The aim of the study is to levels of nicotine and cotinine were elevated the oxidative stress malondialdehyde (MDA) and inflammation as nitric oxide (NO2 and NO3) may possibly be associated with decreased antioxidant enzyme activities and can sensitively indicate the production of reactive oxygen species (ROS). To evaluate the quantitative analysis of nicotine and cotinine levels and the alterations in the selected parameters of antioxidant metabolisms during nitroxidative stress in the saliva of smokeless tobacco consumers. Saliva nicotine and cotinine was measured by HPLC method and nitric oxide, lipid peroxidation and activities of antioxidant enzymes were estimated by spectrophotometric methods. Significant increase in concentrations of nicotine and cotinine levels of saliva in smokeless tobacco users in comparison to controls. Saliva lipid peroxidation was increased in experimental subjects (gutkha group 39.28% and khaini group 25.00%) as compared to controls and nitric oxide in the form of nitrites and nitrates was significantly increased in the saliva of smokeless tobacco users compared to controls. The activity levels of antioxidant enzymes were decreased in the saliva of the smokeless tobacco users in comparison with normal controls. A strong positive correlation of nicotine and cotinine with nitroxidative stress markers in gutkha and khaini users. Increased expression of inducible nitric oxide synthase (iNOS) enzyme leads to intoxication in saliva and indirectly induces inflammation process. Increased production of reactive oxygen species (ROS) and decrease in the activity levels of antioxidant enzymes in the saliva of smokeless tobacco users indicate conspicuous cell and tissue damage.