The Purification and Properties of Lysozyme

Fleming (1) in 1922 reported the occurrence of a bacteriolytic principle, lysozyme, in egg white, tears, and other animal fluids. Since then much work has been devoted to its bacteriological behavior, but the only work of importance on its purification and chemical nature was that of Wolff (2). Wolff precipitated diluted egg white with colloidal iron, evaporated the filtrate to a small volume, precipitated the lysozyme with acetone, and dialyzed the aqueous solution of the precipitate. The last two steps were repeated several times and finally a white powder was obtained, soluble in water, but insoluble in the common organic solvents. It was reported to give no Molisch reaction and to be free of sulfur, phosphorus, and nitrogen, although the biuret reaction was weakly positive. In the present work an attempt was made to obtain the lysozyme from egg white in a purified form and to study its nature. Preparation-The starting material for purification was a dry powder obtained by precipitation of native egg white with 9 volumes of ice-cold acetone. After the mixture had stood cold overnight, the precipitate was filtered off, washed well with acetone and ether, and dried in a heated vacuum desiccator over PZOs. Usually 14 to 15 per cent of the total weight of the egg white was thus obtained as a dry white powder with no loss of potency. Acetone precipitation at the original alkaline pH of egg white renders the greater part of the proteins insoluble, while the lysozyme retains its solubility in aqueous media. In a procedure similar to that of Wolff, 40 gm. lots of the egg white powder were extracted with three 500 cc. portions of 0.9