AbstractAcid nitrophenyl phosphatases from sea urchin eggs and embryos were investigated by gel filtration. Four different forms were found in Hemicentrotus pulcherrimus, and three forms in Anthocidaris crassispina and Pseudocentrotus depressus.The first and second forms (designated AcP‐1 and AcP‐2) had the highest activity in the range of pH 5.6–6.0. The third (designated AcP‐3) had an apparent optimum pH between pH 4.3 and 4.8, and the fourth (designated AcP‐4) showed the maximum activity at pH 3.0. AcP‐1 was much more thermolabile than AcP‐2 and AcP‐3 at 56°C. NaF inhibited AcP‐2, AcP‐3, and AcP‐4 but not AcP‐1. AcP‐1, AcP‐2, and AcP‐3 were observed in the three species, whereas AcP‐4 was not detected in A. crassispina and P. depressus. AcP‐1, AcP‐2, and AcP‐3 were separted by gel filtration.AcP‐1 and AcP‐2 of A. crassispina and H. pulcherrimus were studied in developing embryos. The behavior of these forms in gel filtration changed during development, from unfertilized eggs to the pluteus stage.