Isolation, characterization and phospholipid composition of lamellar bodies and subcellular fractions from dog lung

Abstract

1. 1. Lamellar body fractions from dog lung can be separated by a procedure based on differential centrifugation before ultracentrifugation onto a discontinuous sucrose gradient. This fraction yields about 1 % of total protein from the homogenate. 2. 2. The different fractions obtained in the isolation were assayed for the measurement of four subcellular marker enzymes: β- N-acetylglucosaminidase, acid phosphatase, 5'-nucleotidase and succinate dehydrogenase. 3. 3. Lamellar bodies were not contaminated by mitochondria (0.7 succinate dehydrogenase relative specific activity), whereas high specific hydrolase activities were found ( β- N-acetylglucosaminidase and 5'-nucleotidase were enriched 1.8- and 2.8-fold, respectively). 4. 4. The chemical criterion was established by measuring the specific components of lamellar bodies. The lamellar bodies have the highest phospholipid/protein ratio (0.35); cholesterol/protein ratio (0.15) and the highest phosphatidylglycerol percentages (7.9%). 5. 5. The phospholipid composition of lamellar bodies is distributed among phosphatidylcholine (64.5%), phosphatidylethanolamine (11%), phosphatidylglycerol (7.9%), sphingomyelin (4%), phosphatidylserine and phosphatidylinositol (3%), respectively. The remainder were considered as trace amounts ( < 1%).