HPLC detection method of nisin
A technology of nisin and nisin, which is applied in the field of high-performance liquid chromatography detection of nisin, and can solve the problems of reproducibility, poor accuracy, long time course and difficult integration of sample determination results
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[0065] Solution preparation
[0066] Reference substance solution: Accurately weigh about 10 mg of Nisin reference substance, put it in a 10ml volumetric flask, dissolve it with 0.02N HCl and dilute to the mark, and use it as the reference substance solution.
[0067] Sample solution: Scrape Lactococcus lactis ssp.lactis from the inclined surface and insert it into the seed solution (the formula is 0.5% soybean protein isolate, 0.5% peptone, 0.25% yeast extract, 0.5% beef extract, 0.5% glucose , ascorbic acid 0.05%, Na 2 HPO 4 12H 2 O 1.0%, MgSO 4 ·7H 2 O 0.012%, pH6.8) at 37°C for 20h. Insert the seed liquid into the fermentation broth (the formula of the medium is: 2.0% sucrose, 3.0% yeast extract, 2.0% peptone, 0.02% magnesium sulfate, 1.0% potassium dihydrogen phosphate, 0.2% NaCl, Tween-800.5%, pH 6.8) Ferment at 30°C for 12-14 hours, stop the fermentation, take a certain amount of Nisin fermentation broth, adjust the pH to 2.0 with 0.02NHCL, centrifuge at 12000-140...
Embodiment 1
[0069] Initial mobile phase B: mobile phase A=20:80 isocratic elution for 3 minutes, then mobile phase B continuously increased from 20% to 40% elution for 27 minutes, and mobile phase B returned to 20% in the last 2 minutes ,balance. Under this method, the retention time of the reference substance is 16.38min, the resolution is 1.08, and the number of theoretical plates reaches 16738. See figure 2 with image 3 .
Embodiment 2
[0071] Initial mobile phase B: mobile phase A=20:80 isocratic elution for 3 minutes, then mobile phase B continuously increased from 20% to 40% for 12 minutes, and mobile phase B returned to 20% in the last 2 minutes ,balance. Under this method, the peak eluting time is about 10 minutes, the baseline drifts, and the number of theoretical plates is 12800.
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