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Tobacco plants are notably known for its pesticidal properties, particularly due to its nicotine content. In this study, Nicotiana tabacum L. bio-oil was obtained using pyrolysis technique. The safety of the bio-oil to be used as bioinsecticide was analyzed through acute oral toxicity test by administering 5000 mg bio-oil/kg body weight of female winstar rats that were analogous to humans. It was concluded that the bio-oil was not toxic due to absence of mortality and no significant change in the body weight and behavior of the rats.