Hydroxycinnamic Acid Esters Research Articles

Chemical Characterization of Cider Produced in Hardanger—From Juice to Finished Cider

Our investigation delves into the previously uncharted territory of cider composition from Norway. This study aimed to obtain an overview of the qualitative and quantitative compositions of general chemical parameters, polyphenols (individual and total expressed as gallic acids equivalents), selected esters, and selected C6-alcohols in ciders with the PDO label Cider from Hardanger. In total, 45 juice and cider samples from the fermentation process were collected from 10 cider producers in Hardanger in 2019, 2020, and 2021. Individual sugars, acids, ethanol, and 13 individual phenols were quantified using HPLC-UV/RI. Seven ethyl esters of fatty acids, four ethyl esters of branched fatty acids, ten acetate esters, two ethyl esters of hydroxycinnamic acids, and four C6-alcohols were quantified using HS-SPME-GC-MS. For samples of single cultivars (‘Aroma’, ‘Discovery’, ‘Gravenstein’, and ‘Summerred’), the sum of the measured individual polyphenols in the samples ranges, on average, from 79 to 289 mg L−1 (the lowest for ‘Summerred’ and highest for ‘Discovery’ and ‘Gravenstein’). Chlorogenic acid was the most abundant polyphenol in all samples. Ethyl butyrate, ethyl hexanoate, ethyl octanoate, ethyl decanoate, ethyl isobutyrate, ethyl 2-methylbutyrate, isoamyl acetate, and hexanol were present at concentrations above the odour threshold and contributed to the fruity flavour of the Cider from Hardanger.

Enzymatic Synthesis of Hydroxycinnamic Acid Amides in Water Using the Promiscuous Hydrolase/Acyltransferase PestE from Pyrobaculum calidifontis VA1

AbstractHydroxycinnamic acid amides are believed to have antioxidant, antidiabetic, cytotoxic, anticancer, neuroprotective, and antiinflammatory properties, making them interesting target molecules for potential applications in the food, cosmetics, and pharmaceutical industries. Here, we describe the synthesis of hydroxycinnamic acid amides starting from hydroxycinnamic acid esters and the corresponding amines by using variants of the promiscuous hydrolase/acyltransferase from Pyrobaculum calidifontis VA1 (PestE) in water as a solvent. Up to 97% conversion within two hours at 60 °C was achieved with methyl ferulate and tyramine as substrates. This is a promising, environmentally friendly alternative strategy to established chemical synthesis routes or enzymatic methods using lipases in nonaqueous organic solvents.

Characterization of isogenic mutants with single or double deletions of four phenolic acid esterases in Lactiplantibacillus plantarum TMW1.460

In plants, hydroxycinnamic and hydroxybenzoic acids occur mainly as esters. This study aimed to determine the contribution of individual phenolic acid esterases in Lp. plantarum TMW1.460, which encodes for four esterases: TanA, Lp_0796, Est_1092 and a homolog of Lj0536 and Lj1228 that was termed HceP. To determine which of the phenolic acid esterases present in Lp plantarum TMW1.460 are responsible for esterase activity, mutants with deletions in lp_0796, est_1092, tanB, hceP, or hceP and est_1092 were constructed. The phenotype of wild type strain and mutants was determined with esters of hydroxycinnamic acids (chlorogenic acid and ethyl ferulate) and of hydroxybenzoic acids (methyl gallate, tannic acid and epigallocatechin-3-gallate). Lp. plantarum TMW1.460 hydrolysed chlorogenic acid, methyl ferulate and methyl gallate but not tannic acid or epigallocatechin gallate. The phenotype of mutant strains during growth in mMRS differed from the wild type as follows: Lp. plantarum TMW1.460ΔhceP did not hydrolyse esters of hydroxycinnamic acids; Lp. plantarum TMW1.460ΔtanB did not hydrolyse esters of hydroxybenzoic acids; disruption of est_1092 or Lp_0796 did not alter the phenotype. The phenotype of Lp. plantarum TMW1.460ΔΔhceP/est_1092 was identical to Lp. plantarum TMW1.460ΔhceP. The metabolism of phenolic acids during growth of the mutant strains in broccoli puree and wheat sourdough did not differ from metabolism of the wild type strain. In conclusion, esters of hydroxycinnamic and hydroxybenzoic acids each are hydrolysed by dedicated enzymes. The hydroxycinnamic acid esterase HceP is not expressed, or not active during growth of Lp. plantarum TMW1.460 in all food substrates.

Coordinating caffeic acid and salvianic acid A pathways for efficient production of rosmarinic acid in Escherichia coli

Rosmarinic acid is a natural hydroxycinnamic acid ester used widely in the food and pharmaceutical industries. Although many attempts have been made to screen rate-limiting enzymes and optimize modules through co-culture fermentation, the titer of rosmarinic acid remains at the microgram level by microorganisms. A de novo biosynthetic pathway for rosmarinic acid was constructed based on caffeic acid synthesis modules in Escherichia coli. Knockout of competing pathways increased the titer of rosmarinic acid and reduced the synthesis of rosmarinic acid analogues. An L-amino acid deaminase was introduced to balance metabolic flux between the synthesis of caffeic acid and salvianic acid A. The ratio of FADH2/FAD was maintained via the coordination of deaminase and HpaBC, which is responsible for caffeic acid synthesis. Knockout of menI, encoding an endogenous thioesterase, increased the stability of caffeoyl-CoA. The final strain produced 5780.6 mg/L rosmarinic acid in fed-batch fermentation, the highest yet reported for microbial production. The strategies applied in this study lay a foundation for the synthesis of other caffeic acid and rosmarinic acid derivatives.

THE INFLUENCE OF CHITOSANES ON THE DYNAMICS OF THE CONTENT OF PHENOLIC COMPOUNDS IN TOMATO LEAVES

The aim of this work was to study the influence of chitosans of different molecular weight on the dynamics of the content of phenolic compounds in tomato plants of the 'Zagadka' variety. In the experiment, tomato plants treated with chitosan with a molecular weight of 50-190 kDa and 310-375 kDa were studied. Using the method of high-performance thin-layer chromatography (HPTLC), biochemical profiling of the extracts of the studied plants was performed. Differences in the primary reactions of plants to chitosans of different molecular weights were revealed. Low molecular chitosan caused a significant increase in the content of phenols and, accordingly, increased the antioxidant potential of leaf tissues. The increase in the content of phenols in response to the action of low-molecular-weight chitosan occurs due to the activation of phenylpropanoid synthesis and an increase in the total antioxidant potential, which indicates the mobilization of the plant organism against a potential pathogen. Thus, an hour after the treatment of plants with a solution of low molecular weight chitosan, the content of rutin in the leaves was 3.36 mg/g, and after 18 hours - 5.56 mg/g. The content of chlorogenic and caffeic acids in the leaves increased in 12 hours. In 18 hours, the content of chlorogenic acid was 1.64 mg/g, and caffeic acid was 0.18 mg/g. This may be due to the gradual decomposition of chlorogenic acid into its constituent components under the influence of low molecular weight chitosan, which is accompanied by the release of caffeic acid. A positive correlation was established between the pool of chlorogenic and caffeic acid (r = 0.995; p < 0.06). Therefore, depending on the molecular weight, chitosan causes significant changes in the synthesis of phenylpropanoids and biochemical transformation of complex and simple esters of hydroxycinnamic acids in tomato plants. The high-molecular chitosan solution caused a decrease in the number of compounds with high antioxidant potential in tomato leaves, which indicates the activation of another plant defense system. The content of rutin in leaves treated with high-molecular-weight chitosan after 18 hours was 1.32 mg/g, which is four times less compared to the treatment of plants with low-molecular-weight chitosan. At the same time, in response to treatment with high molecular weight chitosan, the content of caffeic acid in the leaves increased significantly. The decrease in the content of phenols is associated with the isolation of tissues from a potential threat, which is aimed at its neutralization. The result of such a reaction is the oxidation of phenolic compounds, additional lignification and suberinization of cell walls. The activation of various induced immunity reactions in the plant organism already in the first hours after eating low-molecular or high-molecular chitosan indicates the existence of a system of differential recognition of complex bipolymers by plants.

Phenolipids as new food additives: from synthesis to cell-based biological activities

Increasing interest has been shown in phenolic compounds for enhancing food quality, but their hydrophilicity restricts application in lipophilic systems. Therefore, in this study, twelve hydroxycinnamates derivatives (alkyl and steryl esters of sinapic acid (SA), caffeic acid (CA), and ferulic acid [FA]) were synthesised and evaluated for antioxidant and cytotoxic characteristics. CA esters had the highest radical scavenging activity (RSA) analysed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assays. Values of inhibitory concentration (IC50) of synthesised compounds were related to their structure and lipophilicity. The effect of these hydroxycinnamic acid esters on the antioxidant potential of real samples (rapeseed oil, margarine and mayonnaise) was estimated. None of the investigated derivatives significantly affected the viability of the model intestinal cells Caco2, while the octyl esters demonstrated a toxic effect at low concentrations. The synthesised esters exerted cytotoxic and anti-proliferative effects against transformed cell lines (HeLa and A549). Octyl esters were potent anticancer compounds on two human cancer cell lines. The synthesised phenolipids, as valuable and safe antioxidant additives, can find broader applications in the production of fat-based products to prevent oxidation processes, extend their shelf life and improve quality.

Síntesis benigna y sencilla de dímeros de tipo lignano usando peroxidasa cruda de rábano rojo (Raphanus sativus var sativus)

Alkyl esters of hydroxycinnamic acids and vanillin derivatives were used as model compounds in the oxidative coupling reactions catalyzed by a crude enzymatic extract of Raphanus sativus var sativus (red radish). Six products were isolated and characterized: three alkyl esters of hydroxycinnamic acids dimers and three vanilloids dimers. Herein, we report the enzymatic extract of Raphanus sativus var. sativus showed peroxidase activity catalyzing the oxidative coupling of phenols. Vanilloid dimers were coupled in an ortho-ortho way as expected, while alkyl ester dimers were formed through 5-8 and 8-8 bonds via a radical mechanism prior to cyclization

Profiling of Polyphenols and Glucosinolates in Kale and Broccoli Microgreens Grown under Chamber and Windowsill Conditions by Ultrahigh-Performance Liquid Chromatography High-Resolution Mass Spectrometry

Vegetables from the Brassica family are rich in many health-beneficial components, including high levels of polyphenols and glucosinolates (GSLs). However, their exact metabolite compositions are highly affected by cultivating and environmental conditions. Here, we provide the first report of polyphenol and GSL composition of kale and broccoli microgreens grown in growth chamber and windowsill environments. The samples were harvested 10 days after sowing and analyzed using ultrahigh-performance liquid chromatography high-resolution mass spectrometry. Principal component analysis was performed to visualize group clustering, trends, and discriminative ion features. Flavonol derivatives (21 in kale and 7 in broccoli) and hydroxycinnamic acid derivatives (14 in kale and 22 in broccoli) were identified under both growing conditions. Additionally, GSLs, including 7 in kale and 9 in broccoli, were detected. The results demonstrated a clear variation in secondary metabolites between the two growing conditions. The discriminative ion features were hydroxycinnamic acid esters of flavonols and indolic GSLs. Compared to chamber conditions, hydroxycinnamic acid esters of flavonols such as kaempferol-3-disinapoyl-triglucoside-7-diglucoside, kaempferol-3-disinpapoyl-triglucoside-7-glucoside, and kaempferol-3-disinpapoyl-diglucoside-7-glucoside were found to be greater in the windowsill-grown vegetables. The contents of 4-methoxyglucobrassicin and neoglucobrassicin, on the other hand, were greater in the chamber-grown ones. The results suggest that kale and broccoli microgreens grown under commercial- and in-home-grown conditions have different secondary metabolite compositions.

Highly Selective Mitsunobu Esterification of Cellulose with Hydroxycinnamic Acids

AbstractCellulose esters of hydroxycinnamic acids, namely ferulic‐, p‐coumaric‐, and caffeic acid, are synthesized by Mitsunobu esterification applying triphenylphosphine and diisopropyl azodicarboxylate in N,N‐dimethylacetamide/LiCl. The reaction is found to be highly selective and took place at primary hydroxyl groups at position 6 of cellulose without undesired conversion of double bonds and phenolic hydroxyl groups. Molecular structure characterization is performed by FTIR‐ and NMR spectroscopy. The products are soluble in dimethyl sulfoxide and possessed degree of substitution (DS) values of up to 0.5. DS values are determined by two independent methods, namely 1H NMR spectroscopy of peracetylated derivatives and UV spectroscopy of extracted ferulic acid. The synthesis of cellulose hydroxycinnamates is very important to design lignocellulosic model systems and antioxidative bioinspired materials.

Agronomic and Metabolomic Side-Effects of a Divergent Selection for Indol-3-Ylmethylglucosinolate Content in Kale (Brassica oleracea var. acephala).

Brassica oleracea var. acephala (kale) is a cruciferous vegetable widely cultivated for its leaves and flower buds in Europe and a food of global interest as a “superfood”. Brassica crops accumulate phytochemicals called glucosinolates (GSLs) which play an important role in plant defense against biotic stresses. Studies carried out to date suggest that GSLs may have a role in the adaptation of plants to different environments, but direct evidence is lacking. We grew two kale populations divergently selected for high and low indol-3-ylmethylGSL (IM) content (H-IM and L-IM, respectively) in different environments and analyzed agronomic parameters, GSL profiles and metabolomic profile. We found a significant increase in fresh and dry foliar weight in H-IM kale populations compared to L-IM in addition to a greater accumulation of total GSLs, indole GSLs and, specifically, IM and 1-methoxyindol-3-ylmethylGSL (1MeOIM). Metabolomic analysis revealed a significant different concentration of 44 metabolites in H-IM kale populations compared to L-IM. According to tentative peak identification from MS interpretation, 80% were phenolics, including flavonoids (kaempferol, quercetin and anthocyanin derivates, including acyl flavonoids), chlorogenic acids (esters of hydroxycinnamic acids and quinic acid), hydroxycinnamic acids (ferulic acid and p-coumaric acid) and coumarins. H-IM kale populations could be more tolerant to diverse environmental conditions, possibly due to GSLs and the associated metabolites with predicted antioxidant potential.

Effects of hydroxycinnamic acid esters on sweetpotato weevil feeding and oviposition and interactions with Bacillus thuringiensis proteins

Sweetpotato weevil (SPW) pest management is challenging because the pest target is sub-terranean, so the application of pesticides is impractical and usually ineffective. Host plant resistance and the genetic transformation of sweetpotatoes to produce entomotoxic Bt proteins offer potential for environmentally benign pest control. Resistance can be conferred by naturally occurring hydroxycinnamic acids which protect against oviposition by adults, but these compounds are restricted to the root surface so do not protect against the cortex bound larvae where the greatest damage occurs. Resistance could be enhanced if combined with expression of Bt proteins in transformed plants, but interactions between hydroxycinnamic acids and Bt proteins remain unknown. Here the bioactivity of Cry7Aa1 protein and hydroxycinnamic acid esters was evaluated individually and in combination against SPW larvae and mortality determined. Low and high concentrations of hydroxycinnamic acid esters alone caused significantly higher mortality of both weevil species in all experiments compared to the control. SPW larval mortality was greater when tested as a combination of hydroxycinnamic acid esters and Bt protein, but this effect was additive not synergistic. Although we report no evidence of antagonistic interactions, the antifeedant effects of the plant compounds conferring host plant resistance could have reduced consumption of the Bt protein in our assays leading to a lower efficacy when combined. Further work is required to determine whether the toxic effects of Bt proteins function alongside host plant resistance in sweetpotato under field conditions.

New Zileuton-Hydroxycinnamic Acid Hybrids: Synthesis and Structure-Activity Relationship towards 5-Lipoxygenase Inhibition.

A novel series of zileuton-hydroxycinnamic acid hybrids were synthesized and screened as 5-lipoxygenase (5-LO) inhibitors in stimulated HEK293 cells and polymorphonuclear leukocytes (PMNL). Zileuton’s (1) benzo[b]thiophene and hydroxyurea subunits combined with hydroxycinnamic acid esters’ ester linkage and phenolic acid moieties were investigated. Compound 28, bearing zileuton’s (1) benzo[b]thiophene and sinapic acid phenethyl ester’s (2) α,β-unsaturated phenolic acid moiety 28, was shown to be equipotent to zileuton (1), the only clinically approved 5-LO inhibitor, in stimulated HEK293 cells. Compound 28 was three times as active as zileuton (1) for the inhibition of 5-LO in PMNL. Compound 37, bearing the same sinapic acid (3,5-dimethoxy-4-hydroxy substitution) moiety as 28, combined with zileuton’s (1) hydroxyurea subunit was inactive. This result shows that the zileuton’s (1) benzo[b]thiophene moiety is essential for the inhibition of 5-LO product biosynthesis with our hydrids. Unlike zileuton (1), Compound 28 formed two π–π interactions with Phe177 and Phe421 as predicted when docked into 5-LO. Compound 28 was the only docked ligand that showed a π–π interaction with Phe177 which may play a part in product specificity as reported.

Cytotoxic Effect of Vanicosides A and B from Reynoutria sachalinensis Against Melanotic and Amelanotic Melanoma Cell Lines and in silico Evaluation for Inhibition of BRAFV600E and MEK1.

Vanicosides A and B are the esters of hydroxycinnamic acids with sucrose, occurring in a few plant species from the Polygonaceae family. So far, vanicosides A and B have not been evaluated for anticancer activity against human malignant melanoma. In this study, we tested these two natural products, isolated from Reynoutria sachalinensis rhizomes, against two human melanoma cell lines (amelanotic C32 cell line and melanotic A375 cell line, both bearing endogenous BRAFV600E mutation) and two normal human cell lines—keratinocytes (HaCaT) and the primary fibroblast line. Additionally, a molecular docking of vanicoside A and vanicoside B with selected targets involved in melanoma progression was performed. Cell viability was studied using an MTT assay. A RealTime-Glo™ Annexin V Apoptosis and Necrosis assay was used for monitoring programmed cell death (PCD). Vanicoside A demonstrated strong cytotoxicity against the amelanotic C32 cell line (viability of the C32 cell line was decreased to 55% after 72 h incubation with 5.0 µM of vanicoside A), significantly stronger than vanicoside B. This stronger cytotoxic activity can be attributed to an additional acetyl group in vanicoside A. No significant differences in the cytotoxicity of vanicosides were observed against the less sensitive A375 cell line. Moreover, vanicosides caused the death of melanoma cells at concentrations from 2.5 to 50 µM, without harming the primary fibroblast line. The keratinocyte cell line (HaCaT) was more sensitive to vanicosides than fibroblasts, showing a clear decrease in viability after incubation with 25 µM of vanicoside A as well as a significant phosphatidylserine (PS) exposure, but without a measurable cell death-associated fluorescence. Vanicosides induced an apoptotic death pathway in melanoma cell lines, but because of the initial loss of cell membrane integrity, an additional cell death mechanism might be involved like permeability transition pore (PTP)-mediated necrosis that needs to be explored in the future. Molecular docking indicated that both compounds bind to the active site of the BRAFV600E kinase and MEK-1 kinase; further experiments on their specific inhibitory activity of these targets should be considered.

Metabolic Engineering of Saccharomyces cerevisiae for Rosmarinic Acid Production.

Rosmarinic acid is a hydroxycinnamic acid ester commonly found in the Boraginaceae and Lamiaceae plant families. It exhibits various biological activities, including antioxidant, anti-inflammatory, antibacterial, antiallergic, and antiviral properties. Rosmarinic acid is used as a food and cosmetic ingredient, and several pharmaceutical applications have been suggested as well. Rosmarinic acid is currently produced by extraction from plants or chemical synthesis; however, due to limited availability of the plant sources and the complexity of the chemical synthesis method, there is an increasing interest in producing this compound by microbial fermentation. In this study, we aimed to produce rosmarinic acid by engineered baker’s yeast Saccharomyces cerevisiae. Multiple biosynthetic pathway variants, carrying only plant genes or a combination of plant and Escherichia coli genes, were implemented using a full factorial design of experiment. Through analysis of variances, the effect of each enzyme variant (factors), together with possible interactions between these factors, was assessed. The best pathway variant produced 2.95 ± 0.08 mg/L rosmarinic acid in mineral medium with glucose as the sole carbon source. Increasing the copy number of rosmarinic acid biosynthetic genes increased the titer to 5.93 ± 0.06 mg/L. The study shows the feasibility of producing rosmarinic acid by yeast fermentation.

Metagenomic discovery of feruloyl esterases from rumen microflora

Feruloyl esterases (FAEs) are a key group of enzymes that hydrolyze ferulic acids ester-linked to plant polysaccharides. The cow's rumen is a highly evolved ecosystem of complex microbial microflora capable of converting fibrous substances to energy. From direct cloning of the rumen microbial metagenome, we identified seven active phagemids conferring feruloyl esterase activity. The genomic inserts ranged from 1633 to 4143 bp, and the ORFs from 681 to 1359 bp. BLAST search reveals sequence homology to feruloyl esterases and esterases/lipases identified in anaerobes. The seven genes were expressed in Escherichia coli, and the proteins were purified to homogeneity. The FAEs were found to cover types B, C, and D in the feruloyl esterase classification system using model hydroxycinnamic acid esters. The release of ferulic acid (FA) catalyzed by these enzymes was established using natural substrates corn fiber (CF) and wheat insoluble arabinoxylan (WIA). Three of the enzymes were demonstrated to cleave diferulates and hence the capability to break down Araf-FA-FA-Araf cross-links. The wide variation in the sequence, activity, and substrate specificity observed in the FAEs discovered in this study is a confirming evidence that combined actions of a full range of FAE enzymes contribute to the high-efficiency fiber digestion in the rumen microbial ecosystem.

Authentication of black cohosh (Actaea racemosa) dietary supplements based on chemometric evaluation of hydroxycinnamic acid esters and hydroxycinnamic acid amides.

Ester and amide derivatives of hydroxycinnamic acids are found in black cohosh (Actaea racemosa) and other Actaea plants. These two compound groups were evaluated for authentication of black cohosh dietary supplements. The hydroxycinnamic acid esters (HCAE) were profiled by ultra-performance liquid chromatography-photodiode array detection (UPLC-PDA). The hydroxycinnamic acid amides (HCAA) were acquired simultaneously by mass spectrometry-multiple reaction monitoring (UPLC-MRM) mode. In contrast with the traditional HCAE method using 8 compounds, profiles of HCAA using only 4 feruloyl dopamine-O-hexosides was more convenient for peak by peak comparison. Partial least square discriminant analysis (PLS-DA) was applied to both HCAE and HCAA datasets. Authenticated plant samples of five Actaea species were randomly divided into training and test sets to build and validate the two PLS-DA models. Both models provided reasonable estimates for the classification of A. racemosa and other Actaea plant samples. However, HCAA model performs better in sensitivity, specificity, and accuracy. Assessment of supplement samples provided quite different results for the solid and liquid dietary supplement samples, indicating the dosage form could affect the composition of marker compounds. Graphical abstract.

Production of rosmarinic acid with ATP and CoA double regenerating system

Rosmarinic acid (RA), as a hydroxycinnamic acid ester of caffeic acid (CA) and 3,4-dihydroxyphenyllactic acid (3,4-DHPL), is a phenylpropanoid-derived plant natural product and has diverse biological activities. This work acts as a modular platform for microbial production using a two-cofactor (ATP and CoA) regeneration system to product RA based on a cell-free biosynthetic approach. Optimal activity of the reaction system was pH 8 and 30 °C. Total turnover number for ATP and CoA was 820.60 ± 28.60 and 444.50 ± 9.65, respectively. Based on the first hour data, the RA productivity reached 320.04 mg L−1 h−1 (0.889 mM L−1 h−1).

A BAHD hydroxycinnamoyltransferase from Actaea racemosa catalyses the formation of fukinolic and cimicifugic acids.

The nucleotide sequence of a BAHD hydroxycinnamoyltransferase was amplified from Actaea racemosa (Ranunculaceae) and expressed in E. coli. The protein catalysed the formation of cimicifugic acids and thus is named hydroxycinnamoyl-CoA:piscidic acid hydroxycinnamoyltransferase (ArHPT1; cimicifugic acid synthase). Actaea racemosa (syn. Cimicifuga racemosa) is known to contain triterpene lactone glycosides and cimicifugic acids. The latter are esters of various hydroxycinnamic or benzoic acids with piscidic or fukiic acid. Amplification of a nucleotide sequence from A. racemosa, that was already known as HCT1 from an EST approach, and its expression in E. coli resulted in a protein that was able to catalyse the formation of several cimicifugic acids. For the characterisation of this hydroxycinnamoyltransferase (hydroxy)cinnamoyl-coenzyme A thioesters were synthesised as donor substrates and piscidic acid isolated as acceptor substrate. The lowest Km-value with 6.8µM was determined for p-coumaroyl-CoA. More than 30 possible acceptor substrates were tested, but only piscidic acid and putatively fukiic acid were accepted. The apparent Km-value for piscidic acid was 32.3µM. High expression of the hydroxycinnamoyltransferase gene was found in roots, but the content of cimicifugic acids was higher in leaves and flowers than in roots. This work describes for the first time a biosynthetic step in the formation of cimicifugic acids catalysed by a so far uncharacterised hydroxycinnamoyltransferase accepting piscidic acid as acceptor substrate thus being a hydroxycinnamoyl-CoA:piscidic acid hydroxycinnamoyltransferase (ArHPT1; cimicifugic acid synthase).

Photoprotective potential of medicinal plants from Cerrado biome (Brazil) in relation to phenolic content and antioxidant activity

Photooxidative damage affects cellular lipids, proteins and DNA in addition to being involved in the pathobiochemistry of erythema formation, premature skin aging, photodermatoses development and skin cancer. Phenolic compounds, flavonoids and hydroxycinnamic acid esters protect plant tissues against harmful UV-radiation. This study aimed to evaluate the sun protection factor of several Brazilian plant extracts in relation to UVB radiation absorption, which causes skin cancer, and to correlate the findings with their antioxidant activity, as well as with total phenol and flavonoid content. The antioxidant activity of the extracts were evaluated using the DPPH radical scavenging test. The photoprotective effect was evaluated using the methodology developed by Mansur. The antioxidant activity from the extracts showed IC50 values ranging from 4.91 to 132.24 μg/mL when compared to the standard quercetin with an IC50 of 5.01 μg/mL; the phenolic content varied from 3.77 to 57.14 mg GAE/g extract while flavonoid content varied from 1.80 to 5.89 mg EQ/g extract. Almost all extracts showed a potential for UVB radiation absorption in accordance with the ANVISA (Agência Nacional de Vigilância Sanitária) technical regulation for cosmetic sunscreens. The extracts with greater sun protection factors were those originating from Lippia microphylla (SPF = 26.82) and Dimorphandra gardneriana (SPF = 20.12). A sun protection factor of 15 or higher contributes to protect the skin, where the presence of sakuranetin flavonoids and quercetin glycosides contribute to this action.

Two R2R3-MYB proteins are broad repressors of flavonoid and phenylpropanoid metabolism in poplar.

The phenylpropanoid pathway leads to the production of many important plant secondary metabolites including lignin, chlorogenic acids, flavonoids, and phenolic glycosides. Early studies have demonstrated that flavonoid biosynthesis is transcriptionally regulated, often by a MYB, bHLH, and WDR transcription factor complex. In poplar, several R2R3 MYB transcription factors are known to be involved in flavonoid biosynthesis. Previous work determined that poplar MYB134 and MYB115 are major activators of the proanthocyanidin pathway, and also induce the expression of repressor-like MYB transcription factors. Here we characterize two new repressor MYBs, poplar MYB165 and MYB194, paralogs which comprise a subgroup of R2R3-MYBs distinct from previously reported poplar repressors. Both MYB165 and MYB194 repressed the activation of flavonoid promoters by MYB134 in transient activation assays, and both interacted with a co-expressed bHLH transcription factor, bHLH131, in yeast two-hybrid assays. Overexpression of MYB165 and MYB194 in hybrid poplar resulted in greatly reduced accumulation of several phenylpropanoids including anthocyanins, proanthocyanidins, phenolic glycosides, and hydroxycinnamic acid esters. Transcriptome analysis of MYB165- and MYB194-overexpressing poplars confirmed repression of many phenylpropanoid enzyme genes. In addition, other MYB genes as well as several shikimate pathway enzyme genes were downregulated by MYB165-overexpression. By contrast, leaf aromatic amino acid concentrations were greater in MYB165-overexpressing poplars. Our findings indicate that MYB165 is a major repressor of the flavonoid and phenylpropanoid pathway in poplar, and may also affect the shikimate pathway. The coordinated action of repressor and activator MYBs could be important for the fine tuning of proanthocyanidin biosynthesis during development or following stress.