Acori Tatarinowii Rhizoma (ATR), the rhizome of Acorus tatarinowii Schott, is a common traditional Chinese medicine being used clinically for mental disorder. However, other Acorus species herbs are all having the same Chinese name 'Chang Pu', making the confusion in herbal market. Acori Graminei Rhizoma (AGR) and Acori Calami Rhizoma (ACR) are common adulterants of ATR. Here, we aim to provide a comparative analysis between ATR, AGR, and ACR in potentiating neuronal differentiation. Volatile oil, derived from Acorus species, was applied onto cultured PC12 cells, and various parameters were determined: (i) transcriptional activation of neurofilament promoters was determined by the promoter-driven luciferase activity assay; (ii) the neurite outgrowth of PC12 cells was captured and measured; and (iii) the neurofilament expression and its underlying mechanism were analyzed by western blotting. The co-treatment of ATR, AGR, or ACR volatile oil with low concentration of nerve growth factor (NGF) could potentiate the NGF-induced neuronal differentiation in cultured PC12 cells. In addition, application of protein kinase A inhibitor H89 in cultures blocked the induction of neurofilament. Among these three Acorus species, ATR volatile oil showed the highest NGF-induced induction in neurite outgrowth and neurofilament expression, as compared with that of AGR and ACR. Copyright © 2017 John Wiley & Sons, Ltd.