Effects Of Salt Stress On Growth,Ion Response And Metabolites Of Spirulina Platensis

Spirulina platensis is a multicellular spiral prokaryotic cyanobacteria,which is often used as an important nutrient and drug resource because it contains a large amount of biological active substances such as proteins,polysaccharides,vitamins and unsaturated fatty acids.Spirulina has the advantages of relatively fast growth rate,large-scale and commercial cultivation potential,and high photosynthetic efficiency.S.platensis can produces a large number of metabolites such as extracellular polymeric substances(EPS),unsaturated fatty acids,proline,carotenoids,phycocyanin,etc.Studying the changes of physiological responses to S.platensis under salt stress is of great significance for improving the production of biologically active metabolites and revealing salt tolerance mechanisms.At present,the research on salt tolerance mechanism of S.platensis has focused on antioxidant components and metabolites,and there is no uniform and systematic conclusion.At the same time,there are few studies on the regulation mechanism of S.platensis ion in salt stress.Therefore,this paper selected S.platensis as the experimental material,and studies the salt tolerance from four aspects:the growth,ion current,EPS and intracellular metabolic products.The result are as follows:1.Effects of salt stress on the growth of S.platensisDifferent concentrations(0,0.1,0.2,0.3,0.4,0.5,0.6 mol·L-1)of NaCl were used to treat S.platensis and its growth indexes such as OD560nm,chlorophyll,aggregation rate and phycocyanin were detected.The results showed that different NaCl concentrations had effects on the growth of S.platensis,and the low concentration promoted its growth.The promoting effect of 0.1 mol·L-1 was more obvious,and the high concentration(0.3 mol·L-1 or more)inhibited the growth.And low NaCl concentration(0.1,0.2,0.3 mol·L-1)promoted the accumulation of chlorophyll and carotenoids.The aggregation rate showed a significant increase at 1 d of NaCl stress,and the increase rate was 61.6%-193.8%.The aggregation rate in the stable phase was in a stable state,and the low NaCl concentration(0.1,0.2,0.3 mol·L-1)was not significantly different from the control group,which was higher than that of the high concentration NaCl.The phycocyanin in the stable phase(15 d)decreased significantly with salt stress(except 0.6 mol·L-1).2.Effects of salt stress on the ion current of S.platensisThe effect of salt stress on the ion current of S.platensis was studied by non-invasive micro-test technique(NMT).The results showed that Na+ of all parts(end,middle and tail)of CK in the control group showed influx under different treatment time(1 d,6 d,12 d),and increased significantly with time,K+was expressed as efflux.S.platensis maintains intracellular ion balance,regulates water balance,and maintains metabolic activities such as photosynthesis by controlling the efflux of Na+ and K+ ion currents.Ca2+ showed inconsistency in different parts.The results showed that Ca2+ had a greater demand for Ca2+in the middle of S.platensis,which had stronger metabolic activity.Under NaCl stress(0.1,0.6 mol·L-1),Na+showed efflux.As the NaCl concentration increased,the net flow rate of the efflux increased significantly,and with the increase of the stress time,the efflux rate decreased.K+showed efflux in the early stage of stress(1 d,6 d),and the K+efflux rate increased with the increase of salt concentration.In the late stage of stress(12 d),the rate of efflux decreased with the increase of salt concentration,and showed an inflow at 12 d under 0.6 mol·L-1 treatment.Ca2+showed influx in all parts during NaCl stress,and the influx rate increased significantly with the increase of NaCl concentration and stress time.S.platensis can resist salt stress by reducing the efflux ability of Na+and K+under high NaCl concentration stress.The Na+,K+and Ca2+flows in different parts have significant differences,which are mainly in the middle>end>tail.In summary,under salt stress,S.spirulina regulates K+/Na+ and promotes Ca2+influx by inhibiting Na+ influx and K+efflux to improve the ability to resist salt stress.3.Effect of salt stress on the EPS of S.platensisThe response surface methodology was optimized for the ultrasonic extraction conditions of S.platensis B-EPS.The optimal extraction conditions were determined:ultrasonic temperature 52.02?,amplitude percentage 57.88%,ultrasonic time 6.91 min,B-EPS content was 184.78 mg·g-1 under these conditions.Different concentrations of NaCl treatment affected the EPS contents,the total S-EPS range was 410.03-643.35 mg·g-1.The polysaccharide contents of S-EPS increased significantly with the high NaCl concentration(0.4,0.5,0.6 mol·L-1)treatments,and was higher than the protein content.The contents of B-EPS polysaccharide increased with the concentration of NaCl,and the content decreased first and then increased.The total B-EPS ranged was 177.88-255.96 mg·g-1,and the polysaccharide contents increased with the concentration of NaCl.The protein contents decreased first and then increased,and the DNA concentration increased significantly at high NaCl concentration(0.5,0.6 mol·L-1).The results of Fourier transform infrared spectroscopy indicated that B-EPS contained protein(C=O,C-N and N-H),carbohydrate(C-O-C),sugar ring(-C-O),benzene ring-containing substances(C-H)and other groups.S-EPS contains protein(C=O,C-N and N-H),carbohydrates(C-O-C),a sugar ring(-C-O),a fatty substance(C-H or CH2,-CH3)and the like.Under different NaCl concentrations stress,the EPS composition had changed.The S-EPS contained lipids under low NaCl concentration.At any NaCl concentration,benzene rings were detected in B-EPS.4.Effects of salt stress on the antioxidant enzymes and secondary metabolic products of S.platensisThe activities of antioxidant enzymes(SOD,POD,CAT),MDA,proline content and fatty acid content were detected under different concentrations of NaCl.The results showed that NaCl stress enhanced the activity of antioxidant enzymes(SOD,POD,CAT).The three antioxidant enzymes had similar regulation.The three antioxidant enzymes increased significantly under low salt stress,and the increase of antioxidant enzymes decreased under high salt stress.As the concentration of NaCl increased,the contents of MDA and proline increased significantly(p<0.05),and the contents of phenolic compounds decreased.The fatty acid of S.platensis is dominated by even fatty acids.?-linolenic acid(C18:3n6)is the most important fatty acid of S.platensis.NaCl stress has no significant effect on the total amount of fatty acids,and under 0.1 mol·L-1 NaCl stress,the content of oleic acid(C18:1n9c)increased significantly.Based on the above studies,the growth,ion regulation,EPS,antioxidant enzyme system,osmotic adjustment substance and fatty acids were systematically investigated in this thesis.Taken together,this study clarifies the effects of salt stress on the growth and metabolism of S.platensis and their mechanisms.