Extraction And Quantitative Detection Of Extracellular Polymer For Early Warming Of Acidification

Food waste has the characteristics of high organic matter content and high fat content,and is easily acidified during the anaerobic digestion process under high load.However,due to the complexity of the anaerobic digestion system,there are certain limitations in the currently used indicators in the operation of the system.The development of a new effective early warning indicator becomes particularly important.Extracellular polymer(EPS)is a macromolecular polymer formed by microorganisms under certain environmental conditions.It is closely related to the life state of microbial cells and can accurately reflect the state of the system to a certain extent.Based on the above problems,this topic optimized the extraction method of EPS in the simple system anaerobic digestion process,studied the changes in EPS during the anaerobic acidification process,and explored its early warning capability as an early warning indicator for anaerobic acidification.At the same time,fluorescent dyes are used to mark the EPS to achieve quantitative detection of specific substances in the EPS sample.The main results are as follows:(1)By comparing the five commonly used extraction methods,the thermal extraction-CER method is determined to be the best extraction method for simple system EPS.In the design single factor experiment,50℃,60℃,and 70℃ were selected as the three levels of heating temperature;20 min,25 min,and 30 min were the three levels of heating time;80 r/min,120 r/min,and 160 r/min were three levels of stirring rate;2 h,3 h and 4 h were three levels of stirring time.The four-factor three-level orthogonal experiment was designed,and the best extraction conditions for EPS were:heating temperature 60℃,heating time 25 min,stirring rate 80 r/min,stirring time 3 h.(2)In the high F/M acidification system,the traditional indicators of pH value,unit VS gas production and early warning time of VFA are 26 h,24 h and 22 h,respectively.These indicators are quite different due to changes in the reaction system and cannot be separated.As an effective indicator of acidification instability.Using the protein to polysaccharide concentration ratio PN/PS as the acidification warning index about 18 to 20 hours to make an early warning,about 5 hours earlier than other commonly used indicators,can be used as an effective indicator of acidification warning.(3)The positively charged Cy5 molecule is designed to response to the negatively charged EPS by electrostatic interaction.A new absorption peak at 560 nm of the absorption spectrum can be used to indicate the combination of Cy5 and EPS.The response time of the interaction is less than 30 s.The interaction between Cy5 and EPS was verified by absorption spectrum,fluorescence spectrum and Zeta potential method.Furthermore,Cy5 is further used as a probe to detect the specificity of sodium alginate in EPS.According to the standard curve method,the concentration of sodium alginate in EPS was 1.01μg/mL.This probe based method can realize quantitative,rapid and accurate detection of sodium alginate in EPS,which provides theoretical guidance for EPS detection in practical production.