Development and Validation of an HPLC-RI Method for the Simultaneous Determination of DPPC, Palmitic Acid, and Cholesterol in Bovine Pulmonary Surfactant

This study aimed to develop and validate an HPLC-RI method for the simultaneous determination of dipalmitoylphosphatidylcholine (DPPC), palmitic acid, and cholesterol in bovine pulmonary surfactant, a critical treatment for infants suffering from Respiratory Distress Syndrome (RDS). The Iranian surfactant, Beraksurf, was used as the sample. A simple extraction procedure involving methanol dissolution and protein precipitation was employed, followed by HPLC analysis. The method underwent comprehensive validation for various parameters, including system suitability, repeatability (intra-assay precision), intermediate precision, linearity, accuracy, limit of detection (LOD), limit of quantification (LOQ), and robustness. The results demonstrated excellent system suitability. Repeatability and intermediate precision were confirmed with low %RSD values, while linearity was established with correlation coefficients (R²) exceeding 0.99 for all analytes. The method exhibited high accuracy, with recoveries ranging from 97% to 100%, and low LOD and LOQ values, indicating its sensitivity. Robustness testing revealed that minor variations in method parameters did not significantly impact results. This validated RP-HPLC-RID method provides a reliable and efficient approach for the analysis of key surfactant components, contributing to improved quality control in surfactant preparations and enhancing therapeutic strategies for managing RDS in infants.