The purpose of this study was to investigate the effect of subjective fatigue on ball velocity and control ability after long-time repeated pitching among women's softball pitchers. 7 elite college female softball pitchers were recruited to perform a simulated game consisting of 15 fastball pitches per inning up to 10 innings to approach subjective fatigue. Ball velocity, strike ratio and error distance per inning were analyzed. Difference of pitching performance between pre and post subjective fatigue was analyzed using dependent sample z test of nonparametric statistics. Performance of first two innings was shown that the ball velocity was 54.53 ± 2.21 mile/hr, pitching accuracy ratio was 59.05 ± 16.97%, and error distance was 21.91±1.83 cm. Most pitchers felt fatigue after pitching 8 innings, the pitching counts were 120. The ball velocity was drop to 52.90 ± 2.19 mile/hr with significance, and the error distance was significant lager to 31.53±0.40 cm. The conclusion of this study was that ball velocity and control ability were a downward trend with increase of pitching innings. Suggest the softball coaches and players could monitoring the pitching count, or observation pitching accuracy to determine the status of pitcher fatigue. There will be a recommendation for softball coaches and players, we could observe pitcher's fatigue condition through monitoring pitching counts and control ability.