外源咖啡酸和阿魏酸对黑莓汁中花色苷的辅色研究

为增强黑莓汁中花色苷的稳定,添加适量咖啡酸和阿魏酸到黑莓清汁中,采用可见吸收光谱和高效液相色谱-质谱研究其对黑莓花色苷的辅色作用。研究结果表明:黑莓汁中添加咖啡酸和阿魏酸显著增加了花色苷的最大吸收值(Aλmax),最大吸收波长(λmax)红移,说明咖啡酸和阿魏酸对黑莓汁中花色苷产生了辅色作用,辅色效应随时间的延长和咖啡酸、阿魏酸浓度的增加显著增强。HPLC-DAD-MS分析发现,咖啡酸辅色产生了两种新的花色苷衍生物(矢车菊素-3-O-葡萄糖苷-4-乙烯基儿茶酚和矢车菊-3-O-草酸酐酰葡萄糖苷-4-乙烯基儿茶酚),阿魏酸辅色产生了三种新的花色苷衍生物(矢车菊素-3-O-葡萄糖苷-4-乙烯基愈创木酚、矢车菊-3-O-草酸酐酰葡萄糖苷-4-乙烯基愈创木酚和矢车菊-3-O-阿拉伯糖苷-4-乙烯基愈创木酚),这些衍生物均为羟苯基-吡喃花色苷。     

Influence of Side-Chain Substituents on the Position of Cleavage of the Benzene Ring by Pseudomonas fluorescens

Pseudomonas fluorescens was grown on mineral salts media with phenol, p-hydroxybenzoic acid, p-hydroxy-phenylacetic acid, or p-hydroxy-trans-cinnamic acid as sole carbon and energy source. Each compound was first hydroxylated, ortho to the hydroxyl group on the benzene ring, to give catechol, protocatechuic acid (3,4-dihydroxy-benzoic acid), homoprotocatechuic acid (3,4-dihydroxy-phenylacetic acid), and caffeic acid (3,4-dihydroxy-trans-cinnamic acid), respectively, as the ultimate aromatic products before cleavage of the benzene nucleus. Protocatechuic acid and caffeic acid were shown to be cleaved by ortho fission, via a 3,4-oxygenase mechanism, to give beta-substituted cis, cis-muconic acids as the initial aliphatic products. However, catechol and homoprotocatechuic acid were cleaved by meta fission, by 2,3-and 4,5-oxygenases, respectively, to give alpha-hydroxy-muconic semialdehyde and alpha-hydroxy-gamma-carboxymethyl muconic semialdehyde as initial aliphatic intermediates. Caffeic acid: 3,4-oxygenase, a new oxygenase, consumes 1 mole of O(2) per mole of substrate and has an optimal pH of 7.0. The mechanism of cleavage of enzymes derepressed for substituted catechols by P. fluorescens apparently changes from ortho to meta with the increasing nephelauxetic (electron donor) effect of the side-chain substituent.     

Modeling suberization with peroxidase-catalyzed polymerization of hydroxycinnamic acids: cross-coupling and dimerization reactions

An anionic potato peroxidase (EC 1.11.1.7, APP) thought to be involved in suberization after wounding was isolated from slices of Solanum tuberosum in order to elucidate the first steps of dehydrogenative polymerization between pairs of different hydroxycinnamic acids (FA, CafA, CA and SA) present in wound-healing plant tissues. Use of a commercial horseradish peroxidase (HRP)-H2O2 catalytic system gave the identical major products in these coupling reactions, providing sufficient quantities for purification and structural elucidation. Using an equimolar mixture of pairs of hydroxycinnamic acid suberin precursors, only caffeic acid is coupled to ferulic acid and sinapic acid in separate cross-coupling reactions. For the other systems, HRP and APP reacted as follows: (1) preferentially with ferulic acid in a reaction mixture that contained p-coumaric and ferulic acids; (2) with sinapic acid in a mixture of p-coumaric and sinapic acids; (3) with sinapic acid in a mixture of ferulic and sinapic acids; (4) with caffeic acid in a reaction mixture of p-coumaric and caffeic acids. The resulting products, isolated and identified by NMR and MS analysis, had predominantly beta-beta-gamma-lactone and beta-5 benzofuran molecular frameworks. Five cross-coupling products are described for the first time, whereas the beta-O-4 dehydrodimers identified from the caffeic acid and sinapic acid cross-coupling reaction are known materials that are highly abundant in plants. These reactivity trends lead to testable hypotheses regarding the molecular architecture of intractable suberin protective plant materials, complementing prior analysis of monomeric constituents by GC-MS and polymer functional group identification from solid-state NMR, respectively.     

Characterisation of new oligoglycosidic compounds in two Chinese medicinal herbs

A series of caffeic acid derivatives (3,5-dicaffeoyl-quinic acid, 3,4-dicaffeoyl-quinic acid, and 4,5-dicaffeoyl-quinic acid), and the new compound beta,3,4-trihydroxyphenethyl-O-[beta-apiofuranosyl-(1-->4)-alpha- rhamnopyranosyl-(1-->3)]-(4-O-caffeoyl)-beta-glucopyranoside (wedelosin), as well as three known flavonoid glycosides (quercetin 3-O-beta-glucoside, kaempferol 3-O-beta-apiosyl-(1-2)-beta-glucoside, and astragalin or kaempferol 3-O-beta-glucoside) were isolated from the Chinese medicinal herb Wedelia chinensis. Wedelosin showed an inhibitory activity on both the classical and the alternative activation pathway of the complement system. Another Chinese medicinal herb, Kyllinga brevifolia, yielded two known flavonoid glycosides [kaempferol 3-O-beta-apiosyl-(1-2)-beta-glucoside and isorhamnetin 3-O-beta-apiosyl-(1-2)-beta-glucoside], and a new quercetin triglycoside [quercetin 3-O-beta-apiofuranosyl-(1-->2)-beta-glucopyranoside 7-O-alpha-rhamnopyranoside]. The latter compound showed a moderate anti-viral activity.     

Non-natural cinnamic acid derivatives as substrates of cinnamate 4-hydroxylase

Cinnamate 4-hydroxylase (C4H), a monooxygenase in the plant phenylpropanoid pathway, was assayed for its ability to hydroxylate 29 substrate analogues. Nine of the tested analogues with various aromatic side chains, including 3-coumaric acid, were metabolized by C4H. Seven products from these reactive analogues were characterized using LC/MS, 1H NMR and 13C NMR spectroscopic analysis. For example, caffeic acid was the product of 3-coumaric acid. The products 4-hydroxy-2-chlorocinnamic acid and 4-hydroxy-2-ethoxycinnamic acid are novel compounds that have not been previously reported. The kinetic parameters of C4H towards these analogues were determined.     

New phenolic inhibitors of the peroxidse-catalysed oxidation of indole-3-acetic acid

Previously we reported two metabolites of the insecticide carbofuran as persistent inhibitors of the peroxidase-catalysed oxidtion ofindole-3-acetic acid. In searching for more active inhibitors of this type, we have found that 5-hydroxy-2,2-dimethylchromene (β-tubanol), 2′,6′-dihydroxycetophenone oxime, 5-hydroxy-2,2-dimethylchroman, 2′,6′-dihydroxyacetophenone and 2,6-dihydroxybenzoic acid methyl ester were more active than the carbofuran metabolite 7-hydroxy-2,2-dimethyl-3-oxo-2,3-dihydrobenzofuran. Resorcinol, 5-hydroxy-2,2-dimethylchroman-4-one, 3-hydroxy-5-methoxy-2,2-dimethylchroman-4-one and 5-hydroxy-2-methylchrom-4-one were also inhibitory but with less activity. The new inhibitors differed from the well-known phenolic inhibitors such as caffeic acid in inhibition kinetics as demonstrated by the rate of disappearance of indole-3-acetic acid, the rate of formation of the oxidation products, and the transient spectral change in the enzyme.     

Redox cycles of caffeic acid, alpha-tocopherol, and ascorbate: implications for protection of low-density lipoproteins against oxidation

This study addresses the dynamic interactions among alpha-tocopherol, caffeic acid, and ascorbate in terms of a sequence of redox cycles aimed at accomplishing optimal synergistic antioxidant protection. Several experimental models were designed to examine these interactions: UV irradiation of alpha-tocopherol-containing sodium dodecyl sulfate micelles, one-electron oxidations catalyzed by the hypervalent state of myoglobin, ferrylmyoglobin, and autoxidation at appropriate pHs. These models were assessed by ultraviolet (UV) and electron paramagnetic resonance (EPR), entailing direct- and continuous-flow experiments, spectroscopy and by separation and identification of products by HPLC. The alpha-tocopheroxyl radical EPR signal generated by UV irradiation of alpha-tocopherol-containing micelles was suppressed by caffeic acid and ascorbate; in the former case, no other EPR signal was observed at pH 7.4, whereas in the latter case, the alpha-tocopheroxyl radical EPR signal was replaced by a doublet EPR spectrum corresponding to the ascorbyl radical (A*-). The potential interactions between caffeic acid and ascorbate were further analyzed by assessing, on the one hand, the ability of ascorbate to reduce the caffeic acid o-semiquinone (generated by oxidation of caffeic acid by ferrylmyoglobin) and, on the other hand, the ability of caffeic acid to reduce ascorbyl radical (generated by autoxidation or oxidation of ascorbate by ferrylmyoglobin). The data presented indicate that the reductive decay of ascorbyl radical (A*-) and caffeic acid o-semiquinone (Caf-O*) can be accomplished by caffeic acid (Caf-OH) and ascorbate (AH-), respectively, thus pointing to the reversibility of the reaction Caf-O* + AH- <--> Caf-OH + A*-. Continuous-flow EPR measurements of mixtures containing ferrylmyoglobin, alpha-tocopherol-containing micelles, caffeic acid, and ascorbate revealed that ascorbate is the ultimate electron donor in the sequence encompassing transfer of the radical character from the micellar phase to the phase. In independent experiments, the effects of caffeic acid and ascorbate on the oxidation of two low-density lipoprotein (LDL) populations, control and alpha-tocopherol-enriched, were studied and results indicated that alpha-tocopherol, caffeic acid, and ascorbate acted synergistically to afford optimal protection of LDL against oxidation. These results are analyzed for each individual antioxidant in terms of three domains: its localization and that of the antioxidant-derived radical, its reduction potential, and the predominant decay pathways for the antioxidant-derived radical, that exert kinetic control on the process.     

Downregulation of caffeic acid 3-O-methyltransferase and caffeoyl CoA 3-O-methyltransferase in transgenic alfalfa. impacts on lignin structure and implications for the biosynthesis of G and S lignin

Transgenic alfalfa plants were generated harboring caffeic acid 3-O-methyltransferase (COMT) and caffeoyl CoA 3-O-methyltransferase (CCOMT) cDNA sequences under control of the bean phenylalanine ammonia-lyase PAL2 promoter. Strong downregulation of COMT resulted in decreased lignin content, a reduction in total guaiacyl (G) lignin units, a near total loss of syringyl (S) units in monomeric and dimeric lignin degradation products, and appearance of low levels of 5-hydroxy guaiacyl units and a novel dimer. No soluble monolignol precursors accumulated. In contrast, strong downregulation of CCOMT led to reduced lignin levels, a reduction in G units without reduction in S units, and increases in beta-5 linked dimers of G units. Accumulation of soluble caffeic acid beta-d-glucoside occurred only in CCOMT downregulated plants. The results suggest that CCOMT does not significantly contribute to the 3-O-methylation step in S lignin biosynthesis in alfalfa and that there is redundancy with respect to the 3-O-methylation reaction of G lignin biosynthesis. COMT is unlikely to catalyze the in vivo methylation of caffeic acid during lignin biosynthesis.     

Effet de L'acide Caféoylshikimique des Racines du Palmier Dattier sur L'activité et la Production des Enzymes Hydrolytiques de Fusarium oxysporum f. sp. albedinis

Effect of caffeoylshikimic acid of date palm roots on activity and production of Fusarium oxysporum f. sp. albedinis cell wall‐degrading enzymes The caffeoylshikimic acid (CSA), a major phenolic compound of date palm roots, represents one of the resistance factors of the host to Fusarium oxysporum f. sp. albedinis. The CSA was tested at various concentrations (0,25 to 3 µ mol/ml) on the activity and the production of F. oxysporum f. sp. albediniscell wall‐degrading enzymes (CWDE): proteases, cellulases, pectinemethyl‐esterases (PME), polygalacturonases (PG) and polygalacturonate trans‐eliminases (PGTE). The results obtained show that CSA had very little effect on the activity of the various enzymes although it greatly reduced their production. The mycelial growth was also affected by CSA, but this does not explain why only the production of CWDE was noticeably reduced. In order to explain this differential effect of CSA on the activity and production of CWDE, in one group of experiments the effects of the products of hydrolysis of CSA (caffeic acid and shikimic acid) was tested and in another, the effect of the products of CSA (quinones) obtained by tyrosinase oxidation was investigated. The results obtained show that the shikimic acid did not have a significant effect on the activity of the CWDE but presented a weak inhibition of their production.The caffeic acid showed a larger inhibition of the activity of the various CWDE that was more than that of CSA and its inhibiting effect appeared to be more important during their production. The oxidation of CSA by tyrosinase was accompanied by a greater inhibition of the activity of the various CWDE. This inhibition was appreciable in comparison with that observed due to the effect of non‐oxidized CSA on CWDE production. In the same way, oxidation of caffeic acid provoked a greater inhibiting effect on the activity of CWDE than unoxidized caffeic acid. These results suggests that CSA generates products of hydrolysis (in particular caffeic acid) and products of oxidation (quinones) which inhibit the activity of the proteolytic, cellulolytic and pectinolytic enzymes produced by F. oxysporum f. sp. albedinis in the culture medium.     

Phytochemicals from the aerial parts of Ligularia thomsonii and their radical scavenging activity

Phytochemical investigation of the aerial parts of Ligularia thomsonii has led to the isolation of three new phenylpropanoid glucosides ligularoside I (1), ligularoside II (2) and ligularoside III (3) along with nine known compounds; cinnamic acid (4), 3-phenylpropanoic acid (5), 3,4,5-trihydroxybenzoic acid (6), 4-hydroxybenzoic acid (7), p-coumaric acid (8), caffeic acid (9), 3,4-dihydroxybenzoic acid (10), kaempferol 3-O-β-d-glucopyranoside (11) and 3,5-di-O-caffeoylquinic acid (12), hitherto unreported from L. thomsonii. Their chemical structures were elucidated by spectroscopic analysis and chemical transformation. All these compounds were tested for antioxidant activity using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay. Among them, compounds 9, 10 and 12 showed significant antioxidant activity against DPPH radicals with IC₅₀ of 19.6, 23.3 and 8.9 μm, respectively.     

Novel Xanthine Oxidase (XO) inhibitory phenylindanes produced by thermal reaction of caffeic acid

The products from the thermal reaction of chlorogenic and caffeic acids, which is a model process of roasting coffee beans, exhibited xanthine oxidase (XO) inhibitory activity. From caffeic acid, six inhibitory phenylindanes were identified, and a new phenylindane displayed the highest inhibitory activity among them. The activity of these phenylindanes may contribute to XO inhibition-related functions of roasted coffee beverages.     

Extracellular tyrosinase from the fungus Trichoderma reesei shows product inhibition and different inhibition mechanism from the intracellular tyrosinase from Agaricus bisporus

Tyrosinase (EC 1.14.18.1) is a widely distributed type 3 copper enzyme participating in essential biological functions. Tyrosinases are potential biotools as biosensors or protein crosslinkers. Understanding the reaction mechanism of tyrosinases is fundamental for developing tyrosinase-based applications. The reaction mechanisms of tyrosinases from Trichoderma reesei (TrT) and Agaricus bisporus (AbT) were analyzed using three diphenolic substrates: caffeic acid, L-DOPA (3,4-dihydroxy-l-phenylalanine), and catechol. With caffeic acid the oxidation rates of TrT and AbT were comparable; whereas with L-DOPA or catechol a fast decrease in the oxidation rates was observed in the TrT-catalyzed reactions only, suggesting end product inhibition of TrT. Dopachrome was the only reaction end product formed by TrT- or AbT-catalyzed oxidation of L-DOPA. We produced dopachrome by AbT-catalyzed oxidation of L-DOPA and analyzed the TrT end product (i.e. dopachrome) inhibition by oxygen consumption measurement. In the presence of 1.5mM dopachrome the oxygen consumption rate of TrT on 8mM L-DOPA was halved. The type of inhibition of potential inhibitors for TrT was studied using p-coumaric acid (monophenol) and caffeic acid (diphenol) as substrates. The strongest inhibitors were potassium cyanide for the TrT-monophenolase activity, and kojic acid for the TrT-diphenolase activity. The lag period related to the TrT-catalyzed oxidation of monophenol was prolonged by kojic acid, sodium azide and arbutin; contrary it was reduced by potassium cyanide. Furthermore, sodium azide slowed down the initial oxidation rate of TrT- and AbT-catalyzed oxidation of L-DOPA or catechol, but it also formed adducts with the reaction end products, i.e., dopachrome and o-benzoquinone.     

Phenolic compounds from the branches of Eucalyptus maideni

Three new phenolic compounds, eucalmaidin F (1), (3S)-5-guaiacyl-3-hydroxypentanoic acid (2), and 8-β-C-glucopyranosyl-5,7-dihydroxy-2-isobutylchromone (3), were isolated from the branches of E. maideni, together with 30?known compounds, including four phenylpropanoids, three lignans, four phloroglucinol glucosides, five dihydroflavonoids, seven simple phenolic compounds, six terpenoids, and glycerol. The new structures were established by spectroscopic studies (MS, and 1D- and 2D-NMR), chemical degradation, and modified Mosher's method. Compounds 3, guaiacylglycerol, 3-hydroxy-1-(4-hydroxyphenyl)propan-1-one, caffeic acid, (2E)-3-(4-hydroxyphenyl)prop-2-enoic acid, (7'S,8R,8'R)-lyoniresinol, (+)-lyoresinol 3α-O-α-L-rhamnopyranoside, garcimangosone, phlorocetophenone 2'-glucopyranoside, (+)-taxifolin 3α-O-α-L-rhamnopyranoside, (+)-aromadendrin, (+)-taxifolin, resveratrol, piceatannol, 3,4,5-trihydroxyphenol. Tachiaside, gallic acid, macrocapals A und G, and oleuropeic acid were evaluated for their cytotoxicities against five human cancer cell lines. Resveratrol, piceatannol, gallic acid, and macrocapal G exhibited moderate inhibitory effects on human myeloid heukemia HL-60 cell, with IC(50) values of 22.05, 22.05, 7.75, and 31.93?μM, respectively; and only macrocapal G showed inhibitory effect on hepatocellular carcinoma SMMC-7721 cell, with an IC(50) value of 26.75?μM.     

Inhibition by chlorogenic acid of haematin-catalysed retinoic acid 5,6-epoxidation.

Chlorogenic acid (3-O-caffeoylquinic acid) inhibited haematin- and haemoglobin-catalysed retinoic acid 5,6-epoxidation. Some other phenol compounds (caffeic acid and 4-hydroxy-3-methoxybenzoic acid) also showed inhibitory effects on the haematin- and haemoglobin-catalysed epoxidation, but salicylic acid did not. Of the above compounds, caffeic acid and chlorogenic acid were potent inhibitors compared with the other two, suggesting that the o-hydroquinone moiety of chlorogenic acid and caffeic acid is essential to the inhibition of the epoxidation. Although caffeic acid inhibited retinoic acid 5,6-epoxidation requiring the consumption of O2, formation of retinoic acid radicals was not inhibited on the addition of caffeic acid to the incubation mixture. The above results suggest that caffeic acid does not inhibit the formation of retinoic acid radicals but does inhibit the step of conversion of retinoic acid radical into the 5,6-epoxide.     

Purification and characterization of a O-methyltransferase capable of methylating 2-hydroxy-3-alkylpyrazine from Vitis vinifera L. (cv. Cabernet Sauvignon).

An S-adenosyl-L-methionine-dependent O-methyltransferase capable of methylating 2-hydroxy-3-alkylpyrazine (HP) was purified 7,300-fold to apparent homogeneity with an 8.2% overall recovery from Vitis vinifera L. (cv. Cabernet Sauvignon) through a purification procedure including column chromatography on DEAE-Sepharose FF, Ether-5PW, hydroxyapatite, G2000SW(XL), and DEAE-5PW. The relative molecular mass of the native enzyme estimated on gel permeation chromatography was 85 kDa, and the subunit molecular mass was estimated to be 41 kDa on SDS-polyacrylamide gel electrophoresis. The enzyme also methylates caffeic acid. The Vmax for IBHP and caffeic acid were 0.73 and 175 pkatals/mg, respectively, and the respective Km for IBHP and caffeic acid were 0.30 and 0.032 mm. The optimum pH for IBHP (8.5) was different from that for caffeic acid (7.5).     

玉米苗中DIMBOA与几种酚酸类物质抑菌活性比较

本文从室内培养的7日龄玉米幼苗中提取、分离、鉴定了抗性次生化合物丁布(2,4-d ihydroxy-7-m ethoxy-2H-1,4-benzoxazin-3(4H)-one,D IMBOA),并就该物质对玉米纹枯病病原菌立枯丝核菌(Rhizoctonia solani)的活性与三种酚酸类物质(阿魏酸、对羟基肉桂酸和咖啡酸)进行了离体比较研究。结果表明,丁布(D IMBOA),对立枯丝核菌有很强的生物活性,在浓度为50μg/mL时即可抑制立枯丝核菌菌丝的生长,抑制率为18.52%。阿魏酸、对羟基肉桂酸和咖啡酸,这三种酚酸在浓度250μg/mL时对立枯丝核菌菌丝的生长有抑制作用,抑制率分别为26.30%、8.50%和6.30%。不仅如此,丁布与对羟基肉桂酸之间、以及三种酚酸两两组合之间还存在一定的协同作用。在浓度相等的情况下,丁布与对羟基肉桂酸的等量混合液的抑菌率显著高于这两种物质单独存在时的抑菌率之和;同样,对羟基肉桂酸与阿魏酸的等量混合液的抑菌率比单一的对羟基肉桂酸溶液的抑菌率高18.89%,比单一的阿魏酸溶液的抑菌率高13.33%;对羟基肉桂酸与咖啡酸的等量混合液,抑菌率比两者单独试验时分别高9.63%和14.83%;阿魏酸与咖啡酸的混合液,抑菌率比两酸单独试验时分别高11.48%和22.23%。这一结果提示植物体内产生适当比例不同次生化合物的组合对植物抗病性的提高是至关重要的。     

团花树皮的化学成分研究

采用硅胶、MCI和Sephadex LH-20层析方法对团花树皮的化学成分进行分离纯化,运用现代波谱技术鉴定了10个化合物:4-carboxy-3-hydroxy-5-methylphenyl 3-methoxy-4-hydroxy-5-methylbenzoate(1),谷甾醇-3-O-(6’-O-棕榈酰基)-β-D-葡萄糖苷(2),喹诺酸-3-O-α-L-鼠李糖苷(3),clethric acid(4),常春藤苷元(5),钩藤苷元C(6),morolic acid(7),咖啡酸甲酯(8),卡丹宾(9)和3α-二氢卡丹宾(10)。其中化合物1为一个新的酚性成分,化合物2～8首次从该属植物中分离得到。     

Coffee polyphenol caffeic acid but not chlorogenic acid increases 5'AMP-activated protein kinase and insulin-independent glucose transport in rat skeletal muscle

Chlorogenic acid is an ester of caffeic and quinic acids, and is one of the most widely consumed polyphenols because it is abundant in foods, especially coffee. We explored whether chlorogenic acid and its metabolite, caffeic acid, act directly on skeletal muscle to stimulate 5'-adenosine monophosphate-activated protein kinase (AMPK). Incubation of rat epitrochlearis muscles with Krebs buffer containing caffeic acid (≥0.1 mM, ≥30 min) but not chlorogenic acid increased the phosphorylation of AMPKα Thr(172), an essential step for kinase activation, and acetyl CoA carboxylase Ser(79), a downstream target of AMPK, in a dose- and time-dependent manner. Analysis of isoform-specific AMPK activity revealed that AMPKα2 activity increased significantly, whereas AMPKα1 activity did not change. This enzyme activation was associated with a reduction in phosphocreatine content and an increased rate of 3-O-methyl-d-glucose transport activity in the absence of insulin. These results suggest that caffeic acid but not chlorogenic acid acutely stimulates skeletal muscle AMPK activity and insulin-independent glucose transport with a reduction of the intracellular energy status.     

Exogenous caffeic acid inhibits the growth and enhances the lignification of the roots of soybean (Glycine max)

The allelopathic effect of caffeic acid was tested on root growth, phenylalanine ammonia-lyase (PAL) and peroxidase (POD) activities, hydrogen peroxide (H₂O₂) accumulation, lignin content and monomeric composition of soybean (Glycine max) roots. We found that exogenously applied caffeic acid inhibited root growth, decreased the PAL activity and H₂O₂ content and increased the soluble and cell wall-bound POD activities. The p-hydroxyphenyl (H), guaiacyl (G), and syringyl (S) monomers and total lignin (H + G + S) increased in the caffeic acid-exposed roots. When applied in conjunction with piperonylic acid (PIP, an inhibitor of the cinnamate 4-hydroxylase, C4H), caffeic acid equalized the inhibitory effect of PIP, whereas the application of methylene dioxocinnamic acid (MDCA, an inhibitor of the 4-coumarate:CoA ligase, 4CL) plus caffeic acid decreased lignin production. These results indicate that exogenously applied caffeic acid can be channeled into the phenylpropanoid pathway via the 4CL reaction, resulting in an increase of lignin monomers that solidify the cell wall and inhibit root growth.     

Free radical scavengers from Cymbopogon citratus (DC.) stapf plants cultivated in bioreactors by the temporary immersion (TIS) principle

The biomass production of Cymbopogon citratus shoots cultivated in bioreactors according to the temporary immersion (TIS) principle was assessed under different growth conditions. The effect of gassing with CO2-enriched air, reduced immersion frequency, vessel size and culture time on total phenolic and flavonoid content and free radical scavenging effect of the methanolic extracts was measured. From the TIS-culture of C. citratus, seven compounds were isolated and identified as caffeic acid (1), chlorogenic acid (2), neochlorogenic acid (3), p-hydroxybenzoic acid (4), p-hydroxybenzoic acid 3-O-beta-D-glucoside (5), glutamic acid (6) and luteolin 6-C-fucopyranoside (7). The occurrence of compounds 1-7 and their variability in C. citratus grown under different TIS conditions was determined by HPLC. The free radical scavenging effect of the methanolic extract and compounds was measured by the discoloration of the free radical 1,1-diphenyl-2-picrylhydrazyl (DPPH). The main metabolites in 6- and 8-week-old cultures, both in 5 and 10 1 vessels, were chlorogenic acid (2) (100-113 mg%) and neochlorogenic acid (3) (80-119 mg%), while in the cultures with CO2-enriched air and reduced immersion frequency the main compound detected in the extracts was glutamic acid (6) (400 and 670 mg% for the green and white biomass and 619 and 630 mg% for the green and white biomass, respectively). The most active compounds, as free radical scavengers, in the DPPH discoloration assay were caffeic acid (1), chlorogenic acid (2), neochlorogenic acid (3) and the flavonoid luteolin 6-C-fucopyranoside (7).