Oxidative Medicine and Cellular Longevity

Oxidative Medicine and Cellular Longevity / 2017 / Article
Special Issue

Role of Natural Antioxidants from Functional Foods in Neurodegenerative and Metabolic Disorders

View this Special Issue

Research Article | Open Access

Volume 2017 |Article ID 5863523 | 12 pages | https://doi.org/10.1155/2017/5863523

Antioxidant and Hepatoprotective Activities of Polysaccharides from Spent Mushroom Substrates (Laetiporus sulphureus) in Acute Alcohol-Induced Mice

Academic Editor: José C. T. Carvalho
Received07 Aug 2017
Revised11 Oct 2017
Accepted26 Oct 2017
Published21 Dec 2017

Abstract

In order to contribute to the exploitation and utilization of spent mushroom substrates (SMS) of Laetiporus sulphureus, hot-water-extractable polysaccharides (H-SMPS) and enzymatic-extractable polysaccharides (E-SMPS) were successfully isolated from SMS of L. sulphureus. Both H-SMPS and E-SMPS were found to have high reducing power and potential scavenging activities against hydroxyl, DPPH, and superoxide anion radicals. In vivo assays showed that the administration of H-SMPS and E-SMPS has potential hepatoprotective effects against alcohol-induced alcoholic liver disease (ALD), possibly brought about by improving liver function, increasing antioxidant status, and reducing lipid peroxidation. Furthermore, monosaccharide composition analysis showed that fucose may play a vital role in guaranteeing stronger hepatoprotection. These results may provide references for the exploitation of the SMS of L. sulphureus as a source of H-SMPS and E-SMPS, which in turn can be used as functional foods or natural drugs for the prevention of ALD and other liver diseases.

1. Introduction

With the prevalence of poor diet structures in China, increasing alcohol consumption has been demonstrated to be involved in the increasing incidence of alcoholic liver disease (ALD), which is a progressive disease that could subsequently induce hepatitis, hepatic fibrosis, cirrhosis, and liver cancer [13]. Furthermore, ALD could also cause other clinical diseases including hypertension, hyperlipidemia, and atherosclerosis [4, 5]. Alcohol metabolism mainly occurs in the liver, and many studies have indicated that many in vivo enzymes, such as alcohol dehydrogenase (ADH), aldehyde dehydrogenase (ALDH), and cytochrome P4502E1 (CYP2E1), are involved in alcohol metabolism [6, 7]. The toxicity of acute alcohol consumption in the liver is possibly due to the inhibition of ADH and ALDH activities, as well as the increase in CYP2E1 activity, which is accompanied by the excessive formation of reactive oxygen species (ROS) [5, 8]. Increasing amounts of evidence indicate that ROS could oxidize biological membranes, proteins, nucleic acids, and other biological macromolecules, thereby damaging cellular integrity and functionality [9]. At present, the mechanisms underlying acute alcohol metabolism and its metabolic consequences are still poorly understood. The traditional therapies for ALD are mainly abstinence and the administration of nutritional supplements and corticosteroid substances; however, their therapeutic effects have been proven to be poor [10]. Recently, natural substances from edible and medicinal mushrooms have attracted attention as potential drugs in preventing and treating alcohol-induced liver injury [11]. Hence, it seems necessary and significant to explore natural and nontoxic therapeutic strategies for the treatment of ALD.

It has been shown that more than five million tons of spent mushroom substrates (SMS), the cultivation matrix that remains after mushroom cultivations, are produced in China annually with low utilization, causing environmental pollution and resource waste [12]. SMS contains residual mycelia and fruiting bodies of mushrooms; trace elements including Fe, Ca, Zn, and Mg; active enzymes such as cellulose, hemicellulose, and ligninase; and biomacromolecules including polysaccharides, proteins, and lipids [12, 13]. Polysaccharides extracted from SMS have received attention owing to their abundant pharmacological activities including antioxidation, hepatoprotection, and antihyperlipidemia [14, 15]. Additionally, these polysaccharides possess less toxic side effects and are more accessible locally and more environment friendly, compared to synthetic substances [14].

Laetiporus sulphureus, an edible and medicinal fungus belonging to Basidiomycetes, is widely distributed in Asia, Europe, and North America [16]. Documents in literature have indicated that intracellular polysaccharides and exopolysaccharides from L. sulphureus show antioxidant, anti-inflammatory, and antitumor activities [1719]. However, studies on the hepatoprotective activities of polysaccharides isolated from L. sulphureus SMS are scarce. In this work, the hepatoprotective effects and antioxidant activities of hot-water-extractable SMPS (H-SMPS) and enzymatic-extractable SMPS (E-SMPS) from L. sulphureus SMS in acute alcohol-induced ALD mice were investigated.

2. Materials and Methods

2.1. Materials and Chemicals

SMS of L. sulphureus was obtained from Tai’an Agricultural Sciences Academy (Tai’an, China). Kunming strain mice (male) were purchased from Taibang Biologic Products Co. Ltd., Tai’an, China. Monosaccharide standard samples were provided by Sigma Chemicals Co. Ltd., St. Louis, USA The diagnostic kits for analyzing the activities of ADH, ALDH, CYP2E1, superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT), as well as the levels of malondialdehyde (MDA), lipid peroxidation (LPO), total cholesterol (TC), and triglyceride (TG), were purchased from Jiangsu Meibiao Biological Technology Co. Ltd., Jiangsu, China. All other reagents used in this experiment were provided by local chemical suppliers.

2.2. Preparation of H-SMPS and E-SMPS

The SMS of L. sulphureus was dried at 55°C and ground into powder. H-SMPS and E-SMPS were extracted by incubating the SMS in deionized water (90°C, 8 h) and snailase solutions (4% w/v, 37°C, 6 h), respectively. After centrifugation (3000 rpm, 10 min), the supernatant was collected, concentrated, and precipitated by incubation with three volumes of 95% v/v ethanol at 4°C overnight. The precipitate was deproteinated by the Sevage method [20] and dialyzed against deionized water.

2.3. Animal Experiments

Animal procedures were performed in accordance with the institutional ethical guidelines for animals’ welfare of Shandong Agricultural University Committee. Seventy male Kunming strain mice (20 ± 2 g) were housed in an animal room under standardized environmental conditions (temperature of 23 ± 2°C, a relative humidity of 50 ± 5%, and a 12/12 h light/dark cycle) for 5 days with free access to food and water.

After domestication, the mice were randomly divided into seven groups of ten mice each. These groups were the normal control (NC) group, model control (MC) group, positive control (PC) group (received bifendatatum at 150 mg/kg), H-SMPS high-dose group (400 mg/kg), H-SMPS low-dose group (100 mg/kg), E-SMPS high-dose group (400 mg/kg), and E-SMPS low-dose group (100 mg/kg). The mice in the NC and MC groups received isometric normal saline until the 25th day as a blank. On the 26th day, all mice except those in the NC group received alcohol (50% v/v) at a dose of 12 mL/kg by gavages. All mice were sacrificed quickly by euthanasia overnight after inducing toxicity with alcohol. The animal experiment design thought was in accordance with previous literature [6, 21, 22].

Blood samples were collected and centrifuged (10,000 rpm, 10 min, 4°C) to obtain the sera. Alanine aminotransferase (ALT) activity and aspartate aminotransferase (AST) activity in the sera were measured using an automatic biochemical analyzer (BS-380, Shenzhen, China).

The liver was surgically removed, weighed, and homogenized (10% w/v) in phosphate buffer solution (0.2 M, pH 7.4). After centrifugation (3000 rpm, 10 min, 4°C), the supernatant was collected for further biochemical assay. The hepatic activities of ADH, ALDH, CYP2E1, SOD, GSH-Px, and CAT, as well as the levels of MDA, LPO, TC, and TG, were determined using commercial reagent kits following the manufacturer’s instructions. The liver samples used for histopathological observations were soaked in formalin (10% w/v). Thin sections (4-5 μm thickness) were acquired using a microtome and stained with hematoxylin and eosin. Hepatic histopathological changes were observed and photographed using a microscope under ×400 magnification.

2.4. In Vitro Antioxidant Analysis

The reducing powers of H-SMPS and E-SMPS were determined using the methods reported by Oyaizu [23]. The reaction system, which contained 1 mL of either of the polysaccharides (0–800 μg/mL), 2.5 mL of phosphate buffer (0.2 M, pH 6.6), and 1.0 mL of potassium ferricyanide (1% w/v), was kept warm in a water bath at 50°C for 20 min. The reaction was terminated by adding 2 mL of trichloroacetic acid (10% w/v) and 1.2 mL of ferric chloride (0.1% w/v). The absorbance was measured at 700 nm using deionized water as a blank control and butylated hydroxytoluene (BHT) as a positive control.

The scavenging activities of H-SMPS and E-SMPS against hydroxyl radicals were measured according to a reported method [24]. The reaction mixture, composed of 1 mL of ferrous sulfate (9 mM), 1 mL of salicylic acid (9 mM), 1 mL of either of the polysaccharides (0–800 μg/mL), and 1 mL of hydrogen peroxide (8.8 mM), was incubated for 30 min at 37°C. After centrifugation (12,000 rpm, 6 min), the absorbance was measured at 510 nm using deionized water as a blank control and BHT as a positive control. The scavenging rate was calculated using the following formula: where is the absorbance of the blank and is the absorbance of the polysaccharides or BHT.

The DPPH radical scavenging activities of H-SMPS and E-SMPS were measured using the method reported by Cheng et al. [25]. The reaction mixture, which contained either of the polysaccharides (0.2 mL, 0–800 μg/mL) and DPPH solution (0.6 mL, 0.004% w/v in methanol), was stored in the dark for 30 min without moving. The absorbance was measured at 517 nm using deionized water as a blank control and BHT as a positive control. The scavenging rate was evaluated as follows: where is the absorbance of the blank control and is the absorbance of the polysaccharides or BHT.

Superoxide anion radical scavenging activities were determined using the method reported by Wei et al. [26]. The reaction system, composed of 1 mL of either of the polysaccharides (0–800 μg/mL) and 2 mL of Tris-HCl buffer (50 mM, pH 8.2), was incubated for 20 min at 25°C. After incubation, 0.4 mL of 1,2,3-phentriol (5 mM) was added to terminate the reaction. The absorbance was measured at 325 nm using deionized water as a blank control and BHT as a positive control. The scavenging rate was calculated using the following equation: where is the absorbance of the blank and is the absorbance of polysaccharides or BHT.

The IC50 values (μg/mL) were defined as the effective concentrations of the sample at which the radicals were inhibited by 50%.

2.5. Monosaccharide Composition Determination

Monosaccharide compositions of H-SMPS and E-SMPS were analyzed by gas chromatography (GC-2010, Shimadzu, Japan) equipped with a flame ionization detector (FID) according to a reported method [27]. Briefly, 0.1 g of the sample was hydrolyzed with 1.8 mL of trifluoroacetic acid (2 M) at 100°C for 4 h. The resulting hydrolysate was neutralized with 0.6 mL of ammonium hydroxide (12 M), restored with 0.4 mL of sodium borohydride in ammonium hydroxide (2% w/v), and acetylated with 0.6 mL of methylimidazole and 4 mL of acetic anhydride. After centrifugation (3000 rpm, 10 min), 1 μL of the supernatant was injected into a capillary column of Rtx-1 (30 mm × 0.25 mm × 0.25 μm). Monosaccharide components were evaluated based on the standard curves of rhamnose, arabinose, xylose, mannose, galactose, glucose, ribose, and fucose.

2.6. Statistical Analysis

The software SAS was used to perform statistical evaluation. Data are expressed as mean ± SD (standard deviations). One-way ANOVA was performed to analyze the data. Significant differences between experimental groups were determined using Tukey’s tests. was considered as the threshold for statistical significance.

3. Results

3.1. Effects of H-SMPS and E-SMPS on AST and ALT Activities in Sera

The mice in the MC group showed abnormally higher serum AST and ALT activities than the mice in the NC group (, Figure 1), indicating that alcohol-induced liver injury was successfully established in mice. Interestingly, the elevation of AST and ALT activities could be attenuated by supplementation of H-SMPS and E-SMPS, especially at high doses. Compared to mice in the MC group, the serum AST and ALT activities of mice in the H-SMPS high-dose group decreased by 28.29% and 28.0%, respectively, while those of mice in the E-SMPS high-dose group decreased by 36.6% and 50.0% of mice, respectively. In addition, bifendatatum was also observed to decrease the serum enzyme activities. These results demonstrate that both H-SMPS and E-SMPS have the potential to suppress acute alcohol-induced elevation of AST and ALT activities in order to maintain liver function.

3.2. Effects of H-SMPS and E-SMPS on Hepatic Lipid Properties

As shown in Figure 2, the mice in the MC group exhibited higher hepatic TG and TC levels than the mice in the NC group (), indicating that hepatic lipid metabolic disturbance was induced by acute alcohol toxicity. The administration of H-SMPS and E-SMPS attenuated the elevation of TG and TC levels, especially with E-SMPS administered at a high dose. Briefly, the hepatic TG and TC levels in mice treated with a high dose of E-SMPS reached 1.25 ± 0.14 and 3.7 ± 0.18 mmol/mg prot, respectively, which were lower than those observed in the H-SMPS group at the same dose (TG: 1.54 ± 0.11 mmol/mg prot and TC: 4.8 ± 0.16 mmol/mg prot), and were almost equal to those in the NC group (TG: 1.12 ± 0.13 mmol/mg prot and TC: 3.5 ± 0.16 mmol/mg prot). The present results indicate that both H-SMPS and E-SMPS could potentially restore lipid metabolism to its state before acute alcohol-induced toxicity.

3.3. Effects of H-SMPS and E-SMPS on Hepatic ADH and ALDH Activities and CYP2E1 Level

The hepatic ADH and ALDH activities and CYP2E1 levels are shown in Figure 3. Compared with mice in the NC group, the mice in the MC group had decreased ADH and ALDH activities and increased CYP2E1 levels (), indicating that alcohol metabolism was partly suppressed in the alcohol dehydrogenase oxidation system and accelerated in the microsome alcohol oxidation system in response to acute alcohol-induced toxicity. Interestingly, the remarkable elevation in ADH and ALDH activities and reduction in CYP2E1 levels could be observed when H-SMPS and E-SMPS are administered at the tested dosages (). The ADH and ALDH activities of mice treated with a high dose of E-SMPS reached maximum values of 58.9 ± 3.4 and 18.5 ± 1.9 U/mg prot, respectively, which were higher than those of mice in the H-SMPS group (ADH: 55.3 ± 2.8 U/mg prot and ALDH: 15.9 ± 2.1 U/mg prot) at the same dose. Additionally, compared with the CYP2E1 levels in the H-SMPS high-dose group (64.30 ± 2.72 ng/mL), the CYP2E1 levels in the E-SMPS high-dose group were lower (56.73 ± 2.98 ng/mL).

Meanwhile, bifendatatum-treated mice also exhibited an increase in the activities of ADH and ALDH, as well as an attenuation in the CYP2E1 levels when compared with the MC group mice ().

3.4. Effects of H-SMPS and E-SMPS on Hepatic Antioxidative Enzymes

Compared with the mice in the NC group, the mice in the MC group were observed to have an abnormal reduction in hepatic SOD, GSH-Px, and CAT activities (, Figures 4(a)4(c)), indicating that the hepatic antioxidative defense had been damaged by the administration of alcohol. The changes in these parameters were ameliorated by treatment with H-SMPS, E-SMPS, and bifendatatum at all doses (with all ). The hepatic SOD, GSH-Px, and CAT activities in the H-SMPS group were elevated by 30.6%, 36.8%, and 44.6%, respectively, compared with those in the MC group. In the E-SMPS high-dose group, hepatic SOD, GSH-Px, and CAT activities were elevated by 40.8%, 58.4%, and 54.5%, respectively, compared with those in the MC group. Based on the present results, H-SMPS, E-SMPS, and bifendatatum can potentially restore the damaged antioxidative defense.

3.5. Effects of H-SMPS and E-SMPS on Hepatic Lipid Peroxidation

In the MC group, the hepatic lipid peroxidation of MDA and LPO was remarkably increased by 104.8% and 159.4%, respectively, as compared to the NC group (, Figures 4(d)4(e)). This indicates that oxidative stress had occurred in the liver as a result of the administration of alcohol. However, treatment with H-SMPS and E-SMPS significantly mitigated the abnormal increases in MDA and LPO levels (). A high dose of H-SMPS inhibited MDA and LPO levels by 27.9% and 41.0%, respectively, while E-SMPS inhibited MDA and LPO levels by 40.7% and 50.6%, respectively, when compared with levels in the MC group. These results demonstrate that both E-SMPS and H-SMPS could protect the liver against oxidative stress by inhibiting lipid peroxidation. Furthermore, the MDA and LPO levels in the PC group were almost equal to those in the NC group.

3.6. Effects of H-SMPS and E-SMPS on Hepatic Histological Changes

Images of hepatic histological sections are displayed in Figure 5. No obvious histological changes were observed in the livers of mice in the NC group. In contrast, hepatic sections of mice in the MC group showed nucleus contraction, loss of cellular boundaries in the cytoplasm of hepatocytes, necrotic hepatocytes around the central vein, and massive fatty tissues. Interestingly, the hepatic sections of the mice treated with H-SMPS and E-SMPS showed obvious improvement compared to the mice in the MC group. In particular, the mice treated with a high dose of E-SMPS had hepatic sections similar to those from the NC group, indicating that H-SMPS and E-SMPS could protect liver tissue from acute alcohol-induced hepatic injury.

3.7. Antioxidant Activities In Vitro

The reducing power of H-SMPS and E-SMPS increased as the polysaccharide concentration increased from 0 to 800 μg/mL (Figure 6(a)). At a concentration of 800 μg/mL, the reducing power of E-SMPS (1.67 ± 0.08) was higher than those of H-SMPS (1.39 ± 0.06) and BHT (1.11 ± 0.09).

The scavenging activities of E-SMPS and H-SMPS against hydroxyl radicals were shown to be dose dependent (Figure 6(b)). At a concentration of 800 μg/mL, the hydroxyl radical scavenging activities of H-SMPS, E-SMPS, and BHT reached 74.10 ± 4.11%, 86.6 ± 4.52%, and 66.22 ± 3.77%, respectively. Moreover, the IC50 values of H-SMPS, E-SMPS, and BHT were found to be 500.31 ± 2.70, 375.75 ± 2.58, and 564.64 ± 2.75 μg/mL, respectively, indicating that E-SMPS has stronger scavenging activities against hydroxyl radicals than H-SMPS does.

The color of DPPH solutions was reduced with the enhancement of antioxidant activities. H-SMPS and E-SMPS showed observable scavenging activities against DPPH radicals following an increase in the concentrations of the polysaccharides (Figure 6(c)). At a concentration of 800 μg/mL, the DPPH scavenging activity of E-SMPS reached 86.6 ± 4.73%, which was higher than those of H-SMPS (73.88 ± 4.25%) and BHT (61.27 ± 3.41%). The IC50 values of H-SMPS (479.90 ± 2.68 μg/mL), E-SMPS (378.12 ± 2.58 μg/mL), and BHT (604.12 ± 2.78 μg/mL) were consistent with the above results.

The superoxide radical scavenging activities of H-SMPS, E-SMPS, and BHT are shown in Figure 6(d). A positive correlation between scavenging activity and concentration is seen. The scavenging activities of H-SMPS, E-SMPS, and BHT against superoxide radicals were 59.51 ± 2.84%, 67.22 ± 3.11%, and 48.40 ± 2.91%, respectively. Furthermore, the IC50 values of H-SMPS, E-SMPS, and BHT were 674.35 ± 2.83, 553.09 ± 2.72, and 868.62 ± 2.94 μg/mL, respectively, indicating that E-SMPS displays stronger superoxide radical scavenging activity than H-SMPS and BHT.

3.8. Monosaccharide Composition Analysis

The monosaccharide compositions of H-SMPS and E-SMPS were determined by comparing their retention times with those of reference monosaccharides (Figure 7(a)). As seen in Figures 7(b) and 7(c), H-SMPS is composed of arabinose, xylose, mannose, galactose, and glucose, with mass percentages of 4.45%, 10.08%, 6.78%, 17.22%, and 61.37% (Figure 7(b)), respectively. However, E-SMPS was found to be composed of fucose, arabinose, xylose, mannose, galactose, and glucose, with mass percentages of 6.88%, 15.04%, 22.71%, 8.81%, 18.43%, and 28.13% (Figure 7(c)), respectively.

4. Discussion

In recent years, the use of polysaccharides from fungus as natural medicines has gained increasing attention owing to the lack of negative consequences in using them as treatments against many pathological diseases, as compared to chemical synthetic drugs. Several publications have established many polysaccharide extraction methods including hot water, alkaline, acidic, enzymatic, ultrasonic, and microwave extractions [2832]. Increasing amounts of evidence have shown that enzymatic-extractable polysaccharides from edible mushrooms possess high antioxidant and biological activities. The use of these polysaccharides has many other advantages such as high extraction yields, reproducibility of results, low energy consumption, and low pollution [30, 33]. However, few reports on the exploration of the antioxidant and hepatoprotective activities of E-SMPS in alcohol-induced mice have been published.

As an organ that is sensitive to cytotoxicity during chemotherapy, the liver plays important roles in alcohol metabolism [6]. Acute alcohol consumption could induce the formation of poisonous metabolic products, causing abnormalities in alcohol metabolism, thereby leading to liver damage [34]. Clinically, serum enzymes (AST and ALT), hepatic lipids (TC and TG), and enzymes involved in alcohol metabolism (ADH, ALDH, and CYP2E1) are commonly used as biochemical markers for early diagnosis of hepatic injury. Serum ALT and AST activities are elevated when hepatic injury occurs since ALT and AST could leach out of hepatocytes into the blood circulation. This is associated with ballooning degeneration, massive centrilobular necrosis, and cellular infiltration [35, 36]. Furthermore, acute alcohol-induced lipid metabolism disorders, including accumulation of fat and lipochondrion on the surface of hepatocytes, could be reflected in abnormal changes of hepatic TC and TG levels. In vivo, alcohol metabolism mainly consists of two pathways, the alcohol dehydrogenase oxidation system and the microsome alcohol oxidation system, catalyzed by the metabolic enzymes ADH, ALDH, and CYP2E1 [6]. During alcohol consumption, about 90% of the alcohol can be metabolized into acetaldehyde by ADH, which is further metabolized into acetic acid by ALDH, and eventually decomposed into carbon dioxide and water. About 8–10% of alcohol is metabolized by CYP2E1 in the tricarboxylic acid cycle [37, 38]. The activities of ADH and ALDH, two crucial enzymes in regulating alcohol metabolism, could be remarkably suppressed when acute alcohol consumption occurs [5]. The activity of CYP2E1, an important metabolic enzyme in catalyzing exogenous and endogenous compounds that plays a role in hepatic alcohol metabolism, could be increased when alcohol accumulation activates the microsome alcohol oxidation system [34]. In the present study, the ALD mice in the MC group showed serious liver damage as evidenced by significant increases in serum enzyme activities (AST and ALT), hepatic CYP2E1 levels, and hepatic lipid levels (TC and TG), as well as decreases in hepatic ADH and ALDH activities, when compared with the mice in the NC group. The abnormal changes were remedied by the supplementation of H-SMPS and E-SMPS, indicating that polysaccharides from the SMS of L. sulphureus have potential protective effects against toxicity induced by acute alcohol consumption. Furthermore, the hepatoprotective effects of H-SMPS and E-SMPS were confirmed by observation of histopathological sections.

It has been reported that acute alcohol consumption can induce oxidative stress in vivo, leading to the overproduction of ROS. This plays an important role in the development of ALD due to the toxic effects of ROS on lipids, enzymes, nucleic acids, and proteins [3941]. Hence, it seems necessary to increase the scavenging abilities against ROS. Previous literature has indicated that reducing power, which reflects the electron donation capacity of a compound, is one of the most important indicators for evaluating antioxidant activity [42]. The hydroxyl radicals, which are the major type of radicals, have been proven to be involved in the toxicity against biomolecules and in the induction of lipid peroxidation. Hence, there is great potential in discovering natural antioxidant agents with higher hydroxyl radical scavenging activities for the development of treatments against radical-induced diseases [43]. Scavenging activities against DPPH radicals, which are stable free radicals, are a wide indicator and allow a rapid method for assessing antioxidant activities [44]. Superoxide anion radicals, which are relatively weak oxidants, are one of the precursors of singlet oxygen and hydroxyl radicals and, as such, could indirectly activate lipid peroxidation and amplify cellular damage [45]. Based on the present results, both H-SMPS and E-SMPS showed significant reducing power and scavenging activities against hydroxyl, DPPH, and superoxide anion radicals. Previous studies have proposed that antioxidant enzymes, including SOD, GSH-Px, and CAT, are the primary defense in eliminating ROS-induced oxidative stress in vivo [46]. The possible underlying mechanism may be the degradation of superoxides into H2O2 by SOD and the subsequent decomposition to O2 and H2O by CAT and GSH-Px, which results in the suppression of ROS generation [47]. Moreover, accumulated studies have proven that ALD is also connected with lipid peroxidation, which is also regarded as an indicator of oxidative damage [48, 49]. In our in vivo assays, significant decreases in SOD, GSH-Px, and CAT activities as well as remarkable increases in MDA and LPO levels were observed after alcohol consumption, indicating that liver function had been damaged and the antioxidant defense mechanism had been disabled [50]. The polysaccharide supplements significantly remedied the decreased enzyme activities and increased lipid peroxidation, demonstrating that both H-SMPS and E-SMPS have the potential to alleviate acute alcohol-induced liver damage.

The monosaccharide compositions of polysaccharides from Catathelasma ventricosum [46], Lentinus edodes [12], T. albuminosus [27], and Flammulina velutipes [14] were found to be different from the monosaccharide compositions of H-SMPS and E-SMPS from the SMS of L. sulphureus determined in the present study. This difference may be attributed to the differences in strains, culture methods, and extraction conditions used. Furthermore, previous studies have demonstrated that the biological functions of polysaccharides were mainly determined by their monosaccharide compositions [51]. Only fucose could be observed in E-SMPS, indicating that fucose may play an important role in conferring higher biological activities. Additionally, Schneider et al. also revealed that fucose plays a vital role in maintaining biological functions such as immunoregulation and anticancer activities in mammals [52].

5. Conclusions

The in vitro and in vivo analyses in this study demonstrate that both H-SMPS and E-SMPS confer effective hepatoprotection against acute alcohol-induced ALD, possibly by reducing oxidative stress. E-SMPS showed superior effects, indicating that enzymatic hydrolysis has a potential effect on enhancing bioactivities. The results may provide a mechanistic basis for the use of polysaccharides from L. sulphureus as a potential natural and functional food supplement for the prevention and alleviation of ALD and its complications.

Abbreviations

ADH:Alcohol dehydrogenase
ALT:Alanine aminotransferase
ALD:Alcoholic liver disease
ALDH:Aldehyde dehydrogenase
AST:Aspartate aminotransferase
BHT:Butylated hydroxytoluene
CAT:Catalase
CYP2E1:Cytochrome P4502E1
E-SMPS:Enzymatic-extractable polysaccharides
FID:Flame ionization detector
GSH-Px:Glutathione peroxidase
H-SMPS:Hot-water-extractable polysaccharides
LPO:Lipid peroxidation
MC:Model control
MDA:Malondialdehyde
NC:Normal control
PC:Positive control
ROS:Reactive oxygen species
SD:Standard deviations
SMS:Spent mushroom substrates
SOD:Superoxide dismutase
TC:Total cholesterol
TG:Triglyceride.

Conflicts of Interest

The authors declared no conflicts of interest.

Acknowledgments

This work was supported by grants from the Mushroom Technology System of Shandong Province (SDAIT-07-05 and SDAIT-07-04) and Shandong Agricultural Fine Breed Project (2014lz058).

References

  1. J. Wang, Y. Zhang, R. Liu, X. Li, Y. Cui, and L. Qu, “Geniposide protects against acute alcohol-induced liver injury in mice via up-regulating the expression of the main antioxidant enzymes,” Canadian Journal of Physiology and Pharmacology, vol. 93, no. 4, pp. 261–267, 2015. View at: Publisher Site | Google Scholar
  2. J. Altamirano and R. Bataller, “Alcoholic liver disease: pathogenesis and new targets for therapy,” Nature Reviews Gastroenterology & Hepatology, vol. 8, no. 9, pp. 491–501, 2011. View at: Publisher Site | Google Scholar
  3. P. Mathurin and R. Bataller, “Trends in the management and burden of alcoholic liver disease,” Journal of Hepatology, vol. 62, no. 1, pp. S38–S46, 2015. View at: Publisher Site | Google Scholar
  4. C. S. Lieber, “Alcohol and the liver: 1994 update,” Gastroenterology, vol. 106, no. 4, pp. 1085–1105, 1994. View at: Publisher Site | Google Scholar
  5. C. S. Lieber, “Ethanol metabolism, cirrhosis and alcoholism,” Clinica Chimica Acta, vol. 257, no. 1, pp. 59–84, 1997. View at: Publisher Site | Google Scholar
  6. D. C. Wen, X. Y. Hu, Y. Y. Wang et al., “Effects of aqueous extracts from Panax ginseng and Hippophae rhamnoides on acute alcohol intoxication: an experimental study using mouse model,” Journal of Ethnopharmacology, vol. 192, pp. 67–73, 2016. View at: Publisher Site | Google Scholar
  7. Y. Y. Chen, C. L. Zhang, X. L. Zhao, K. Q. Xie, and T. Zeng, “Inhibition of cytochrome P4502E1 by chlormethiazole attenuated acute ethanol-induced fatty liver,” Chemico-Biological Interactions, vol. 222, pp. 18–26, 2014. View at: Publisher Site | Google Scholar
  8. M. Jin, A. Ande, A. Kumar, and S. Kumar, “Regulation of cytochrome P4502e1 expression by ethanol: role of oxidative stress-mediated pkc/jnk/sp1 pathway,” Cell Death & Disease, vol. 4, no. 3, article e554, 2013. View at: Publisher Site | Google Scholar
  9. D. Wu and A. I. Cederbaum, “Oxidative stress and alcoholic liver disease,” Seminars in Liver Disease, vol. 29, no. 02, pp. 141–154, 2009. View at: Publisher Site | Google Scholar
  10. B. Gao and R. Bataller, “Alcoholic liver disease: pathogenesis and new therapeutic targets,” Gastroenterology, vol. 141, no. 5, pp. 1572–1585, 2011. View at: Publisher Site | Google Scholar
  11. H. Wang, G. Li, W. Zhang, C. Han, X. Xu, and Y. P. Li, “The protective effect of Agaricus blazei murrill, submerged culture using the optimized medium composition, on alcohol-induced liver injury,” Biomed Research International, vol. 2014, Article ID 573978, 9 pages, 2014. View at: Publisher Site | Google Scholar
  12. H. J. Zhu, K. Sheng, E. F. Yan, J. J. Qiao, and F. Lv, “Extraction, purification and antibacterial activities of a polysaccharide from spent mushroom substrate,” International Journal of Biological Macromolecules, vol. 50, no. 3, pp. 840–843, 2012. View at: Publisher Site | Google Scholar
  13. C. Paredes, E. Medina, R. Moral et al., “Characterization of the different organic matter fractions of spent mushroom substrate,” Communications in Soil Science and Plant Analysis, vol. 40, no. 1-6, pp. 150–161, 2009. View at: Publisher Site | Google Scholar
  14. L. Lin, F. Y. Cui, J. J. Zhang et al., “Antioxidative and renoprotective effects of residue polysaccharides from Flammulina velutipes,” Carbohydrate Polymers, vol. 146, pp. 388–395, 2016. View at: Publisher Site | Google Scholar
  15. L. Wang, N. Xu, J. J. Zhang et al., “Antihyperlipidemic and hepatoprotective activities of residue polysaccharide from Cordyceps militaris SU-12,” Carbohydrate Polymers, vol. 131, pp. 355–362, 2015. View at: Publisher Site | Google Scholar
  16. J. Petrović, J. Glamočlija, D. S. Stojković et al., “Laetiporus sulphureus, edible mushroom from Serbia: investigation on volatile compounds, in vitro antimicrobial activity and in situ control of Aspergillus flavus in tomato paste,” Food and Chemical Toxicology, vol. 59, no. 8, pp. 297–302, 2013. View at: Publisher Site | Google Scholar
  17. R. G. Jayasooriya, C. H. Kang, M. J. Seo, Y. H. Choi, Y. K. Jeong, and G. Y. Kim, “Exopolysaccharide of Laetiporus sulphureus var. miniatus downregulates LPS-induced production of NO, PGE2, and TNF-α in BV2 microglia cells via suppression of the NF-κB pathway,” Food and Chemical Toxicology, vol. 49, no. 11, pp. 2758–2764, 2011. View at: Publisher Site | Google Scholar
  18. M. Y. Lung and W. Z. Huang, “Production, purification and tumor necrosis factor-α (TNF-α) release capability of exopolysaccharide from Laetiporus sulphureus, (Bulliard: Fries) Bondartsev & Singer in submerged cultures,” Process Biochemistry, vol. 46, no. 2, pp. 433–439, 2011. View at: Publisher Site | Google Scholar
  19. M. Y. Lung and W. Z. Huang, “Antioxidant properties of polysaccharides from Laetiporus sulphureus in submerged cultures,” African Journal of Biotechnology, vol. 11, no. 23, pp. 6350–6358, 2012. View at: Publisher Site | Google Scholar
  20. A. M. Staub, “Removal of proteins from polysaccharides methods,” Carbohydrate Chemistry, vol. 5, no. 5, pp. 5-6, 1965. View at: Google Scholar
  21. S. Song, S. Li, N. Su, J. Li, F. Shi, and M. Ye, “Structural characterization, molecular modification and hepatoprotective effect of melanin from Lachnum YM226 on acute alcohol-induced liver injury in mice,” Food and Function, vol. 7, no. 8, pp. 3617–3627, 2016. View at: Publisher Site | Google Scholar
  22. M. Wang, P. Zhu, C. Jiang, L. Ma, Z. Zhang, and X. Zeng, “Preliminary characterization, antioxidant activity in vitro and hepatoprotective effect on acute alcohol-induced liver injury in mice of polysaccharides from the peduncles of Hovenia dulcis,” Food and Chemical Toxicology, vol. 50, no. 9, pp. 2964–2970, 2012. View at: Publisher Site | Google Scholar
  23. M. Oyaizu, “Studies on products of browning reactions: antioxidative activities of products of browning reaction prepared from glucosamine,” Japanese Journal of Nutrition, vol. 44, no. 6, pp. 307–315, 1986. View at: Publisher Site | Google Scholar
  24. C. C. Winterbourn and H. C. Button, “Hydroxyl radical production from hydrogen peroxide and enzymatically generated paraquat radicals: catalytic requirements and oxygen dependence,” Archives of Biochemistry and Biophysics, vol. 235, no. 1, pp. 116–126, 1984. View at: Publisher Site | Google Scholar
  25. N. Cheng, N. Ren, H. Gao, X. Lei, J. Zheng, and W. Cao, “Antioxidant and hepatoprotective effects of Schisandra chinensis pollen extract on CCl4-induced acute liver damage in mice,” Food and Chemical Toxicology, vol. 55, no. 3, pp. 234–240, 2013. View at: Publisher Site | Google Scholar
  26. Q. Wei, X. H. Ma, S. S. Zhang, and S. C. Liu, “Antioxidant activities and chemical profiles of pyroligneous acids from walnut shell,” Journal of Analytical and Applied Pyrolysis, vol. 88, no. 2, pp. 149–154, 2010. View at: Publisher Site | Google Scholar
  27. H. J. Zhao, S. S. Li, J. J. Zhang et al., “The antihyperlipidemic activities of enzymatic and acidic intracellular polysaccharides by Termitomyces albuminosus,” Carbohydrate Polymers, vol. 151, pp. 1227–1234, 2016. View at: Publisher Site | Google Scholar
  28. L. Zheng, M. Liu, G. Y. Zhai, Z. Ma, L. Q. Wang, and L. Jia, “Antioxidant and anti-ageing activities of mycelia zinc polysaccharide from Pholiota nameko SW-03,” Journal of the Science of Food and Agriculture, vol. 95, no. 15, pp. 3117–3126, 2015. View at: Publisher Site | Google Scholar
  29. G. L. Song and Q. Z. Du, “Structure characterization and antitumor activity of an αβ-glucan polysaccharide from Auricularia polytricha,” Food Research International, vol. 45, no. 1, pp. 381–387, 2012. View at: Publisher Site | Google Scholar
  30. J. J. Zhang, G. Y. Meng, C. Zhang et al., “The antioxidative effects of acidic-, alkalic-, and enzymatic-extractable mycelium zinc polysaccharides by Pleurotus djamor on liver and kidney of streptozocin-induced diabetic mice,” BMC Complementary and Alternative Medicine, vol. 15, no. 1, pp. 440–452, 2015. View at: Publisher Site | Google Scholar
  31. J. Tang and Y. Q. Ma, “The application of ultrasonic technology in the Auricularia auricula polysaccharide extractions,” Food and Machinery, vol. 21, no. 1, pp. 28-29, 2005. View at: Google Scholar
  32. L. S. Fan, S. H. Zhang, and X. G. Wu, “Studies on microwave assisted extractions of Auricularia auricula polysaccharides,” Food and Fermentation Industries, vol. 31, no. 10, pp. 142–144, 2005. View at: Google Scholar
  33. N. Xu, Z. Gao, J. Zhang et al., “Hepatoprotection of enzymatic-extractable mycelia zinc polysaccharides by Pleurotus eryngii var. tuoliensis,” Carbohydrate Polymers, vol. 157, pp. 196–206, 2017. View at: Publisher Site | Google Scholar
  34. R. Brůha, K. Dvorák, M. Dousa, J. Petrtýl, and T. Svestka, “Alcoholic liver disease,” Prague Medical Report, vol. 110, no. 3, pp. 181–190, 2009. View at: Google Scholar
  35. J. Ozer, M. Ratner, M. Shaw, W. Bailey, and S. Schomaker, “The current state of serum biomarkers of hepatotoxicity,” Toxicology, vol. 245, no. 3, pp. 194–205, 2008. View at: Publisher Site | Google Scholar
  36. G. J. Huang, J. S. Deng, S. S. Huang, Y. Y. Shao, C. C. Chen, and Y. H. Kuo, “Protective effect of antrosterol from Antrodia camphorata submerged whole broth against carbon tetrachloride-induced acute liver injury in mice,” Food Chemistry, vol. 132, no. 2, pp. 709–716, 2012. View at: Publisher Site | Google Scholar
  37. C. S. Lieber, “Hepatic, metabolic and toxic effects of ethanol: 1991 update,” Alcoholism Clinical and Experimental Research, vol. 15, no. 4, pp. 573–592, 1991. View at: Publisher Site | Google Scholar
  38. A. I. Cederbaum, “Alcohol metabolism,” Clinics in Liver Disease, vol. 16, no. 4, pp. 667–685, 2012. View at: Publisher Site | Google Scholar
  39. J. G. O'Grady and R. Williams, “Classification of acute liver failure,” Lancet, vol. 342, no. 8873, p. 743, 1993. View at: Publisher Site | Google Scholar
  40. R. S. O’Shea, S. Dasarathy, and A. J. McCullough, “Alcoholic liver disease,” Hepatology, vol. 51, no. 1, pp. 307–328, 2010. View at: Publisher Site | Google Scholar
  41. C. Zhang, Z. Gao, C. L. Hu et al., “Antioxidant, antibacterial and anti-aging activities of intracellular zinc polysaccharides from Grifola frondosa SH-05,” International Journal of Biological Macromolecules, vol. 95, pp. 778–787, 2016. View at: Publisher Site | Google Scholar
  42. J. Liu, J. Luo, H. Ye, and X. Zeng, “Preparation, antioxidant and antitumor activities in vitro of different derivatives of levan from endophytic bacterium Paenibacillus polymyxa EJS-3,” Food and Chemical Toxicology, vol. 50, no. 3-4, pp. 767–772, 2012. View at: Publisher Site | Google Scholar
  43. B. Halliwell and J. M. C. Gutteridge, “Role of free radicals and catalytic metal ions in human disease: an overview,” Methods in Enzymology, vol. 186, no. 1, pp. 1–85, 1990. View at: Google Scholar
  44. C. L. Ke, D. L. Qiao, D. Gan, Y. Sun, H. Ye, and X. X. Zeng, “Antioxidant activity in vitro and in vivo of the capsule polysaccharides from Streptococcus equi subsp. zooepidemicus,” Carbohydrate Polymers, vol. 75, no. 4, pp. 677–682, 2009. View at: Publisher Site | Google Scholar
  45. X. Liu, B. Zhou, R. Lin et al., “Extraction and antioxidant activities of intracellular polysaccharide from Pleurotus sp. mycelium,” International Journal of Biological Macromolecules, vol. 47, no. 2, pp. 116–119, 2010. View at: Publisher Site | Google Scholar
  46. Y. Liu, J. Sun, S. Rao, Y. Su, and Y. Yang, “Antihyperglycemic, antihyperlipidemic and antioxidant activities of polysaccharides from Catathelasma ventricosum in streptozotocin-induced diabetic mice,” Food and Chemical Toxicology, vol. 57, no. 12, pp. 39–45, 2013. View at: Publisher Site | Google Scholar
  47. D. Yao, W. Shi, Y. Gou et al., “Fatty acid-mediated intracellular iron translocation: a synergistic mechanism of oxidative injury,” Free Radical Biology & Medicine, vol. 39, no. 10, pp. 1385–1398, 2005. View at: Publisher Site | Google Scholar
  48. V. Vande, J. Sedman, and A. Russin, “Lipid analysis by vibrational spectroscopy,” European Journal of Lipid Science Technology, vol. 103, pp. 15–25, 2001. View at: Google Scholar
  49. Y. Cui, Q. Ye, H. Wang, Y. Li, W. Yao, and H. Qian, “Hepatoprotective potential of Aloe vera polysaccharides against chronic alcohol-induced hepatotoxicity in mice,” Journal of the Science of Food and Agriculture, vol. 94, no. 9, pp. 1764–1771, 2014. View at: Publisher Site | Google Scholar
  50. Y. F. Sun, X. B. Yang, X. S. Lu, D. Y. Wang, and Y. Zhao, “Protective effects of Keemun black tea polysaccharides on acute carbon tetrachloride-caused oxidative hepatotoxicity in mice,” Food and Chemical Toxicology, vol. 58, pp. 184–192, 2013. View at: Publisher Site | Google Scholar
  51. Q. Han, Q. Yu, J. Shi, C. Xiong, Z. Ling, and P. He, “Structural characterization and antioxidant activities of 2 water-soluble polysaccharide fractions purified from tea (Camellia sinensis) flower,” Journal of Food Science, vol. 76, no. 3, pp. C462–C471, 2011. View at: Publisher Site | Google Scholar
  52. M. Schneider, E. Al-Shareffi, and R. S. Haltiwanger, “Biological functions of fucose in mammals,” Glycobiology, vol. 27, no. 7, pp. 601–618, 2017. View at: Publisher Site | Google Scholar

Copyright © 2017 Huajie Zhao et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


More related articles

963 Views | 408 Downloads | 5 Citations
 PDF  Download Citation  Citation
 Download other formatsMore
 Order printed copiesOrder

Related articles

We are committed to sharing findings related to COVID-19 as quickly and safely as possible. Any author submitting a COVID-19 paper should notify us at help@hindawi.com to ensure their research is fast-tracked and made available on a preprint server as soon as possible. We will be providing unlimited waivers of publication charges for accepted articles related to COVID-19. Sign up here as a reviewer to help fast-track new submissions.