Efficient Dissolution Of Penicillin Mycelium And Enzymatic Hydrolysis Of Mycelium Protein

More than 2 million tons of antibiotics fermentation residues(AFRs)were produced in china every year,which were hazardous solid waste,and how to safely dispose and utilize them is attracting attention.AFRs are mainly composed of antibiotic mycelium.Antibiotic mycelium are rich in organic substances,such as protein,which accounts for about 50%in dry mycelium.The author's team proposed a promising suggestion,in which antibiotic mycelium are dissolved firstly,and then prepare amino acids,and use them to fermentation process.In this paper,the dissolution of penicillin mycelium by Alkali-thermal method,Acid-thermal method and low temperature NaOH/urea solution system were studied,and the distribution of mycelium protein,the amino acid composition and content in mycelium solution were determined.The enzymatic hydrolysis of penicillin mycelium protein in the solution was investigated,and the optimal enzymatic hydrolysis process was established.The main research contents and conclusions are as follows:(1)The dissolution of penicillin mycelium by Alkali-thermal method,Acid-thermal method and low temperature NaOH/urea solution system were studied.respectively.The results showed that the solvent compositions,the mass ratio of the mycelium to the solvent,the reaction temperature and time had significantly effects on dissolution of penicillin mycelium.Alkaline heating method showed the best performance on dissolution rate,the performance order was:Alkaline-thermal method>low temperature NaOH/urea solution system>the acid-thermal method.The optimal dissolution conditions in alkali-thermal method were as follows:sodium hydroxide concentration 4 wt%,mass ratio of solution to mycelium 5:1,temperature 50? and dissolving time 30 min.The mycelium dissolution rate reached 69.5%.The capacity of mycelium in the solution was 141.3 g/L,in which the protein and polysaccharide accounted for 25.3%and 9.5%to the mycelium mass,respectively.The optimal dissolution parameters of the low temperature NaOH/urea solution system were as follows:the solvent system composition 10 wt%NaOH and 12 wt%urea solution,mass ratio of solution to mycelium 8:1,temperature-8?,and dissolving time 50 min.The mycelium dissolution rate reached 69.3%.The mycelium capacity was 88.9 g/L in the solution,in which the protein and polysaccharide accounted for 23.5%and 15.3%to the mycelium mass,respectively.Compared with the alkaline-thermal method,the polysaccharide content in the low-temperature NaOH/urea solution system increased by 5.8 percentage point.(2)On the basis of penicillin mycelium dissolution by the Alkali-thermal method,the distribution characteristics of mycelium protein in the solution were investigated by isoelectric point precipitation,and the amino acid composition and content in the mycelium solution were determined by PITC pre column derivatization method.The isoelectric points pH of protein in the mycelium solution was at pH 2-11.At pH 4,pH 7 and pH 9,the protein precipitation ratio were 77.7%,30.5%and 18.3%respectively.That indicated the isoelectric point of mycelium protein at pH 4 and pH 7 were significantly higher than that at pH 9.16 kinds of amino acids were detected in mycelium solution,and aspartic acid,glutamic acid,leucine,alanine and glycine accounted for 20.59%,13.20%,10.91%,8.11%and 7.15%of the total amino acids,respectively.(3)The enzymatic hydrolysis of penicillin mycelium protein was explored in this solution using enzyme as catalyst,such as Alcalase(A),Bromelain(B)Neutrase(C),and Acid protease(D).The results showed that the enzymes types,the ratio of enzyme to protein,temperature and time had obviously effect on enzymatic hydrolysis of penicillin mycelium protein.In single enzyme catalyzed hydrolysis process,Alcalase showed the best performance,the hydrolysis degree of protein reached 31.43%at following conditions,the mass ratio of Alcalase to protein as 6%,pH 10,temperature 50? and reaction time 3 h.The experiment results demonstrated that the complex enzyme constituted of Alcalase and Bromelain as catalyst gave a better performance on enzymatic hydrolysis degree than Alcalase as catalyst.The hydrolysis degree of mycelium protein reached 42.73%at the optimal conditions,the mass ratio of complex enzyme to protein as 9%(A:B=2:1),pH 10,temperature 50? and reaction time 3 h.The hydrolysis degree of mycelium protein was increased by 11.3%percentage point than the best single-enzyme method.The HPLC analysis measured 16 amino acids in the enzymatic hydrolysate,such as glutamic acid,aspartic acid,glycine,leucine and alanine,in which the ratio was 15.47%,11.94%,9.41%,9.23%,9.22%,respectively.