Marine fungi are a promising source of novel and biologically active natural products for drug discovery (Duarte et al., 2012; Xu et al., 2015). In the last decades, the number of marine-derived natural products has steadily increased (Debbab et al., 2010), of which the nitrogen-containing alkaloids play a special role in the natural products field (Joule, 2016). Azaphilones are a novel class of alkaloids with different biological activities including antimicrobial (Laakso et al., 2003; Kanokmedhakul et al., 2006), nematicidal (Dong et al., 2006), antioxidative (Quang et al., 2004), cytotoxic (Wang et al., 2015), anti-tau aggregation (Paranjape et al., 2015), anti-parasitic (Suzuki et al.,1999), anti-inflammatory (Cheng et al., 2012), and antiviral effects (Osmanova et al., 2010). In our ongoing search for new bioactive azaphilones (Li et al., 2014; Ma et al., 2015; Xiao et al., 2012), we have focused on an alga-derived fungal strain, Penicillium sp. ZJ-27, which was collected from sea water of Zhanjiang Mangrove National Nature Reserve in Guangdong Province, China. Four new azaphilones (1–4), along with four known analogues (5–8), were isolated from Penicillium sp. ZJ-27. Herein, we report the isolation, structural elucidation and bioactivity of compounds 1–8.