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We established an efficient methodology for the determination of those analytes in spiked nasal mucosa and nasopharyngeal swabs from real human samples. A simple and quick protein precipitation method was used for sample extraction, using acetonitrile. RP-HPLC\/DAD method was performed using an Exsil 100 ODS C18 (250\u2009\u00d7\u20094.6\u00a0mm, 5\u00a0\u03bcm) column with an acetonitrile: water (70:30 v\/v) solvent system at a flow rate of 0.7 mL\/min. A photodiode array detector was applied at 240\u00a0nm. A good separation of the three proposed analytes with a short run time of 10\u00a0min was noted. Our method was validated according to FDA guidelines for bioanalytical validation methods. Calibration curves were linear in nasal mucosa samples at concentration ranges of 8\u2013125, 10\u2013100, and 10\u2013125\u00a0\u00b5g\/mL, with average recoveries\u2009\u00b1\u2009SD of 101.56%\u00b10.39, 102.45%\u00b10.86, and 104.61%\u00b14.52 for AZT, FP, and OXY; respectively. The results of precision and accuracy are within acceptable limits. According to stability assays, the three analytes under investigation were stable throughout sample preparation, storage, and injection. Our method was applied to real nasopharyngeal swabs. It shows that the results of the swabs were not affected by gender or age. Good recoveries with low % RSD were observed: 99.03% \u00b1 0.75, 100.02% \u00b1 0.94, and 100.94% \u00b1 1.98 for both genders, and 100.45% \u00b1 0.96, 100.69% \u00b1 1.08, and 100.32% \u00b1 1.53 for different ages for AZT, FP, and OXY; respectively. Moreover, the amount of those drugs in the nasal mucosa was observed for seven hours, and a constant concentration with a low% RSD was noted for the first four hours. Therefore, this method can be applied to monitor the therapeutic dose in the nasal mucosa for the determination of those analytes.<\/jats:p>","DOI":"10.1038\/s41598-024-82387-7","type":"journal-article","created":{"date-parts":[[2025,2,4]],"date-time":"2025-02-04T15:06:30Z","timestamp":1738681590000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Validated HPLC method for simultaneous determination of azelastine hydrochloride fluticasone propionate and oxymetazoline in nasal mucosa and nasopharyngeal swabs from real human samples"],"prefix":"10.1038","volume":"15","author":[{"given":"Neven N.","family":"Mikawy","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nancy","family":"Magdy","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marwa H.","family":"Mohamed","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amira M.","family":"El-Kosasy","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,2,4]]},"reference":[{"key":"82387_CR1","doi-asserted-by":"crossref","unstructured":"Frew, A. 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