Abstract:
OBJECTIVE To analyze three enzymatic peptide components TD1 (LEEDLERSEERL), TD2 (EEKDKALQNAEGEVAAL) and TD3 (MILPTGASSF) in centipedes by UFLC-MS/MS method for comparing the differences of peptide components in different species of centipedes and identification of centipedes.
METHODS UFLC-MS/MS of the three peptide components was performed on an XSelect HSS T3 (4.6 mm×150 mm, 3.5 μm) column and the mobile phase was acetonitrile -0.1% formic acid solution with gradient elution. A triple quadrupole mass spectrometer detector with electrospray ionization (ESI) and multiple reaction monitoring (MRM) in positive ion mode were used for signal acquisition. The samples were subjected to orthogonal experimental methods to optimize the protein extraction method, and the final extraction method was determined by sonicating the protein lysate for 60 min to obtain the centipede protein extraction solution, which was enzymatic hydrolyzed and desalted before injecting. Three peptide components were determined in 20 batches of centipedes and 8 batches of other animal medicines and the results were compared.
RESULTS TD2 and TD3 were not detected in earthworm, stiff silkworm, leech, ground beetle and scorpion. TD1 and TD2 were more different in S. subspinipes mutilans L.Koch and S. subspinipes dehaani Brandt and S. mojiangica Zhang et chi, and TD3 was more different in S. subspinipes mutilans L.Koch and S. negrocapitis Zhang et Wang and S. subspinipes multidens Newport.
CONCLUSION This method can be used to distinguish the S. subspinipes mutilans L. Koch from those of earthworm, stiff silkworm, leech, ground beetle and scorpion. By comparing the differences of three peptides, Scolopendra subspinipes mutilans L.Koch can be distinguished with other four common non-pharmacopoeial species, providing a reference for quality evaluation and basic research of centipedes.