| Pickles are traditional lactic acid fermented foods in our country and accepted widely for its unique flavor and nutritional value.Artificial inoculation of lactic acid bacteria in pickles can not only reduce time of fermentation and improve safety,stability and quality,but also help to reduce costs and adjust it to industrialized,standardized,large-scale production and control.In this study,lactic acid bacteria with excellent characteristics of fermentation were selected,and their special culture mediums were optimized through response surface methodology(RSM).The effects of lactic acid bacteria on texture and flavor of pickles was studied as well,and the fermentation process was optimized to improve quality and flavor of pickles while reducing fermentation time.The main findings are as follows:(1)LAB were isolated from Sichuan Laotan sauerkraut,Guizhou sour soup and fermented greens,the strains with good ability of acid producing and tolerance,resolving nitrite and tolerating salt were screened out,their bacteriostasis,passage stability,enzyme activity and freeze-drying survival rate were studied as well.The results showed that based on the studies including traditional morphological,physiological and biochemical characteristics,two LAB were isolated from 56 strains.The results showed that the strains 3-2-5-1’ and P-3-3-2 had good fermentation performance,the pH and total acid were 3.76,2.13%and 3.63,2.17%respectively after 48h fermentation;the degradation rate of nitrite was 82.99%and 83.43%,respectively.The antibacterial ability of 3-2-5-1’ and P-3-3-2 were strong,the antibacterial circle of E.coli,S.aureus and Salmonella were 18.45mm,17.42mm,15.65mm and 15.26mm,9.26mm,12.34mm respectively.They showed a good heredity stability by stability study,and the freeze-dried survival rates of which were 81.43%and 85.37%;the result of amino acid decarboxylase test was negative.3-2-5-1’ were identified as Lactobacillus plantarum and P-3-3-2 were identified as Lactobacillus fermentum by 16S rDNA sequence analysis.(2)The lactic acid bacteria proliferation medium was optimized.On the basis of single factor test of basal medium,nitrogen source,carbon source and growth factor,the key factors affecting the proliferation of two strains were determined by Plackett-Burman design,and the factor level was optimized by response surface method.The optimal whey medium of 3-2-5-1’,which was as follows:Whey powder 10.29%,Sugar 2.03%,Sodium citrate 0.52%,Soy protein isolate powder 1%,CaCO3 0.3%and Carrot juice 8%,was obtained rest on the analysis of regression model.Using the optimal medium to culture 3-2-5-1’ for 24h at 30℃,the viable cell numbers was 2.63×109CFU/mL.The best medium formula of P-3-3-2 was:Whey powder 5.84%,CaCO3 0.32%,Carrot juice 8.33%,Soy protein isolate powder 1%,Fructo-oligosaccharide 1%and K2HPO4 0.3%.The number of viable cells reached 2.69×109CFU/mL after culturing for 24h at 30℃.It was proved that the optimization results were accurate and reliable,and the response surface method was feasible to optimize the fermentation condition of lotus root,it had certain practical application value.(3)Effects of LAB fermentation on texture and flavor of lotus root pickles.Comprehensively analyzed various indexes,the best proportion of strains were confirmed to be 3-2-5-1’:P-3-3-2:Y430 = 1:1:1.In this study,the effects of fermentation conditions and supplementary materials on texture and flavor of lotus root pickles were studied by single factor experiment.A scheme of orthogonal test,which includes 3 factors and 3 levels,was designed.Temperature,salinity,inoculation were chosen to be factors,and total acid,hardness,sensory evaluation were chosen as evaluation indexes.The optimum conditions were as follows:Inoculation 10%,Temperature 25 ℃,Salinity 2%.Results of three indexes from verification experiment were:total acid 1.021%,hardness 43.ON,sensory evaluation 86.According to the PB test,the main factors of excipient addition were the dosage of sugar,pepper and spice.The results showed that the optimal dosage of these additive were 1.83%,3.61 ‰,1.84%respectively.The sensory score of the lotus root pickles made by this recipe was 92.The optimization results of supplementary materials was validated,the indexes of the pickles were hardness 49.2N,chromaticity 77.65,pH 3.49,total acid 1.09%and sensory score 90.(4)Different fermentation process were comparative studied and results showed that inoculation fermentation was better than natural fermentation.Inoculation fermentation was beneficial to the production and accumulation of lactic acid,the pH value was 3.2 and the total acid content was 1.292%after 9 days of fermentation at 25℃.The reductive VC of inoculation fermentation was lower than that of natural fermentation on the early stage of fermentation,and it was opposite later.The amino nitrogen kept a high content after its equilibrium which was 0.1%.Through the whole fermentation,the texture and color were maintained at a high level,the hardness was 39.8N and the color was 72.Inoculation fermentation also had a lower content of the ’nitrite’,around 0.6mg/kg.The number of LAB was 4×108 CFU/mL after 9 days of inoculation fermentation,the total number of colonies,molds and coliform bacteria were lower than natural fermentation in whole fermentation process.The number of yeast was at a low level,lower than natural fermentation except for the first day.The main volatile compounds of natural and inoculation fermented lotus roots were determined by GC-MS method.The volatile flavor substances detected by the two fermented methods were 69 and 54,respectively,of which 27 were the same.The main volatile flavor components of lotus roots were ethers,alcohols and alkenes.Among them,ethers accounted for 61.89%and 60.98%,made a greater contribution to the flavor for both.Results also showed that inoculation fermented lotus roots contains more alcohols,aldehydes,hydro carbons,acids,alkenes,sulfur compounds and less esters,ethers,ketones than natural fermentation.In terms of aroma species and relative content,inoculation fermentation was better than natural fermentation.The determination of biogenic amines in two kinds of pickles were conducted by HPLC.In addition to spermidine,the contents of tryptamine,putrescine,histamine and tyramine in natural fermented lotus root pickles were higher than that in inoculation fermentation,and the total content of natural fermentation was higher as well.Spermidine was the most important biogenic amine in lotus root pickles,the average content was 123.14mg/kg,followed by histamine,putrescine and histamine.The level of tyramine was the lowest(6.58mg/kg).According to FDA provisions,two types of pickles were within the safety standard range. |
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