_____________________________________________________________________________________________________ *Corresponding author: E-mail: sagartrivedi2000@gmail.com; Journal of Pharmaceutical Research International 33(60A): 233-240, 2021; Article no.JPRI.79564 ISSN: 2456-9119 (Past name: British Journal of Pharmaceutical Research, Past ISSN: 2231-2919, NLM ID: 101631759) Stability Indicating HPLC Method for Estimation of Thymoquinone in Nasal Simulated Fluid: Method Development and Validation Sagar Trivedi a* , Abhidha Deshpande b , Veena Belgamwar a , Vidyadevi Bhoyar a and Kamlesh Wadher b a Department of Pharmaceutical Sciences, Rashtrasant Tukadoji Maharaj Nagpur University Campus, Nagpur, Maharashtra, India. b Department of Pharmaceutics, Smt. Kishoritai Bhoyar College of Pharmacy, Kamptee, Nagpur, Maharashtra, India. Authors’ contributions This work was carried out in collaboration among all authors. All authors read and approved the final manuscript. Article Information DOI: 10.9734/JPRI/2021/v33i60A34480 Open Peer Review History: This journal follows the Advanced Open Peer Review policy. Identity of the Reviewers, Editor(s) and additional Reviewers, peer review comments, different versions of the manuscript, comments of the editors, etc are available here: https://www.sdiarticle5.com/review-history/79564 Received 09 October 2021 Accepted 18 December 2021 Published 19 December 2021 ABSTRACT Introduction: A simple rapid and precise HPLC method was developed for estimation of TH in nasal simulated fluid and stability was assessed in various stressed conditions. Methods: Chromatographic separation of TH in nasal simulated fluid was done using HPLC AS- 4050 coupled with Jasco UV 2075 Plus detector, Jasco LC-Net 11/ADC valve, Jasco PU-2080 pump and hypersil gold C18 (250x6x5 μm) column, ChromNAV 2.0 Chromatography Data System software with mobile phase as acetonitrile: water (65:35) and acetonitrile: NSF (60:40) at a flow rate of 1ml/min and having run time of 10 min with loop volume of 20 μl and detection wavelength of 252 nm. The method was validated for accuracy, precision, linearity, specificity, and sensitivity in accordance with ICH (Q2B) guidelines. Results: The results of all the validation parameters were found to be within the acceptable limits. The calibration plots were linear over the concentration ranges from 2 to 14μg/ml. The accuracy and precision were found to be between 97.04±0.112 to101.081±0.0191and ≤2% for three drugs. Developed method was successfully applied for the determination TH in nasal simulated fluid and recovery was found to be >98% for three drugs. The degradation products produced as a result of stress studies did not interfere with drug peak. Original Research Article