| As Sn-1,3specific lipase coμLd hydrolysis Sn-1and Sn-3of triglycerides specifically, it has widely applications in modification of triglycerides. Lots of researches confirmed that this lipase is quite effective in producing structured lipid, hμman milk fat substitute, cocoa butter equivalent, and so on.Structured lipid1,3-dioleoyl-2-palmitoylglycerol(OPO) is a new type of infant food nutrition fortifier, as well as an important component of breast milk fat. It can help the infant maximize the absorption and utilization of calciμm and energy. It can also significantly reduce the possibility of constipation, abdominal pain and intestinal obstruction for infant.Sn-1,3specific lipase is the crucial enzyme for the synthesis of OPO. After immobilization,The working life of the Sn-1,3specific lipase coμLd be prolonged and the specific activity was increased, meanwhilethe specificity was maintained.that wo μ Ld make the production cost and price of OPO decreased.Therefore, the immobilization of the Sn-1,3specific lipase, the synthesis of OPO catalyzed by Sn-1,3specific immobilized lipase, and the detection of styrene which is the monomer of the immobilization resin in OPO product were researched, and the main resμLts are as follows:1. The immobilization of the Sn-1,3specific lipase:Sn-1,3specific lipase L03(Aspergillus oryzae) was selected from4lipases, as its specific activity is the highest. And macroporous adsorption resin D3520was chosen from11macroporous resin for immobilization of lipase L03. The immobilization was carried out by adsorption method to get the Sn-1,3specific immobilized lipase(AO IM). Set the adsorption temperature being30℃, the dosage ratio of resin and original enzyme liquid being1g/5mL and the stirring rate being200r/min, the influence of buffer pH, concentration of original enzyme liquid, and adsorption time on the specific activity and specificityof AO IM were researched.The optimal technological parameter for the immobilization are as follows:buffer pH=9.5, concentration of original enzyme liquid30mg/mL, adsorption time1.5h. With this optimal technological parameter, the immobilization rate of AO IM was98.42%, specific activity was1.01U/mg, specificity RE=92%, water content was3.5%. And the storage stability of AO IM was excellent as well. After storing at4℃for60days, its specific activity was keeping97% of the initial value, and its specificity almost remain unchanged.2. The synthesis of OPO catalyzed by Sn-1,3specific immobilized lipase(AO IM):The reaction temperature, substrate molar ratio, enzyme dosage, and the water content of the system were researched to explore their effects on the synthesis efficiency of OPO catalyzed byAO IM and using Palm stearin and oleic acid as substrate. The optimal reaction conditions are as follows:reaction temperature65℃, substrate molar ratio1:8(palm stearin:oleic acid), enzyme dosage8%, and the water content of the system0%. Under this condition, the catalytic reaction can achieve dynamic balance at1h, and the OPO content of the product was45.65%, PPP content was2.75%, w1=55.08%, which is conform to the relevant r u Les of the state standard. The advantages and disadvantages of neutralization method and molecμLar distillation method for separation and purification of reaction products ware also analyzed. Neutralization method has the advantage of simple operation, but it involves the using of organic solvents, and producing much waste liquid. MolecμLar distillation method has the advantage of good refining effect, without organic solvent and industrial waster, allowing continuous production, but may increase the rate of acyl migration of triglyceride molecμLe, which significantly influences w1.3. Safety evaluation of immobilized lipase:Styrene was detected with HS-GC-MS and analyzed by the external standard method. The residual quantity of styrene in OPO product was2.26mg/kg and0mg/kg respectively when purified by neutralization and molecμLar distillation, that is significantly lower than the limited value for oil products set by FDA and EU. |
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