Lycium barbarum Polysaccharides Protects against Strenuous

Exercise-induced Oxidative Damage in Rats

Lantao Liu and Weiqiang Zhang

†

Department of Physical Education, Central South University Changsha City, Hunan Province, 410083, China

Keywords: Lycium Barbarum Polysaccharides, Oxidative Damage, Exhaustive Running Exercise, Rats.

Abstract: Lycium barbarum contains a variety of nutrients and bioactive ingredients, which has multiple biological

and pharmacological effects. In the theory of Chinese medicine, Lycium barbarum as a common Chinese

herbal medicine can be used for the treatment of many diseases. The aim of the current study was to

evaluate the protective effects of Lycium barbarum polysaccharides (LBPs) on strenuous exercise-induced

oxidative damage in rats. The animals were divided into one control group (treated with distilled water) and

three LBPs groups (treated with 50, 100 and 200 mg/kg LBPs, respectively). After 28 days of treatment, the

exhaustive running exercise was performed, followed by the relevant biochemical parameter analysis. the

datas revealed that LBPs increases exhaustive running times, and levels of superoxide dismutase (SOD),

glutathione peroxidase (GPx), reduced glutathione (GSH), glutathione reductase (GR), and catalase (CAT)

in liver. LBPs decreases the levels of creatine kinase (CK), myoglobin (Mb), tumor necrosis factor-α (TNF-

α) and interleukin 1β (IL-1β) in serum, and also the levels of oxidized glutathione (GSSG),

malondialdehyde (MDA) and 8-hydroxy-2’-deoxyguanosine (8-OHdG) in liver. These results suggest that

LBPs has protective effects on oxidative damage induced by strenuous exercise.

1 INTRODUCTION

Reactive oxygen species (ROS) are the chemically

active oxygen free radicals and the substances that

can be converted to the free radicals, which are

generated during the aerobic metabolism of cells.

ROS mainly includes oxygen free radicals and some

non-free radicals substances such as hydrogen

peroxide (H

), hydroperoxide (ROOH), etc.

(Kurutas 2016). Under normal physiological

conditions, the body's antioxidant defense systemcan

remove ROS, maintaining a dynamic balance.

However, strenuous exercise can increase oxygen

intake, accompanied by the generation of ROS in

various tissues by mean of different ways, which

may exceed the capacity of defense system, resulting

in increased oxidative stress. Exercise-induced

endogenous ROS were produced by a variety of

sources, including mitochondrial respiratory chain

pathway, xanthine oxidase reaction pathway,

neutrophils respiratory burst pathway, hemoglobin

oxidation reaction pathway, and so on. It has been

reported that increased oxidative stress can bring

about various levels of oxidative damage to various

substances that make up cell tissue such as lipids,

sugars, proteins and DNA (Jówko et al. 2011).

Growing evidences show that exogenous

antioxidants from food and natural products

supplementation can be an effective means to cut

down exercise-induced oxidative damage (Chen et

al. 2013).

Lycium barbarum (L. barbarum) is a perennial

woody plant, mainly grown in some provinces of

northern China, such as Inner Mongolia, Ningxia

and Hebei. The fruits of L. barbarum, known as

wolfberry and Goqi, have been widely used as

traditional herbs and supplements more than 2500

years. In 2002, the fruits of L. barbarum were

identified as both food and medicine items by the

Chinese government departments. Many types of

components, such as polyphenols, polysaccharides,

alkaloids, carotenoids, vitamins, amino acids,

aminoethanesulfonic acids, and fatty acids in fruits

of L. barbarum have been isolated and identified,

which have some biological and health-related

activity (Tang et al. 2015). Numerous studies have

suggested that L. barbarum polysaccharides (LBPs)

is the most important ingredients of L. barbarum to

play many biological activities (Liu et al. 2015).

LBPs account for about 5 - 8% of the dry weight of

Liu, L. and Zhang, W.

Lycium barbarum Polysaccharides Protects against Strenuous Exercise-induced Oxidative Damage in Rats.

DOI: 10.5220/0011316100003444

In Proceedings of the 2nd Conference on Artiﬁcial Intelligence and Healthcare (CAIH 2021), pages 255-260

ISBN: 978-989-758-594-4

255

fruits of L. barbarum, which is a

heteropolysaccharide containing protein, and

generally consists of 6 - 8 monosaccharides, 18

amino acids and a variety of trace elements. The

molecular weight is in the range of 24 to 241 kDa

(Chen et al. 2009). Modern pharmacological studies

indicated LBPs has multiple pharmacological and

biological functions, including anti-diabetic, anti-

hypoxia, anti-fatigue, hypolipidemic,

antihypertensive, anti-aging, anti-cancer, analgesic,

immune regulation and liver protection effects.

Especially, this compound exhibited strong

antioxidant activities in vitro by inhibiting different

types of free radicals (DPPH, ABTS, superoxide

anion and hydroxyl radical), reducing power

activities and metal ion chelating capability (Li and

Zhou 2007). Animal experiments also indicated that

LBPs can significantly lower lipid peroxidation and

improve antioxidant enzyme activities (Zhao et al.

2015). Based on the antioxidant activities of LBPs, it

can be hypothesized that strenuous exercise-induced

oxidative damage in animal model can be prevented

by LBPs pretreatment. Thence, the research was

implemented to investigate whether LBPs

administration could prevent strenuous exercise-

induced oxidative damage in rats.

2 EXPERIMENTAL

2.1 Plant Material

The dried fruits of L. barbarum were collected in

Zhong-ning County of Ning Xia Huizu Autonomous

Region and provided by Qinian Biological

Technology Co., Ltd. (Yinchuan, China). The plant

samples were authenticated by a biologist in the

college of chemistry and chemical engineering,

Central South University (Changsha, China). The

voucher specimen was laid in plants herbarium of

Central South University.

2.2 Chemicals and Reagents

Commercial diagnostic kit for creatine kinase (CK)

was provided by Suzhou Comin Technology Co.

(Suzhou, China). Commercial diagnostic kits for

reduced glutathione (GSH), oxidized glutathione

(GSSG) and glutathione reductase (GR) were

provided by Beyotime Biotechnology Institute

(Haimen, China). Commercial diagnostic kits for

catalase (CAT), superoxide dismutase (SOD),

malondialdehyde (MDA), and glutathione

peroxidase (GPX) were provided by Jiancheng

Research institutions (Nanjing, China). ELISA kits

for myoglobin (Mb) were provided by Huamei

Biological Engineering Co. (Wuhan, China). ELISA

kits for tumor necrosis factor-α (TNF-α), interleukin

1β (IL-1β) and 8-hydroxy-2’-deoxyguanosine (8-

OHdG) were provided by Huijia Biological

Technology Co. (Xiamen, China).

2.3 Experimental Animals

Male Wistar rats (weight 180 - 200 g) adapt to the

environment and diet for one week before the

experiment. During the experiment, four rats were

placed in an individual plastic cage under standard

feeding conditions (temperature of 22 ± 2 °C,

relative humidity of 50 ± 15%, and 12 h light: 12

hours dark cycle circulation). Animals ingested

commercial rodent food and free drinking purified

water. This animal experiment was approved by the

Ethics Committee of Central South University.

2.4 Preparation of L. barbarum

Polysaccharides

L. barbarum polysaccharides (LBPs) were extracted

according to the previously published method in the

literature (Zhao et al. 2005), and have been slightly

adjusted. Briefly, the dried samples were crushed to

fine powder with electric mill and passed the 200

mesh sieve. Then the powder was refluxed twice

with petroleum ether (1 h every time) to remove the

lipid, and then refluxed twice with 80% ethanol (1 h

every time) to remove the small molecule sugar. The

residue was extracted with 10 volumes of distilled

water at 90 °C for three times (2.5 h every time).

The filtrate from combined and filtered water

extracts was concentrated in a rotary evaporator

under reduced pressure at 50 °C. Then the

concentrate was centrifuged (3000 rpm, 15 min), and

the supernatant was mixed with 4 volumes of 95%

ethanol and stockpiled overnight at 4 °C. The

precipitation was washed in order with anhydrous

ethanol, acetone and ether, and the reagent was

evaporated. The resulting precipitate was dispersed

in distilled water, dialyzed and lyophilized to afford

crude polysaccharides.

2.5 Experimental Design

Animals had one week adaption period, and after

that, they were divided into four groups, each

consisting of 8 rats. LBPs were given to the rats at

doses of 0, 50, 100 and 200 mg/kg and the four

groups were accordingly named as the control (C)

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group, the low-dose LBPs treatment (LBPL) group,

the medium-dose LBPs treatment (LBPM) group

and the high-dose LBPs treatment (LBPH) group.

LBPs were dissolved in 1.0 mL distilled water and

administered by oral gavage one time per day lasting

for 28 days. After 21 days, the rats were introduced

to the motor-driven treadmill (WI78059, Shanghai

Yuyan Scientific Instrument Co., Ltd.) and made to

run at 15 m/min and a 0° grade for 15 min one time

per day lasting for 7 days to accommodate running

exercise. At the final day of experiment (the 28th

day), the incremental running exercise to exhaustion

was conducted using methods previously described

(Huang et al. 2013) with some modifications. The

rats were introduced into a treadmill, started running

at 15 m/min and 0 °grade for 10 min, then at 20

m/min and 0 °grade for 10 min, and finally at 30

m/min and 10 ° grade to exhaustion. Exhaustion is

defined when the rats can't continue running on the

treadmills after 12 s of continuous electric shock,

and the exhaustive running time was measured.

2.6 Analysis of Biochemical

Parameters

After exhaustive running exercise, the rats were

sacrificed by decapitation under ether anesthesia.

Blood was collected and centrifuged (3000 rpm, 15

min) at 4 °C to obtain serum for CK, Mb, TNF-α

and IL-1β analysis. The liver samples were

immediately isolated, weighed, and homogenized for

GSH, GSSG, SOD, CAT, GPX, GR, MDA and 8-

OHdG determinations. The levels of CK, Mb, TNF-

α, IL-1β, SOD, CAT, GPX, GR, GSH, GSSG, MDA

and 8-OHdG were determined using commercial

assay kits and abiding by the procedures advised by

manufacturers.

2.7 Statistical Analysis

All data were presented as mean ± standard

deviation (SD), and SPSS software is used for

Statistical analysis.

3 RESULTS AND DISCUSSION

3.1 Effects of LBPs on the Exhaustive

Running Times

Figure 1: Effects of LBPs on the exhaustive running times.

According to Figure 1, the exhaustive running time

of different doses of LBP groups (LBPL, LBPM and

LBPH) were significantly longer than that of C

group (p < 0.05). Compared with LBPL group,

exhaustive running time of LBPH group was

significantly prolonged (p < 0.05). Compared with

LBPM group, the exhaustive running time of LBPH

group was significantly prolonged (p < 0.05). The

above data showed that LBPs had a strong anti-

fatigue effect.

3.2 Effects of LBPs on the CK and Mb

in Serum

Strenuous exercise leads to increased muscle

membrane permeability or muscle cell damage,

causing creatine kinase (CK) and myoglobin (Mb)

and other proteins to escape from the cell and into

the blood circulation.

According to Figure 2, the CK and Mb levels of

the different doses of LBP groups (LBPL, LBPM

and LBPH) were significantly reduced compared

with the C group (p < 0.05). The CK levels of the

LBPH groups, as well as the Mb levels of the LBPM

and LBPH groups were significantly reduced

compared with the LBPL group (p < 0.05). The CK

and Mb levels of the LBPH groups were

significantly reduced compared with the LBPM

group (p < 0.05). The above data showed that LBPs

might prevent muscle damage or promote rapid

regeneration of damaged muscle

Lycium barbarum Polysaccharides Protects against Strenuous Exercise-induced Oxidative Damage in Rats

257

Figure 2: Effects of LBPs on the CK and Mb in serum.

3.3 Effects of LBPs on the TNF-α and

IL-1β in Serum

Previous studies have shown that strenuous exercise

can induce proinflammatory cytokines and

pleiotropic cytokine secretion to increase. TNF-α

and IL-1β are inflammatory cytokines secreted by

monocyte-macrophages and have proinflammatory

effects, which could also stimulate the production of

pleiotropic cytokine IL-6.

Figure 3: Effects of LBPs on TNF-α and IL-1β in serum.

According to Figure 3, the TNF-α and IL-1β

levels of LBPL, LBPM and LBPH were

significantly reduced compared with the C group (p

< 0.05). Compared with the LBPL group, the TNF-α

and IL-1β levels of the LBPH groups were

significantly reduced (p < 0.05); Compared with the

LBPM group, the IL-1β levels of the LBPH groups

were significantly reduced (p < 0.05). The above

data showed that LBPs could attenuate strenuous

exercise-induced inflammatory responses.

3.4 Effects of LBPs on the SOD, CAT,

GPX and GR in Liver

Antioxidant enzymes play an important protective

role in exercise-induced free radical oxidative

damage, and the decrease in the activity of these

enzymes means that the tissue is more susceptible to

free radical damage (Lee et al. 2009).

Figure 4: Effects of LBPs on the SOD, CAT, GPX and GR

in liver.

According to Figure 4, the SOD and GPX levels

of the LBPL, LBPM and LBPH groups; the CAT

and GR levels of the LBPM and LBPH groups were

significantly prolonged compared with the C group

(p < 0.05). Compared with the LBPL group, the

SOD, CAT, GPX and GR levels of the LBPM and

LBPH groups were significantly prolonged (p <

0.05). Compared with the LBPM group, the CAT,

GPX and GR levels of the LBPH groups were

significantly prolonged (p < 0.05). The above data

showed that LBPs could up-regulate the expression

of antioxidant enzymes to prevent strenuous

exercise-induced oxidative damage.

3.5 Effects of LBPs on the GSH and

GSSG in Liver

Glutathione is a tripeptide consisting of glutamine,

cysteine and glycine, which has two forms of

reduced (GSH) or oxidized (GSSG) (Masella et al.

2005). As a main intracellular antioxidant, GSH

plays an important role in preventing exercise-

induced oxidative damage by removing free radicals

and preventing the accumulation of hydroperoxides.

Under the action of GPx, GSH can reduce the H

to produce H

O, while GSH is oxidized to GSSG in

cells. GSSG also produces GSH under the catalysis

of GR. Strenuous exercise can cause severe

oxidative stress, leading to accumulation of GSSG

and reduction of GSH. GSH depletion can increase

the formation of hydroxyl radicals, which would

further lead to DNA damage (Muñoz et al., 2010).

Therefore, depletion of GSH in tissues has been used

as a sensitive index of exercise-induced ROS

production.

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Figure 5: Effects of LBPs on the GSH and GSSG in liver.

According to Figure 5, compared with the C

group, the GSH levels of the LBPL, LBPM and

LBPH groups were significantly prolonged (p <

0.05); the GSSG levels of the LBPM and LBPH

groups were significantly reduced (p < 0.05).

Compared with the LBPL group, the GSH levels of

the LBPH groups were significantly prolonged (p <

0.05); the GSSG levels of the LBPM and LBPH

groups were significantly reduced (p < 0.05). The

above data showed that LBPs are adequate

protection against exercise-induced ROS generation.

3.6 Effects of LBPs on the MDA and

8-OHdG in Liver

MDA is the end product of peroxidative

decomposition of polyenic fatty acids and has been

often used as a marker of lipid peroxidation. 8-

OHdG is one of the major products of DNA

oxidation. When ROS attacks the guanine in DNA,

it causes deoxyguanosine oxidation to form 8-

OHdG. 8-OHdG can be measured with high

sensitivity and it is extensively investigated in

human and animal exercise studies, and is thus used

as a biomarker of oxidative DNA damage (Hamurcu

et al. 2010).

According to Figure 6, the MDA and 8-OHdG

levels of the LBPL, LBPM and LBPH groups were

reduced compared with the C group (p < 0.05).

Compared with the LBPL group, the MDA and 8-

OHdG levels of the LBPM and LBPH groups were

significantly reduced (p < 0.05). Compared with the

LBPM group, the MDA levels of the LBPH groups

were significantly reduced (p < 0.05). The above

data showed that LBPs could attenuate lipid

peroxidation and oxidative DNA damage induced by

exhaustive exercise.

Figure 6: Effects of LBPs on the MDA and 8-OHdG in

liver.

4 CONCLUSIONS

The results of this study provide strong evidence that

LBPs have protective effects on oxidative damage

induced by strenuous exercise in rats due to

increased the levels of SOD, CAT, GPX, GR and

GSH in liver, simultaneously decreased the levels of

CK, Mb TNF-α and IL-1β in serum, and the levels

of GSSG, MDA and 8-OHdG in liver. The

protective effects on oxidative damage of LBPs was

dose-dependent in rats, which might be related to the

per se antioxidant activities of LBPs.

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