New LC-(ESI)-HRMS Method for Determination of N-Nitrosodimethylamine in Metformin Demonstrates Trolox as a Potent Nitrosamine Inhibitor

摘要: Considering the high risk of nitrosamine impurities in pharmaceutical products, this pilot study was designed to investigate the efficiency of the selected antioxidants as inhibitors of nitroso dimethyl amine (NDMA) in metformin products. For this study, a new sensitive LC-HRMS method for NDMA analyses has been developed on the EC-C18 column with shorter elution time using a combination of 0.1% formic acid containing methanol and water, and mass spectrometric tuning as ESI positive/SIM mode. The developed method demonstrated the LOD and LOQ values of 0.3 pg/ml and 1.0 pg/ml, respectively. The linearity was established between 0.1 ng/ml to 15000 ng/ml with an r2 value of 0.9994. The intra-day and intra-day precision for the range between 50 and 10000 ng/ml were below 2% RSD whilst the recovery was between 97.3%-101.9%. For NDMA attenuation experiments, sodium nitrite was used as an NDMA inducer through nitrosation at the dimethyl amino group of metformin in both solution and solid phase studies. The antioxidants such as alpha-tocopherol, ascorbic acid, and Trolox were tested as NDMA attenuators in metformin pellets under vulnerable storage conditions. The developed method quantified the levels of NDMA in experimental samples, and the results were treated statistically. The solution (12 h) and solid phase studies (3 months) on NMDA generation by nitrosation at the dimethyl amino group of metformin revealed a positive correlation (r2= 0.9672) between sodium nitrite residue and NDMA formation (p< 0.001). Evaluation of antioxidants as NDMA inhibitors revealed the significant (p< 0.001) NDMA attenuation …