Extraction And Purification Of Spirulina Polysaccharides And Analysis Of Their Antioxidant Activities

Spirulina is a kind of prokaryote-blue green algae. Spirulina contains a lot of proteins, polysaccharides, unsaturated fatty acids and a variety of trace elements, so it has great prospects as a new raw material of health care products. Numerous studies show that Spirulina polysaccharide can enhance the immune system, has anti-oxidation, anti-aging, anti-viral, anti-tumor and lower blood sugar effects, it has a great value in the fields of the agriculture, medicine, cosmetics, health products and environment and etc. In this paper, we optimized the cultivation medium for the production of Spirulina polysaccharides, and studied the extraction and purification methods of Spirulina polysaccharides, also identified its physical-chemical properties and its antioxidant activity in vitro. The main results are as follows:1. By single factor test, we determined the optimal concentration of different nutrition in medium for the production of polysaccharides. The concentration of NaHCO3, NaCl, K2HPO4, NaNO3 and K2SO4 is 21.0 g/L,1.0 g/L,0.3 g/L,1.0 g/L 0.60 g/L separately.2. Using the response surface methodology, we optimized the cultivation medium for the production of Spirulina polysaccharides. we found that the content of intracellular polysaccharide was most significantly affected by NaNO3, followed by NaHCO3, K2HPO4, and the least is K2SO4. The best medium components are:NaHCO3 21.6 g/L, K2HPO4 0.30 g/L, NaNO3 1.40 g/L and K2SO4 0.60 g/L.Under optimal conditions, the content of Spirulina intracellular polysaccharide is (19.5± 0.15)%.3. We compared a variety of extraction methods of Spirulina polysaccharides according to its yield. The optimum extraction conditions of ultrasonic broken method:ultrasonic power 500 W, working 2 s, stop 5 s, the total working time 10 min, temperature protection 0 ℃. Hot water extraction method: pH12,80 ℃ hot water for 4 h. Ultrasonic cleaning extraction method: 300 W,40 min,45 ℃. The yield of polysaccharides was the highest by the mix method:firstly ultrasonic broken:pH10, ultrasonic 10 min, then 70℃ hot water extraction for 4 h.4.Using the response surface methodology, we optimized the mix method for the extraction of Spirulina polysaccharides. The optimal condition was:ultrasonic power 510 W, ultrasonic time 10.61 min, water extraction temperature 70 ℃, extraction time 4.1 h, the polysaccharide extraction rate was high about (6.34±0.015)%.5. Spirulina polysaccharide was purified by ion exchange chromatography and other methods. We obtained relative pure polysaccharides from crude products of Spirulina cells and cultivation medium after alcohol precipitation, dialysis and a series of purificationprocedures. The obtained polysaccharide products was further purificated by DEAE-52 ion-exchange chromatography. Using NaCl gradient elution, we finally got six groups of polysaccharides, which are named as SPS1. SPS2, SPS3, SPS4, SPS5 and SPS6. Each SPS was relatively uniform for the elution peak in Sephadex G-100 column chromatography showed symmetric unimodal.6. We used a variety of chemical methods to identify the six groups, such as repeated freezing and thawing experiment, molish reaction, iodine reaction, ferric chloride reaction, ninhydrin reaction, and examination of coomassie brilliant blue reaction. The results show that SPS1. SPS2, SPS3, SPS4, SPS5 and SPS6 are all polysaccharides.7. The antioxidant effect of Spirulina polysaccharides was identified through 3 kinds of free radicals scavenging reactions, such as superoxide anion,. With the increase of the purity of the Spirulina polysaccharide, the antioxidant effect was also increased. As for Spirulina intracellular polysaccharides, SPS3 had the strongest ability of removing O2- and R-, reached (88.58±2.4)%and (78.4±1.3)%respectively. And SPS2 could remove DPPH by (92.03±2.3)%. As for the Spirulina extracellular polysaccharides, SPS6 is the strongest to remove O2- by (81.13±1.3)%. SPS4 is the strongest to remove R- by (69.71±0.9)%. SPS5 is the strongest to remove DPPH by (86.84±1.4)%. Over all, the antioxidant activity of intracellular polysaccharides was stronger than that of extracellular polysaccharide...