Comprehensive analysis of vitamin D3 impurities in oily drug products using supercritical fluid chromatography–mass spectrometry

Abstract

AbstractVitamin D3, an essential micronutrient, often requires supplementation via medicines or food supplements, which necessitate quality control (QC). This study presents the development of a method for detecting and quantifying seven impurities of vitamin D3 in oily drug products using supercritical fluid chromatography–mass spectrometry (SFC‐MS). Targeted impurities include two esters of vitamin D3 and five non‐esters including four that are isobaric to vitamin D3. Firstly, a screening study highlighted the Torus 1‐AA column and acetonitrile modifier as adequate for the separation, followed by optimization of the SFC conditions. Secondly, make‐up solvent composition and MS settings were optimized to reach high sensitivity. For both the separation and MS response, the screening design of experiments proved useful. Lastly, a fast saponification and liquid–liquid extraction method was developed, enabling efficient sample cleanup and impurities recovery from the complex oily matrix. The SFC‐MS method suitability was assessed in two validation studies. The first study employed the ICH Q2 guideline for impurity limit test to demonstrate method specificity and establish a limit of detection (LOD) and a limit of quantification (LOQ) at 0.2% and 0.5%, respectively, for ester impurities. The second study conducted a comprehensive quantitative assessment for three non‐ester impurities using a total error approach, determining method validity through accuracy profiles. The validated method exhibited reliable performance across impurity concentrations from 0.1% to 2.0%, with estimated LODs ranging from 2 to 7 ng/mL. This study further promotes SFC‐MS as a valuable, versatile, and green tool for routine pharmaceutical QC.