Sulfated polysaccharides from Cyclocarya paliurus reduce H2O2-induced oxidative stress in RAW264.7 cells
Introduction
The reactive oxygen species (ROS), including superoxide radical, hydroxyl radical, hydrogen peroxide, can cause oxidative damage to DNA, proteins and lipids [1]. The oxidative stress, mainly caused by ROS, was demonstrated to be the principal cause of many chronic diseases, such as aging, cancer, Alzheimer, diabetes and cardiovascular diseases [2]. Many studies had shown that the ROS was produced from metabolism or exogenous intake, and caused damage to human tissue indirectly by cellular dysfunction and death. Over the past few years, there has been an increasing attention to antioxidants from natural products, such as flavone, polyphenol and polysaccharides without the side effect and toxicity [3], [4], [5].
Polysaccharide is a kind of natural macromolecule substances [6], [7], possessing extensive biological activities, such as antioxidant, anti-tumor and immunoregulation [8], [9], [10]. It has been reported that many polysaccharides have the activities that scavenge DPPH free radical, hydroxyl radical and hydrogen peroxide [11], protect mice macrophages RAW264.7 against oxidative damage induced by hydrogen peroxide [12]. Moreover, the effect of superoxide dismutase (SOD), one of the representative antioxidant enzymes, in scavenging radicals and reducing oxidative damage could be improved by polysaccharides has been reported [13].
The activities of polysaccharides are closely related to their structure. In the past decades, there was a burst of concentration on the study of structure modification in polysaccharides due to low toxicity and high bioactivities, such as sulfated, phosphorylated, carboxymethylated and acetylated modification. Sulfated polysaccharides could enhance the antioxidant activity, in which the hydroxyl groups are replaced by sulfated groups [14]. And it is well known that the biological activities of polysaccharides could be affected by sulfated modification because of the change of structure including the degree of substitution and molecular weight [15]. Furthermore, a number of sulfated polysaccharides obtained by chemical synthesis have been recently demonstrated to play an important role in resistance to oxidative stress. For example, the sulfation derivatives from Artemisia sphaerocephala polysaccharide had better antioxidant activities compared to native one in vitro [16]. And the polysaccharides produced by Enterobacter cloacae Z0206 exhibited stronger protective effects in RAW264.7 murine macrophages against H2O2-induced oxidative damage [17]. Many studies have suggested that sulfated polysaccharides are potent antioxidants and may be applied to functional food and pharmaceutical industries [18], [19].
Cyclocarya paliurus (Batal.) Iljinskaja, commonly named sweet tea tree, widely grows in southern China, such as Jiangxi, Hunan and Zhejiang Province. With high nutritional values and health functions, the leaves of C. paliurus have been widely used in China. The extraction, purification, chemical composition, structural characterization and some biological activities of the polysaccharides from the leaves of C. paliurus (CP) have been reported by our laboratory [20], [21]. Chemical analysis showed that CP was a heteropolysaccharide and contained 8.44% of protein, 17 kinds of amino acids and 18 mineral elements [21], [22], [23]. In addition, CP was found to exert significant antioxidant activities [3], [21]. However, to the best of our knowledge, there are no references available on the protective effect of polysaccharides from C. paliurus in oxidative stress-induced cell injury.
In the present study, two different sulfated polysaccharide derivatives (S-CP1–4 and S-CP1–8) were conducted with chlorosulfuric acid–pyridine in formamide, and the scanning electron microscope (SEM) was used to observe the surface change in polysaccharides morphology. The effects of sulfated polysaccharides and native polysaccharides on the RAW264.7 cells viability were compared by CCK-8 method. The intracellular contents of SOD and MDA were investigated. The objective was to evaluate the protective effect of sulfated derivatives against oxidative stress in RAW264.7 cells, and investigate the probability of improving antioxidant activity of CP through sulfated modification.
Section snippets
Materials and reagents
C. paliurus leaves were purchased from Jiangxi Xiushui Miraculous Tea Industry Co. (Jiangxi, China). Chlorosulfonic acid, pyridine and 1,1-diphenyl-2-picrylhydrazyl (DPPH) were purchased from Sigma Chemical Co. (St. Louis, MO, USA). BCA protein content, SOD and MDA assay kits were obtained from Beyotime Institute of Biotechnology (Shanghai, China). Murine macrophage cell line RAW264.7 cells were purchased from the Type Culture Collection of Chinese Academy of Sciences (Shanghai, China). RPMI
The physicochemical properties and structural features of CP, S-CP1–4 and S-CP1–8
According to our previous work, the CP was confirmed to be a kind of glycoprotein [27], including 60.62% total carbohydrate, 7.57% protein and 16.14% uronic acid. After sulfated modification, the significant change could be found in two sulfated derivatives. The DS of S-CP1–4 and S-CP1–8 were 0.42 ± 0.04 and 0.12 ± 0.02, respectively, indicating that the sulfation was successful. The total sugar content in S-CP1–4 and S-CP1–8 was decreased to be 39.41 ± 0.63% and 49.71 ± 0.56%, and the content of
Conclusions
In this study, the water-soluble polysaccharide extracted from C. paliurus was modified successfully by the CSA–Pyr method, and two kinds of sulfated derivatives were obtained. The results showed that sulfated modification might reduce oxidative stress in RAW264.7 cells by increasing SOD activity and inhibiting lipid peroxidation. All the sulfated derivatives could inhibit the lipid peroxidation, and S-CP1–4 exhibited stronger promotion in the production of SOD. Then their antioxidant
Conflict of interest
The authors declare that there is no conflict of interest.
Acknowledgements
The authors gratefully acknowledge the financial supports by National Natural Science Foundation of China (Nos. 31471702, 31201297, 31130041; 31201326), the National Key Technology R&D Program of China (No. 2012BAD33B06), the Natural Science Foundation of Jiangxi Province, China (No. 20152ACB21004) and the Objective-Oriented project of State Key Laboratory of Food Science and Technology, Nanchang University, China (No. SKLF-ZZA-201301).
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2022, International Journal of Biological MacromoleculesCitation Excerpt :Sulfate content (−OSO3H groups) in the molecules of SPs is considered the major feature in the molecules of SPs associated with protective effects against oxidative damage [108]. Modified oversulfated SPs from the plant Cyclocarya paliurus are more protective against H2O2-induced oxidative stress on RAW264.7 cells than their normal counterparts (natural SPs) [108]. When comparing SPs with similar MW but differing in their sulfate content, the antioxidant activity increases as increasing the level of sulfate content, as demonstrated with SPs from seaweed Porphyra yezoensis, Gracilaria lemaneiformis, Sargassum fusiform, and Undaria pinnatifida on damaged HK-2 cells (proximal kidney tubule epithelial cell line) [109,110].