| Microalgae are a kind of unicellular organism,which are widely distributed in ecosphere.Microalgae have been drawn more attention in many fields because of their high biomass,high light efficiency,ease of cultivation and abundant metabolites.Antioxidant activities were selected as evaluation criteria to screen a microalgal strain from nature,which possess high antioxidant activities,and then the identification of its species,chemical constituents and scale-up cultivation were analyzed.The results were showed as below:To exploit indigenous microalgae with potential for antioxidant activity,ten microalgal strains were isolated from fresh and sea waters in Hainan Province according to their high biomass,strong antioxidant capacity and antioxidant enzymes activity.The strains were then identified based on their morphology,18s rDNA and ITS sequence analysis.Five were identified as Dictyosphaerium sp.1A10,Desmodesmus armatus(D.armatus)B38,Masaia oloidia(M.oloidia)2Fg,Chlorella vulgaris(C.vulgaris)B43 and Chlamydomonas chlamydogama(C.chlamydogama)WC11.Dictyosphaerium sp.1A10 showed the highest superoxide dismutase(SOD)activity(935.60±12.65 U/g tissue),Chlorella sorokiniana(C.sorokiniana)XS04 showed the highest catalase(CAT)activity(122.09±4.11 U/g tissue)and D.armatus B38 showed the highest glutathione peroxidase(GPX)activity(107.56±8.01 U/g tissue).Dictyosphaerium sp.1A10 revealed the highest DPPH radical scavenging capacity(99.60±10.00%)and oxygen radical absorbance capacity(621.02±10.73?mol Trolox/g),D.armatus B38 showed the highest ferrous ion chelating capacity(92.88±6.88%)and hydroxyl radical-scavenging capacity(97.101±1.76%),Scenedesmus ecornis(S.ecornis)G2 showed the highest ABTS scavenging capacity(92.11±4.99%),M.oloidia 2Fg showed the highest ferric reducing antioxidant power(505.07±18.23 ?mol FeSO4/g),Desmodesmus sp.WC08 showed the highest superoxide radical-scavenging capacity(83.5±3.24%)and Micractinium reisseri(M.reisseri)C67 showed the highest anti-lipid peroxidation capacity(82.72±3.42%).Although,Dictyosphaerium sp.1A10 and D.armatus B38 could be the most appropriate candidate for antioxidants,the biomass of Dictyosphaerium sp.1A10(1.97±0.23 g/L)was much higher than the biomass of D.armatus B38(1.38±0.37 g/L),therefore,Dictyosphaerium sp.1A10 was chosen for further analyses.Chemical constituents of Dictyosphaerium sp.1A10,including separation and purification of natural products and nutrient components,were analyzed.The results of nutrient components showed that Dictyosphaerium sp.1A10 was rich in carbohydrates(19.15%),grease(24.56%),ash(12.43%),fiber(12.31%)and protein(18.55%).The energy of Dictyosphaerium sp.1A10 was 371 Kcal.The contents of linoleic(15.3%)and linolenic(8.5%)among fatty acids were high,and there were 18 kinds of amino acids in Dictyosphaerium sp.1A10,which took up 10.16%of dry microalgae.And the content of essential amino acids accounted for 3.2%of that of total amino acid,which proved to be high in score of ratio coefficient of amino acid(SRCAA).The contents of VBi(2.15 mg/100g),VB3(6.25 mg/100g),VB6(1.31 mg/100g),VA(2.37 mg/100g)and VE(5.28 mg/100g)were highest among seven kinds of vitamins.Besides,magnesium(3517.26 mg/kg),calcium(813.27 mg/kg),iron(301.78 mg/kg)and potassium(9426.87 mg/kg)were highest among mineral elements.In terms of natural products,Dictyosphaerium was extracted sequentially by petroleum ether and ethyl acetate.Further separation and detection of Dictyosphaerium extractions were extracted by silica gel column chromatography,high-performance liquid chromatography,gas chromatography-mass spectrometry(GC-MS).According to the physicochemical properties and nuclear magnetic resonance spectroscopy(NMR),the structures of the compounds were determined.Ten compounds were identified and the antioxidant activities assays were conducted.Experimental results showed that ten compounds were(+)-3-hydroxy-ionone,ryegrass,loliolide,5-hydroxyl-3,4-dimethyl-5-pentyl-2(5-1-1)furanone,pheophorbide a ethyl ester,cholesterol,3?-hydroxy-5?,6?-epoxy-7-megastigmen-9-ketone,24-perhydroxyl-24-vinyl cholesterol,stigma-5,24(28)-diene-3?-D-alcohol and cholesterol-4-ene-6?-alcohol-3ketone.All compounds had antioxidant activities except pheophorbide a ethyl ester.To exploit new biofuel resource and optimize cultivation conditions of a fresh water microalgal strain Dictyosphaerium sp.1A10 for high biomass and high lipid accumulation,the medium and natural conditions were fully utilized.NaHCO3,glycine and 360 ?m/s·m2 were the best carbon source,nitrogen source and light density,respectively.The optimized cultivation conditions using response surface method(RSM)for Dictyosphaerium sp.1A10 were as follows:0.19 mmol/L of NaHCO3,30.00 mmol/L of glycine,0.1 g/L of K2HPO4 and 100 L/h of aeration for maximal biomass(2.25 g/L);0.94 mmol/L of NaHCO3,17.60 mmol/L of glycine,0.02 g/L of K2HPO4 and 100 L/h of aeration for maximal lipid content(41.76%);0.19 mmol/L of NaHCO3,14.86 mmol/L of glycine,0.1 g/L of K2HPO4 and 98.1 L/h of aeration for maximal lipid yield(0.74 g/L).Meanwhile,fed-batch was the best feeding strategy in outdoor scale-up cultivation.These results suggest that the optimized conditions could achieve the high-intensity and high-oil cultivation of Dictyosphaerium sp.1A10.Through the investigation of antioxidant components of Dictyosphaerium sp.1A10,lipids and polysaccharides were further sperated and purified to obain the depurated lipids and polysaccharides.Essence was made up of multiple natural ingredients without any synthetic antiseptic and cosolvent. |
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