Mobile phase solvents and reagents for LC-MS biopharmaceutical applications

Published: 26-Oct-2022

Fisher Chemical Optima grade LC-MS solvents are high purity and meet all the needs for LC-MS analysis

Biopharmaceutical drugs are a unique class of therapeutic proteins that have far more complex structures than small molecule drugs. Liquid chromatography (LC) coupled with mass spectrometry (MS), LC-MS, has become increasingly popular for characterising biopharmaceutical drugs. LC-MS is widely used for characterising the primary structure of protein drugs such as amino acid sequencing for peptides/proteins, profiling of process-related impurities such as host cell proteins (HCPs), disulfide bond mapping, N-glycan profiling of monoclonal antibodies (mAbs), and nucleotide sequencing for oligonucleotides. 

At the primary structure level, the protein post-translational modifications (PTMs) that arise at different stages of manufacturing and storage can impact drug efficacy and safety; thus, PTMs are classified as critical quality attributes (CQA) that should be monitored and controlled. Combining the power of LC separation with the sensitivity and specificity of MS detection, LC-MS is the tool of choice for peptide mapping to detect multiple PTMs in a single analytical run, i.e., the multi-attribute method (MAM) workflow.

A critical component of the MAM workflow is to separate peptides of various lengths from the digested therapeutic protein. This is typically achieved by employing a long LC gradient of water and acetonitrile with formic acid added as a modifier. Mobile phase solvents and reagents must be of high purity with low organic impurities to wnsure no interference with detection by electrospray (ESI) MS. 

Thermo Fisher Scientific offers two grades of LC-MS solvents. Fisher Chemical Optima grade LC-MS solvents are high purity and meet all the needs for LC-MS analysis. Pre-made LC-MS blends are also available in Fisher Chemical Optima grade and are precisely mixed with lot-to-lot consistency and tested by LC-MS, making these blends convenient for use in biopharma QC labs. The Thermo Scientific UHPLC-MS grade solvents are a higher grade with more stringent specifications to meet the most sensitivity-demanding analyses. Both LC-MS and UHPLC-MS grades of solvents and blends have been fully tested to ensure low organic impurities to minimise interference and ion suppression. 

Another important quality attribute of our high-purity LC-MS regents is the extremely low levels of metal impurities. Trace metal impurities can cause poor chromatography peak shape, peak tailing, and diminished recovery via the metal-ion mediated adsorption on the LC column media. The alkali metal ions such as sodium (Na+) and potassium (K+) are electrostatically attracted to the negatively charged carboxyl group of the C-terminal amino acid in peptides/proteins and the polyanionic backbone of oligonucleotides. The Na+ and K+ adducts decrease the overall detection sensitivity and complicate the peptide identification in LC-MS analysis. Minimising the metal contamination in mobile phase solvents and regents has become ever more critical with the growing demand for higher LC-MS sensitivity. 

All Thermo Fisher Scientific LC-MS grade solvents and reagents are subject to strict control of trace metal contamination during the production process and thoroughly tested to stringent specifications with ICP-MS. A comparison of the four most common alkali and alkaline contaminant metal impurities in pre-blended LC-MS grade 0.1% formic acid in water from three competitors are listed along with the Fisher Chemical Optima offering in Table 1. As shown, the Fisher Chemical Optima pre-blended mobile phase has the lowest allowable overall metal ions present. 

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