Integrated LC-MS/MS and Proteomics Analysis Reveals the Chemical and Proteinaceous Composition of Andrographolide Sulfonate

Objective This study aimed to enhance the safety profile of Andrographolide Sulfonate Injection (ASI) by characterizing these impurities and supporting the refinement of quality control standards for andrographolide sulfonate (AS). Methods We employed a combination of UHPLC-QE-Orbitrap-MS for the tentative identification of related compounds and a proteomic approach for residual protein analysis. The protein analysis workflow involved extraction, quantification, tryptic digestion, desalting, and nanoLC-MS/MS analysis. The resulting MS data were searched against an Andrographis paniculata protein database using Peaks Studio software. Results Our analyses led to the putative identification of 27 related compounds. Furthermore, 12 residual proteins were newly identified, among which the protein A0A4Y5UXR9_ANDPA was found to be present at a significantly higher level than the others. Conclusions The findings provide critical data on the composition of AS, which is essential for advancing the quality control and ensuring the clinical safety of ASI. This study also offers a scientific basis for evaluating the necessity of ultrafiltration in its manufacturing process.