384 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS nique at constant pressure shows that in the sys. tems cetyl alcohol- sodium cetyl sulfate, cholesterol- sapchin and cholesterol-sodium cetyl sulfate, 1:1 complexes definitely Interaction I½odmcj to Complexes b•tw•n HAEMOGLOBIN and C225ULPHAToe pH =10 20øC $urfac• Pressure = 2 8,1yn½s/cm 9 I'lc• mø•Tn/700co 2('3 40 6• 1oo 80 ._• •J O•4o Time in minut• Figure 3. exist irrespective of the pressure of expansion and concentration of the penetrating substance in the under- lying solution (22). The influence of salts in the under- lying solution is very important. Harkins and his co-workers, using distilled water, were unable to re- peat the results of Schulman and Stenhagen (2) for the system cetyl alcohol-sodium cetyl sulfate, w•ich had been obtained on M/S00 phos- phate buffer at pH 8. They ascribed the discrepancy to defects in the earlier workers' technique, but Schulman and Matalon (6) showed that injection of phosphate beneath a mixed film of cetyl alcohol and sodium cetyl sulfate on distilled water caused an immediate increase in surface pressure. However, it was also found that during com- pression of a mixed film, higher pressures were attained, due to closer packing in the film, than dur- ing spontaneous penetration. BIOLOGICAL APPLICATIONS The concepts described above have important biological applica- tions. In the first place, Schulman andRideal showed (1) that hemolytic activity is closely associated with the equilibrium penetration pres- sure of the dissolved substance when penetrating a cholesterol monolayer, the order of activity being the same as that given above for the latter phenomenon. Some hemolytic sub- stances, such as the long-chain alco- hols, have a strong dispersing ac- tion on protein films, and it seems that hemolysis is also due to the penetrating substance displacing the protein portion of the cell wall. The specificity mentioned earlier, namely that of cholesterol with sap- chin or long-chain sulfates, and long-chain hydrocarbons polyethyl- ene oxides or alcohols for protein films can be used to distinguish be- tween the components of cells. Thus, lysis of cells can be obtained by penetration of the available cholesterol portion by cholesterol- penetrating substances or dispersion of the. available protein portion by protein-penetrating or -dispersing : substances. A protein monolayer collapses at a surface pressure of about 16 dynes/cm., and so substances that

lower the sin-face tensoin by about i•i•!i::•: 20 dynes/cm. will lyse cells if pro- rein is available and if the agent is in su•cient concentration to lower !i'll •':the surface tension to that extent. The cholesterol-active substances are effective in concentrations below 10 -a per cent and so, are active at concentrations which lower the sur- face tension approximately 5 dynes/ cm. or less. Substances which adsorb on to protein monolayers, and do not penetrate, are all agglutinating agents, as they render the hydro- philic surfaces of the cells hydro- phobic. The chemistry of the cell sin'- faces can thus be established by the nature of the substances which at- tack them. Further, some cells can be shown to be resistant to both these types of agent, and conse- quently to be composed of chitin or other polysaccharide material. It is also of interest that active sub- stances in sublytic doses can induce a permeability of .the cell to mate- rials which are not normally taken up or given out by the cell. PENETRATION AND COMPLEX-FORMATION IN MONOLAYERS 385 SUMMARY The penetration and adsorption phenomena may be summarized under the following headings: (a) constant area, (b) variable area and pressure, (c) constant pressure. 1. kFeak _ nteraction. (a) If the surface pressure of the soluble agent is •r•, and the collapse surface pres- sure of the film-forming molecules is *rv, then if •r• •rv, displacement of the film material will take place, if there is no association by polar forces between the two molecules. The resultant surface pressure will be •r 8. (b) Ejection of a compound in a mixed film will take place at its collapse pressure if there is no polar interaction. 2. Strong Interaction. (a) If there is strong association by van der Waals forces between the film- forming molecules and the soluble molecules in the underlying solu- tion, the resultant surface pressure of the mixed film will approach •rs q- •rv, well above the collapse pressure of either of the compo- nents. (b) No ejection occurs from a mixed monolayer of the two asso- ciated molecules, but a 1:1 com- plex of very high collapse pressure is formed. Ejection of the excess of dissolved molecules from the 1:1 complex monolayer can be followed experimentally. Phase diagrams obeying a two-dimensional phase rule can be plotted for these mixed monolayers. 1 and 2 (c). The percentage in- crease in area of a monolayer on penetration by soluble molecules from the underlying solution, at constant pressure and below the re- sultant equilibrium surface pressure of the monolayer at constant area, is directly related to the ratio of the molecular areas of the interacting species at their collapse pressures. Thus, a cholesterol monolayer with an area of 40 A.2 per molecule on being penetrated by sodium cetyl sulfate (20 A.2) will expand by half