Chinese Journal of Chromatography ›› 2018, Vol. 36 ›› Issue (7): 615-620.DOI: 10.3724/SP.J.1123.2018.01026

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Methylamidation and normal phase chromatography with silica-hydride-based stationary phase for N-glycosylation characterization of monoclonal antibody drugs

WANG Yao1, LIANG Gaodao1, HAN Qing1, HU Xun1, ZHANG Qiwei2, HE Zhenyu1   

  1. 1. Wuhan Centers for Disease Prevention & Control, Wuhan 430015, China;
    2. Jianghan University, Wuhan 430056, China
  • Received:2018-01-16 Online:2018-07-08 Published:2014-06-03
  • Supported by:

    General Program of Health and Family Planning Commission of Hubei Province(No. WJ2017M198).

Abstract:

Monoclonal antibodies (mAbs) are efficacious therapeutic agents against various complex diseases. N-glycosylation characterization of mAbs is based primarily on fluorescence derivatization coupled with hydrophilic interaction liquid chromatography (HILIC). However, this method has some drawbacks. Presently, the N-glycosylation of mAbs was analyzed by methylamidation and silica-hydride-based normal phase chromatography (SiH-NPC). Samples were analyzed by liquid chromatography-mass spectrometry after enzymatic cleavage, methylamidation, and purification. SiH-NPC had some advantages compared to HILIC. It offered a novel separation mechanism that allowed for high resolution using salt-free mobile phases, avoiding contamination of the mass spectrometer. SiH-NPC may be applicable for rapid analysis. Its structure was more stable than that of HILIC, and it provided a long service life. In combination with the sialic acid derivatization, SiH-NPC presented a significant advantage in the analysis of sialylated glycans as well as isomeric oligosaccharides by liquid chromatography-mass spectrometry. The approach has potential applications in the biopharmaceutical industry.

Key words: N-glycans, liquid chromatography-mass spectrometry(LC-MS), methylamidation, monoclonal antibodies(mAbs), silica-hydride

CLC Number: