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Putative Identification and Adduct Formation of Reference Materials for Cotton Secondary Metabolite using UPLC-ESI-MS Analysis

Li Shezeng1, 2, Niu Luxin2, Li Bochao3, Chen Xiuye2, Ma Ping2*, Ma Zhiying1*   

  1. 1. North China Key Laboratory for Crop Germplasm Resources of Education Ministry, Hebei Agricultural University, Baoding, Hebei 071001, China; 2. Integrated Pest Management Center of Hebei Province, Key Laboratory of IPM on Crops in Northern Region of North China, Ministry of Agriculture and Rural Affairs/Plant Protection Institute, Hebei Academy of Agricultural and Forestry Sciences, Baoding, Hebei 071000, China; 3. Department of Biomedical Engineering, University of Southern California, Los Angeles, CA 90098, USA
  • Received:2020-06-12 Revised:2020-07-23 Accepted:2020-07-23 Online:2020-07-27 Published:2020-07-27

Abstract: [Objective] The aim this study is to establish an identification method for cotton metabolites in ultra-performance liquid chromatography-electrospray ionization-mass spectrometry (UPLC-ESI-MS) analysis, and to investigate the adduct types, dominant adducts and appropriate ESI ion modes of cotton secondary metabolites under the determined UPLC and ESI conditions. [Method] UPLC-ESI-MS was employed to analyze 18 cotton metabolite standards, Online XCMS software was used to extract the nontargeted mass spectrum data, and MATLAB software was used to prepare calculation programs for the identification method of cotton metabolite standards. [Result] A high-throughput identification method of cotton secondary metabolite, named POSid and NEGid separately for the positive and negative ESI modes, was established based on the calculated accurate molecular weight. In the determined UPLC condition and positive and negative ESI mode, 14 cotton metabolite standards were correctly identified. There found 6 adducts including [M+H]+, [M+Na]+, [M+NH4]+, [2M+NH4]+, [2M+Na]+ and [2M+H]+ in positive ion mode, and 8 adducts including [M-H]-, [2M-H]-, [M+Cl]-, [M+FA-H]-, [3M-H]-, [M+Na-2H]-, [M-H2O-H]- and [M+TFA-H]- in negative ion mode, while 1 to 6 adducts were observed in the mass spectrum of a single standard, and each metabolite standard had a dominant adduct of a preference for ESI mode. Melibose was suitable for ESI positive ion mode detection, gossypol was suitable for both ion modes detection; and 12 compounds were suitable for negative ion mode detection due to mass spectrum signals of their dominant adduct stronger in negative ion mode than positive ion mode. [Conclusion] Based on the accurate molecular weight, the established identifying method can identify the 18 cotton metabolite standards with their nontarget mass spectrometry data. Under the determined UPLC and ESI condition, the dominant adducts of cotton secondary metabolites have the preference of ESI mode. These results provide technical and theoretical data support for further study of cotton metabolomics.

Key words: cotton metabolite, ultra-performance liquid chromatography-electrospray ionization-mass spectrometry (UPLC-ESI-MS), mass spectrum identification, adduct