Product Number: P109004
English Name: Pretomanid Impurity 4
English Alias: (R)-2-nitro-6-((3-(trifluoromethoxy)benzyl)oxy)-6,7-dihydro-5H-imidazo[2,1-b][1,3]oxazine
CAS Number: 2222639-62-3
Molecular Formula: C₁₄H₁₂F₃N₃O₅
Molecular Weight: 359.26
Product Advantages：
High purity and chiral confirmation：HPLC purity ≥99.0%, with chiral configuration confirmed by chiral HPLC (optical purity ≥99.5%) and circular dichroism (CD), meeting ICH Q6A requirements for chiral impurity reference standards.
Excellent stability：Stable for 36 months when stored at -20°C in the dark, with a degradation rate <0.8% after 14 days at room temperature in solution (e.g., methanol-acetonitrile system), suitable for long-term quality monitoring and complex experimental conditions.
Dual isomer specificity (regio- and chiral)：Clearly defined as a dual isomer impurity with (R)-configuration and a 3-position benzyloxy group, enabling precise tracking of risks from failed chiral induction and insufficient regioselectivity in Pretomanid synthesis.
Applications：
Chiral and regioisomeric impurity detection：Used for chiral LC-MS/MS detection of Impurity 4 in Pretomanid APIs, controlling its content ≤0.1% in accordance with ICH Q3A standards to ensure compliance with regulatory requirements for both chiral and regioisomeric impurities.
Multi-dimensional synthesis process optimization：In benzylation and chiral cyclization reactions, monitoring impurity content (e.g., reducing impurity from 1.3% to 0.1% when changing the chiral catalyst to (S)-binaphthol) synchronously optimizes regioselectivity and chiral induction efficiency.
Comprehensive analytical method validation：Serves as a dual isomer impurity reference standard for developing multi-dimensional detection methods, such as 2D chiral HPLC (first dimension separates chiral isomers, second dimension separates regioisomers), enabling precise quantification of complex impurities.
Toxicological difference assessment：Provides samples for evaluating the toxicity of chiral-regioisomeric synergy, facilitating in vitro receptor binding assays and in vivo teratogenicity tests to meet FDA requirements for in-depth stereoisomer safety assessment.
Background Description：
Pretomanid Impurity 4 is a dual isomer impurity introduced during Pretomanid synthesis due to insufficient stereoselectivity in chiral cyclization reactions and regiisomerization in benzylation. Its (R)-configuration is the enantiomer of the parent drug's (S)-configuration, and the 3-position benzyloxy group further alters the molecular conformation, potentially significantly affecting the drug's biological activity and toxicity. According to ICH M7(R1) and Q6A guidelines, such dual isomer impurities require independent quality control and toxicological evaluation. Current industry standards set the individual impurity limit at ≤0.1%.
Research Status：
Breakthrough in detection technology：Chiral LC-MS/MS combined with a tandem cyclodextrin chiral column (e.g., Chiralpak IA) and C18 analytical column uses multi-step gradient elution to achieve simultaneous separation of chiral and regioisomeric impurities, with a detection limit (LOD) as low as 0.002ppm, providing a new method for trace dual isomer impurity analysis.
Formation mechanism analysis：Impurity formation is closely related to the enantioselectivity of chiral catalysts (e.g., increased impurity formation with (R)-proline derivatives) and the steric hindrance of benzylation reagents (larger steric hindrance of 3-substituted benzyl chloride leading to reduced regioselectivity). Introducing chiral Lewis acid catalysts (e.g., Ti(OiPr)₄ complex with (S)-tartrate) and microwave-assisted reactions (100W, 40°C) can reduce impurity formation by over 95%.
Safety evaluation progress：In vitro cytotoxicity tests show that the impurity has an IC₅₀ three times higher than the parent drug against THP-1 macrophages, but in vivo embryo-fetal development toxicity tests (rats) showed no teratogenicity at doses ≤200mg/kg. Combined risk-benefit analysis suggests a limit of ≤0.1%.
China
