Preparation Of Enterobacter Sakazakii Standard Material In Milk Powder

enterobacter sakazakii will cause serious neonatal meningitis, bacteremia, enterocolitis; moreover,it could cause neurological disorders and serious sequelae, Mortality rate is over 20% -50%. enterobacter sakazakii has aroused world wide attention.The experiment studied preparation method of enterobacter sakazakii standard material in milk powder. In this study, we used a standard material specifications (JJG 1006-94) as the criterion, and The experiment complied with Specification of "detection of enterobacter sakazakii from dehydrated powdered milk - Part 1: Isolation and enumeration" ,which is from the Republic of China's Entry-Exit Inspection and Quarantine trade standards.Because a optimal protectant is important for freeze-dry preservation of enterobacter sakazakii standard material,therefore the study design a test agent to select the most suitable protectant .three protectants were chosed in the test: milk (20g milk +80mL sterile water), nutrition broth 100 mL +7.5 g glucose, 0.1 mol / L phosphate buffer 100 ml + L- monosodium glutamate 3 g + Lactose 5 g + Polyvinylpyrrolidone 6g.. at fist, add the enterobacter sakazakii into the three protectants , put them into ampoule bottles and freeze them at -70℃for one day;then freeze-dry the ampoule bottles and proceed biochemistry test,;in the end,we detect the number of enterobacter sakazakii in those ampoule bottles at regular intervals.The results indicate that dried skimmed milk is the most suitable protectant for enterobacter sakazakii.in addition,keep on detecting the number of enterobacter sakazakii which use dried skimmed milk as the protectant to test its stability.Add enterobacter sakazakii into the nutrient broth overnight ,the concentration is always around 108,so dilute it to 105 and imbibe 2ml bacilli liquid,then add it into milk (40 g skim milk +160 ml sterile water); secondly,put the milk into sterile porcelain , pre-freeze at -70℃for 18-24h and freeze-drying for 48-60 h;then put content in homogeneous bag and flap them in uniform; take out 10g milk into 90 ml sterile water, measured The number of bacteria and According to it, dilute the milk with sterile skim milk ; the bacteria will eventually be diluted to 100g skim milk contains only a few hundred enterobacter sakazakii, the milk are subpackaged into the smallest unit(10g)and vacuumized;in the end,we detect those standard material's stability and uniformity.The results show that its stability and uniformity are in line with the standard material, including the F values of two uniformity test were 1.759 and 0.548.We carry out the uncertainty of assessment after the preparation of the standard material. to. There are so many uncertainties could affect Preparation of standard material, in this study,we assess the uncertainty which has an important impact on the uncertainty, they are uncertainties of colony counting, the standard material valuation methods, electronic balance. the uncertainty of colony counting is 0.06429; the uncertainty of standard material valuation methods is 11.508; electronic balance's compositive uncertainty U is 1.42518, the expanded uncertainty for 2.79335.In addition,This study detect biochemical characteristics of enterobacter sakazakii standard material which being pre-freezed and freeze-dried.the results shows that its biochemical characteristics have not changed.