A live show in the stomach: Monitoring in vitro and in vivo human gastric digestion with MRI

A better understanding of the gastric digestion of solid food will allow for a more thorough exploration of the relationship between food properties and physiological mechanisms underlying digestion of nutrients. The breakdown of food structures has mainly been studied via in vitro models. However, these results have been verified only to a limited extent in vivo. This limitation necessitates the investigation of the potential use of non-invasive approaches to bridge the link between in vivo and in vitro digestion research. Thus, this thesis aimed to investigate the potential of magnetic resonance (MR) techniques in monitoring gastric digestion of solid food in static, (semi-)dynamic in vitro models and in humans.

The work began with a well-controlled in vitro static digestion model in Chapter 2. We explored the use of time-domain nuclear magnetic resonance (TD-NMR) and magnetic resonance imaging (MRI) to monitor the gastric digestion of whey protein (solution and gel). During digestion, free amino groups (-NH2 groups) and protein concentrations in the supernatant were measured. Transverse relaxation time (T2) values of the digestion mixture were determined by TD-NMR and MRI, and transverse relaxation rate (R2 = T2-1) was calculated. Subsequently, relative amplitudes (TD-NMR) for different T2 values and T2 distribution (MRI) were determined. For the solution, protein concentration and T2 did not change during digestion. For the gels, water in both supernatant and gel phase could be discriminated on the basis of their T2 values. During digestion, R2 of the supernatant correlated positively with the protein (-NH2 groups) concentration in the supernatant. MRI T2-mapping showed similar associations between R2 of supernatant and protein (-NH2 groups) concentration. Thus, R2 was shown to be a useful marker to monitor in vitro gastric digestion of whey protein gels and TD-NMR measurements contributed to interpreting MRI data.

TD-NMR results from Chapter 2 showed that water transportation (namely swelling) took place during digestion and may be of great importance for digestion rate. Therefore, we investigated the effect of swelling on gastric digestion of protein gels in Chapter 3. Whey protein gels with NaCl concentrations of 0-0.1 M were used as model foods. Young’s modulus, swelling ratio, acid uptake and digestion rate of the gels were measured. Pepsin transport was monitored by confocal laser scanning microscopy using green fluorescent protein (GFP), which has a similar size as pepsin. We observed that an increase of NaCl in gels corresponded with increased Young’s modulus, reduced swelling and slower digestion. Additionally, a reduction of acid transport was observed, as well as a reduction of GFP both at the surface and in the gels. This shows that swelling affects digestion rate not only by enhancing acid diffusion but also by modulating partitioning of pepsin at the food-gastric fluid interface and thereby the total amount of pepsin in food particle. This perspective on swelling provides new insight for designing food with a specific digestion rate for targeted dietary demands.

The work in Chapter 2 was performed under static conditions. Thus, further work was conducted in Chapter 4. We developed a novel MRI-compatible semi-dynamic gastric simulator (MR-GAS) that includes gastric secretion, emptying and mixing, and applied it to investigate the potential of relaxation rates in monitoring digestion. During protein gel digestion, pH and protein hydrolysis were measured. R2 and R1 (= T1-1) of the supernatant were measured by time-domain nuclear magnetic resonance (TD-NMR) and MRI. With TD-NMR, 99% of the variance in R2 and 96% of the variance in R1 could be explained as a function of protein concentration and [H+]. With MRI, the explained variances were 99% for R2 and 60% for R1. From these analyses, the obtained equations enabled the prediction of protein concentration and pH with measured R2 and R1 values. This shows that MR-GAS model may be used in a clinical MRI scanner to monitor gastric digestion under in vitro dynamic circumstances, by measuring R2 and R1. These results underscored the potential of MRI to monitor nutrient hydrolysis and pH changes in in vivo studies. Therefore, in Chapter 5, we conducted a human randomized cross-over trial in which we assessed the effect of food hardness and protein content on gastric emptying and additionally investigated the application of the T1 and T2 to monitor in vivo gastric digestion.

The trial was conducted with 18 healthy males provided with three gels differing in hardness and protein content: a soft gel with low protein content (Soft-LP), a hard gel with low protein content (Hard-LP), and a hard gel with high protein content (Hard-HP). Before and after ingestion, abdominal MRI scans and appetite and well-being ratings were obtained until t = 85 min after the start of ingestion. At t = 100 min participants ate an ad libitum lunch. Overall, gastric content volume was different among the treatments: High-HP < Soft-HP < Soft-LP. Mean T2 and T1 of the measured stomach content decreased after ingestion from baseline and then gradually increased from 15 min onwards. The treatments resulted in different T1 and T2 values: Hard-HP < Soft-HP < Soft-LP, although not all the time points differed significantly. The high protein content was the main factor in delaying gastric emptying and high hardness was the secondary factor. T1 and T2 measurements can provide extra information on the dilution and digestion taking place in the stomach. This study suggests the potential of MRI parameters for providing more insights on in vivo digestion, and their results may contribute to linking in vitro and in vivo digestion research.

Finally, Chapter 6 discusses findings from in vitro models to in vivo human trials in this thesis. It provides an overview of the application of MR techniques to measure gastric digestion, the added value of MRI measurements for digestion research, and the effects of food properties on gastric digestion. To conclude, MR techniques can provide molecular-level and quantitative information on protein hydrolysis in solid food through T1 and T2 measurements. Moreover, the findings from this thesis can aid in informing in vitro and in silico models and bridging the link between in vitro and in vivo digestion research.