Optimization Of The Key Enzymes Involved In The Efficient Liquefaction And Saccharification Of High Concentration Corn Starch

This research is supported by the National Science and TechnologyFoundation in the Twelfth Five-year Plan (2012BAD34B00), and theResearch Project of the Key Techniques in Production of High Quality CornSyrup (20130305026NY). In this study, the corn starch solution (50%, w/w)was prepared as the experimental material, and the high performance liquidchromatography (HPLC) was employed for determination. The optimalparameters for enzymatic liquefaction and saccharification of highconcentration corn starch were obtained by a series of research methods,such as the single factor experiment, the response surface optimizationexperiment and the quadratic orthogonal rotary regression experiment. Theultrasonic treatment was also taken into account. Based on the obtainedoptimal parameters for the enzymes, the influence of ultrasonic pretreatmentwith the optimized treatment time and ultrasonic power on the compositionof starch syrup was analyzed by the HPLC method. The main results are asfollows:(1) In research of the key enzymes involved in the efficient liquefactionand saccharification of high concentration corn starch, the influence of the treatment time, temperature, dosage of medium-temperature α-amylase andheat-stable α-amylase, and the amount of the CaCl2on the liquefaction ofcorn starch solution of high concentration was studied. Taking DE value asthe measuring index, I established a liquefaction method for corn starch athigh concentration. The50%(w/w) corn starch solution was added with14u/g medium-temperature α-amylase and0.2%(w/w) of anhydrous calciumchloride for liquefaction at70℃for23minutes. The following step wasthe addition of12u/g heat-stable α-amylase for liquefaction at94℃for18minutes. This product was qualified for the next saccharification step. Thiscombining use of these two enzymes makes the corn starch solution easy tostir up and mix well. In addition, the viscosity of the high-concentrationcorn starch solution was effectively reduced.(2) In research of the optimization of key enzymes, the cooperativeroles of the amyloglucosidase, β-amylase and pullulanase in saccharificationwere studied in the liquefied products. Through the single factorexperiments and a quadratic orthogonal rotary regression experiment, theregression equation is DE=81.73353+1.12622X1+0.88671X2+0.38926X3-0.099023X1X2+0.11772X1X3-0.24515X2X3+1.11873X221+0.39588X2+0.51361X23. The optimal process parameters of the enzymes were obtainedto establish a saccharification method for corn starch at high concentration.The saccharification was conducted by adding β-glucoamylase (331.8u/g),pullulanase (0.354AUSP) and myloglucosidase (59.5u/g) at60℃for40h. (3) In determination of the saccharified products and pratical researchof the ultrasonic-assisted enzymatic hydrolysis of high concentration cornstarch, the high performance liquid chromatography (HPLC) method wasemployed to determine the content of main components of the obtained cornstarch syrup. The calibration curves andregression equations for glucose,maltose and maltotriose, respectively were obtained by means of theexternal reference method. Thus the content of glucose, maltose andmaltotriose in the syrup product was598mg/mL,20.5mg/mL, and6.5mg/mL respectively. Based on these results, the influence ofultrasound-assisted hydrolysis of corn starch was also investigated. Takingthe DE value as the measure index, the technological parameters, thetreatment time and ultrasonic power, were optimized via single-factor tests.After comparing the products with/without ultrasonic treatment, it is foundthat the ultrasonic could improve the reaction rate of saccharification, whichcontributes to the increase of starch sugar yield. The results provide atheoretical guide for the industrial production of starch sugar.