Screening Of Nitrite-lowering Lactic Acid Bacteria In Fermented Capsicum And Study On Its Nitrite-lowering Mechanism

Fermented Capsicum is a special processed food in Hunan..Naturally fermented capsicum is mainly fermented by lactic acid bacteria attached to the surface of capsicum,but the process of Capsicum fermentation will produce nitrite,which will cause food safety risks.In this paper,fermented Capsicum in different areas of Hunan Province was used as raw material to analyze the correlation between p H value,total acid,salt content and nitrite in fermented capsicum samples.Then,the lactic acid bacteria with high efficiency in degrading nitrite were screened out from the samples,and their prebiotic characteristics and degradation mechanism were studied.The following conclusions were obtained:(1)The nitrite content in 65 fermented Capsicum samples collected ranged from 0.55to 21.42 mg/kg,with an average of 6.29 mg/kg.Among them,the nitrite content of three fermented Capsicum samples exceeded the national nitrite limit standard for cured vegetables,which were 20.1 mg/kg,20.65 mg/kg and 21.42 mg/kg,respectively.There were potential food safety hazards.There was a very significant negative correlation between salt content and total acid content in fermented Capsicum(P<0.01),a significant negative correlation between nitrite content and total acid content(P<0.05),and a very significant positive correlation between nitrite content and p H value(P<0.01).(2)Six strains of high-efficiency Nitrite-Degrading lactic acid bacteria,H3D,L13-13,X14-1,X14-7,Z1-6 and YY-6,were screened out from the samples,and the nitrite degradation rates were 79.3%,86.3%,85.4%,87.7%,80.9%and 83.9%in 24 h fermentation,respectively.It was identified as Lactobacillus plantarum H3D,Lactobacillus pentosus L13-13 and X14-1,Lactobacillus delbrueckii X14-7,Lactobacillus brevis Z1-6,Lactobacillus casei YY-6.(3)In the simulated gastrointestinal tolerance test,the survival rates of Lactobacillus brevis Z1-6 and Lactobacillus pentosus X14-1 were lower,and Lactobacillus delbrueckii X14-7 and Lactobacillus casei YY-6 showed better simulated gastrointestinal environmental tolerance.In hydrophobicity test,Lactobacillus plantarum H3D,Lactobacillus delbrueckii X14-7 and Lactobacillus pentosaceus X14-1 had strong hydrophobicity and might have strong cell adhesion.Six lactic acid bacteria showed different degrees of resistance in antibiotic susceptibility test,among which Lactobacillus delbrueckii X14-7 and Lactobacillus casei YY-6 were intolerant to multiple antibiotics with the highest safety;Lactobacillus brevis Z1-6 could tolerate multiple antibiotics with the risk of drug resistance gene transfer.In the amino acid decarboxylase test,Lactobacillus brevis Z1-6 lysine decarboxylase reacted positively with histidine decarboxylase,and Lactobacillus casei YY-6tyrosine decarboxylase reacted positively,which was not suitable for fruits and vegetables fermentation and probiotic lactic acid bacteria research(4)Under nitrite stress culture,Lactobacillus delbrueckii X14-7 was less affected by nitrite,and the acid production capacity of Lactobacillus brevis Z1-6 increased,while the rest strains were inhibited.Nitrite degradation rates of Lactobacillus delbrueckii X14-7,Lactobacillus brevis Z1-6,Lactobacillus pentosus L13-13 and Lactobacillus pentosus X14-1increased while those of Lactobacillus H3D and Lactobacillus casei YY-6 decreased when cultured at constant p H value in the fermenter.The results of nitrite reductase assay showed that Lactobacillus delbrueckii X14-7,Lactobacillus brevis Z1-6,Lactobacillus pentosus L13-13 and Lactobacillus pentosus X14-1 had higher nitrite reductase activity.In the metal ion catalyzed nitrite reductase test,Cu²⁺,Fe²⁺and Fe³⁺all had the promoting effect on the nitrite reductase activity of the above four lactic acid bacteria metabolism,among which Cu2+had the strongest promoting effect,so it was speculated that the nitrite reductase extracted from the four lactic acid bacteria might all be copper-type nitrite reductase.(5)Lactobacillus delbrueckii X14-7,which has excellent ability to degrade nitrite and probiotic characteristics in vitro,was selected as the test bacteria for transcriptional sequencing under nitrite stress.Transcription data showed a total of 1152 differentially expressed genes,589 up-regulated genes and 563 down-regulated genes.Differentially expressed genes were mainly enriched in carbon metabolism pathway,glycolysis and gluconeogenesis pathway,microbial metabolism pathway under different environments,pentose phosphate pathway,fructose and mannose metabolism,amino acid biosynthesis pathway and other pathways,but not in nitrogen metabolism pathway.Through functional analysis of differentially expressed genes,we found that a large number of redox protein genes were up-regulated under nitrite stress.Among them,the copper oxyreductase encoded by LDB?RS02850 is structurally similar to the copper nitrite reductase and may be a potential nitrite reductant protein gene.