In vitro ¹H MT and CEST MRI mapping of gastro-intestinal milk protein breakdown

Protein digestion is commonly studied using in vitro models. Validating these models with more complex in vivo observations remains challenging, in particular due to the need for non-invasive techniques. Here, we explore Magnetization Transfer (MT) and Chemical Exchange Saturation Transfer (CEST) MRI for non-invasive monitoring of protein solubilization and hydrolysis during static in vitro digestion using skim milk (SM). We measured CEST spectra of unheated and heated SM during gastric digestion, from which the relative amount of soluble proteins/peptides was estimated by calculating the asymmetric MT ratio (MTR_asym). We also obtained semi-solid volume fractions (v_ss), MT ratio (MTR) and MTR_asym from the same measurement, within 1.3 min. The MTR_asym area increased with gastric digestion, due to solubilization of the initially-formed coagulum, yielding a mean difference of 20 ± 7% between unheated and heated SM (p < 0.005). The v_ss and MTR decreased during gastric digestion and can be used to monitor changes in the coagulum, but not for assessment of soluble proteins/peptides. The MTR_asym increased for heated SM during gastro-intestinal digestion, proving sensitive to protein solubilization and hydrolysis, and is suitable for monitoring protein hydrolysis at later digestion stages. Future steps will include similar MT and CEST studies under dynamic conditions.