Extractive Technology And Physical Degradation Of Polysaccharides From Porphyra Yezoensis

Porphyra is an impotant and economical alga in China.Its polysaccharides possess multi-pharmaceutic activities,and extractive and value-added technologies of polysaccharides from porphyra are key problems in the deep processing.Preparation technology,physicochemical property,ultrasonic degradation（kinetics and modification）,microwave degradation and its non-thermal effects of polysaccharide from Porphyra yezosensis（PSPY） were studied.The detail contents and conclusions are as follows.The contents of protein,fat and total sugar in P.yezoensis were determined.The Plackett-Burman design method was used to the screening of the main impact factors on the extraction rate of P.yezoensis polysaccharide,and results are extraction temperature （P=0.0017）,extraction time（P=0.0043）,ratio of sample to solvent（P=0.0180）.Extracting technology of polysaccharide was designed and carried out in the method of quadratic regression rotation design.The optimal condition with rather higher rate（purity） for extraction of polysaccharide are determined as follows:ratio of sample to solvent 1.16:100（1.16:100）（g/mL）,extraction time 4.7（1.3） h and extraction temperature 100（73.2）℃.In the present test,the order for affecting rate in the extraction is extraction temperature＞ratio of sample to solvent＞extraction time,and so the corresponding order for purity is ratio of sample to solvent＞extraction time＞extraction temperature.The regression model was effective and predictability, which was confirmed by experimental verification.PSPY was obtained in the optimal condition（with rather higher rate）,the rate is 19.5%,purity is 94.5%,sulfated group is 7.2%,3,6-AG is 12.3%,and monosaccharide composition is Gal、Xyl、Fuc、Ara,in the molar ratio of 46:2:1:1,respectively.The intrinsic viscosity of PSPY in distilled water at 25℃is 3.38dL/g.,Test with maillard reaction,browing is rather higher in acid condition than in the alkaline one,the least is in neutral condition,this may be affect by hydrolysis of PSPY.Equivalent point of PSPY is 7.15.PSPY has inhibition retrogradation of starch.Effects of PSPY on tumor cells growth were investigated.Results are as follows:in rather low concentration（＜200μg/mL）,PSPY has a certainty promotion on cell 293,but in rather high concentration（＞200μg/mL） PSPY has a certainty inhibition on cell 293.PSPY also has a certainty inhibition on cell SGC-7901and 95D,the highest inhibition activity as much as 20%.As cell U937 is concerned,PSPY has markedly promotion effect,and in concentration 1000μg/mL,the promotion activity is up to 26.82%.Antioxidation activity of PSPY was also studied.EC₅₀ of scavenging superoxide anion radical and scavenging hydroxyl radical is 0.18mg/mL and 1.32mg/mL,respectively,and good （r＞0.95） logarithmic activity-dose relationship was found here.Three homogeneity fractions were obtained,and molecular weight is 2.2×10⁵ Da、3.9×10⁵ Da and 6.1×10⁵ Da,sulfated group content is 7.4%、10.5%and 7.9%,monosaccharide composition in the molar ratio of PSPY3（Gal:Ara:Fuc,25.8:1.1:1）,PSPY4（Gal:Ara,27:1）, PSPY5（Gal）,respectively.Ultrasound degradation of PSPY was systematic studied.The Mark-Houwink equation was also extablished firstly,then,augmentation of power and enhance temperature can promote the degradation,and effect of concentration on the degradation is not monotony,the rather higher degradation is at concentration of 0.75g/dL,these were concluded from single factor investigation.Remarkable exponential relation between reaction rate constant and reaction time was found from dynamics equation established in different temperature conditions.Activation energy of the degradation is 52.13kJ/mol,which was estimated according to Arrhenius and dynamics equations. There is exponential relation between intrinsic viscosity of PSPY and reaction time, which was validated from theoretical derivation.Further study,as structure of UD-PSPY（Ultrasound degradation of PSPY） is concerned,distribution of molecular weight was notability altered and moved to the rather lower.As for anti-tumor activity of cell SGC-7901 and U937,the activities were observably improved,but the effects on activities for 95D and 293 were not too obviously changed.As about antioxidation, scavenging superoxide anion radical(UD-PSPY:EC₅₀=0.14mg/mL,PSPY:EC₅₀= 0.20mg/mL) and scavenging hydroxyl radical(UD-PSPY:EC₅₀=0.59mg/mL,PSPY: EC₅₀=1.32mg/mL) were also notedly improved.Three homogeneity fractions（F2,F3 and F4） were obtained from UD-PSPY, molecular weight is 2.1×10⁵Da,3.1×10⁵ Da and 4.0×10⁵ Da,sulfated group content is 7.6%,10.3%and 11.7%,monosaccharide composition in the molar rato of（Gal、Ara、Fuc）F2（48.5:1.2:1）、F3（39.0:1.1:1） and F4（36.4:1.1:1）,respectively.It has a linear backbone of alternating[G-A]and[G-L6S]in F2,[G-A]and[G-L6S]and[G-A2M]in F3,[G-A]and[G-L6S]and[G-A2or6M]in F4,which were analyzed by FT-IR and NMR spectroscopies.The order of activity scavenging superoxide anion radical is F4＞F3＞F2＞Vc,activity scavenging hydroxyl radical is F2＞F3＞F4＞Mannitol,chelating ability is EDTA＞F4＞F3＞F2,reducing power is Vc＞F2＞F3＞F4,respectively.Decrease of molecular weight was dominant reason for increases on activities of scavenging hydroxyl radical and reducing power of F1,F2 and F3,and increase of sulfate content in molecule was dominant reason for increases on activities of scavenging superoxide anion radical and chelating ferrous ions.Parameters of the microwave reaction instrument were mensurated.The apparatus log time is about 4s,and there is no markedly difference in rate of temperature increasing was found between volumes between 20mL to 40mL,which was confirmed by t test check.The non-thermal effects in microwave reaction were investigated by constant temperature-compare methord adopting MR as model reaction.Non-thermal effect（accelerating the MR,without changing the reaction path） in microwave reaction was found in the present study,which was authenticated and confirmed by browning, spectrum of ultraviolet-visible scanning,spectrum of synchronous fluorescent scanning and GC-MS analyses.There is no degradation for microwave on PSPY,but it can accelerate degradation of H₂O₂ on PSPY.As content of H₂O₂ is concerned,increasing of H₂O₂ content can promote the degradation,but up to 2%and 4%,the increasing action is very weak.Increasing microwave anode current can promote the degradation of H₂O₂ on PSPY.Non-thermal effect in microwave reaction was also validated in the degradation of H₂O₂ on PSPY.