| The naturally fermented soybean pastes could bring a series of problems, including inferior technology, long fermentation periods, small production scales, low efficiencies, low utilization rates of materials, and unguaranteed product safety. All these contribute to the great hidden danger in the final products. In the present study, soybean pastes were made using Aspergillus oryzae huniang 3.042. Changes of several key indexes, the characteristic flavor compounds and microorganism composition were analyzed during the natural fermentation. The key flavory compound and key microorganisms were identified and a strategy on the multi inoculation and multi-temperature fermentation was adopted. The fermentation period decreased and the flavors were well maintained.Changes of the principle components of the soybean paste were investigated during the natural fermentation. Total acids and pH rapidly changed at the beginning two to four weeks, and then gradiently changed and reached 3.48% and 4.55 at the 16th week. Reducing sugar content arrived at 27.6% at the first week, and then decreased and stabilized at 8.0% at the 8th week. Changes of free amino acids content were consistent with that of amino-nitrogen. It increased rapidly at the beginning four weeks, and then slowly increased and amino-nitrogen content arrived at 1.78% at the 16th week. The fresh amino acid content was 38.9mg/g and accounted for 28.89%.SPME-GC-MS was introduced to analyze the changes of volatile flavor compounds. It increased fastly after eight weeks and kept rising after sixteen weeks. SPME-GC-O was used to analyze flavor-active compounds in soybean paste. HEMF, linoleic acid ethyl ester, ethyl phenylacetate, phenylethanol and furfuraldehyde were recognized as key flavor compounds. Esters, 2,3-butanediol, acetic acid, benzaldehyde, phenylacetaldehyde and 2,6-dimethyl pyrazine also played an important role in the flavors of soybean paste.Changes of bacteria and yeasts during the fermentation process were investigated. The results revealed that the trends of bacteria quantities were in consistent with that of yeasts. The quantities increased rapidly in the first two weeks. The total quantity of bacteria was 108CFU/g, and lactobacilli quantity reached 0.9×10~7CFU/g. The quantities of bacteria and lactobacilli decreased gradually after two weeks. The quantity of yeasts had a relatively slow decrease. The maximum quantity of yeast arrived at 1.5×10~6CFU/g at the fourth week and then it began to decrease. PCR-DGGE combined with traditional culture method was adopted to analyze the changing composition of microorganisms during the fermentation and fifteen strains showed high tolerance to high-salt environment. Lactobacillus plantarum 630-MO-115, Pediococcus halophilus 630-YN-27, Tetragenococcus halophilus 625-MN-50 and Zygosaccharomyces rouxii 625-YO-125 were confirmed as the key microorganisms for inoculation fermentation.Contents of total acids and amino-acid nitrogen of soybean paste were compared during the fermentation at 40℃at different salt contents. The salt contents were 6%, 8%, 10%, 12% and 14%, respectively. 10% was confirmed as the proper salt content. Regularities of pH, total acids, amino-acid nitrogen, and the quantities of lactobacilli and yeasts were investigated at 20℃, 25℃, 30℃, 35℃, 40℃and 45℃, respectively. The results indicated that 30℃and 35℃were proper for lactobacilli and yeasts.Four different temperature patterns were introduced during the soybean paste fermentation. The temperature was kept at 20℃for the first stage and turned to 30℃or 40℃when the amino-acid nitrogen content reached 1.4%. The temperature was kept at 40℃for the first stage and turned to 20℃or 30℃when the amino-acid nitrogen content reached 1.4%. Changes of pH, total acids and fresh amino acids contents and cont of key flavor compounds in the final products were studied at the four different temperature patterns. The results showed that 40℃→30℃pattern needed the shortest fermentation period and the key flavor content was a little lower than others.The symbiotic relationships between Lactobacillus plantarum 630-MO-115, Pediococcus halophilus 630-YN-27, Tetragenococcus halophilus 625-MN-50 and Zygosaccharomyces rouxii 625-YO-125 were investigated and a strategy on the multi inoculation and multi-temperature fermentation was adopted. A response surface analysis was carried out to optimize the fermentation process. Fermentation temperatures, inoculation amount for lactobacilli and yeasts, and the period for yeasts inoculation were selected as independent variables with three levels for each variable. Sensory evaluation scores of soybean paste were used as responses. The final optimized conditions were 32.4℃for fermentation temperature, 2.87×106CFU/g lactobacilli inoculation and 4.55×105CFU/g yeasts inoculation. Yeasts were added when pH fell to 5.29. The predictive value by the model was 90.8±0.7. Sensory evaluation score for the soybean paste made under this optimized condition was 91.2, which coincided with the predictive value. The fermentation period decreased from 180 days to 50 days.Principal components were compared between the naturally fermented soybean paste and that produced under the optimized fermentation condition. Soybean paste under the optimized fermentation condition contained less total acid and reducing sugar. Amino-acid nitrogen content increased to 1.80%. The fresh amino acids increased from 38.90mg/g to 47.72mg/g. In addition, the amounts of key flavor compounds had significant increases. In a word, soybean paste under the optimized condition had better qualities.
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