Embryo development is a complex and tightly controlled process. Nanoparticle injury can affect normal development and lead to malformation or miscarriage of the embryo. However, the risk that these nanoparticles may pose to reproduction is not clear. In this study, chitosan nanoparticles (CSNP) of near uniform size, in the range of 100 nm, were synthesized and confirmed by a particle size analyzer and transmission electron microscopy. Morulae-stage embryo exposure to CSNP during in vitro culture caused blastocyst complications that had either no cavity or a small cavity. Furthermore, CSNP-treated embryos showed lower expression of not only trophectoderm-associated genes but also pluripotent marker genes. When blastocysts developed in both media with and without CSNP were transferred to recipients, the percentage of blastocysts resulting in viable pups was significantly reduced. These detrimental effects are linked to the reduction of total cell numbers, enhanced apoptosis, and abnormal blastocoels forming at the blastocyst stage, indicating that CSNP treatment might have long-term adverse biological effects in view of pregnancy outcome.