| CPC A61K 35/747 (2013.01) [A61K 9/0053 (2013.01); A61P 21/00 (2018.01); C12N 1/205 (2021.05); C12R 2001/25 (2021.05)] | 11 Claims |
|
1. A microbial agent, comprising an inactivated strain of L. plantarum Nice-02, preserved in CCTCC (China Center for Type Culture Collection) on Apr. 21, 2023 with a biological preservation number of CCTCC NO: M 2023571;
wherein the microbial agent is prepared by a method comprising:
S1: activation culture: performing plate streaking on the L. plantarum Nice-02 into an activation culture medium to obtain a pure strain;
S2: primary seed solution preparation: picking and placing a single colony of the L. plantarum Nice-02 prepared in S1 into a first culture solution at 28 to 30° C. for stationary culture for 12 to 14 h to obtain a primary seed solution;
S3: secondary seed solution preparation: inoculating the primary seed solution into a second culture solution according to an inoculation amount of 0.1 to 5% for stationary culture at 30° C. to 40° C. for 10 to 12 h to obtain a secondary seed solution;
S4: tertiary seed solution preparation: inoculating the cultured secondary seed solution into a seeding tank containing a third culture solution under an aseptic condition at an inoculation amount of 0.5 to 5%, and performing stir culture at a stirring speed of 20 to 50 r/min, a temperature of 30° C. to 40° C., a pH value of 7.0 and a tank pressure of 0.03 to 0.06 MPa to obtain a tertiary seed solution after the culture for 10 to 12 h;
S5: L. Plantarum primary fermentation: inoculating the tertiary seed solution into a fermentation tank containing a sterilized culture solution at an inoculation amount of 0.5 to 5%, a temperature of 32° C., a stirring speed of 40 to 50 r/min to be cultured for 5 to 6 h;
S6: L. plantarum secondary fermentation: after 5 to 6 h, lowering the fermentation temperature to 30° C., accelerating the stirring speed to 50 to 70 r/min, keeping the tank pressure at 0.05 MPa and the natural pH, and performing fermentation for 3 to 4 h;
S7: L. plantarum tertiary fermentation: after 8 to 10 h, keeping the stirring speed unchanged, raising the fermentation temperature to 35° C., keeping the tank pressure at 0.05 MPa, keeping the pH value at 6.5, and performing fermentation for 2 h;
S8: L. plantarum quaternary fermentation: after 10 to 12 h, decelerating the stirring speed to 30 to 40 r/min, raising the fermentation temperature to 40° C., keeping the tank pressure at 0.05 MPa and the pH value at 6.5, and performing fermentation for 1 to 2 h to obtain a fermentation solution; and
S9: L. plantarum quinary fermentation: after 11 to 14 h, keeping the stirring speed unchanged, raising the temperature to 45° C., adding sterile lipoteichoic acid accounting for 0.05 to 0.2% of the volume of the fermentation solution into the fermentation tank, lowering the temperature to 20° C., and keeping the state for 30 min;
wherein the microbial agent is obtained by performing high-temperature inactivation and spray drying the fermentation solution prepared in S9.
|