Influence of select emulsifying salts on pepsin-mediated casein hydrolysis of acidified skim milk.

In milk, casein proteins are oriented into spherical micellar structures. During gastric digestion, casein is destabilized via the pepsin-induced cleavage of κ-casein, forming a clot that certain populations (e.g., infants, elderly) have a difficult time digesting. Emulsifying salts (ES), such as sodium citrate (SC) and sodium hexametaphosphate (SHMP), are used in some dairy products (e.g., protein-fortified beverages, infant formula) and serve several beneficial functions including protein stabilization. As an investigative study, this research aims to determine whether SC or SHMP addition (1-100 mM) impact the pepsin-mediated hydrolysis of casein proteins by monitoring the interaction between milk proteins and pepsin in 2 distinct acidification stages: (1) high pH, reflecting infant or early-stage adult digestion, and (2) low pH, representing late-stage adult digestion. The high pH stage (pH 6.0), represented by the initial interaction of milk within the infant or adult gastrointestinal tract, is characterized by protein destabilization and clotting. During this stage, rheology was used to determine clotting kinetics, protein clots were periodically extracted and weighed, and gel electrophoresis was used to monitor κ-casein hydrolysis. The low pH stage (pH 2.2) corresponds to when the milk system has reached typical gastric acidity, and the rate of protein hydrolysis during this stage was monitored by gel electrophoresis. During the high pH stage, the control acidified skim milks formed a strong clot with a relative strength of 304.9 ± 40.6 Pa after 30 min. However, the acidified skim milks containing 10 or 100 mM SC failed to form any pepsin-induced clot, reflected by consistently low storage modulus values (<0.70 Pa) and lack of clot separation. In the low pH stage, pepsin-induced hydrolysis occurred in all samples, yet the 100 mM SC-containing sample had a reduced level of casein hydrolysis after 15 min. In contrast, samples with SHMP (10 and 100 mM) had severely inhibited hydrolysis rates compared with the control skim milks, reflecting SHMP-induced pepsin inactivation. Specifically, all of the casein was hydrolyzed in the control skim milks after 60 min, while 40.5 ± 14.5% and 43.5 ± 6.7% intact casein remained in the samples containing 10 mM and 100 mM of SHMP, respectively. These results indicate that SC and SHMP impact the pepsin-induced hydrolysis of acidified skim milk systems, and these differences are dependent on ES type and concentration. By resisting clotting, ES addition might allow for a faster transit rate through the gastric digestion phase and offer options for populations that struggle to metabolize casein clots, but there is a need to further investigate digestion and nutrient absorption using more advanced digestion protocols (e.g., INFOGEST).