Bioconversion Of Corn Stover To Lactic Acid By Thermotolerant Microbial Consortium

Lactic acid（LA）is an important platform chemical due to its significant applications in a wide range of fields including the food,pharmaceutical,cosmetic,leather,and chemical industries.Low-cost non-food lignocellulosic feedstocks from agro-industrial,forestry and municipal sources are considered attractive raw materials with the advantages of ready availability,sustainability,and renewability.The detoxification procedure was generally followed in the pretreated corn stover（CS）to remove inhibitor compounds.The microbial consortium shows great robustness against environmental disturbances and microbial invasion.Importantly,consortium can use less renewable and complex feedstocks displaying a higher bioconversion efficiency than monocultures,requiring less expensive purification processes.Firstly,the enrichment and selection of cattle stomach-dervied microbial consortia domesticated to degrade corn stover hydrolyzates（CSH）to produce LA at different temperatures was investigated.Bacterial 16S r RNA gene amplicon sequencing analyses indicated that the three microbial consortia were taxonomically distinct and Enterococcus became dominant at high temperature.The dominant genus of DUT37 was Escherichia/Shigella（69.22%）and Enterococcus（29.49%）.The dominant genus of DUT45and DUT47 were Enterococcus（96.58%）and Enterococcus（97.79%）.The final LA titer of42.64 g/L,43.91 g/L,and 43.56 g/L were obtained by microbial consortium DUT37,DUT45and DUT47 from non-detoxified CSH,respectively.Secondly,adaptive evolution was developed to obtain microbial consortium DTU50,with the high tolerance of temperature and lignocellulose-derived inhibitors,and co-utilization of glucose and xylose.Bacterial 16S r RNA gene amplicon sequencing analyses indicated that DUT50 contained Enterococcus（93.66%）,Lactobacillus（1.05%）,Bacillus（0.87%）and others（4.42%）.Thirdly,the key influential factors in SSF process,including dry biomass and enzyme loading,corn steep liquor powder concentration and pre-hydrolysis time were optimized.The LA concentration and yield reached 71.04 g/L and 0.49 g/g stover at 50^oC and p H 5.5 using the cellulase of 35 FPU/g stover,20 g/L corn stover liquid powder（CSLP）and pre-hydrolysis time of 4 h from 20%（w/v）H₂SO₄-pretreated and non-detoxified CS under non-sterile condition.Finally,mini consortia were isolated by series dilution of the original consortium DUT50to evaluate the potential interaction in original consortium.The performance of mini consortia showed the abilities of xylose utilization and LA production decreased with the increasing of dilution.Synthetic microbial consortium was constructed usingtwo isolated strains of E.faecium from DUT50.Itexhibited lower LA formation than original DUT50,while higher than the isolated strains.Based on the above results,it indicated that low abundance of Lactobacillus and Bacillus have non-negligible positive effect on LA production from non-detoxified CS.The unique properties of the adapted microbial consortium DUT50 were demonstrated by its excellent thermo-and inhibitors-tolerance during SSF process of lignocellulose utilization,which shorted the pretreatment process,reduced waste water and saved the costs.The result provided a promising strategy for bioconversion of lignocellulose to bio-based chemicals.