In angiosperms, the mature seed consists of embryo, endosperm, and a maternal plant-derived seed coat (SC). The SC plays a role in seed filling, protects the embryo, mediates dormancy and germination, and facilitates the dispersal of seeds. SC properties have been modified during the domestication process, resulting in the removal of dormancy, mediated by SC impermeability. This study compares the SC anatomy and histochemistry of two wild (JI64 and JI1794) and two domesticated (cv. Cameor and JI92) pea genotypes. Histochemical staining of five developmental stages: 13, 21, 27, 30 days after anthesis (DAA), and mature dry seeds revealed clear differences between both pea types. SC thickness is established early in the development (13 DAA) and is primarily governed by macrosclereid cells. Polyanionic staining by Ruthenium Red indicated non homogeneity of the SC, with a strong signal in the hilum, the micropyle, and the upper parts of the macrosclereids. High peroxidase activity was detected in both wild and cultivated genotypes and increased over the development peaking prior to desiccation. The detailed knowledge of SC anatomy is important for any molecular or biochemical studies, including gene expression and proteomic analysis, especially when comparing different genotypes and treatments. Analysis is useful for other crop-to-wild-progenitor comparisons of economically important legume crops.

Main conclusion: We showed that wild pea seeds contained a more diverse combination of bioactive GAs and had higher ABA content than domesticated peas. Although the role of abscisic acid (ABA) and gibberellins (GAs) interplay has been extensively studied in Arabidopsis and cereals models, comparatively little is known about the effect of domestication on the level of phytohormones in legume seeds. In legumes, as in other crops, seed dormancy has been largely or entirely removed during domestication. In this study, we have measured the endogenous levels of ABA and GAs comparatively between wild and domesticated pea seeds during their development. We have shown that wild seeds contained more ABA than domesticated ones, which could be important for preparing the seeds for the period of dormancy. ABA was catabolised particularly by an 8´-hydroxylation pathway, and dihydrophaseic acid was the main catabolite in seed coats as well as embryos. Besides, the seed coats of wild and pigmented cultivated genotypes were characterised by a broader spectrum of bioactive GAs compared to non-pigmented domesticated seeds. GAs in both seed coat and embryo were synthesized mainly by a 13-hydroxylation pathway, with GA₂₉ being the most abundant in the seed coat and GA₂₀ in the embryos. Measuring seed water content and water loss indicated domesticated pea seeds´ desiccation was slower than that of wild pea seeds. Altogether, we showed that pea domestication led to a change in bioactive GA composition and a lower ABA content during seed development.

The seed coat serves as the primary protective barrier, offering mechanical and chemical defense for the embryo. It contains various metabolites, including phenolic compounds, which can be oxidized by polyphenol oxidase (PPO) to form oligomers. In this study, we heterologously expressed a 515 amino acid protein derived from wild pea (Pisum elatius), omitting its N-terminal signal sequence, and analyzed its biochemical properties. The recombinant PeaPPO required sodium dodecyl sulfate (SDS) for activation and exhibited activity between pHs 5.2 and 7.0, peaking at pH 6.0 with 0.25 mM SDS. Tropolone and its isomer thujaplicin were the most effective inhibitors. PeaPPO catalyzed reactions with seed coat-derived substrates, displaying activity toward phenols, catechols, and pyrogallols, with the highest affinity for catechols. Principal component analysis of LC-MS/MS-derived phenolic profiles demonstrated that PPO+ and ppo- genotypes differ significantly in their accumulation of PPO substrates and inhibitors. These findings confirm that PeaPPO possesses both monophenolase and catechol oxidase activities, identifying it as a tyrosinase.

Pollen is a cornerstone of life for plants. Its durability, adaptability, and complex design are the key factors to successful plant reproduction, genetic diversity, and the maintenance of ecosystems. A detailed study of its chemical composition is important to understand the mechanism of pollen–pollinator interactions, pollination processes, and allergic reactions. In this study, a multimodal approach involving Fourier transform infrared spectrometry (FTIR), direct mass spectrometry with an atmospheric solids analysis probe (ASAP), matrix-assisted laser desorption/ionization (MALDI) and ultra-high-performance liquid chromatography–mass spectrometry (UHPLC-MS) was applied for metabolite profiling. ATR-FTIR provided an initial overview of the present metabolite classes. Phenylpropanoid, lipidic, and carbohydrate structures were revealed. The hydrophobic outer layer of pollen was characterized in detail by ASAP-MS profiling, and esters, phytosterols, and terpenoids were observed. Diacyl- and triacylglycerols and carbohydrate structures were identified in MALDI-MS spectra. The MALDI-MS imaging of lipids proved to be helpful during the microscopic characterization of pollen species in their mixture. Polyphenol profiling and the quantification of important secondary metabolites were performed by UHPLC-MS in context with pollen coloration and their antioxidant and antimicrobial properties. The obtained results revealed significant chemical differences among Magnoliophyta and Pinophyta pollen. Additionally, some variations within Magnoliophyta species were observed. The obtained metabolomics data were utilized for pollen differentiation at the taxonomic scale and provided valuable information in relation to pollen interactions during reproduction and its related applications.