Study On The Extraction And Modification Of Active Components From Krill Antarctic By Multi-liquid Phase Enzyme Catalysis System

The biological resources of Antarctic krill(Euphausia superba)are huge,and the nutritional value is high.Astaxanthin,krill protein and lipids combined with n-3 PUFAs are contained in krill,which has attracted more and more attention in food,medicine and health products.However,most of the current traditional extraction methods can only extract the mixture of active ingredients in krill.The separation of the active ingredients is difficult,and the extraction efficiency is not high,which severely limits its application.Therefore,the development of efficient and green separation and extraction methods is a research hotspot in this field.The three-liquid phase system is a new type of enzyme catalytic system with fast reaction rate and simultaneous separation of reaction materials.In this paper,different three-liquid phase systems are constructed in combination with enzymatic reactions.Explored the active ingredients in Antarctic krill are simultaneously extracted and separated using protease.The active components in krill oil were modified with Lecitase Ultra phospholipase,and the microstructure of the system was further discussed.The main research results of this paper are as follows:A three-liquid phase system suitable for enzymatic extraction of active ingredients in krill was constructed.The system and hydrolytic protease were screened,and papain with better catalytic effect was obtained.The n-hexane/PEG-400/Na₂SO₄system is the best three-liquid system combined with papain to be established to separate and extract the active components from Antarctic krill simultaneously.The influence of the concentration of phase-forming substances and reaction time on the enzymatic hydrolysis reaction in this system was explored.When the concentrations of PEG-400 and sodium sulfate are 20%and14%,respectively,after 4 hours of reaction,phospholipids,astaxanthin and glycerides were almost all extracted into the top phase contains oil.The amino acids of the hydrolysate are mainly distributed in the middle and bottom phase.Protease is mainly recovered in the middle phase,and the recovery rate of enzyme activity can reach 90.07%.The relative enzyme activity is still more than 80%after 8 batches of repeated use of the enzyme-rich middle phase.A three-liquid phase system suitable for phospholipase Lecitase Ultra was constructed and screened.Through modification of phospholipids,the enrichment of n-3 PUFAs in lipids and the effective separation of astaxanthin and n-3 PUFAs were achieved.The study found that after the reaction of the three-liquid phase system with PEG-600/Na₂SO₄as the phase-forming substance,an emulsion layer rich in n-3 PUFAs can be formed between theupper and middle phases,that is,a multi-liquid system is formed.The influence of the concentration of phase-forming substances and different p H values on the catalytic efficiency is investigated.When the content of Na₂SO₄and PEG-600 are 18%and 19%,and the p H of the system is 6,the content of n-3 PUFAs is increased from 20%to approximately 55%.Phospholipase is recovered in the middle phase and the recovery rate can reach 95.06%,and the recovery rate of phosphorus can reach about 90%after 2h of the reaction.The reuse of the enzyme-rich mid-phase is further explored,and it is found that the relative enzyme activity of the mid-phase after repeated 8 batches is still 80%,and the n-3 PUFAs content is over 38%.The effects of oil-water system and multi-liquid phase system on the catalytic efficiency of enzymatic reactions were compared.The results show that the reaction efficiency in the multi-liquid phase system has been improved,but the improvement of the interfacial enzymes is more obvious.In order to study the cause of this phenomenon,the microstructure of the system was explored with Malvern particle size analyzer and laser confocal.It is found that the particle size distribution in the multi-liquid system is smaller than that of the traditional oil-water system.Observed with a confocal microscope,a new emulsion structure will be formed in the multi-liquid phase system.In the emulsion structure,due to the presence of amphiphilic phospholipids,the oil is more dispersed in the system and the catalytic efficiency of interfacial enzymes is improved.