Charagondla Krishnaiah, Mukherjee Partha S, Ginjupalli Kalyani, Hu Bing, Kaur Simrat, Thanki Paresh, Tailor Tejas, Prajapati Bhavin, Ramdas Saroj
Amicus Therapeutics Inc, 47 Hulfish Street, Princeton, NJ, 08542, USA.
SGS Health Science North America-Canada, Mississauga, Canada.
Pharm Res. 2025 Jun 17. doi: 10.1007/s11095-025-03875-7.
This study aims to develop and validate a highly sensitive LC-MS/MS method for quantifying eight nitrosamine (NA) impurities NDMA, NMBA, NDEA, NEIPA, NDIPA, NMPA, NDPA, and NDBA in a small molecule commercial Active Pharmaceutical Ingredients (API) and Drug Product (DP) capsules used for the treatment of Fabry rare disease in global patients.
The method utilized gradient separation on a C18 column, ensuring optimum resolution between NAs and internal standards. Elution was monitored at precursor and product ions. The method was validated according to ICH Q2 (R1) guidelines, assessing specificity, linearity, recovery, and repeatability in both API and capsule matrices.
Quantification of NAs ranged from 0.0215 to 0.780 ppm in relation to a 20.5 mg/mL sample concentration. Linearity coefficients ranged from 0.99 to 1.00. The Detection Limit (DL) was 0.154-0.560 ng/mL, and the Quantitation Limit (QL) was 0.438-1.590 ng/mL. Mean recoveries for all NAs were between 90 and 107%, and repeatability was under 15%. An Analytical Target Profile (ATP) concept was successfully employed to monitor Analytical Method Performance Characteristics (AMPC).
The validated LC-MS/MS method proved effective for quantifying NAs (NDMA, NMBA, NDEA, NEIPA, NDIPA, NMPA, NDPA, and NDBA) in both API and capsule matrices, with results consistently below the Quantitation Limit (QL) in confirmatory testing. This approach supports the elimination of routine NA testing, adhering to FDA and EMA's conservative Acceptable Intake (AI) limits, ensuring safety and regulatory compliance for global patients.
本研究旨在开发并验证一种高灵敏度的液相色谱-串联质谱（LC-MS/MS）方法，用于定量分析用于治疗全球法布里罕见病患者的小分子商业活性药物成分（API）和药品（DP）胶囊中的八种亚硝胺（NA）杂质，即N-亚硝基二甲胺（NDMA）、N-亚硝基甲基苄胺（NMBA）、N-亚硝基二乙胺（NDEA）、N-亚硝基乙基异丙基胺（NEIPA）、N-亚硝基二异丙基胺（NDIPA）、N-亚硝基甲基丙基胺（NMPA）、N-亚硝基二丙基胺（NDPA）和N-亚硝基二丁基胺（NDBA）。
该方法采用C18柱进行梯度分离，确保了NA与内标之间的最佳分离度。在前体离子和产物离子处监测洗脱情况。该方法根据国际人用药品注册技术协调会（ICH）Q2（R1）指南进行验证，评估了API和胶囊基质中的特异性、线性、回收率和重复性。
在20.5 mg/mL的样品浓度下，NA的定量范围为0.0215至0.780 ppm。线性系数范围为0.99至1.00。检测限（DL）为0.154 - 0.560 ng/mL，定量限（QL）为0.438 - 1.590 ng/mL。所有NA的平均回收率在90%至107%之间，重复性低于15%。成功采用分析目标轮廓（ATP）概念来监测分析方法性能特征（AMPC）。
经过验证的LC-MS/MS方法被证明可有效定量分析API和胶囊基质中的NA（NDMA、NMBA、NDEA、NEIPA、NDIPA、NMPA、NDPA和NDBA），在确证性测试中结果始终低于定量限（QL）。这种方法支持取消常规的NA检测，符合美国食品药品监督管理局（FDA）和欧洲药品管理局（EMA）保守的可接受摄入量（AI）限值，确保全球患者的安全和法规合规性。
