Eco‐Friendly and Highly Sensitive Liquid Chromatography Mass Spectrometric Method for Trace Level Quantification of N ‐Nitroso Dorzolamide in Dorzolamide Drug Substance and Ophthalmic Formulations

ABSTRACT Nitrosamine contaminants in pharmaceutical products are recognized for their considerable genotoxic potential, highlighting the need for ultra‐sensitive and dependable analytical techniques. This research presents an innovative and environmentally friendly liquid chromatography–mass spectrometry method for the ultra‐trace detection of N ‐nitroso dorzolamide ( N ‐DRZ), a suspected impurity in dorzolamide‐based APIs and ophthalmic solutions. The acceptable intake value and control limit was derived using Carcinogenic Potency Category Approach for N ‐DRZ. The study employs synthesis and structure confirmation by NMR spectroscopic techniques, streamlined sample preparation, and effective chromatographic separation using a Waters Cortecs T3 C18 column, and detection via negative electrospray ionization in multiple reaction monitoring mode, targeting the transitions m / z 352.03 → 215.98 (quantifier) and 352.03 → 78.78 (qualifier). Method validation followed ICH Q2(R2) standards, demonstrating excellent linearity ( R 2 > 0.999), strong specificity, high precision, and a notably low detection limit of 0.03 µg/g. The method's green profile was evaluated using Green Analytical Procedure Index (GAPI), Analytical GREEnness (AGREE), Eco‐Scale, and Blue Applicability Grade Index (BAGI) assessments, all confirming its environmental advantages over traditional methods. Application to commercial dorzolamide formulations confirmed compliance with regulatory requirements, highlighting the method's reliability and high sensitivity, and reinforcing its importance for public health and safety through sustainable impurity monitoring and robust pharmaceutical quality assurance.