Document Type : Original Article


Micro Labs Ltd., API R&D Centre, Bommasandra Jigani Link Road, KIADB INDL Area, Bommasandra, Bangalore-560105, Karnataka, India.


A new LC-MS based method was developed to detect three potential genotoxic impurities namely, N',N'''-(4,6-dichloropyrimidine-2,5-diyl)bis[N,N-dimethyl(imidoformamide)]} (Impurity-I), {N-(2-amino-4-chloro-6-{[(1S,2R)-2-(hydroxymethyl)cyclopent-3-en-1-yl]amino}pyrimidin-5-yl)formamide} (Impurity-II) and {(1R,5S)-5-[(2,5-diamino-6-chloropyrimidin-4-yl)amino]cyclopent-2-en-1-yl}methanol (Impurity-III), at low level in Abacavir sulfate, a nucleoside reverse transcriptase inhibitor. It utilizes Zorbax phenyl hexyl column (150 mm x 4.6 mm, 3.5 μm) with electrospray ionization in selected ion monitoring (SIM) mode for quantitation of these PGIs. The method is able to quantify Impurity-I at 0.74 ppm, Impurity-II and Impurity-III at 0.73 ppm with respect to 5.0 mg/mL of Abacavir sulfate. The method was found to be linear in the range of LOQ to 150% of Toxicological Threshold Concentration level of 2.5 ppm. The correlation coefficients of PGI’s obtained were >0.999 in each case. The method accuracy of these PGI’s is in the range between 88.7-115.0%. The method is sensitive, specific, linear, accurate, precise and meets the criteria of validation as per International Conference on Harmonization.

Graphical Abstract

A Simple and Sensitive Method for the Ultra Trace Determination of Potential Genotoxic Impurities in Abacavir Sulfate by LC-MS


Main Subjects

Raman N.V.V.S.S., Prasad A.V.S.S., Reddy K.R. J. Pharm. Biomed. Anal., 2011, 55:662
Robinson D.I. Org. Process Res. Dev., 2010, 14:946
Guideline on the limits of genotoxic impurities, EMA guidance, EMEA/CHMP/QWP/251344/2006.
Guideline for Assessment and Control of DNA Reactive (Mutagenic) Impurities in
Pharmaceuticals to Limit Potential Carcinogenic Risk, M7, ICH, 2014.
Gallant J.E. J. Clin. Virol., 2002, 25:317
Moyle G., Boffito M., Fletcher C., Higgs C., Hay P.E.,  Song I.H., Lou Y., Yuen G.J., Min S.S., Guerini E.M. Antimicrob. Agents. Chem., 2009, 53:1532
Muller L., Mauthe R.J., Riley C.M., Andino M.M., De antonis D., Beels C., De George J., De knaep A.G.M., Ellison D., Fagerland J.A., Frank R., Fritschel B., Galloway Sh., Harpur E., Humfrey C.D.N., Jacks A.S., Jagota N., Mackinnon J., Mohan G., Ness D.K., O’Donovan M.R., Smith M.D., Vudathala G., Yotti L. Regul. Toxicol. Pharmacol., 2006, 44:198
Ashby J., Tennant R.W. Mutat. Res., 1988, 204:17
Reddy A.V.B., Jaafar J., Umar K., Majid Z.A., Aris A.B., Talib J., Madhavi G. J. Sep. Sci. 2015, 38:764
Prakash A., Teotia A.K., Farooqi J.A., Singh G.N. Ind. J. Chem., 2016, 55(B):213
Seshachalam U., Haribabu B., Chandrasekhar K.B. J. Sep. Sci., 2007, 30:28
Kurmi M., Sahu A., Singh S. J. Pharm. Biomed. Anal., 2017, 134:372
ICH Q2 (R1), Validation of Analytical Procedures: Text and Methodology, 2005.