The delivery of probiotic microorganisms as food additives via oral administration is a straightforward strategy to improve the intestinal microbiota. To protect probiotics from the harsh environments in the stomach and small intestine, it is necessary to formulate them in biocompatible carriers, which finally release them in the ileum and colon without losing their viability and functions. Despite major progresses in various polymer-based formulations, many of them are highly heterogeneous and too large in size and hence often "felt" by the tongue. In this study, we established a new formulation for probiotics Lactobacillus rhamnosus GG (LGG) and systematically correlated the physicochemical properties of formulations with the functions of probiotics after the delivery to different gastrointestinal compartments. By reducing the stirring speed by 1 order of magnitude during the emulsification of polyalginate in the presence of xanthan gum, we fabricated microparticles with a size well below the limit of human oral sensory systems. To improve the chemical stability, we deposited chitosan and polyalginate layers on particle surfaces and found that the deposition of a 20 nm-thick layer is already sufficient to perfectly sustain the viability of all LGG. Compared to free LGG, the colony-forming units of LGG in these formulations were by factors of 10 larger in stomach fluid and 10 larger in small intestine fluid. The metabolic functionality of LGG in polymer formulations was assessed by measuring the amount of lactate produced by LGG in a human gastrointestinal simulator, showing 5 orders of magnitude larger values compared to free LGG. The obtained results have demonstrated that the minimal formulation of LGG established here boosts not only the viability but also the metabolic functionality of probiotics throughout oral uptake, passage through the gastrointestinal tract, and delivery to the ileum and colon.

The accumulation of phosphatidylcholine (PC) in the intestinal mucus layer is crucial for the protection of colon epithelia from the bacterial attack. It has been reported that the depletion of PC is a distinct feature of ulcerative colitis. Here we addressed the question how PC interacts with its binding proteins, the mucins, which may establish the hydrophobic barrier against colonic microbiota. In the first step, the interactions of dioleoylphosphatidylcholine (DOPC) with two mucin preparations from porcine stomach, have been studied using dynamic light scattering, zeta potential measurement, and Langmuir isotherms, suggesting that mucin binds to the surface of DOPC vesicles. The enthalpy of mucin-PC interaction could be determined by isothermal titration calorimetry. The high affinity to PC found for both mucin types seems reasonable, as they mainly consist of mucin 2, a major constituent of the flowing mucus. Moreover, by the systematic variation of net charges, we concluded that the zwitterionic DOPC has the strongest binding affinity that cannot be explained within the electrostatic interactions between charged molecules.

Coating of mesoporous silica carriers with dioleoylphosphatidylcholine allowed triggering of the selective delivery of functional enzymes by lipolysis under simulated intestinal conditions.

The oral uptake of probiotic microorganisms as food additives is one widely used strategy to sustain and improve the homeostasis of intestinal microbiota that protect the intestinal epithelia from attack by pathogenic bacteria. Once delivered to the ileum and colon, probiotics must adhere and form colonies on mucus that coats the surface of intestinal epithelial cells. Although an increasing amount of knowledge about the genetic and molecular level mechanisms of probiotics−mucus interactions has been accumulated, little is known about the physicochemical aspects of probiotics−mucus interactions under physiological shear in intestines. In this study, we established well-defined models of intestinal epithelial cell monolayers based on two major constituents of gut epithelia, enterocytes and goblet cells. First, the formation of a polarized cell monolayer sealed by tight junctions was monitored by transepithelial electrical resistance over time. The establishment of tight junctions and secretion of mucus proteins (mucin) was confirmed by immunofluorescence staining. In the next step, we measured the elasticity of cell monolayer surfaces by indentation using particle-assisted atomic force microscopy. The effective elastic modulus of goblet cell-like cells was 30 times smaller compared to that of enterocyte-like cells, which can be attributed to the secretion of a 3 μm thick mucin layer. As probiotics, we used Lactobacillus rhamnosus GG (LGG), which is one of the most widely used strains as food additives. To investigate the dynamic adhesion of LGG to the intestine model surface, we transferred the epithelial cell monolayer into a microfluidic chamber. A distinct difference in dynamic adhesion between two cell types was observed, which could be attributed to the difference in the mucin expression amount. Remarkably, we found that the dynamic LGG adhesion is enhanced by the increase in shear stress, showing a maximum binding efficiency at 0.3 Pa. Finally, we examined the persistence of LGG adhesion by a stepwise increase in the shear stress exerted on adherent LGG, demonstrating that LGG could withstand high shear stress even beyond that of physiological stress. The obtained results present a large potential to quantitatively understand the influence of engineered foods and probiotics on the homeostasis of microbiota on the surface of intestinal epithelia.