Preparation Of Polysaccharide Bacteriostatic Protein Complex Film And Initial Applications

The advantages of biological methods to control postharvest disease had gradually exceed those traditional preservation methods,the development of anti-bacterial control products had become a major in the field of preservation of post harvest fruits and vegetables.However,the application of biocontrol microbiologies was limited to the direct action of antagonistic microbial bacteria or biological antimicrobial agents on pathogens and fruits,which is difficult to maintain effectiveness and industrialization.In this paper,the antibacterial protein extract preliminarily isolated and purified from the culture supernatant of antagonistic microbial membrane Pichia membranaefaciens preserved in the laboratory was blended with food grade membraneforming materials to prepare composite membranes.The optimal formulation of the composite membrane was determined by single factor experiment and response surface experiment,and the effect of antibacterial protein extract on the performance of the composite membrane was studied.The inhibitory effect of the antibacterial protein film on the pathogen Alternaria in vitro and in vivo was studied.The antibacterial protein composite film was preliminarily applied to broccoli fruit,and the effects of the antibacterial protein composite film on the physiological quality and fresh-keeping effect of broccoli were studied.The main conclusions are as follows:(1)Through single factor experiment and response surface experiment,the influence order of each film-forming material on the water vapor transmission coefficient of composite membrane is determined: glycerol addition > carrageenan proportion > corn starch addition > carrageenan solution concentration.The optimum formula of film formation was determined.The concentration of corn starch was 6%(w/v),the concentration of carrageenan solution was 0.4%(w/v),the addition of glycerol was 1.0%(w/v),and the proportion of carrageenan was 50%.At this time,the water vapor transmission coefficient of the composite film was 0.73.(2)The addition of bacteriostatic protein extract had a significant effect on the color,transparency and mechanical properties of the composite membrane.The greater the concentration of bacteriostatic protein,the more the color of the composite membrane changed to yellow.When the protein extract was diluted to 16 times,that is,when the concentration of protein extract was 0.625 mg/ml,the B~* values of the basic composite membrane and bacteriostatic membrane were 20.4000 and 20.6667 respectively,and there was no significant difference in yellowness between them(p >0.05).The addition of protein extract decreased the tensile strength(TS)and increased the elongation at break(EAB)of the composite membrane,which changed the composite membrane from brittleness to flexibility.When the concentration of protein extract was 10 mg/ml,TS decreased by 51% and EAB increased by 43%.The transparency of the composite membrane will decrease with the increase of the concentration of bacteriostatic protein extract.When the protein extract is diluted to 16 times,that is,when the concentration of protein extract is 0.625 mg/ml,there is no significant difference between the two(p>0.05).At this time,the transparency of the composite membrane is 76%.In addition,the protein extract had no significant effect on the thickness,water vapor transmission coefficient and expansion rate of the composite membrane.Through Fourier transform infrared spectroscopy analysis,the protein extract has good compatibility with the basic film-forming materials,and crosslinking may occur,which is manifested in the change of hydrogen bond.(3)The results of plate confrontation test and drug containing plate test showed that the protein extract had a significant inhibitory effect on hyphal expansion of Alternaria(p<0.05);The film-forming materials had no obvious inhibitory effect on Alternaria,and the film-forming materials did not interfere with the antibacterial function of protein extract.The inhibition rate of protein extract against Alternaria reached 95% on the second day of culture,and decreased to 74% on the seventh day of culture.The results showed that the inhibitory effect of antibacterial membrane solution on hyphal expansion of Alternaria was the most significant in the period of 2-6 d.The minimum inhibitory concentration of protein extract against Alternaria was 0.1667 mg/ml.In addition,on the broccoli fruit,the antibacterial membrane solution still had a significant inhibitory effect on the growth and reproduction of Alternaria.(4)The indexes such as chromaticity,chlorophyll content,vitamin C and weight loss rate of broccoli fruit during storage were studied.The results showed that the antibacterial protein composite membrane could effectively delay the yellowing phenomenon of broccoli fruit,effectively reduce the weight loss rate of broccoli to a certain extent,delay the decomposition of chlorophyll and the decrease of vitamin C content,maintain the good sensory quality of broccoli and increase the storage time.