Rapid Determination of Endogenous 20-Hydroxyecdysone in Plants on MALDI-TOF/TOF Mass Spectrometry via Chemical Labeling Based on Boronate Affinity

20-Hydroxyecdysone(20E)derived from plants has a wide range of physiological and pharmacological ef ects on animals and humans,and rapid and sensitive methods for screening of the endogenous 20E in plants are thus required.Herein,a matrixassisted laser desorption/ionization time-of-f ight tandem mass spectrometry(MALDI-TOF/TOF MS)method is described for rapid and sensitive determination of endogenous 20E in plants.It is based on the use of the(3-(acridin-9-ylamino)phenyl)boronic acid(AYPBA)as the mass tag to assist the MS and tandem MS(MS^(n))analysis of 20E on MALDI-TOF/TOF MS.Good linearity was obtained with a determination coef cient(R^(2))larger than 0.99 in the range of 0.025–2.5μΜ.The limit of detection(LOD)was 2.4 fmol.Acceptable precision and accuracy were gained by intra-and inter-day analysis with relative standard deviations less than 19.5%and relative recoveries ranging from 85.7 to 105.2%.In addition,the AYPBA labeled 20E produced abundant characteristic fragment ions under the high energy collision-induced dissociation,which facilitated the identif cation of 20E by MS^(2)analysis on MALDI-TOF/TOF MS.Using the method,we enabled the identif cation and quantif cation of endogenous 20E in four herbs including Cyanotis arachoidea,Achyranthes bidentata,Spinacia oleracea and Chenopodium quinoa willd.,demonstrating the feasibility of the proposed method for screening of the endogenous 20E in plants.

Simultaneous determination of indole metabolites of tryptophan in rat feces by chemical labeling assisted liquid chromatography-tandem mass spectrometry

As the connecting part of diet and host physiology,intestinal microbes can convert the ingested diet into a huge number of physiologically active small molecules.Indole metabolites of tryptophan are precursors or signal molecules for many biologically active substances,which are involved in serotonin and microbial catabolism pathways.To understand the influence of tryptophan metabolism in the intestinal environment on the neurological and immune systems at the molecular level,it is important to establish a high-coverage analytical method to comprehensively analyze the metabolites involved in tryptophan metabolism.However,due to a small molecular weight and poor response during mass spectrometry analysis,as well as weak retention on the reversed-phase chromatography,determination of indole metabolites of tryptophan is challenging.Here,we proposed a method for the simultaneous determination of 20 indole metabolites of tryptophan in a single run on reversed-phase chromatography by chemical labeling coupled to liquid chromatography-tandem mass spectrometry analysis.4-(Dimethylamino)benzaldehyde(DMAB)was used for the labeling of indole metabolites of tryptophan,which could significantly improve the detection sensitivities and retention of these metabolites on reversed-phase chromatography.With the developed method,we realized the sensitive detection and comprehensive analysis of 15 endogenous indole metabolites of tryptophan in rat feces samples with functional dyspepsia intervention by acupuncture.The developed method offers a useful tool for studying tryptophan metabolism-related diseases.