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Adherence interactions between milk proteins and human intestinal surface layer components : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy, Massey University, Palmerston North, New Zealand
Recent research suggests a number of food-derived proteins may be used as orally delivered
functional components. The native structure is often vital to their activity and requires
protection during the digestive process. Nutrient vehicles are used as protective envelopes and
as a mechanism for targeting specific sites of activity, e.g. the small intestine. This study
evaluated molecules which adhere to one or more in vitro models of three human intestinal
surface layers. Successful candidates could then be incorporated into nutrient vehicles,
promoting adhesion to the surface layers and resulting in prolonged retention of the active
ingredient at the site of action or absorption.
To identify molecules that adhere to models of the intestinal surface, an adhesion protocol was
developed to screen the proteome of whole milk, skim milk and whey for candidate proteins.
Molecules adhering to model layers of the human gastrointestinal tract (intestinal epithelial
cells, mucin or bacteria with the propensity to form a biofilm) were screened by SDS-PAGE
analysis and identified by mass spectrometry and Western blot. The binding behaviour of
selected proteins was further investigated by flow cytometry. The combined results showed that
milk and whey proteins exhibit different binding affinities to the models of individual surface
layers. a-Lactalbumin was found to adhere to a model of the intestinal epithelial cells, while ß-
lactoglobulin showed binding to the protective mucin layer. Lactoferrin and various components
of immunoglobulins showed highest binding affinity to bacteria. Finally, IgM appeared to
adhere to all three tested model layers of the human gastrointestinal surface. Least binding was
observed to the intestinal epithelial cells in culture. The validity of the developed adhesion
protocol was demonstrated by replicating adhesion of immune-related proteins, lactoferrin and
immunoglobulins, to bacterial cells.
This work reveals new important characteristics of milk-derived proteins in their ability to
adhere to models of the gastrointestinal surface. These may be further utilised in site-specific
targeting of functional foods.