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The aim of this study was to examine the effect of Shengmai-san (SMS) on learning and memory impairment induced by scopolamine (1 mg/kg, i.p.) in mice. The passive avoidance task (PAT) and Morris water maze (MWM) test served as the behavioral models for testing memory. To elucidate the mechanism of its cognitive-enhancing activity, the effects of SMS (2, 4, or 8 g/kg) on activities of acetylcholinesterase (AChE) and antioxidant enzymes and levels of acetylcholine (ACh) and nitrite were evaluated in brain homogenate. Tacrine (THA) (10 mg/kg, p.o.) was used as a reference drug. SMS (4 or 8 g/kg) significantly prevented scopolamine-induced impairments as measured by the PAT and MWM (probe trial session). SMS (4 or 8 g/kg) also significantly reduced the oxidative-nitrative stress, as evidenced by decreased malondialdehyde and nitrite levels and by its prevention of decreases in glutathione and superoxide dismutase levels. The activity of AChE was decreased in scopolamine-treated mice but was inhibited significantly by SMS treatment (4 or 8 g/kg) in both salt- and detergent-soluble fractions of brain homogenates. Further SMS treatment (4 or 8 g/kg) significantly increased the ACh levels in the brain homogenate to a level similar to that observed in THA treatment. Thus, the significant cognitive enhancement observed after 7 days of administration of SMS is closely related to the strong antioxidant properties of SMS in addition to its inhibition of brain AChE activity. These findings stress the critical impact of SMS on higher brain functions such as learning and memory.