Isolation and identification of embryo-derived platelet-activating factor in mice

We isolated a platelet-activating factor (PAF) derived from mouse embryos and identified it by bioassay utilizing washed rabbit platelets. We also tried further characterization of the factor by high-performance liquid chromatography (HPLC) and electron-impact mass spectrometry (EI-MS). 1. In the thin-layer chromatography (TLC) of the medium after a two day culture of 30 embryos at the two-cell stage, no obvious spot appeared at the site of 1-O-acetyl-2-O-alkyl-SN-glyceryl-3-phosphocholine (AGEPC). But a small, pale spot appeared at the site of lyso-AGEPC. 2. A remarkable aggregation activity was present at the site of the Rf and appearance time that was almost the same as for AGEPC in both TLC and HPLC. However, detection of the substance by UV absorption (203nm) in HPLC was unsuccessful. 3. The PAF activity shown in the culture medium was suspected to be 10(-10 - 10(-11) M/embryo and it was not inhibited by pretreatment with indomethacin and ADP scavenger. 4. Embryo-derived PAF had an HPLC appearance time 1-2 min. longer than that of synthetic AGEPC (C16). EI mass spectrum of embryo-derived PAF was partly inconsistent with that of synthetic AGEPC (C16). These results confirm the evidence of PAF release from mouse embryos and show the possibility that a far greater amount of its lyso-derivative is present in the culture medium. Embryo-derived PAF is presumed to be AGEPC or a substance resembling AGEPC in its chemical structure and some structural difference may exist between embryo-derived PAF and synthetic AGEPC (C16).