Objective: To analyze the primary structure of human interferon α2b related proteins by using human interferon α2b related proteins detection method combined with mass spectrometry, and to identify related proteins with different retention times. Methods: Agilent ZORBAX C18(250 mm×4.6 mm, 300, 5 μm) column was used and mobile phase A was 30% acetonitrile(0.2% TFA), mobile phase B was 80% acetonitrile(0.2% TFA). The flow rate was 0.5 mL·min-1, column temperature was room temperature. Electrospray positive ion mode with MS mode scaning was used. Capillary voltage was 2 500 V, cone voltage was 40 V, desolvation temperature was 350 ℃, source temperature was 120 ℃, desolvation gas flow was 800 L·h-1, scan scope was 400-4 000 m/z, collision voltage was 40 V and MS acquisition time was 20-60 min. Results: The molecular weight of the related proteins was consistent with the theoretical relative molecular mass. Peak 3 was the main peak, which relative molecular mass were 19 264.81 and 19 395.93. This peak was human interferon α2b and N-terminal methionine human interferon α2b. The peak 1 with relative molecular mass of 19 280.72, was oxidized human interferon α2b; and the peak 2 with relative molecular mass of 19 412.25 was (oxidation+N-terminal methioninylation) human interferon α2b. Peak 4 was human interferon α2b with relative molecular mass at 19 265.13. Peak 5 was (oxidation+N-terminal acetylation) human interferon α2b with the relative molecular mass of 19 322.81. Peak 6 was N-terminal acetylation of human interferon α2b with the relative molecular mass of 19 306.79. Conclusions: The main peak are human interferon α2b and N-terminal methionine human interferon α2b and the related proteins with retention time in front of the main peak were identified as oxidized and (oxidized+N-terminal methioninylation) modified human interferon α2b. The related proteins with retention time behind the main peak were (oxidation+N-terminal acetylation) and N-acetylated modified human interferon α2b.
ZHU Qiu-mei, LIU Yu-lin, YU Lu, WANG Jiang-lin, LIANG Guan-qiao, SUN Rui-xin, LIU Lin-lin, LIU Jing-hui
. Analysis the primary structure of human interferon α2b related proteins based on 2020《Pharmacopoeia of the People’s Republic of China》 related proteins detection method combined mass spectrometry[J]. Chinese Journal of Pharmaceutical Analysis, 2022
, 42(6)
: 1096
-1100
.
DOI: 10.16155/j.0254-1793.2022.06.23
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