Study On The Preparation Of Calcium Lactate From Fermented Eggshell By Immobilized Enterococcus Mundtii

In the production process,most eggshells were discarded as waste.Such treatment method would easily cause the remaining egg white and other materials to produce microorganisms,which would not only cause environmental pollution,but also cause waste of eggshell resources.Studies had shown that eggshells could be used in a variety of fields such as medicine,chemical industry,and nutrition enhancement,and the development and utilization of eggshells had become a current research hotspot.Therefore,in order to realize the resource utilization of eggshell,this study took eggshell as raw material and immobilized cell fermentation technology as the basis,explored the physical and chemical indexes of different embedding carriers and their effects on the yield of calcium lactate in fermentation broth,and determined the optimal embedding carrier.The influence of eggshell powder in the fermentation broth on the mechanical strength stability of the immobilized cells was investigated by structural characterization Based on the single factor experiment,the process conditions of shake flask fermentation were optimized by response surface experiments.Based on the technological conditions of shake flask fermentation,an expansion culture of a 5 L fermentation tank was carried out,and the parameter settings for preparing eggshell source calcium lactate in the fermenter were explored.The main findings were as follows:1. In this experiment,the effects of sodium alginate?SA?,sodium alginate-polyvinyl alcohol?SA-PVA?and sodium alginate-activated carbon?SA-C?on the preparation of calcium lactate from eggshell by immobilized Enterococcus mundtii cells were studied.Investigated and compared the mechanical strength,mass transfer coefficient,embedding rate,calcium lactate production performance,and reusability stability index of different embedding carriers to determine the most suitable embedding carrier,and related performance of the most suitable embedding carrier was determined.The results showed that SA-C carrier was the most suitable embedding carrier,and the highest yield of calcium lactate was 101.02±1.83 g/L,and the high yield was maintained after 18 times of repeated fermentation,with a tendency to shorten the fermentation period after 10 times of repeated fermentation.The results of scanning electron microscopy showed that after repeated fermentation for 18 times,a large number of bacterial cells still gathered inside the SA-C embedded carrier,and the internal structure strength was still good,maintaining high-density cell fermentation,which may be the main reason for its tendency to shorten the fermentation cycle after repeated fermentation for 10 times.Compared with free cells,the operating stability,temperature,p H and storage stability of the cells were significantly improved in a certain range.2. The performance of SA-C immobilized cells was explored by measuring Fouriertransform infrared spectroscopy?FTIR?and thermogravimetric analysis?TGA?.The swelling ratio,X-ray diffraction?XRD?and X-ray energy spectroscopy?EDS?explored the effect of eggshell powder in the fermentation broth on the mechanical strength stability of the immobilized cells.TGA results showed that the addition of activated carbon improved the thermal stability of the immobilized cells.The results of the swelling rate showed that the presence of eggshell powder in the fermentation broth effectively prevented the immobilized cells from swelling with water,and the swelling rate was too high,which easily caused the mechanical strength of the immobilized cells to weaken.In addition,the results of XRD and EDS showed that the presence of egg shell powder in fermentation broth could effectively provide calcium ions(Ca²⁺)to maintain the mechanical stability of immobilized cells.3. Enterococcus mundtii cells were embedded with the optimal sodiumalginate-activated carbon as the optimal embedding carrier.On the basis of single factor,the optimal process conditions for the production of calcium lactate from eggshell were optimized by response surface experiments.The response surface experiment results showed that the optimal fermentation conditions were:sodium alginate concentration of3%,activated carbon concentration of 1%,eggshell powder content of 11g,inoculation amount of 1.6%,and the influence of each factor on the yield of calcium lactate in fermentation broth was successively:eggshell powder>inoculation amount>sodium alginate>activated carbon.Under these conditions,the yield of calcium lactate in the fermentation broth reached 105.52±1.07 g/L.4. Based on the optimized process conditions of shake flasks,an expanded cultureof a 5 L fermenter with eggshell-derived calcium lactate from immobilized Enterococcus mundtii cells was carried out to explore the effects of parameters such as aeration rate,stirring speed,eggshell powder content,inoculum and initial glucose concentration on the yield of calcium lactate in the fermentation broth.As a result,the optimal fermentation tank parameters were set as ventilation rate 1.2v v^-1m^-1,stirring speed 180rpm,eggshell powder content 6%,inoculum content 4%and initial glucose concentration80 g/L.Under this condition,the yield of calcium lactate in the fermentation broth reached 98.38±2.12g/L.Compared with free cell fermentation?calcium lactate g/L h:1.32?under the same fermentation condition,the yield of calcium lactate in immobilized cell fermentation?calcium lactate g/L h:1.46?was higher.