| Fermented fruit juice by lactic acid bacteria contains rich nutritional and functional components,which can regulate the gastrointestinal tract,improve the immunity,and prevent the aging of human.Hence,fruit juice fermentation has gained much attention in the deep-processing of fruits and vegetables.However,there are some problems in juice fermentation by lactic acid bacteria at an industrial scale due to the unstable quality,slow growth of microorganism,and unclear metabolic mechanism,etc..Recently,many studies have reported that low frequency ultrasound can promote the growth and metabolism of microorganisms in fermentation.Studying the effect of low frequency ultrasound on the growth and metabolism of Lactobacillus plantarum in fruit juice and its mechanism can provide theoretical foundation and technical support for the application of low frequency ultrasound in the field of fruit and vegetable fermentation.In this thesis,the ultrasound conditions for promoting the growth and metabolism of Lactobacillus plantarum was optimized by investigating the changes in microorganism growth and metabolism during the juice fermentation by Lactobacillus plantarum,and the underlying mechanism was explored by transcriptome and proteomic analysis.The main results are as follows:1.The effects of low frequency ultrasound on the number of colonies,the content of sugars,organic acids,phenolics,and antioxidant activity during the fermentation of apple juice and strawberry juice by L.plantarum BNCC337796 were compared.Compared with strawberry juice,Lactobacillus plantarum BNCC337796 grew better in apple juice.Ultrasound promoted the growth of L.plantarum BNCC337796 in both apple and strawberry juice.At the 32nd hour of fermentation,the number of L.pdantarum in apple juice treated by ultrasound was 0.15 LogCFU/mL higher than that in the control group(p<0.05).The effect of ultrasound on the metabolism of sugars,organic acids and phenolics was more significant during apple juice fermentation.2.To analysis the effect of ultrasound conditions on growth,basic metabolism and phenolics conversion during the apple juice fermentation by lactic acid bacteria,a two-factor completely randomized trial was performed with growth phases(lag,logarithmic,and stationary phases)and power density(58.3 W/L,93.6 W/L)as independent variables.The results showed that the number of viable cells was significantly increased by ultrasound at lag and logarithmic phases.58.3 W/L ultrasound at lag phase and 93.6 W/L ultrasound at logarithmic phase were more prominent,which increased the number of viable bacteria by 0.43 and 0.31 LogCFU/mL,respectively(without futher incubation).Ultrasound increased the content of lactic acid.The lactic acid content in apple juice treated by 58.3 W/L ultrasound at lag phase and 93.6 W/L ultrasound at logarithmic phase was 21.8%and 26.3%higher than that of the control,respectively(without more incubation).The promotion effect on the bacteria growth and lactic acid synthesis disappeared when the growth of bacteria entered stationary phase,indicating that the ultrasound treatment mainly improved the growth and lactic acid synthesis rate of L.plantarum BNCC337796.Ultrasound also enhanced the metabolism of malic acid and had significant effects on other organic acids and amino acids.Ultrasound at lag phase and logarithmic phase significantly increased the contents of caffeic acid and catechin,decreased the contents of chlorogenic acid,proanthocyanin B2 and gallic acid during the fermentation.Ultrasound treatment can improve the antioxidant activity of fermented apple juice.After fermentation to 32.5h,the ABTS free radical scavenging capacity and ferric ion reducing power of apple juice with 93.6W/L ultrasound treatment at logarithmic phase were 17%and 8.1%higher than that of the control,respectively.Ultrasound at logarithmic phase also enhanced the membrane permeability of L.plantarum,especially with a power density of 93.6W/L.The effect of ultrasound at stationary phase was negligible.Considering the effects on the growth,metabolism and antioxidant activity of L.plantarum during the fermentation of apple juice,58.3 W/L ultrasound at lag phase and 93.6 W/L ultrasound at logarithmic phase were selected for further investigation.3.The genes of L.plantarumBNCC337796 under ultrasound treatment and normal culture(as a control)was analyzed by transcriptome sequencing.The results showed that 58.3 W/L ultrasound at lag phase improved the glycolysis of L.plantarum and inhibited the synthesis of tryptophan and arginine,which promoted the synthesis of energy and reduced its consumption,thus enabling more energyto be used for the growth and metabolism of L.plantarum.Under 93.6 W/L ultrasound treatment at logarithmic phase,the enhanced synthesis of folic acid increased the proliferation of L.plantarum,the up-regulated expression of pentose phosphate pathway provided more energy and raw materials for cell proliferation,while the down-regulated expression of gene related to ribosome protein means the reduction of ribosome synthesis and energy consumption.The down-regulation of genes related to the synthesis of fatty acids might change the composition of fatty acids in the cell membrane and improve the membrane permeability of L.plantarum.padR was up-regulated under 58.3 W/L ultrasound at lag phase,which was the transcriptional inhibitor ofpadA(phenolic acid decarboxylase),while padA was down-regulated under 93.6 W/L ultrasound treatment at logarithmic stage.The result indicated that the decarboxylation of hydroxycinnamic acid was inhibited by ultrasound treatment at both 58.3 W/L and 93.6 W/L ultrasound treatments.Under 93.6 W/L ultrasonic treatment at logarithmic phase,the up-regulated expression of tannin enzyme tanL and gallic acid decarboxylase activity related-gene UbiD could promote the conversion from tannins into gallic acid,and further the decarboxylation of gallic acid into pyrogallic acid.In addition,the up-regulated expression of lp2953 coding esterase could promote the hydrolysis of phenolics containing ester,like chlorogenic acid,and release more phenolic acids,such as coumaric acid,ferulic acid and caffeic acid.4.The differentially expressed proteins of L.plantarumBNCC337796 under ultrasound treatment and normal culture(as a control)was analyzed by iTRAQ Under 58.3 W/L ultrasound at lag phase,28 proteins were up-regulated and 37 proteins were down-regulated.Under 93.6 W/L ultrasound at logarithmic phase,149 proteins were up-regulated and 175 proteins were down-regulated.KEGG enrichment analysis showed that the biosynthesis of terpenes was significantly enriched under 58.3 W/L ultrasound at lag phase.Ribosomes,purine and pyrimidine metabolism,homologous recombination,oxidative phosphorylation and RNA degradation were significantly enriched under 93.6 W/L ultrasound at logarithmic phase.Similar to transcriptome analysis,the significantly down-regulated expression of ribosomal proteins inhibited the synthesis of ribosomal protein under 93.6 W/L ultrasound at logarithmic phase.The up-regulated expression of ribonucleoside-diphosphate reductase(nrdE)and DNA polymerase ?(polC),and the down-regulated expression of DNA-directed RNA polymerase(rpoZ)could enhance the DNA biosynthesis.Meanwhile,the homologous recombination and oxidative phosphorylation provided support for the correct DNA replication. |
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