In screening for acetylcholinesterase (AChE) inhibitors and cytotoxic constituents from medicinal plants, a bioactivity-guided isolation of the alkaloidal extract of the whole plant of Palhinhaea cernua led to the isolation of six lycopodium alkaloids. Their structures were elucidated by spectroscopic methods (IR, MS and NMR). The absolute configuration of 1 was established by computational methods. To our best knowledge, compound 1 was a rare cernuane-type alkaloid bearing an oxygen bridge between C-12 and C-15 in Lycopodiaceae. Among these alkaloids, compound 5 was the most active against AChE with IC50 value of 1.9 mu M, and the new compound 1 showed weak inhibitory activity with IC50 value of 102 mu M. Moreover, the new compound 1 and the known one 3 inhibited A549 cell line with IC50 values of 12.6 and 18.4 mu M, respectively. Meanwhile, preliminary structure-activity relationships were discussed in this study. (C) 2014 Phytochemical Society of Europe. Published by Elsevier B.V. All rights reserved.