The Mycelium Of Pingxiang "Cuocai" And The Preparation Of Solid Starter Strains

Microbial physiological and biochemical identification and modern molecular biology has been used to study the microbial strains in Cuocai.Advantage strains were selected to produce liquid and solid starter cultures.Finally,pH value,crude protein,total sugar,total acid and volatile flavor substance during the process of artificial fermentation and natural fermentation have been compared.The advantage strains were selected from the samples,in order to produce products with stable,health and safety quality and flavors were similar to traditional pingxiang Cuocai.The main conclusions were as follows:1.Seven advantage strains were isolated from natural fermentation Pingxiang Cuocai and they were numbered as J-1,J-2,Q-1,G-1,G-2,G-3 and G-4.Physiological and biochemical analysis as well as molecular biology techniques was used to identify the mycelium.It was shown that advantages yeasts in Cuocai were J-1 strain(Issatchenkia orientalis)and J-2 strain(Issatchenkia occidentalis).Advantages lactic acid bacterias were Q-1 strain(Enterococcus faecalis),G–1strain(Lactobacillus plantarum),G-2 and G-4 strains(Lactobacillus buchneri)and G-3 strain(Lactobacillus pentosus).2.Growth characteristics of the advantage strains of lactobacillus were researched.Among the five strains of lactic acid bacteria,G-1,G-2,G-3 and G-4had better viability at temperature of 35?,and the optimum growth temperature for Q-1 was 30?.Growth curves of five lactic acid bacteria strains were all on the rise from 0 to 18 h and kept stability after 18 h.Among them,lactobacillus bacteria had the highest density and the OD 600 nm reached 0.785.All lactic acid bacterias had good resistance to low pH features and the Lactobacillus plantarum and lactobacillus pentose had better resistence then other three strains.After producing acid test and antagonism test,Lactobacillus plantarum was chosen as the bacteria starter cultures for Cuocai.3.It was showed that the optimum growth temperature of the two strains of yeast J–1and J-2 were both at 30?.Growth curve peak of two strains of yeast appear at 18 h,Issatchenkia orientalis was slightly higher than Issatchenkia occidentalis.Two strains of yeast had similar low pH tolerance ability and cell density increases with the increase of pH in a certain range.Based on the physical properties,Issatchenkia orientalis was chosen as the yeast starter cultures for Cuocai.4.Starter culture strains had been selected based on sensory quality and the best ratio of compound strains of lactic acid bacteria and yeast used for pure breed fermentation was 1:3,the inoculation amount was 1%,fermentation time was 50days.5.It was showed that the best culture medium formula for Lactobacillus plantarum was:glucose 2.71%,yeast powder,0.45%,beef extract 0.6%.The optimal culture conditions were as fellowed:initial pH 6,inoculated quantity 4%,rotary shaker speed 80 r/min,culture temperature at 35?.After growth for 18 h,materials were added every 4 hours and the additive amount was 15 g/L and bacteria were harvest at 48 hours after adding material for six times.The ratio of carbon and nitrogen in fill material was 5:1.In this condition the highest Lactobacillus plantarum bacteria density was obtained.The total number of colonies can reach to 9.28 x 10~9 cfu/mL.The best culture medium formula for Issatchenkia orientalis was:glucose 4.06%,yeast powder 4.06%,potassium dihydrogen phosphate 0.39%.The optimal culture conditions were:initial pH 6 of culture,2%inoculated quantity,rotary shaker speed 120 r/min,culture temperature 30?.After growth for 18 h,materials were added every 4 hours and the additive amount was 25 g/L and bacteria were harvest at 48 hours after adding material for six times.The ratio of carbon and nitrogen in the filled material was 5:1.In this condition the highest Issatchenkia orientalis cell density could be obtained.The total number of colonies can reach to 1.71 x 10~9 cfu/mL.6.The best centrifugal condition of Lactobacillus plantarum and Issatchenkia orientalis were 4000 r/min,20 min and 5000 r/min,20 min,respectively.Protective agents were added to protect thallus from the destruction produced by vacuum freeze drying.Protective agents were selected by the single factor test and orthogonal test.The optimal protective agent were monosodium glutamate 5 g/100 mL,trehalose 10 g/100 mL,skim milk,10 g/100 mL,manganese sulfate,0.7 g/100 mL for Lactobacillus plantarum.Trehalose and skim milk were 10 g/100 mL respectively,glutamate 7 g/100 mL,sucrose 15 g/100 mL for Issatchenkia orientalis.7.The comparison indexes of artificial fermentation and natural fermentation Cuocai showed that:all indexes of artificial fermentation Cuocai were similar to those of natural fermentation,the artificial fermentation showed significantly reduceofnitritecontent.Simultaneousdistillationextraction-gas chromatography-mass spectrometry method had been used to detect the organic volatile flavor substances in Cuocai,it showed that the mainly eight volatile flavor substance in Cuocai were alcohols,aldehydes,alkanes class material substance,esters,ketones,heterocyclic compounds,sulfur and acid.