Human follicular fluid (FF) forms the microenvironment of the developing oocyte and has an important influence on embryo development. Although some published report showed their results of associations between antioxidant activity of FF and embryo quality, there was no information on the lower limit of antioxidant activity in FF which had negative impact on embryo quality. The antioxidant activity of 407 FF samples from 52 infertile patients undergoing IVF were collected and determined by alpha-diphenyl-beta-picrylhydrazyl (DPPH) radical scavenging and reducing power assay. The lower limit of DPPH radical scavenging percentage and reducing power were determined to be 18.3% and 0.257, respectively. It means FF may adversely affect the embryo development when the DPPH radical scavenging ability or reducing power below the lower limit. This estimated lower limit was further validated in FF samples of embryo grading I-IV. This cut off value of antioxidant activity of FF is expected to assist embryologists and clinicians in predicting development of embryo from infertile patients undergoing in vitro fertilization (IVF).