Nutrients Consumption Patterns In Lactobacillus Acidophilus KLDS 1.0738

Lactobacillus acidophilus is widely used in the production of fermented food as a probiotic preparation and fermentation agent.It is considered the third generation of yogurt fermentation bacteria.Compared with foreign countries,the study of fermented dairy products in China started relatively late and research of high-concentrated direct vat set starter is still very weak.Direct vat set starter used in Chinese enterprises is dependent on import basically.High-cell-density culture is one of the key technologies to prepare direct vat set starter,and an important research direction to improve the level of technology of high-cell-density culture.The objective of this study is to investigate the nutrients consumption patterns of Lactobacillus acidophilus dyring culture,and improve the theoretical basis of the nutrients consumption patterns of Lactobacillus acidophilus in the high-cell-density culture process in China.This study provides scientific theory support of designing the excellent culture medium.In this study,Lactobacillus acidophilus KLDS 1.0738 was used as the target strain.Researched growth kinetics of the bacteria during the culture in the chemical defined medium with pH of fermentation system controlled.On this basis,the nutrients consumption patterns in the medium were studied on the key points during growth.The consumption patterns of glucose and vitamins were analyzed by high performance liquid chromatography(HPLC).The consumption patterns of amino acids were studied by amino acid analyzer.The consumption patterns of purine and pyrimidine bases were detected by reversed phase high performance liquid chromatography(RP-HPLC).The consumption of 2’-deoxyguanosine was measured by high performance liquid chromatography tandem mass spectrometry and the consumption patterns of inorganic ions in the medium was analyzed by flame atomic absorption spectrometry(FAAS).The growth curve of Lactobacillus acidophilus KLDS 1.0738 was divided into the lag phase(0～2h,μ1=0.04 h-1)),the initial logarithmic growth phase to the mid-logarithmic growth phase(2～7h,μ2=0.07 h-1),the mid-logarithmic growth phase to the terminallogarithmic growth phase(7～12h,μ3=0.11 h-1)and stable period(12～14h,μ4=0.01 h-1).The total consumption of glucose in the logarithmic growth phase was the highest during the growth of Lactobacillus acidophilus KLDS 1.0738,and the consumption accounting for 80.21%of the total consumption.The consumption of amino acids in μ3=0.11 h-1 was the highest(except Lys),accounting for 76.611%of the total consumption.The most consumed amino acids were aspartic,glycine,alanine and glutamic(Asp,Gly,Ala,Glu)among them,accounting for 59.065%of the total consumption.The consumption amount of cysteineand and methionine(Cys,Met)were the lowest,accounting for 0.781%of the total consumption.The consumption rates of Asp,Tyr,Lys and Arg were all more than 50%,and the consumption of Arg accounted for the largest percentage of its addition,with 55.552%of the total consumption.The consumption of vitamin B6 was the highest in the growth of Lactobacillus acidophilus KLDS 1.0738,and the consumption rate of folic acid was the highest.The concentration of riboflavin in the culture medium increased gradually,and the consumption of nicotinamide in μ3=0.11 h-1 was the highest.The consumption of pantothenate in μ2=0.07 h-1 was the highest.The consumption rate of uracil was highest.The consumption of 2’-deoxyguanosine was the lowest.The consumption of guanine was the highest.The consumption rate of thymine and adenine nucleotides was 81.463%and 69.054%,respectively.Mg2+ and Mn2+ consumed less than the others,and the consumption of Fe2+ was the lowest,while the consumption rate was the highest.The maximum consumption of ions were K+ and PO43-.At different growth rates,the consumption rate of nutrients of the unit bacteria is also different.This study found that the consumption rates of glucose,amino acids,calcium pantothenate,guanine and PO43+ and K+ of the unit bacteria were much higher compared to other periods at μ4=0.01 h-1.The strain was in stable phase,and the number of bacteria did not rise obviously at this point.These components with high consumption rates may be used to maintain the activity of the strain and resist osmotic pressure and acid stress mainly.In this study,the consumption rates of most nutrients components of the unit bacteria at μ1=0.04 h-1 were higher than the rates at μ2=0.07 h-1 and μ3=0.11 h-1.They decreased and reached the minimum at μ2=0.07 h-1 or μ3=0.11 h-1 with the increase of μ.The consumption rates of some nutrients(glucose,calcium pantothenate,adenine nucleotides,2’-deoxyguanosine and Mg2+ and Fe2+)were minimum at μ3=0.11 h-1.For example,the consumption rate of glucose decreased from 94.138±4.71 mmol/gDW at μ1=0.04 h-1 to 66.778±3.33 mmol/gDW at μ3=0.11 h-1,indicating that the utilization rate of these nutrients by strain increased gradually and reached the maximum at μ3=0.11 h-1.Utilization rates of amino acids and most vitamins,inorganic ions and purine and pyrimidine bases of per unit cell were minimum when μ2=0.07 h-1,such as the consumption rate of amino acids reduced from 6.264±0.018 mmol/gDW at μ1=0.04 h-1 to 3.39±0.010 mmol/gDW at μ2=0.07 h-1,and then increased to 27.998±0.073 mmol/gDW at μ3=0.11 h-1,which indicated that the utilization rates of these nutrients when μ3=0.11 h-1 was lower than that of μ2=0.07 h-1.Some of them were used to resist acid stress and osmotic pressure in the fermentation broth but did not use for growth completely.When μ1=0.04 h-1,the strain was in the lag phase and the number of bacteria changed little.The high consumption rate of unit bacteria was used to adjust the cell status and prepare for the logarithmic growth phase mainly.The result of study showed that the large consumption of nutrients of Lactobacillus acidophilus KLDS 1.0738 during culture inhibited the growth of strain,therefore,the concentrations of nutrients with high consumption should be improved in the medium.The initial concentrations of nutrients would also effect the growth of strain so that we should increase the initial concentration.The high initial nutrient may have an adverse effect on the growth of the strain.Depending on the nutrients consumption patterns of Lactobacillus acidophilus KLDS 1.0738 during culture,selecting the appropriate concentrations or forming an effective culture mechanism to add nutrients with high consumption or consumption rate in order to reduce the effects of excessive nutrients or low initial concentration.The result showed that the consumption forms of nutrients during the culture of Lactobacillus acidophilus KLDS 1.0738 were different,and the requirement of nutrients concentrations in different growth stages were different,too.This study provided the theoretical basis for thenutrients addition of high-cell-density culture.