The production of an acid glycoprotein at fertilization was confirmed to occur in the egg of Arbacia punctulata. Chemical analyses of such "fertilization product" and jelly coat substances were carried out. These substances are both glycoproteins containing hexose, hexosamine, sulfate, inorganic cations and 17 amino acids. However, the differences in the respective amounts of their constituents show that the fertilization product, which is believed to be the perivitelline space material, is a different substance from jelly. The release of this substance is detected as the formation of "fertilization acid." The results of the analyses also suggest that the membrane-hardening substance is a protein or protein-rich substance.