Nitrosamines　and　nitrosoureas　are　two　families　of　N-nitroso　compounds　(NNCs)　with　a　common　nitroso　group　connected　to　the　nitrogen　atom;　however,　the　two　types　of　analogs　often　exhibit　distinct　bioactivities　when　they　have　similar　substructures　in　the　side　chains.　Nitrosamines　are　usually　considered　to　be　potent　carcinogens　to　various　organs　and　species,　whereas　nitrosoureas　can　inhibit　cancer　cell　growth.　This　distinction　between　the　two　types　of　compounds　complicates　our　understanding　of　the　structurebioactivity　relationship　of　NNCs.　In　this　work,　the　mechanisms　for　the　formation　of　DNA　interstrand　crosslinks　induced　by　nitrosoureas　and　nitrosamines,　which　are　an　important　DNA　damage　related　to　the　cytotoxicity　of　NNCs,　were　compared　using　the　density　functional　theory　method.　1,3-Bis-(2-chloroethyl)-1-nitrosourea　(BCNU)　and　the　bifunctional　metabolite　of　diethylnitrosamine,　N-nitrosoethyl-(2-hydroxyethyl)amine　sulfonate　(NEHEAS),　were　used　as　the　computational　models.　Four　pathways　for　the　formation　of　GC　crosslinks　induced　by　BCNU　and　NEHEAS　were　compared.　The　results　show　that　the　crosslinking　mechanisms　initiated　by　alpha-alkylation　are　energetically　feasible　for　both　BCNU　and　NEHEAS,　and　the　decompositions　of　BCNU　and　NEHEAS　represent　the　rate-limiting　steps.　However,　for　the　crosslinking　mechanisms　initiated　by　beta-alkylation,　BCNU　and　NEHEAS　exhibit　different　preferences.　The　energy　barrier　for　the　decomposition　of　the　BCNU-induced　beta-alkylation　product　is　much　higher　than　that　of　NEHEAS.　Such　an　energy　barrier　may　inhibit　the　mechanism　of　BCNU　initiated　by　beta-alkylation.　Consequently,　it　is　postulated　that　BCNU　and　NEHEAS　induce　DNA　interstrand　crosslinks　via　different　mechanisms,　which　may　distinguish　the　bioactivity　of　the　two　types　of　NNCs.　Copyright　(c)　2012　John　Wiley　&　Sons,　Ltd.