Purification of phospholipase B from Penicillium notatum by hydrophobic chromatography on palmitoyl cellulose

Phospholipase B (lysolecithin acyl-hydrolase, EC 3.1.1.5) from the mycelia of Penicillium notatum (Institute for Fermentation, Osaka, Japan; No.4640) was adsorbed from a crude solution to palmitoyl cellulose. Adsorption was efficient at pH 4 at low ionic strength (10 mM buffer), and at pH 4-9 at higher ionic strength (1-2M NaCl in 10 mM buffer). The adsorbed enzyme was eluted from the cellulose with a suitable detergent, such as Adekatol SO-120, Triton X-100, or deoxycholate. As an application of this procedure, the enzyme was purified from an extract of the mycelia by column chromatography on a palmitoylated textile (palmitoylated gauze) with an overall recovery of 59% and a 38-fold increase in specific activity. By subsequent column chromatographies on Amberlite XAD-2, Sephadex G-100 and G-150, and DEAE-Sephadex A-50, the enzyme was purified about 4,000-fold to a nearly homogeneous state from a mycelium extract with an overall recovery of 37%.