ABSTRACT
The aim of the present investigation was to demonstrate an approach involving use of liquid chromatography (LC) and liquid chromatography-mass spectrometry (LC–MS) to separate, identify and characterize very small quantities of degradation products (DPs) of acebutolol without their isolation from the reaction mixtures. The drug was subjected to oxidative, hydrolytic, thermal and photolytic stress conditions as per International Conference on Harmonization (ICH) guideline Q1A(R2). Among all the stress conditions the drug was found to be labile in hydrolytic (acidic & basic) and photolytic stress conditions, while it was stable in water-induced hydrolysis, oxidative and thermal stress conditions. A total of four degradation products were formed. A C18 column was employed for the separation of all the DPs on a gradient mode by using high-performance liquid chromatography (HPLC). All the DPs were characterized with the help of their fragmentation pattern and the masses obtained upon LC–MS/MS and MSn analysis. All the hitherto unknown degradation products were identified as 1-(2-(2-hydroxy-3- (isopropylamino)propoxy)-5-(amino)phenyl)ethanone (DP-I), N-(4-(2-hydroxy-3-(isopropylamino) propoxy)-3-acetylphenyl)acrylamide (DP-II), 1-(2-(2-hydroxy-3-(isopropylamino)propoxy)-5-(hydroxymethylamino) phenyl)ethanone (DP-III) and 1-(6-(2-hydroxy-3-(isopropylamino)propoxy)-2,3-dihydro- 2-propylbenzo[d]oxazol-5-yl)ethanone (DP-IV). Finally the in-silico carcinogenicity and hepatotoxicity predictions of the drug and all the DPs were performed by using toxicity prediction softwares viz., TOPKAT, LAZAR and Discovery Studio ADMET. The results of in-silico toxicity studies revealed that acebutolol (0.967) and DP-I (0.986) were found to be carcinogenic, while acebutolol (0.490) and DP-IV (0.437) were found to be hepatotoxic.
ABSTRACT
Zidvovudine (AZT) is a nucleoside analogue reverse transcriptase inhibitor (NRTI), a class of anti-retroviral drug. A stability-indicating assay method for AZT was developed in line with ICH guideline. Successful separation of AZT and its degradation products was achieved by gradient elution mode on reverse phase C18 column using 10 mM ammonium acetate: acetonitrile as the mobile phase at 0.8 mL/min flow rate, 25 μL injection volume, 30 °C column temperature and 285 nm detection wavelength. Two major acid degradation products were identified and characterized by liquid chromatography–electrospray ionization mass spectro-metry (LC–ESI/MS/MS) and accurate mass measurements. The probable mechanisms for the formation of degradation products were identified based on a comparison of the fragmentation pattern of the [M + H] + ions of AZT and its degradation products. One of the degradation products, DP-1, was isolated by semi-preparative high performance liquid chromatography (HPLC) using Waters XBridge Prep C18 (250 mm×10 mm, 5 μm). Degradation products showed higher toxicity compared to the drug in some models assessed by TOPKAT software. The method validation was performed with respect to robustness, specificity, linearity, precision and accuracy as per ICH guideline Q2 (R1).