SKIN IRRITATION BY ANIONIC SURFACTANTS 61 0 0.05 0.10 0.15 0.20 0.25 Substrate Concentration(M) Figure 22. Inhibition of acid phosphatase caused by C•2AS shown by initial rate-substrate concentration curves. Measurement of the SH releasing ability (Figure 21) revealed that like common anionic surfactants (15), these compounds also show increased releases of SH amounts with increasing alkyl chain length and that the magnitude is significantly higher in and C•6MAP than in C•2 and C•4AS (16). Based on the protein denaturating effects observed for keratin powder and contrary to our expectation from its low irritancy, it is suggested that, in general, these compounds have a strong capacity to denature protein. 1001 t i 1• 4 1• 3 I• 2 g / ml 5O •o Sur fact ant Concentration Figure 23. Inhibitory effect of monoalkyl phosphate monosodium salts on acid phosphatase as a function of the concentration.

62 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS ENZYME INHIBITION We have reported (15) an inhibitory effect of various common anionic surfactants with different alkyl chain lengths on acid phosphatase, one of the lysosomal enzymes, and suggested that although inhibition and irritation is not related, surfactants with a certain level of irritating effect seem to have a marked inhibitory effect on acid phosphatase. Therefore, evaluation of the inhibitory ability by MAP-type of surfac- tants was also required for the clarification of the causative factors responsible for low irritation. Figure 22 exhibits initial rate-vs.-substrate concentration curves of acid phosphatase in the presence of the varying concentrations of C•2AS. Judging by both decreased Vmax and non-changed Km, C•2AS has a non-competive inhibitory effect on this enzyme. The inhibitory activity of MAP surfactants on acid phosphatase shown by percentage of inhibition plotted against the additive surfactant concentrations (Figure 23) indicates that C•SF and C•2MAP have a similar inhibitory ability to CuAS, whereas the effect of C•4, C•0 and CsMAP mono-sodium salts is at a significantly lower level (16). These findings indicate that, in general, these compounds possess almost the same inhibitory potential on acid phosphatase as common anionic surfactants except soap, indicating a similar order to protein denaturing effect described in SH releasing experiments. LYSOSOME LABILIZING EFFECT In addition to protein denaturation, the membrane damaging effect of surfactants may play an important role in evoking skin roughness or irritation (20,23). Since MAP is a phosphate derivative and the main component of cell or organelle membrane is phospholipid, it seems important to clarify if there is differential interaction with membrane between MAP and typical anionic surfactants like C•2AS. Therefore, the labilizing effect of these surfactants (10) was also investigated using subcellularly isolated lysosome-rich fraction from guinea pig skin (16). Thus, 1 ml of lysosome preparation which was obtained as supernatant fraction after centrifugation at 700 g • X 1• 5. g/ ml • rl• 5x 1• 4 • 150 '"1 O0 ........... ............. •0 Ci2AS C• 14• C10• C 12•P C 14• Na TEA TEA Figure 24. Labilizing effect of monoalkyl phosphates on guinea pig epidermal lysosome-rich fraction in comparison with C•2AS and C•2-14EO.