The studies of model biological membranes consisted of phospholipid 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) were carried out by means of the Langmuir monolayer technique using subphases containing chitosan (Ch), titanium (IV) oxide (TiO2), hyaluronic acid (HA) or mixture of them. The aim was to determine the effect of individual components of subphase and their respective combinations on behavior of the DPPC membrane. The systems were tested at room temperature (20 °C) and at a natural pH of about 4.8, which was close to the pH of the human skin (4.7–5.6). The surface pressure–area per molecule (π–A) isotherms were obtained. Their analysis showed that all substances studied affected the phospholipid membrane which was revealed in the changes of mean molecular area, compression modulus and pressure of the liquid-expanded/liquid-condensed (LE/LC) phase transition. The results were discussed in terms of the nature and strength of mutual interactions. The more profound effect was found at low surface pressures at which the monolayers occurred in more expanded state. However, at the surface pressure corresponding to that of biological membranes, systems had very similar parameters compared to model DPPC isotherm.