The application of Orbitrap high resolution mass spectrometry in the development and quality control of biopharmaceuticals

From early research and development, characterization analysis, process optimization to CMC and QC analysis, there are complexities and uncertainties in each link of biopharmaceutical products The thermo fly high-resolution mass spectrometry biopharmaceutical solution provides a stable and accurate analysis method to guarantee your drug declaration and listing 1 Protein characterization analysis 01 molecular weight determination of primary structure analysis based on the molecular weight measured by Orbitrap ultra-high resolution mass spectrometry is closer to the actual value, and can accurately identify components with similar quality [1,2], so as to characterize protein with more strict standards The amino acid sequence of biological drugs is the material basis of their biological activity and clinical efficacy The first condition is to ensure that the sequence is consistent with the expectation The post-translational modification of amino acids may affect their affinity and biological activity The identification of the type and location of the modification can be used as an important index for CQA evaluation of biological drugs Figure 1 Orbitrap can obtain more accurate first-order mass and second-order abundant fragment ions of polypeptide Through Orbitrap ultra-high resolution mass spectrometer, it can obtain more accurate first-order mass and abundant second-order fragment information of polypeptide, which provides strong support for amino acid and post translation modification characterization of biological drugs The glycosylation has an important effect on the efficacy, stability and immunogenicity of biological drugs In the face of oligosaccharides with extremely complex structure, semefei has assembled superior products to solve this problem Scheme 1: complete solution for free sugar LC-MS analysis: Classic LC-MS analysis process, release sugar chain, mark sugar chain, quantify sugar chain by fluorescence, and identify sugar chain structure by high-resolution mass spectrometry Scheme II: complete solution of free sugar ic-ms analysis for sialic acid protein with high content, the labeling process of 2Ab will lead to desialization [3], while the scheme of combination of ion chromatography and mass spectrometry of semefei's superior products does not need sugar derivatization The complex sample pretreatment process can realize the direct quantitative and qualitative analysis of undeveloped sugars and reduce the degradation of sialic acid in the labeling process Hdx-ms is a high-level structure analysis method Hdx-ms has a low requirement for samples It does not need crystallization and solution state It has a high tolerance for interfering substances such as buffer salts and excipients It can ensure the determination of proteins or protein complexes in natural non denatured state It can realize the determination of proteins or protein complexes from Detailed information is obtained at the level of integrity, local level and single amino acid level, which makes the information of the advanced structure of Zui terminal protein more comprehensive and accurate Figure 2 Complete solution of hydrogen deuterium exchange mass spectrometry II The presence of HCPs in host cells of biological drugs mainly affects the safety and effectiveness of drugs Based on the high sensitivity and wide dynamic range of Orbitrap, high-throughput qualitative and quantitative analysis of unknown HCPs can be realized Compared with the ELISA method, the development cycle is shorter, both qualitative and quantitative Figure 3 shows the complete solution for very low HCPs Fig 3 Accurate qualitative and quantitative analysis of HCPs with low content based on LC-MS / Ms 3 Bioanalysis of biopharmaceuticals For macromolecular biopharmaceuticals, the quantitative method based on mass spectrometry has the advantages of simultaneous qualitative and quantitative, high specificity and short development cycle compared with the ligand binding assays method Compared with the triple quadrupole mass spectrometry, it has the advantages of Besides the comparable quantitative ability, it can monitor the advantages of biotransformation in vivo The complete solution for the quantification of complete protein level and peptide level of biological drugs is shown in Figure 4, and its quantitative limit and linear range are far higher than the actual demand Figure 4 A complete solution for bioanalysis of recombinant biopharmaceuticals based on LC-MS and LC-MS / Ms 4 Multi quality attribute method workflow mam 1 workflow: HR mam is a workflow with high resolution and high quality accuracy, which realizes the comprehensive characterization and monitoring of product quality attributes from R & D to quality control (QC) 2 software: including 2 software: biopharma finder finds out key quality attributes, chromeleon realizes instrument control, data acquisition and analysis, and has strong compliance Three elements: three elements: quality attribute representation; quality attribute monitoring and unknown component detection Zui finally realizes a set of scheme which can realize the detection results of many traditional analysis methods, and really simplify the complexity Figure 5 A complete solution of multiple key quality attributes (hr-mam) based on Orbitrap ultra-high resolution mass spectrometry At present, an international innovative biopharmaceutical giant has used HR mam in the quality control stage [4], and Zui, the US regulatory agency, recently published an evaluation of MAM method on AC [5], greatly promoting the process of high resolution mass spectrometry used in the practical application of biopharmaceuticals in production quality control This paper summarizes the complete solution of biopharmaceutical based on Orbitrap ultra-high resolution mass spectrometer, which is efficient, fast and accurate, from drug characterization to production process, Wei helps the new drug market Reference materials: [1] He j, Su D, ng C, et al High resolution accurate mass specification enabling in depth characterization of in vivo biotransformations for impact antibody drug associations [J] Analytical chemistry, 2017, 89 (10): 5476-5483 [2] Schachner L, Han g, Dillon m, et al Characterization of chain pairing variants of bispecific IgG expressed in a single host cell by high-resolution native and denaturing mass spectrometry[J] Analytical chemistry, 2016, 88(24): 12122-12127 [3] Szabo Z, Thayer J R, Reusch D, et al High performance anion exchange and hydrophilic interaction liquid chromatography approaches for comprehensive mass spectrometry-based characterization of the N-glycome of a recombinant human erythropoietin[J] Journal of proteome research, 2018, 17(4): 1559-1574 [4] Rogers R S, Nightlinger N S, Livingston B, et al Development of a quantitative mass spectrometry multi-attribute method for characterization, quality control testing and disposition of biologics[C]//MAbs Taylor & Francis, 2015, 7(5): 881-890 [5] Rogstad S, Yan H, Wang X, et al Multi-Attribute Method for Quality Control of Therapeutic Proteins[J] Analytical Chemistry, 2019, 91 (22): 14170-14177 Scan the QR code below to obtain the whole industry solution of Symantec, or pay attention to the public Hao of "Symantec chromatography and mass spectrometry China" for more information+