Method for detecting viable count in pre-coated feed microecological preparation product

Abstract

The invention relates to a method for detecting viable count in a pre-coated feed microecological preparation product. The method comprises the following steps: (1), culturing free bacteria, carryingout gradient dilution to count the viable count, centrifuging a proper amount of bacterial liquid, and carrying out resuspension with sterile water to obtain a bacterial suspension for later use; (2),taking the bacterial liquid obtained in the (1), carrying out gradient dilution, reacting with an MTT solution to obtain a precipitate, dissolving the precipitate with DMSO, and carrying out colorimetry by using a microplate reader to measure a light absorption value; (3), preparing a standard curve by using the viable count of the free bacteria obtained in the (1) and the light absorption valuemeasured in the (2); (4), crushing and sieving a pre-coated sample, adding the crushed and sieved pre-coated sample into a conical flask added with glass beads, adding a capsule breaking solution, andbreaking the capsule to obtain a bacterial suspension; (5), carrying out ultrasonic treatment on the bacterial suspension to obtain a discrete bacterial suspension, centrifuging the discrete bacterial suspension, adding DMSO for resuspending, performing diluting to different gradients, and measuring a light absorption value in the same (2); and (6), calculating the viable count in a pre-coated microecological preparation according to the standard curve obtained in the (3).

Description

technical field [0001] The invention belongs to the technical field of detecting the number of viable microorganisms, and relates to a method for detecting the number of viable bacteria in pre-coated microecological preparation products for feeding. Background technique [0002] Microencapsulated culture of probiotics before fermentation means that free bacteria are encapsulated in capsules using microencapsulation technology before fermentation, and then fermented and cultured. Compared with free cultured probiotics, the strains have faster growth Advantages, effectively improve the tolerance of strains to high copper and gastrointestinal fluids, play a very good role in protecting microbial strains, ensure the play of their physiological functions, and solve the problem of insufficient potency of probiotics as feed additives Stability issue. [0003] A mature probiotic counting method should have the characteristics of good parallelism and good reproducibility of the coun...

AI Technical Summary

Problems solved by technology

However, the traditional MTT method cannot uniformly disperse the tightly bound bacteria in the microcapsules without destroying its activity, and cannot accurately detect the number of live bacteria wrapped in the microcapsules. CN101620188A discloses a method using tetrazolium blue method to quickly The method for measuring the total number of viable bacteria in luminescent bacteria uses the MTT method to detect the number of viable bacteria at different concentrations to make a standard curve, and then uses the MTT method to detect the number of viable bacteria in the fermentation broth. However, this method is suitable for bacterial liquid, and the measurement parameters are not perfect. Bacterial activity tightly combined in microcapsules; CN102053123A discloses a method for detecting the activity of microencapsulated cells. Although the method uses the MTT method to detect microencapsulated cells, it does not consider the method of breaking the microcapsules. damage to the bacterial cells, and there is no corresponding relationship between the adenylate energy charge and the number of viable cells. Therefore, the conventional MTT method is not suitable for the detection of the number of viable bacteria in pre-coated probiotics products

[0005] In summary, there is no suitable and mature detection method in the field of detection of the number of live bacteria in pre-coated microecological preparations

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