Objectives: To identify the unilateral effect of VOCs on cotinine metabolism. Methods: Measurements of 13,652 participants included sociodemographic characteristics and serological indicators (VOCs and cotinine). Correlation between VOCs and cotinine and the effect of VOCs on cotinine metabolism were analyzed by Spearman analysis and multivariable logistic regression analysis. Results: Among smokers infected with HIV | HBV | HCV with the largest exposure dose to tobacco, the intensity of the association between VOCs and cotinine was the strongest. High concentrations of 1,2-Dichlorobenzene (OR: 1.101, CI: 1.069–1.134), Benzene (OR: 1.354, CI: 1.123–1.632), 2,5-Dimethylfuran (OR: 1.054, CI: 1.027–1.082) and Nitrobenzene (OR: 1.347, CI: 1.039–1.746) and low concentration of Carbon Tetrachloride (OR: 0.760, CI: 0.661–0.873) in blood might be independent risk factors leading to the increase of serum metabolite cotinine. Conclusions: The influence intensity of four VOCs (including 1,2-dichlorobenzene, benzene, carbon tetrachloride, 2,5-dimethylfuran and nitrobenzene) on serum cotinine metabolism has a dose-response relationship, that is, the higher the tobacco exposure, the stronger the influence intensity. In addition, this study used molecular epidemiology to indirectly provide new ideas and views on the harm of smoking for smokers.