Effects of Chlorella vulgaris polysaccharides accumulation on growth characteristics of Trachemys scripta elegans

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Abstract

The present study investigated the effects of the accumulated polysaccharides in Chlorella vulgaris microalgae on the growth characteristics of Trachemys scripta elegans. Sodium alginate was used to prepare immobilized C. vulgaris, and the antioxidant effects of the accumulated polysaccharides in it were determined using Caenorhabditis elegans as a model. We determined the specific growth rates of T. s. elegans (10 in each group) and their levels of non-specific immune-related indexes (including alkaline phosphatase; total superoxide dismutase; catalase; malondialdehyde). Under optimal culturing conditions, the accumulated polysaccharide content in C. vulgaris reached 32.7% (dry weight). Polysaccharides from C. vulgaris significantly improved the hydrogen peroxide-induced oxidative stress resistance and resulted in the enhancement of stress resistance-related antioxidant enzymes, including total superoxide dismutase and catalase (p < 0.05). The accumulated polysaccharides in C. vulgaris were heteropolysaccharides comprising rhamnose, ribose, arabinose, xylose, 2−deoxy−D−glucose, mannose, glucose, galactose, and glucosamine with a molar ratio of 0.26: 0.62: 0.21: 0.10: 0.08: 0.18: 1.00: 0.42: 0.17. Compared with the control group with common feeds, suspended and immobilized C. vulgaris with higher accumulated polysaccharide levels had a positive effect on the specific growth rate of the T. s. elegans (p < 0.05). Further, the suspended and immobilized C. vulgaris with higher accumulated polysaccharide levels significantly increased serum alkaline phosphatase, total superoxide dismutase and catalase activity (p < 0.05) and decreased serum malondialdehyde levels of T. s. elegans (p < 0.05).

Introduction

China's aquaculture industry is growing dramatically in recent years and accounts for 60.5% of global aquaculture production [1]. However, there are various problems, such as disease and stresses, which results in a weakened immune system in intensive farming. Therefore, antibiotics have been used to control various diseases in aquaculture industries [2]. The extensive management of antibiotics in the past breeding process has made not only drug-resistant bacteria-infested in the environment but also brought great dangers to the safety of aquatic products and human health.

The growing aquaculture industry requires non-antibiotic products to reduce the risk of bacteria developing and spreading antibiotic resistance [3]. Microalgae are promising antibiotic substitutes for aquatic animals in aquatic farming. As a natural bait, microalgae are rich in amino acids, polyunsaturated fatty acids and growth-promoting factors [4], which are essential nutrients for the aquatic animals [5].

At present, studies on microalgae mainly include the culture technology and preservation, bioactivity such as antioxidant capacity or immunity of model organisms. Besides, it was found that microalgae were beneficial to the growth of the prawn, which the activities of amylase, trypsin and the content of long-chain polyunsaturated fatty acids in prawn were significantly improved or raised [6]. What is more, as a supplement to fish (Nile tilapia Oreochromis niloticus) feed, it has protective and therapeutic effects on arsenic poisoning [7]. As a feed or feed additives, microalgae not only have the potential to replace the use of antibiotics, but also can improve the growth characteristics of cultured animals owing to their high nutritional value.

Microalgae polysaccharides have been widely recognized for their good antioxidant effect [8,9], which are the basis of new feed. At present, studies on the antioxidant capacity of microalgae in vitro have been well developed. It has been proved that brown [10], red [11] and green [12] microalgae have excellent antioxidant properties in vitro. However, few studies have been done on the antioxidant effect of microalgae polysaccharides in vivo.

Chlorella vulgaris is a kind of unicellular green microalgae, studies on its culture conditions, nutritional components, and physiological activities have been carried out as early as 1960s. There are many studies on the physiological functions of C. vulgaris, such as antioxidant and anti-inflammatory activities [13], antitumor effect [14], immune modulation [15], antibacterial and antiviral [16], anti-ageing [[17], [18], [19]], hypoglycaemic and hypolipidaemic effect [[20], [21], [22], [23], [24]]. Therefore, we chose C. vulgaris as our experimental subject because of its wide physiological functions, amazing reproduction rate and easy to expand culture. C. vulgaris polysaccharides have many biological functions, such as chelating heavy metal [25], protecting the intestine [23,26] and liver [27], even improving the fibromyalgia symptoms [28], thus it is very necessary and urgent to optimize the accumulation condition of C. vulgaris polysaccharides for producing higher content of polysaccharides. At present, it has been found that light and temperature of the culture environment have important effects on the accumulation of microalgae polysaccharides. In this paper, PS II was used as the index to explore the most suitable external conditions for the production of C. vulgaris polysaccharides.

Microalgae have a more suitable production environment and dense biomass after immobilization, and their advantages in water purification have been widely confirmed [29]. However, it is rare to use immobilized microalgae as feed for aquaculture animals. Therefore, the objectives of this study are (1) to study the potential of C. vulgaris for its polysaccharides accumulation at different temperatures and light intensities; (2) to investigate the antioxidant effect of polysaccharides from C. vulgaris using Caenorhabditis elegans damaged with hydrogen peroxide as a model organism; (3) to explore the effects of suspended and immobilized C. vulgaris on improving the growth characteristics of T. s. elegans.

Section snippets

Cultivation of C. vulgaris

C. vulgaris was obtained from HuaDan Agricultural Products Co., Ltd., Hangzhou, China. The culture medium of C. vulgaris was BG11 solid medium [1.5 g NaNO3, 0.04 g K2HPO4, 0.075 g MgSO4·7H2O, 0.036 g CaCl2·7H2O, 0.02 g Na2CO3, 0.006 g citric acid, 0.006 g ferric citrate, 1 mL trace element solution (2.86 g H3BO4, 1.81 g MnCl2·4H2O, 0.222 g ZnSO4, 0.39 g Na2MoO4, 0.079 g CuSO4·5H2O, 49.4 g Co(NO3)2·6H2O, 1000 mL deionized water, 20.0 g agar]. C. vulgaris was purified by multi-line separation

Effect of light intensity and culture temperature on the content of C. vulgaris polysaccharides

Photosynthesis of C. vulgaris under different light intensity and the temperature was shown in Table 2.

Light intensity and culture temperature are two important factors affecting the growth of C. vulgaris. Temperature can change the photosynthesis and respiration of C. vulgaris by affecting the activity of related enzymes [32], and the total sugar accumulation in C. vulgaris will be influenced by the change of key enzymes activity, especially sucrose synthase (SS), sucrose phosphate synthase

Discussion

Light is an important factors that affect the accumulation of polysaccharides in microalgae. Within a certain range, an increase in light intensity can increase cell volume [45] as well as accelerate cell division and photosynthesis rate in microalgae [46]. However, the metabolism in microalgae is affected when the light intensity exceeds the intensity required for photosynthesis [47]. In the present study, C. vulgaris polysaccharides could accumulate up to 32.7% under the conditions of

Conclusion

In summary, light intensity and culture temperature are two important factors affecting the growth of C. vulgaris and polysaccharide accumulation. Polysaccharides from C. vulgaris (CVPP) could accumulate up to 32.7% under the conditions of 65 μmol·m−2·s−1, 28 °C in our experiment. CVPP was concluded as a heteropolysaccharide comprised of rhamnose, ribose, arabinose, xylose, 2−deoxy−D−glucose, mannose, glucose, galactose and N−acetyl−D−(+)−glucosamine with a molar ratio of 0.26: 0.62: 0.21:

Acknowledgments

This work was financially supported by the program National Science and Technology Program during the Twelfth Five-year Plan Period in rural areas (No. 2013BAD10B02) and granted from the Key Laboratory of Marine Food Quality and Hazardous Controlling Technology of Zhejiang Province.

Compliance with ethical standards

We solemnly declare that all applicable international, national, and/or institutional guidelines for the care and use of animals were followed.

Declaration of competing interest

The authors declare that there are no conflicts of interest associated with this manuscript.

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