The forms of vitamin B₁₂ compounds in fertilized and unfertilized eggs were separated and identified by cellulose membrane electrophoresis, paper chromatography or micro-HPLC with a microbiological determination method. The results were as follows. 1) The main forms of vitamin B₁₂ compounds in fertilized egg were CN- B₁₂, Ado B₁₂ and HSO₃^-- B₁₂, while those in unfertilized egg were CN- B₁₂ and HSO₃^-- B₁₂. 2) The difference between the forms of vitamin B₁₂ compounds in fertilized and unfertilized eggs was Ado B₁₂ content, which was higher in fertilized egg. This may suggest that a new metabolic pathway including Ado B₁₂ dependent enzyme system is developed in fertilized egg by the incubation after fertilization.