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CN-121975652-A - Probiotics-bacterial cellulose composite membrane and preparation method and application thereof

CN121975652ACN 121975652 ACN121975652 ACN 121975652ACN-121975652-A

Abstract

A probiotic bacteria-bacterial cellulose composite membrane, a preparation method and application thereof comprise lactobacillus plantarum (Lactobacillus plantarum) LCC-605 with a preservation number of CCTCC No. M2016491, a biological membrane, a metabolite and a metazoan thereof, and lactobacillus plantarum (Acetobacter xylinum) and lactobacillus plantarum (Lactobacillus plantarum) LCC-605 are subjected to in-situ co-culture to form the lactobacillus plantarum biological membrane-based bacterial cellulose membrane and related metabolites thereof. The invention not only provides a bacterial cellulose membrane for encapsulating probiotics to enhance the survival rate and stability of the probiotics, but also provides an ideal starter for preparing fermented black soybean milk with excellent texture, high viable count and excellent storage stability.

Inventors

  • LI CHENGCHENG
  • JIANG ZHUO

Assignees

  • 南京林业大学

Dates

Publication Date
20260505
Application Date
20251215

Claims (10)

  1. 1. The probiotics-bacterial cellulose composite membrane is characterized by being formed by co-culturing lactobacillus plantarum (Lactobacillus plantarum) LCC-605 with the preservation number of CCTCC M2016491 and acetobacter xylinum (Acetobacter xylinum) in situ and sequentially carrying out an aerobic culture stage and an anaerobic culture stage, wherein the composite membrane comprises a matrix formed by bacterial cellulose produced by the acetobacter xylinum, the lactobacillus plantarum LCC-605 loaded and embedded in the matrix and a biological membrane formed by the lactobacillus plantarum LCC-605.
  2. 2. The probiotic-bacterial cellulose composite membrane according to claim 1, wherein the aerobic culture stage is under the condition that aerobic culture is performed for 2 to 7 days at 30 to 37 ℃ in a mixed system containing acetobacter xylinum fermentation broth and lactobacillus plantarum LCC-605, and the anaerobic culture stage is under the condition that a membranous material formed after the aerobic culture is placed in a liquid culture medium and anaerobic culture is performed for 24 to 48 h at 30 to 37 ℃.
  3. 3. The probiotic-bacterial cellulose composite membrane according to claim 2, wherein in the aerobic culture phase, the volume percentage of the acetobacter xylinum fermentation liquid in the mixed system is 5-10%, and the initial OD600 value of lactobacillus plantarum LCC-605 is 0.01-0.1.
  4. 4. The probiotic-bacterial cellulose composite membrane according to claim 1, wherein the loading of lactobacillus plantarum LCC-605 in the composite membrane is not lower than 1.0 x 10 8 CFU/g.
  5. 5. A method for preparing the probiotic-bacterial cellulose composite membrane according to any one of claims 1 to 4, which is characterized by comprising the steps of (a) preparing an activated fermentation broth of acetobacter xylinum and an activated bacterial suspension of lactobacillus plantarum LCC-605 respectively, (b) mixing the acetobacter xylinum activated fermentation broth obtained in the step (a) with the activated bacterial suspension of lactobacillus plantarum LCC-605, performing first-stage standing or shaking culture under aerobic conditions until a bacterial cellulose membrane is formed, (c) transferring the bacterial cellulose membrane obtained in the step (b) into an anaerobic environment, replacing the original culture broth with a new liquid culture medium, and performing second-stage anaerobic standing culture, (d) taking out and cleaning the membrane obtained after the anaerobic culture in the step (c) to obtain the probiotic-bacterial cellulose composite membrane loaded with lactobacillus plantarum LCC-605 and a biological membrane thereof.
  6. 6. The method according to claim 5, wherein in the step (b), the aerobic culture is performed at 30 to 37℃for 2 to 7 days, and in the step (C), the anaerobic culture is performed at 30 to 37℃for 24 to 48 h.
  7. 7. The method according to claim 5 or 6, wherein in step (c), the new liquid medium is MRS liquid medium, and the medium is replaced 1-2 times during the anaerobic stationary culture.
  8. 8. A starter culture comprising the probiotic-bacterial cellulose composite membrane of any one of claims 1-4.
  9. 9. Use of a starter according to claim 8 for the preparation of a fermented food product, wherein the starter is inoculated into a plant-based milk for fermentation.
  10. 10. Use of a probiotic-bacterial cellulose composite film according to any one of claims 1 to 4 for the preparation of medical devices, pharmaceutical products, skin care products, microecologics, feeds or pet foods.

Description

Probiotics-bacterial cellulose composite membrane and preparation method and application thereof Technical Field The invention belongs to the technical field of microorganisms, and particularly relates to a probiotic-bacterial cellulose composite membrane, and a preparation method and application thereof. Background Probiotics are susceptible to multiple environmental stresses during processing, storage (e.g. temperature fluctuations, extreme pH, oxidative stress, etc.), resulting in a dramatic decrease in their survival and activity (Xu et al, 2022). Therefore, it is important to develop a technology capable of maintaining a high survival rate of probiotics. The embedding technology is proved to be one of the most effective strategies for guaranteeing the survival rate of probiotics through physical isolation and chemical protection, so that the development of the embedding delivery technology of the probiotics has important practical significance for expanding the application space of the probiotics. Currently, a variety of encapsulation strategies have been used to protect probiotics, such as microencapsulation, emulsification, electrospinning, and spray drying (de Oliveira Filho et al., 2025). Among these, microencapsulation is a common and easy to implement method, forming a gel network structure by loading probiotics into a protective matrix. Sodium Alginate (SA), a natural anionic polysaccharide, is widely used for probiotic encapsulation (Li et al, 2011; razavi et al, 2021; torp et al, 2022; zhang et al, 2021) because of its safety, low cost and high solubility properties, often cross-links with Ca 2+ to form a gel. In addition, SA encapsulation also allows for controlled release of probiotics in the intestinal tract (Li et al, 2024). Although microencapsulation can improve the survival rate of probiotics in severe environments, the protective effect can be further optimized by enhancing the stability of the probiotics themselves. In recent years, biofilm encapsulation or biofilm in combination with other encapsulation strategies have been of interest for their protective effect (Yang et al, 2024; mgomi et al, 2024). Bacteria are often accumulated in the form of biological membranes in nature to resist adverse conditions such as extreme pH, high temperature, oxidative stress, and even antibiotics （Cheow & Hadinoto, 2013; Hu et al., 2019; Vega-Sagardía et al., 2018; Flemming & Wuertz, 2019; Motta et al., 2021; Sadiq et al., 2020）. In addition, probiotics have been developed into functional fermented dairy products, however, probiotics often suffer from problems in the fermentation process, such as long fermentation time, low density, poor viability of the probiotics in the fermented dairy product, insufficient viability of the probiotics in the fermented dairy product, and inability of the probiotic starter to be recycled, etc., resulting in low production efficiency, high production cost and unstable product quality in the fermentation process (Hu et al, 2019). Bacterial cellulose is a biopolymer produced by some aerobic bacteria (Moon et al 2011; ullah et al 2016), with unique physicochemical properties such as natural nanostructure, hydrophilicity, high porosity, high water retention, mechanical strength, crystallinity, biocompatibility, etc. (Li et al 2022). Moreover, bacterial cellulose is widely used in dairy products, can be used as a stabilizer and an emulsifier in the production of the dairy products, can be used as a fat substitute in products such as cheese and the like, and can be used as an additive of functional foods. It can also be used as a carrier for enzymes, bioactive substances and probiotics for developing dairy products with health care functions, and also has applications in food packaging (P ł oska et al., 2023). The probiotics are packaged in bacterial cellulose, so that the probiotics are covered with a layer of protective cover, and can be isolated from the outside under certain conditions, a microcosmic steady-state environment (Frakolaki et al, 2020) is provided for the probiotics, and the probiotics can resist the external bad environment and simultaneously maintain the growth of the probiotics. In addition, the bacteria cellulose coated probiotics have biological safety, so that the fermentation time is shortened, the fermentation density is improved, and the repeated recycling of the fermenting agent can be realized. Thus, the encapsulation of probiotics with bacterial cellulose is a good choice. Plant-based dairy products (e.g., soy milk, rice milk, coconut milk) are effective in supporting probiotic growth and maintain high survival rates during fermentation and storage. According to market research data, the global plant-based yoghurt market is expected to continue to grow at an annual growth rate of 18.9% (Liu et al, 2023). Bean products have become an important product in the plant-based market because of their suitability for vegetarians and lactose intolerant p