Isolation and characterization of degradation products of moxidectin using LC, LTQ FT-MS, H/D exchange and NMR

Abstract

This study aimed to evaluate the degradation profile and pathways, and identify unknown impurities of moxidectin under stress conditions. During the experiments, moxidectin samples were stressed using acid, alkali, heat and oxidation, and chromatographic profiles were compared with known impurities given in European Pharmacopeia (EP) monograph. Moxidectin has shown good stability under heat, while reaction with alkali produced 2-epi and ∆2,3 isomers (impurities D and E in EP) by characteristic reactions of the oxahydrindene (hexahydrobenzofuran) portion of the macrocyclic lactone. Two new, previously unreported, unknown degradation products, i.e. impurity 1 and impurity 2, detected after acid hydrolysis of moxidectin (impurity 2 was also observed to a lesser extent after oxidation), were isolated from sample matrices and identified using liquid chromatography, NMR, high-resolution FT-ICR MS, and hydrogen/deuterium exchange studies. FTMS analysis showed accurate mass of molecular ion peaks for moxidectin at m/z 640.38412, impurity 1 at m/z 656.37952 and impurity 2 at m/z 611.35684, giving rise to daughter ions traceable up to the seventh levels of MS(n) experiments and supporting the proposed structures. Both unknown impurities along with moxidectin were fully characterized by (1)H, (13)C, 1D HMBC and 2D (NOESY, COSY and HSQC) NMR experiments. The interpretation of experimental data positively identified impurity 1 as 3,4-epoxy-moxidectin and impurity 2 as 23-keto-nemadectin. The identification of new impurities and correlation of their chromatographic profiles with the EP method is very useful to establish the stability profile of moxidectin and its preparations, as well as add value to the forthcoming moxidectin finished product European Pharmacopeia monographs.

Synthesis and Immunological Evaluation of Virus-Like Particle-Milbemycin A3/A4 Conjugates

“…The latter was treated with succinic anhydride, resulting in a selective formation of milbemycin hemisuccinate containing a four atom-long linker (milbemycin-L4, M-L4). The selected experimental conditions were sufficiently mild and the possible formation of known side products such as C(2)-epimer, Δ2,3-isomer [ 15 ] or aromatized species [ 16 ] was not observed. To create milbemycins with a longer linker, compound M–L4 was amidated with propargylic amine [ 17 ] and the resulting product reacted with triethylene glycol-derived azido-acid in copper(I)-catalyzed azide-alkyne cycloaddition reaction.…”

Structural characterization of product ions by electrospray ionization and quadrupole time‐of‐flight mass spectrometry to support regulatory analysis of veterinary drug residues in foods. Part 2: Benzimidazoles, nitromidazoles, phenothiazines, and mectins

“…Moxidectin loses the aliphatic side chain (‐C 7 H 12 O) from the protonated molecule thereby producing the ion at m/z 528 (Product Ion I, #6), as described by Awasthi et al . Further fragmentation led to the ion at m/z 498 (Product Ion II, # 6), and the ion at m/z 199 resulted from fragmentation of the macrocyclic lactone …”

“…In addition, the selected product ions obtained from the sodiated adduct of ivermectin (m/z 329, #5) are consistent with the aglycon allylic-lactone cleavage previously reported for ESI-MS/MS fragmentation of mectin adduct ions. [13] Moxidectin loses the aliphatic side chain (-C 7 H 12 O) from the protonated molecule thereby producing the ion at m/z 528 (Product Ion I, #6), as described by Awasthi et al [14] Further fragmentation led to the ion at m/z 498 (Product Ion II, # 6), and the ion at m/z 199 resulted from fragmentation of the macrocyclic lactone. [14] For emamectin and eprinomectin, the three selected product ions are from the saccharide portion of the molecule, observed at m/z 302, 158, and 126 ( Table 4 (#3-4) are in agreement with the product ions reported for mectins by Alberts-Shönberg et al [15] Saccharide product ions were also selected for avermectin (m/z 145 and 113, #1), doramectin (m/z 145 and 113, #2), ivermectin [16] (m/z 183, #5) and selamectin (m/z 145 and 113, #7).…”

“…Thus, few reports have described fragmentation pathways or proposed product ion structures for the compounds of interest, and we revise structures published for benzimidazoles, [7] nitroimidazoles, [8,9] phenothiazines, [10][11][12] and mectins. [13][14][15][16] High-resolution mass spectrometry (HRMS) is a valuable approach to the characterization of product ions for the screening and identification of pesticides, veterinary drugs, and other analytes. [17,18] To characterize the product ions of interest, it is necessary to calculate ion formulae from the experimental data collected, and hence HRMS is useful.…”

Novel degradation products of argatroban: Isolation, synthesis and extensive characterization using NMR and LC-PDA-MS/Q-TOF

“…ICH Q3A(R2) and Q3B(R2) recommend the characterization of impurities/degradation products that are present at a level greater than the identification threshold in a drug substance or drug product [3] , [4] , [5] , [6] , [7] , [8] , [9] . The study of formation of impurities/degradants in the drug substance, their isolation and characterization is very important since it can help to understand the degradation pattern of the drug substance.…”