In this paper various factors that effected on separation of mono and di substituted nitro toluene compounds was investigated by gas chromatography method and optimum separation condition was reported for 2-nitrotolouen (2NT), 4-nitrotolouen (4NT), 2,3-dinitrotolouen (2,3DNT), 2,4-dinitrotolouen (2,4DNT), 2,6-dinitrotolouen (2,6DNT) and 3,4-dinitrotolouen (3,4DNT) compounds. Optimum carrier gas flow rate and injection volume was 20-30 ml.min-1 and 0.3 t1, respectively. Isothermal separation method wasn’t accepted for given good results and then programming mode (T1=80 OC, t1=6S, scan rate=30, T2=240, t2=6s) was used.