The aim of the study was to implement a gastric digestion step using recombinant human gastric lipase (rHGL) in an in vitro pediatric gastro-intestinal digestion model to achieve a physiologically relevant gastric contribution to total gastro-intestinal lipid digestion. A commercial infant formula (NAN Comfort stage 1 (NAN1)) with 3.4% lipid and an in-lab prepared oil-in-water emulsion, emulsified with soy phosphatidylcholine (SPCemul), with 3.5% lipid (oil-blend containing Akonino NS, MEG-3 and ARASCO oils) were subjected to in vitro gastro-intestinal digestion. To achieve a physiologically relevant level of gastric digestion, 50 min of in vitro gastric digestion, using either 0, 3.75 or 7.5 TBU mL-1 rHGL, was followed by 90 min of in vitro intestinal digestion, using either 0 or 26.5 TBU mL-1 pancreatic triglyceride lipase (PTL) from porcine pancreatin. The digestion of the substrates was assessed using titration-based quantification supported by HPLC-ELSD analysis. In vitro gastric digestion of NAN1 and SPCemul with either 3.75 or 7.5 TBU mL-1 rHGL contributed with 10-27% of the total gastro-intestinal digestion, corresponding to the reported contribution in human infants. At the end of the gastro-intestinal digestion (t = 140 min), the combined lipolytic effect of rHGL and PTL was additive during digestion of SPCemul, but not for the digestion of NAN1, as all lipase activity combinations resulted in a similar degree of NAN1 digestion. The effect of gastric digestion with rHGL on total digestion therefore appeared to be substrate dependent. To conclude, a gastric digestion step using rHGL resulting in physiologically relevant gastric contribution to the observed gastro-intestinal digestion was successfully implemented into an in vitro pediatric gastro-intestinal digestion model.