Abstract

A high-density dairy fermentation protocol and composition for Limosilactobacillus reuteri LR08 are disclosed across liquid bovine dairy substrates ranging from skim milk to whole milk, A2 milk, and half-and-half (up to ~18% butterfat), as well as milk/heavy cream blends, scaled from small-batch home preparation (1 quart / 1 liter) up to industrial commercial bioreactor volumes. The method eliminates standard dairy fermentation rate bottlenecks, physical whey separation, oxidative stress, packaging inflation, equipment sterilization overhead, and airborne contamination by integrating modular biochemical and bioprocess mechanisms:

  1. A decoupled dual-fructan system comprising long-chain branched Blue Agave inulin (hydrocolloid matrix builder) and short-chain linear fructooligosaccharides (Inulin FOS, metabolic substrate);
  2. Enzymatic soy peptone supplementation to supply bioavailable oligopeptides and bypass cell-wall proteinase (PrtR) cleavage, reducing lag phase and shortening fermentation time to approximately 20 hours;
  3. Optional Reduced Glutathione (GSH, 0 mg to 2000 mg) addition to lower Oxidation-Reduction Potential (eH) and protect cell membranes when redox control is desired;
  4. Optional pure glycerol (0 g to 30 g) supplementation to drive conversion to reuterin (3-hydroxypropionaldehyde) via glycerol dehydratase when in situ antimicrobial synthesis is desired;
  5. Heterofermentative gas suppression, wherein the composition is free of added fermentable simple carbohydrates, monosaccharides, disaccharides, trisaccharides, sugar alcohols, and metabolic intermediates capable of serving as substrates for heterofermentative CO2 generation via the phosphoketolase pathway (including glucose, dextrose, fructose, galactose, sucrose, maltose, lactose, maltodextrin, corn syrup solids, and polyols), suppressing CO2 off-gassing to enable zero-headspace fermentation in flexible bags and rigid vessels; and
  6. Closed-system in-pouch thermal processing and low-contamination corner-snip inoculation, wherein unheated pre-ferment is sealed in a flexible polymer bag, submerged in a circulating water bath for thermal denaturation (eliminating equipment sterilization requirements), cooled in-bag to ≤100°F, inoculated through a snipped bag corner to minimize airborne exposure, re-sealed, and flex-mixed without oxygen incorporation.  

    • Patent Classifications (CPC / IPC):

      Enter the Cooperative Patent Classification (CPC) codes to ensure searchability by USPTO, EPO, and WIPO examiners:

      A23C9/123, A23L33/135, C12N1/20

    • Keywords / Tags:

      Limosilactobacillus reuteri, LR08, Dairy Fermentation, Inulin, Soy Peptone, Reuterin, Glutathione, Defensive Publication, Prior Art, Non-Patent Literature

    • Cross-Reference / Secondary Identifiers (Optional Field):

      Add the Zenodo DOI cross-reference note:

      Simultaneously published and archived on Zenodo under DOI: https://doi.org/10.5281/10.5281/zenodo.22016184

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