As part of our ongoing search for bioactive constituents from natural Korean medicinal resources, a bioassay-guided fractionation and a chemical investigation of the methanolic extract from the rhizomes of Acorus gramineus resulted in the isolation and identification of two alkaloids, including a new aporphine-type alkaloid, named gramichunosin, and a known pyrrole alkaloid. Their structures were determined by a combination of 1D and 2D nuclear magnetic resonance (NMR) spectroscopic analysis and high resolution mass spectrometry (HRMS). This is the first report of alkaloids from A. gramineus. The compounds isolated showed antiproliferative activities against A549, SK-OV-3, SK-MEL-2 and HCT-15 cell lines with EC50 values in the range of 7.46-45.23 mu M. Moreover, the anti-neuroinflammatory activities of these compounds were determined by measuring the nitric oxide (NO) levels in the medium using murine microglia BV-2 cells. Compound named gramichunosin inhibited NO production in lipopolysaccharide-stimulated BV-2 with IC50 values of 7.83 mu M.