Purification of lamellar bodies from human amniotic fluid

Lamellar bodies were purified from human amniotic fluid obtained from pregnancies at 17, 31, and 40 weeks' gestation by means of Sephacryl S-300 column chromatography. Fractions were labeled with diphenylhexatriene and two peaks of phospholipid were identified at each gestational age. The size of peak I increased relative to that of peak II with advancing gestational age. Further study of the two peaks from 40-week pregnancies showed that peak I contained lamellar bodies that could be identified by electron microscopy. The ratio of the concentration of protein to phospholipid for peak I varied from 0.2 to 1.1. There were four proteins which could be identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (130K, 80K, 50K, and 20K), and there was 90-degree scattering of 400 nm light. Peak II also contained phospholipid. However, bilayer structures could not be visualized by electron microscopy, and there was no 90-degree scattering of 400 nm light. Peak II had a much higher ratio of protein to phospholipid (approximately 100) and two broad bands of protein on sodium dodecyl sulfate electrophoresis (80K, 50K). The anisotropy of diphenylhexatriene in both peaks decreased with advancing gestational age. The anisotropy in peak I was always lower than that in peak II, indicating that the microenvironment in peak I was more fluid. Peak I may represent mature surfactant and peak II, precursors of surfactant.