The rigorous design of preclinical experimental studies of candidate neuroprotectants for the treatment of acute ischemic stroke is crucial for the success of subsequent randomized clinical trials. The efficacy of Ginkgo biloba extracts (GBEs) in complex mixtures for the treatment of acute ischemic stroke remains unclear. In this preclinical randomized controlled trail (pRCT), the effects of a novel (n)GBE containing pinitol versus traditional (t)GBE without pinitol were evaluated on the mouse models of acute transient and permanent stroke, separately. The sample size, an important aspect of study design, was calculated based on our experimental data. Mice with ischemia that were induced by transient middle cerebral artery occlusion (tMCAO) or permanent distal middle cerebral artery occlusion (pdMCAO), were treated with vehicle, nGBE, tGBE, or pinitol alone by tail-vein injection. Our results showed that nGBE significantly reduced infarct size in mice with tMCAO compared with vehicle-treated control mice. Both nGBE and tGBE significantly reduced infarct size in mice with pdMCAO compared with the vehicle-treated controls. None of the three treatments rescued weight loss or prevented the neurological deficits in either the tMCAO- or pdMCAO-model mice. These findings suggest that nGBE, which includes all of the components of tGBE and pinitol, is neuroprotective in two ischemic stroke models. Additional studies of complex GBE mixtures for stroke treatment compared to single component medications are undergoing evaluation.