Clinical adverse reactions to ampicillin sodium are closely related to its impurities and degradation products. Several unknown degradation products have been detected in the degradation samples of ampicillin sodium. Therefore, a sensitive and accurate method is required to rapidly identify unknown degradation products.


Ampicillin sodium was subjected to forced degradation under hydrolytic (acidic and alkaline), hot, photolytic, and humid stress conditions. A combination of liquid chromatography/hybrid triple quadrupole linear ion trap mass spectrometry (LC/QqLIT-MS) and liquid chromatography/hybrid quadrupole time-of-flight mass spectrometry (LC/QqTOF-MS) was used to identify unknown degradation products. The analysis was achieved with gradient elution on an Agilent Zorbax SB-C18 column using ammonium acetate (1 mmol/L, pH 3.5) and acetonitrile.


A total of 19 degradation products and impurities, including five novel degradation products, were identified and characterized in the forced conditions. The novel degradation products were separately identified as (Z)-2-amino-N-((2-oxo-3-phenyl-2,3,6,7-tetrahydro-1H-1,4-diazepin-5-yl)methylene)-2-phenylacetamide (m/z 349), 2-(1-(2-amino-2-phenylacetamido)-2-((carboxy(phenyl)methyl)amino)-2-oxoethyl-5,5-dimethyl-4,5-dihydrothiazole-4-carboxylic acid (m/z 499), (E)-2-(((3,6-dioxo-5-phenyl-1,6-dihydropyrazin-2(3H)-ylidene)methyl)amino)-3-mercapto-3-methylbutanoic acid (m/z 348), 5-(amino(phenyl)methyl)-7-formyl-2,2-dimethyl-2,3-dihydroimidazo[5,1-b]thiazole-3-carboxylic acid (m/z 332), and 2-(1-(2-amino-2-phenylacetamido)-2-((2-(((4-carboxy-5,5-dimethylthiazolidin-2-yl)methyl)amino)-2-oxo-1-phenylethyl)amino)-2-oxoethyl)-5,5-dimethylthiazolidine-4-carboxylic acid (m/z 673).


LC/QqTOF-MS allowed us to obtain more accurate, richer information than LC/QqLIT-MS for the qualitative analysis of unknown compounds. Forced degradation studies could provide us with the data needed to understand the degradation pathways and intrinsic stability of drugs, and to simultaneously validate the feasibility of the analytical procedure. Copyright © 2014 John Wiley & Sons, Ltd.