In the present study, we have demonstrated that a novel synthetic chemical JSH-21 of N¹-Benzyl-4-methylbenzene-1,2-diamine could inhibit nitric oxide (NO) production in lipopolysaccharide (LPS)-activated macrophages RAW 264.7. The JSH-21 showed an IC50 value of 9.2 uM on the LPS-induced NO production. Furthermore, JSH-21 attenuated LPS-induced mRNA and protein levels of inducible NO synthase (iNOS) in the cells, as well as inhibited LPS-induced iNOS promoter activity. These results indicates hat the compound could down-regulate iNOS expression at the transcription level. Since NF-kB activation is a key mechanism in the expression of LPS-inducible iNOS gene, we further examined whether JSH-21 could affect LPS-induced NF-kB activation. JSH-21 inhibited LPS-induced nuclear import of NF-kB p65 in macrophages RAW 264.7 and sequentially prevented NF-kB transcriptional activity. However, JSH-21 was not effective in a LPS-induced degradation of cytoplasmic IkB-alpha in the cells. These results suggest that JSH-21 could inhibit LPS-induced NF-kB activation targeting nuclear import of NF-kB, an event downstream IkB degradation. Taken together, this study may provide a pharmacological potential of JSH-21 in the NO- or NF-kB-associated inflammatory disorders.