The kinetics of
hemolysis of pig erythrocytes by
digitonin was continuously monitored by a
potassium selective
electrode. The following minimal mechanism of
hemolysis was postulated, based upon kinetic measurements: 2 D + E (1)in equilibrium E x D2 (2)leads to (E x D2) (3)leads to (E x D2) where D denotes a
digitonin molecule and E a specific
digitonin binding-site on the membrane. The first step (1) represents a rapid reversible combining of
digitonin with specific binding-sites on the membrane. The second step (2) is prelytic, related to the time required for bound
digitonin molecules to alter the membrane structure so much that
hemolysis may take place; this step has a transition temperature at 26 degrees C, probably related to the "melting" of specific membrane structures at that temperature. The third step (3) is hemolytic, and comprises the changes within the
digitonin-altered membrane during
hemolysis; it is stongly influenced by temperature. Lowering of temperature slows down the rate of
hemolysis and increases the quantity of
digitonin required to obtain a fixed extent of
hemolysis. It appears that two molecules of
digitonin combine with a single binding-site on the outer face of the membrane in a
digitonin-membrane complex.