Multiple Reaction Monitoring LC‐MS/MS Quantification of N‐Nitrosamine Drug Substance Related Impurities of Clonidine in Drug Substance and in Tablet Dosage Form

ABSTRACT N‐nitrosamine drug substance related impurities (NDSRIs) present recent concerns for manufacturers of both drug substances and products. The regulatory authorities prioritize understanding the chemical properties, potential formation, and control methods of these impurities due to their adverse effects. Clonidine is a medication for hypertension and attention deficit hyperactivity disorder, characterized by a secondary amine in its structure and a propensity to generate NDSRIs. Its chemical structural evaluation reveals a possible risk for the formation of two separate NDSRIs, namely, N‐nitroso and N‐dinitroso clonidine. The carcinogenic/mutagenic category was established using carcinogenic potency categorization approach method, and the corresponding acceptable intake values were implemented. A liquid chromatography–tandem mass spectrometry method in a multiple reaction monitoring mode was established to quantify both impurities in its drug substance and tablet dosage. A Poroshell C18 column of 150 × 4.6 mm, and with particle size of 2.7 μm was employed at a temperature of 30°C. A gradient elution method opted with water and acetonitrile as the mobile phases, containing formic acid as a buffering agent. Method validation conducted as per regulatory standards and demonstrated superior specificity, linearity, accuracy, precision, and robustness. As a result, this method is regarded as suitable for its intended purpose.