Maganti Sivaji, Duvvuri Suryakala, Pericharla Prudhvi Raju, Bellam Ravi Kumar, Shaik Ameer Khan
Department of Chemistry, GITAM Deemed to be University, Visakhapatnam, Andhra Pradesh 530045, India; Eugia Pharma Research Centre, A division of Eugia Pharma Specialities Limited, Pashamylaram Sangareddy (Dist.), Telangana 502 307, India.
Department of Chemistry, GITAM Deemed to be University, Visakhapatnam, Andhra Pradesh 530045, India.
Ann Pharm Fr. 2025 Sep;83(5):890-906. doi: 10.1016/j.pharma.2025.03.007. Epub 2025 Apr 1.
In the present study, we developed and validated a sensitive, accurate, robust and simple RP-HPLC method for quantification of related substances of vasopressin (VPS) injection. The development phase prioritized the optimization of wavelength, mobile phase composition and column selection to enhance separation and sensitivity for the analytical evaluation of VPS injection. By considering peak symmetry, resolution and retention time, chromatographic conditions of VPS injection were established. The YMC PACK ODS AM (100×4.6) mm, 3μm was employed in the study. Mobile phase A was prepared with 0.113M NaHPO.HO (pH 3.0) while mobile phase B was a 50:50 (v/v) ratio of acetonitrile and water, pumped through a flow rate of 1.0mL/min. VPS injection was exposed to thermal, photolytic, acid, base and peroxide degradation conditions and these were analysed by the current method. The method was validated following the guidelines set by the International Council for Harmonization guideline (ICH). The linearity studies demonstrating that a correlation coefficient value is more than 0.999 for VPS and its related substances. The detection and quantification limits for all related substances were found to be 0.01% and 0.05%, respectively. All the impurities exhibited consistent recoveries ranging from 85 to 105%. The unknown impurities were eluting at RRT 1.23 and RRT 1.25 in the RS methodology (HPLC-UV) of VPS injection are identified and assessed the impurity levels. Furthermore, aggregate profiles and secondary structure analysis studies were carried out using FTIR and LC-HRMS, comparing the VPS formulation samples with the reference listed drug (RLD) (Vasostrict) samples across various diluents.
在本研究中，我们开发并验证了一种灵敏、准确、稳健且简便的反相高效液相色谱法（RP-HPLC），用于定量分析血管加压素（VPS）注射液的有关物质。方法开发阶段着重优化波长、流动相组成和色谱柱选择，以提高分离度和灵敏度，用于VPS注射液的分析评估。通过考虑峰对称性、分离度和保留时间，确定了VPS注射液的色谱条件。本研究采用YMC PACK ODS AM（100×4.6）mm，3μm色谱柱。流动相A由含有0.113M NaHPO·H₂O（pH 3.0）的溶液配制而成，流动相B为乙腈和水按50:50（v/v）比例混合而成，流速为1.0mL/min。将VPS注射液置于热、光解、酸、碱和过氧化物降解条件下，并用本方法进行分析。该方法按照国际协调会议（ICH）指南进行验证。线性研究表明，VPS及其有关物质的相关系数值大于0.999。所有有关物质的检测限和定量限分别为0.01%和0.05%。所有杂质的回收率均在85%至105%之间。在VPS注射液的对照品溶液（HPLC-UV）方法中，未知杂质在相对保留时间（RRT）1.23和RRT （此处原文有误，推测可能是1.25）处洗脱，对其进行了鉴定并评估了杂质水平。此外，使用傅里叶变换红外光谱（FTIR）和液相色谱-高分辨质谱（LC-HRMS）进行了聚集体图谱和二级结构分析研究，将VPS制剂样品与各稀释剂中的参比制剂（RLD）（Vasostrict）样品进行了比较。
