Biochemical changes after cold acclimation in Nordic red clover (Trifolium pratense L.) accessions with contrasting levels of freezing tolerance.

The ability to tolerate low freezing temperatures is an important component of winter survival and persistence of red clover. Cold acclimation (CA) allows plants to acquire higher levels of freezing tolerance. However, the biochemical responses to cold and the importance of such changes for the plant to acquire adequate freezing tolerance have not been investigated in red clover of Nordic origin, which has a distinct genetic background. To shed light on this, we selected five freezing tolerant (FT) and five freezing susceptible (FS) accessions and studied the effect of CA on the contents of carbohydrates, amino acids, and phenolic compounds in the crowns. Among those compounds which increased during CA, FT accessions had higher contents of raffinose, pinitol, arginine, serine, alanine, valine, phenylalanine, and one phenolic compound (a pinocembrin hexoside derivative) than FS accessions, suggesting a role for these compounds in the freezing tolerance in the selected accessions. These findings, together with a description of the phenolic profile of red clover crowns, significantly add to the current knowledge of the biochemical changes during CA and their role in freezing tolerance in Nordic red clover.

A comparative metabolomics investigation of flavonoid variation in faba bean flowers.

INTRODUCTION: Faba bean (Vicia faba L.) flowers are edible and used as garnishes because of their aroma, sweet flavor and attractive colors. Anthocyanins are the common plant pigments that give flowers their vivid colors, whereas non-anthocyanin flavonoids can serve as co-pigments that can modify the color intensity of flowers. OBJECTIVES: To explore the polyphenol diversity and differences in standard and wing petals of faba bean flowers; and identify glycosylated flavonoids that contribute to flower color. METHODS: Flower standard and wing petals from 30 faba bean genotypes (eight color groups with a total of 60 samples) were used for polyphenol extraction. Samples were analyzed using a targeted method and a semi-untargeted analysis using liquid chromatography-high resolution mass spectrometry (LC-HRMS) combined with photodiode array (PDA) detection. Compound Discoverer software was used for polyphenol identification and multivariate analysis. RESULTS: The semi-untargeted analysis guided by the PDA detected 90 flavonoid metabolites present in faba bean flower petals. Ten anthocyanins largely influenced the flower colors, but other flavonoids (63 flavonols and 12 flavones) found with variable levels in different flower color groups appeared to also influence color, especially in mixed colors. CONCLUSION: Analysis of the different colored faba bean flowers confirmed that the color variation between the flowers was mainly controlled by anthocyanins in brown, red and purple-red flowers. Of the other flavonoids, multiglycosylated kaempferols were abundant in white and brown flowers, monoglycosylated kaempferols were common in red and purple-red flowers, and quercetin and apigenin glycosides were abundant co-pigments in purple-red flowers.

Mass Spectrometry-Based Untargeted Metabolomics Reveals the Importance of Glycosylated Flavones in Patterned Lentil Seed Coats.

Lentil seed coats are rich in antioxidant polyphenols that are important for plant defense and have potential as valorized byproducts. Although biochemical differences among lentil seed coat colors have been previously studied, differences among seed coat patterns remain largely unexplored. This study used mass spectrometry-based untargeted metabolomics to investigate polyphenol differences among lentil seed coat patterns to search for biochemical pathways potentially responsible for seed coat pattern differences. Comparing patterned with non-patterned green lentil seed coats, 28 significantly upregulated metabolites were found in patterned seed coats; 19 of them were identified as flavones. Flavones were virtually absent in non-patterned seed coats, thereby strongly suggesting a blockage in their flavone biosynthetic pathway. Although the black pattern is not readily discernible on black seed coats, many of the same flavones found in green marbled seed coats were also found in black seed coats, indicating that black seed coats likely have a marbled pattern.

Water-soluble phenolic compounds and their putative antioxidant activities in the seed coats from different lentil (Lens culinaris) genotypes.

The seed coat is a major byproduct of lentil processing with potential as a sustainable source of antioxidant polyphenols. Profiles of water-soluble phenolic compounds and antioxidant activities of seven genotypes of lentil which includes both normal-tannin and low-tannin seed coats were investigated. Antioxidant activities were assessed using four antioxidant assays, and phenolic compounds were quantified using liquid chromatography mass spectrometry (LC-MS). Total phenolic content (TPC) varied significantly among genotypes and ranged between 1519 ± 140 and 6502 ± 154 µg/g. Thirty phenolic compounds were identified with kaempferol tetraglycoside, catechin-3-glucoside and procyanidins being the dominant compounds in normal-tannin seed coats. Kaempferol tetraglycoside predominated (80-90%) in low-tannin seed coats. Antioxidant activities strongly correlated with TPC (r > 0.93) with a 6-9 times higher activity in normal-tannin than that of low-tannin lentils. Without flavan-3-ols and procyanidins, low-tannin seed coat may not exert strong antioxidant potential, whereas normal-tannin seed coat contains water-extractable natural phenolic compounds with promising antioxidant potential.

An Untargeted Metabolomics Approach for Correlating Pulse Crop Seed Coat Polyphenol Profiles with Antioxidant Capacity and Iron Chelation Ability.

Pulse crop seed coats are a sustainable source of antioxidant polyphenols, but are typically treated as low-value products, partly because some polyphenols reduce iron bioavailability in humans. This study correlates antioxidant/iron chelation capabilities of diverse seed coat types from five major pulse crops (common bean, lentil, pea, chickpea and faba bean) with polyphenol composition using mass spectrometry. Untargeted metabolomics was used to identify key differences and a hierarchical analysis revealed that common beans had the most diverse polyphenol profiles among these pulse crops. The highest antioxidant capacities were found in seed coats of black bean and all tannin lentils, followed by maple pea, however, tannin lentils showed much lower iron chelation among these seed coats. Thus, tannin lentils are more desirable sources as natural antioxidants in food applications, whereas black bean and maple pea are more suitable sources for industrial applications. Regardless of pulse crop, proanthocyanidins were primary contributors to antioxidant capacity, and to a lesser extent, anthocyanins and flavan-3-ols, whereas glycosylated flavonols contributed minimally. Higher iron chelation was primarily attributed to proanthocyanidin composition, and also myricetin 3-O-glucoside in black bean. Seed coats having proanthocyanidins that are primarily prodelphinidins show higher iron chelation compared with those containing procyanidins and/or propelargonidins.

Do Faba Bean Genotypes Carrying Different Zero-Tannin Genes (zt1 and zt2) Differ in Phenolic Profiles?

Faba bean is a cool season grain legume that produces seeds with a high protein content. Seed coat tannins limit its use in food and feed. A low-tannin phenotype is controlled by either of two unlinked recessive genes zt1 and zt2. Liquid chromatography-mass spectrometry was used to characterize phenolic profiles of seed coat and flower tissue of three faba bean genotypes: CDC Snowdrop (zt1 gene), Disco/2 (zt2 gene), and ILB 938/2 (tannin-containing). For both tissues, clear differences in phenolic profiles of ILB 938/2 were observed in comparison to both low-tannin lines. Although seed coat phenolic profiles of zt1 and zt2 genotypes were similar, distinct differences were evident in flower tissue, suggesting that the gene action results in some different end products of the phenolic biosynthetic pathway. These distinctive compounds could be used as biochemical markers to distinguish between low-tannin phenotypes.

Polyphenol profile comparisons of seed coats of five pulse crops using a semi-quantitative liquid chromatography-mass spectrometric method.

INTRODUCTION: Pulse crops are nutritious and therefore widely grown. Pulse seed coats are typically discarded, despite their high content of polyphenols that are known for their antioxidant properties and health benefits. A better understanding of polyphenol diversity and biochemical pathways will ultimately provide insight into how polyphenols are linked to health benefits, which will help to better utilise these seed coats. OBJECTIVES: To explore polyphenol profiles among seed coats of diverse genotypes of five pulse crops using a targeted liquid chromatography mass spectrometry (LC-MS) method. METHODS: Four genotypes of each of common bean, chickpea, pea, lentil and faba bean seed coats were selected for analysis. Following extraction, polyphenols were quantified using LC-MS. RESULTS: An LC-MS method was developed to quantify 98 polyphenols from 13 different classes in 30 min. The low-tannin seed coats had the lowest concentrations of all polyphenols. Chickpea and pea seed coats had the most similar polyphenolic profiles. The black common bean showed the most diverse seed coat polyphenol profile, including several anthocyanins not detected in any of the other seed coats. CONCLUSION: The LC-MS method reported herein was used to show polyphenol diversity within several polyphenol classes among the pulse crop seed coats. Detected in all seed coats, flavonols and hydroxybenzoic acids appear well-conserved in the edible Fabaceae. The presence of anthocyanins, flavan-3-ols and proanthocyanins in the coloured seed coats suggests that unique divergent branches were introduced in the flavonoid biosynthetic pathway, possibly in response to environmental stressors.