cn w -J 0 a w (jn w -J w CY. o 2 3 4 5 TIME (min) FIG. 2. Release of penicillinase from noninduced Staphylococcus aureus by means of the Nossal bacterial cell disintegrator. Samples were shaken for periods not exceeding 30 sec at an initial temperature of 5 C. Internal heating necessitated rechilling (15 min) after each interval of shaking. The 10-ml aqueous cell suspension contained 0.5 g washed organisms (dry weight) to which were added 8 g acid-washed 75-M glass beads (Superbrite 150,

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... ometrically by the method of Henry and Housewright (J. Biol. Chem. 167:559, 1947), and of total protein, as measured colorimetrically by the method of Lowry et al. (J. Biol. Chem. 193:265, 1951). Release of enzyme appeared to parallel release of total protein until about 50% of both were liberated. Appearance of enzyme in the supernatant fluid increased rapidly with shaking time for the first 2 min. At that time the content was 80 to 85% of that obtained in 5 min. Further shaking yielded little more enzyme, possibly because of thermal inactivation or absorption of liberated enzyme onto the glass beads (Kogut et al., Biochem. J. 62:319, 1956). nesota Mining and Manufacturing Company). Supernatant cell-free fluid was obtained by low speed centrifugation (1,000 X gfor 30 min). O O = Penicillinase activity of supernatant fluid; A--A = protein content of supernatant fluid. Reference points are shown for 50% of the enzyme activity and protein content of the original whole cell suspension.