Deep Annotation of Hydroxycinnamic Acid Amides in Plants Based on Ultra-High-Performance Liquid Chromatography–High-Resolution Mass Spectrometry and Its In Silico Database

Abstract

Hydroxycinnamic acid amides (HCAAs), diversely distributed secondary metabolites in plants, play essential roles in plant growth and developmental processes. Most current approaches can be used to analyze a few known HCAAs in a given plant. A novel method for comprehensive detection of plant HCAAs is urgently needed. In this study, a deep annotation method of HCAAs was proposed on the basis of ultra-high-performance liquid chromatography−highresolution mass spectrometry (UHPLC−HRMS) and its in silico database of HCAAs. To construct an in silico UHPLC− HRMS HCAAs database, a total of 846 HCAAs were generated from the most common phenolic acid and polyamine/aromatic monoamine substrates according to possible biosynthesis reactions, which represent the structures of plant-specialized HCAAs. The characteristic MS/MS fragmentation patterns of HCAAs were extracted from reference mixtures. Four quantitative structure-retention relationship (QSRR) models were developed to predict retention times of mono-trans-HCAAs (aromatic amines conjugates), mono-trans-HCAAs (aliphatic amines conjugates), bis-HCAAs, and tris-HCAAs. The developed method was applied for identifying HCAAs in seeds (maize, wheat, and rice), roots (rice), and leaves (rice and tobacco). A total of 79 HCAAs were detected: 42 of them were identified in these plants for the first time, and 20 of them have never been reported to exist in plants. The results showed that the developed method can be used to identify HCAAs in a plant without prior knowledge of HCAA distributions. To the best of our knowledge, it is the first UHPLC−HRMS database developed for effective deep annotation of HCAAs from nontargeted UHPLC−HRMS data. It is useful for the identification of novel HCAAs in plants.

Hydroxycinnamic Acids and Their Derivatives in Broa, a Traditional Ethnic Maize Bread

“…The different isomers observed in the present work correspond to the different possibilities of double bonds configuration on FA and pCA moieties. In plants, the cis isomers are less common [ 4 ] and, in reversed-phase chromatography, they elute earlier than trans isomers [ 31 ], presenting lower peak areas as well ( Figure 3 ). These compounds were not detected in neither the analysed wheat nor rye flours.…”

“…These compounds were not detected in neither the analysed wheat nor rye flours. Indeed, it is known that maize contains large amounts of conjugated putrescine and spermidine, when comparing to other cereals, such as rice and wheat [ 31 ].…”

“…The monoconjugate N -coumaroyl spermidine (21) was identified in broas , probably formed from the hydrolysis of DCS during processing. The monoconjugates feruloyl and p -coumaroyl putrescine were not detected either in maize flours or broas , although they have been described in maize grains [ 31 , 43 ]. A possible explanation could be that the extraction procedures used by other authors (80% of methanol with 1% of HCl and methanol/isopropanol/water, 8/1/1), could have led to the hydrolysis of more complex HCAAs, liberating the described monoconjugates.…”

“…Compounds 40 and 46 (CFS) were also detected in the commercial wheat and rye flours SF and compounds 39 and 57 (DFS) were also detected in wheat flour SF. These compounds have been recently described in maize grains [ 31 ].…”

Diagnostic fragmentation‐assisted mass spectral networking coupled with in silico dereplication for deep annotation of steroidal alkaloids in medicinal Fritillariae Bulbus

“…[6][7][8][9][10] In addition to the high resolution and sensitivity, MS offers multistage fragmentation data, from which more structural information can be obtained. 11 However, the annotation of PNPs based on untargeted MS data still relies on the differentiation of chemical scaffolds by spectral matching with limited reference databases.…”