Nitrosamine impurities are potentially genotoxic which are considered under cohort of concern as per ICH M7 guidelines and need to be controlled at trace levels during quantification in drug substances and drug products for safe human consumption. Recent regulatory requirements also suggest the need to have highly sensitive analytical methods for the accurate quantification of Nitrosamine impurities. In this paper we have presented simple, rapid and ultra-sensitive LC-MS/MS method for six potential genotoxic nitrosamine impurities: N-Nitroso dimethyl amine (NDMA), N-Nitroso diethyl amine (NDEA), N-Nitroso Ethyl Iso propylamine (NEIPA), N-Nitroso-Nmethyl-4-aminobutyric acid (NMBA) N-Nitroso diisopropylamino (NDIPA) and N-Nitroso dibutyl amine (NDBA) with a LOQ of 0.004 ppm. Chromatographic separation is achieved using Zorbax SB C18 150 × 3.0 mm, 3.5 µ column with 0.1% formic acid in water as mobile phase A and 0.1% formic acid in methanol as mobile phase B at a flow rate of 0.3 ml/min using gradient mode of elution at a total run time of 18 minutes. Six nitrosamine impurities are successfully ionized and quantified in positive mode of atmospheric pressure chemical ionization (APCI) using multiple reaction monitoring (MRM). Method validation is performed as per ICH guidelines evaluating the limit of quantification and detection and found to give good S/N ratios with good linearity range of 0.002 -2 ppm with regression coefficient >0.99 for all the six nitrosamine impurities. Method recoveries are established using three-step sample preparation protocol and are found to be satisfactory within 80% -120%. The method can be used routinely applied for the detection of Nitrosamines in Telmisartan at a concentration of 1.5 ng/ml (0.03 ppm with respect to telmisartan concentration of 50 mg/ml).

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Tolmetin (TMT, CAS 26171-23-3) is a non-steroidal anti-inflammatory drug (NSAID) indicated for the relief of signs and symptoms of osteoarthritis, rheumatoid arthritis and juvenile rheumatoid arthritis. As TMT causes gastro-intestinal side effects like other NSAIDs, its nonacidic prodrug amtolmetin guacil (AMG, CAS 87344-06-7) was synthesized. AMG has similar NSAID properties like TMT with additional gastroprotective property. The aim of this study was to investigate whether TMT and AMG are differentially metabolised in rat and human plasma (fresh and acidified) and liver microsomes. TMT was found to be stable in all the matrices tested viz., rat and human plasma (fresh and acidified) and liver microsomes. AMG was found to be stable only in acidified rat and human plasma. On the contrary, in fresh human plasma and human liver microsomes AMG was rapidly converted to two metabolites, which were subsequently identified as MED5 and MED5 methyl ester, without yielding any intact TMT. However, in rat fresh plasma and liver microsomes, AMG formed MED5 (predominant) and TMT. To corroborate the in vitro findings, in vivo pharmacokinetics (PK) studies were done following separate dosing of AMG in both rats and humans. In rats, the PK data substantiated that following oral administration of AMG it will be converted to TMT resulting in similar PK parameters observed for TMT when it was administered alone. In humans, however, AMG yields low levels of TMT which substantes the in vitro results. Levels of AMG were not detectable in the plasma. These results confirm the species differences in the in vitro and in vivo metabolism and disposition of AMG. More research work to further explore and understand AMG metabolism in humans is required.

A simple, rapid, and highly sensitive LC-MS/MS method has been developed for the simultaneous and trace level quantification of underivatized boronic acids in lumacaftor active pharmaceutical ingredient. Chromatographic separation of boronic acids and lumacaftor achieved using Agilent Poroshell HPH C18 150 × 4.6 mm 2.7 µ column with 0.1% ammonia in water as mobile phase A and 100% acetonitrile as mobile phase B at a flow rate of 0.25 ml/min. Gradient elution was used with a total method run time of 14 minutes. Boronic acids were successfully ionized and quantified without derivatization using electrospray ionization in negative mode using tandem quadrupole mass spectrometry in multiple reactions monitoring mode. Method validation was performed as per ICH guidelines with good linearity over the concentration range of 0.05 ppm to 5 ppm of Lumacaftor test concentration for both the boronic acids with a correlation coefficient of >0.99. Recoveries were found good at different concentration levels and within the range of 80%-120%. The developed method can be successfully used for the routine quantification of boronic acids at a concentration level of 20 ng/ml (1 ppm with respect to 20 mg/ml lumacaftor).