens provided
the highest antioxidant activity with 4.94 µmole of TE/g sample.
Total phenolic content of mushrooms
Phenols are known as important botanical constituents due
to their scavenging ability provided by the hydroxyl groups [13].
Phenolic compounds may have direct contribution to antioxidative
action [14]. Phenolic compounds were reported to be associated with
antioxidant activity and play important roles in the stabilizing of lipid
peroxidation [15]. Natural phenolics are able to provide antioxidative
function through various ways, such as intercepting singlet oxygen,
decomposing primary products of oxidation, preventing continued
hydrogen abstraction from substances, etc [16]. In addition, total
polyphenols were considered as the major naturally occurring
Figure 1: Antioxidant activities (A) and total phenolic contents (B) of various antioxidant compounds in the wild edible mushrooms [17].
extracts from mushrooms B. edulis, B. luridus and B. aereus.
The results of TPC values of mushrooms were shown in Table
3. The TPC of the samples extracted with water and hot water were
much higher than those extracted with acetone and ethanol. Among
the four different solvents, water extraction provided the highest TPC.
There were no significant differences between acetone extracts and
ethanol extracts; they presented in low amounts. Meanwhile, there
were significant (p < 0.05) differences between different mushrooms.
In organic extracts, Boletus pinophilus provided the highest amount
of TPC with 0.91 mg GAE/g in acetone extracts and 0.62 mg GAE/g
in ethanol extracts. However, comparing to other mushroom
samples, it only provided medium TPC in aqueous extracts with
4.26 mg GAE/g in water extracts and 2.86 mg GAE/g in hot water
extracts. In inorganic extracts, water extract and hot water extracts
of Ophiocordyceps sinensis had the highest amount of TPC with 10.82
mg GAE/g and 9.44 mg GAE/g. The distribution of TPC in different
extraction solvents depended on species of the mushrooms.
The correlations of phenolic levels and antioxidant capacities
in triplicate aqueous extracts were shown in Figure 2. According to
Figure 2: Correlation between antioxidant activities and TPC values of water the results, the overall trend was that the antioxidant capacities were
extracts and hot water extracts positively related to the phenolic levels. However, as the values of the

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correlation analysis were 0.31 and 0.24 for water extracts and hot 6. Reshetnikov SV, Wasser SP and Tan KK. Higher basidiomycetes as a source
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Austin J Nutri Food Sci - Volume 2 Issue 1 - 2014 Citation: Wang Y, Xu B. Distribution of Antioxidant Activities and Total Phenolic Contents in Acetone, Ethanol,
Submit your Manuscript | www.austinpublishinggroup.org Water and Hot Water Extracts from 20 Edible Mushrooms via Sequential Extraction. Austin J Nutri Food Sci.
Xu et al. © All rights are reserved 2014;2(1): 5.

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