The aim of this study was to evaluate the effects of probiotic pretreatment on the alteration and recovery of gut microbiota after bowel preparation and its correlation with minor complications. This was a randomized, double-blind, placebo-controlled pilot trial that included participants 40–65 years of age. Participants were randomly provided probiotics (active group) or placebo (placebo group) for 1 month before the colonoscopy and their feces collected. A total of 51 participants were included in the present study (26 in the active group and 25 in the placebo group). In the active group, the microbial diversity, evenness, and distribution were not significantly changed between before and after bowel preparation, but did change in the placebo group. The number of gut microbiota that decreased after bowel preparation in the active group was lower than in the placebo group. On the seventh day after colonoscopy, the gut microbiota in the active group was restored to almost the same level as before bowel preparation. In addition, we identified that several strains were assumed as key microbiota in early colonization and some taxa were increased only in the active group after bowel preparation. In multivariate analysis, taking probiotics before bowel preparation was identified as a significant factor for decreasing the duration of minor complications (odds ratio 0.13, 95% confidence interval 0.02–0.60, p = 0.027). Probiotic pretreatment had benefits on the alteration and recovery of gut microbiota and possible complications after bowel preparation. Probiotics may also aid in the early colonization of key microbiota.