Milk fat globule membrane conditions the reactivity and enzymatic susceptibility of milk fat. The use of bovine membrane extracts to formulate infant formulas more biomimetic of human milk has been suggested in the recent literature 1,2 . A comparison of the physico-chemical behavior of human (HMM) and bovine (BMM) milk membrane extracts and their interaction with gastric lipase is lacking to assess whether their interfacial behavior is similar. This comparison is undertaken in the present project using complementary biophysical tools based on Langmuir film characterization as tensiometry, ellipsometry, Brewster angle (BAM) and atomic force (AFM) microscopies in the presence or in absence of gastric lipase. Milk membrane extracts were obtained from either a pool of mature human milk (n=5) or bovine butterserum and were analyzed using GC and HPLC.Compression isotherms indicated a higher compressibility of HMM than BMM. However, both extracts presented close collapse pressures (~45 mN/m). Despite the presence of liquid condensed (LC) domains (AFM and BAM) from low to highest surface pressure in the two extracts, their morphological aspect and growth differed strongly in relation with their specific lipid composition (GC and HPLC). As reported for BMM 3 , the gastric lipase got adsorbed toward liquid expanded phase (LE) and at several height levels in HMM. However its insertion did not impact on LC domain morphology as observed in BMM. Despite differences in their physico-chemical properties, both extracts present close interfacial reactivity in gastric conditions. Such biophysical Investigation of ‘lipase-complex lipid’ interactions will be broaden to other milk membrane extracts (equidae) interesting for human milk substitution