Callus Induction from Insect Galls by Dryocosmus kuriphilus on Chestnut Tree

Dehydrodicaffeic acid dilactone (DDACD) was found in a cultured mushroom by screening for catechol-O-methyltransferase inhibitors. The enzyme which converts two molecules of caffeic acid to DDCAD has been extracted from the mushroom and purified and the enzyme reaction has been studied. It was markedly inhibited by reducing agents, such as NADPH, NADH, glutathione and ascorbic acid but stimulated by Fe³⁺, Fe²⁺, Co²⁺, Ni²⁺, Cu²⁺, Cu⁺ and Zn²⁺ ions. Sodium diethyldithiocarbamate and sodium cyanide known to be copper chelating agents inactivated the enzyme, but activity was restored by addition of Cu²⁺ or Cu⁺. Although the enzymic reaction did not occur under anaerobic conditions, ¹⁸O-oxygen was not incorporated into DDCAD. o-Diphenol oxidase catalyzed DDCAD formation from caffeic acid and the DDCAD-forming enzyme catalyzed the formation of DOPAchrome from DOPA. Thus, the DDCAD-forming enzyme is a type of o-diphenol oxidase. Peroxidase and hydrogen peroxide produced DDCAD from caffeic acid.

On the other hand, DDCAD was non-enzymatically synthesized from caffeic acid in the presence of CuCl₂ in 64% yield. In both enzymic and non-enzymic syntheses, both (+)- DDCAD and (?)-DDCAD were produced.     

Tissue Browning of Potato Tubers Induced by Gamma Irradiation

A considerable browning was observed especially in cortex tissue and along xylem of potato tubers harvested at Sakai in Osaka Prefecture, after irradiation with 10, 20 and 50 krad doses of cobalt-60 gamma rays. This phenomenon was accompanied by the marked increase in polyphenol content and peroxidase activity, and the transient increase in o-diphenol oxidase activity. Total reducing compounds in the tissue were also increased by gamma irradiation.

The browning phenomenon depended on the storage period from the harvest to gamma irradiation treatment. The browning and the transient increase in o-diphenol oxidase activity were completely suppressed in the case of tubers irradiated 3 months after harvest.

There was no significant change in α-amylase activity in all tubers tested.     

The Inhibitory Effect of some Antibiotics on Increase in o-Diphenol Oxidase Activity during Incubation of Sliced Sweet Potato Tissue

The marked increase in o-diphenol oxidase activity which developed in incubating slices of sweet potato roots was suppressed by administration of actinomycin D, puromycin and blastcidin S. This suggests that the rise in enzyme activity resulted from de novo synthesis of enzyme protein during incubation. The formation of component III of o-diphenol oxidase which occured in response to cutting, was strongly inhibited by supplying the above chemicals.     

A STUDY ON INCREASE IN O-DIPHENOL OXIDASE ACTIVITY DURING INCUBATION OF SLICED SWEET POTATO TISSUE

The increase in o-diphenol oxidase activity and polyphenol contentwas investigated in slices excised from sweet potato roots.o-Diphenol oxidase activity increased in a sigmoidal fashionover a 100 hour period. The increase in polyphenols occurredover a shorter period of time and was evident before an increasein o-diphenol oxidase activity could be detected. Thus, it seemedthat the increase in polyphenol content might be involved inthe enhancement of o-diphenol oxidase activity. However, theabove correlation was not found in different kinds of experimentincluding pretreatment with either vacuum infiltration or wetconditioning. (Received October 14, 1965; )     

Determination of Copper Content in o-Diphenol Oxidase by Neutron Activation Analysis

A polyphenol oxidase (o-diphenol oxidase) [o-diphenol: O₂ oxidoreductase E. C. 1.10. 3.1] from sweet potato named component II_b was highly purified. The copper content of this enzyme was measured by neutron activation analysis. Samples were analyzed with or without chemical separation after neutron irradiation. The copper content of the enzyme was determined to be 0.27%, and the minimum molecular weight of this enzyme was caluculated to be 23,500.     

Antibiotic potential of plant growth promoting rhizobacteria (PGPR) against Sclerotium rolfsii

High performance liquid chromatographic (HPLC) analysis of culture filtrates of plant growth promoting rhizobacteria (PGPR) and medium of inhibitory zone of interaction of Sclerotium rolfsii with PGPR, viz. Pseudomonas aeruginosa, Pseudomonas fluorescens 4, Pseudomonas fluorescens 4 (new) and Pseudomonas sp. varied from sample to sample. In all the culture filtrates of PGPRs, P. aeruginosa had nine phenolic acids in which ferulic acid (14.52 μg/ml) was maximum followed by other phenolic acids. However, the culture filtrates of P. fluorescens 4 had six phenolic acids with maximum ferulic acid (20.54 μg/ml) followed by indole acetic acid (IAA), caffeic, salicylic, o-coumeric acid and cinnamic acids. However, P. fluorescens 4 culture filtrate had seven phenolic acids in which salicylic acid was maximum (18.03 μg) followed by IAA, caffeic, vanillic, ferulic, o-coumeric and cinnamic acids. Pseudomonas sp. also showed eight phenolic acids where caffeic acid (2.75 μg) was maximum followed by trace amounts of ferulic, salicylic, IAA, vanillic, cinnamic, o-coumeric and tannic acids. The analysis of antibiosis zone of PGPRs showed fairly rich phenolic acids. A total of nine phenolic acids were detected in which caffeic acid was maximum (29.14 μg/g) followed by gallic (17.64 μg/g) and vanillic (3.52 μg/g) acids but others were in traces. In P. aeruginosa, antibiosis zone had seven phenolic acids where IAA was maximum (3.48 μg/g) followed by o-coumeric acid (2.08 μg/g), others were in traces. The medium of antibiosis zone of P. fluorescens 4 and P. fluorescens 4 new had eight phenolic acids in which IAA was maximum with other phenolic acids in traces.     

Purification and Characterization of Polyphenol Oxidase from Glandular Trichomes of Solanum berthaultii

Type A glandular trichomes of the wild potato (Solanum berthaultii Hawkes) entrap insects by rapidly polymerizing the trichome contents after breakage by insect contact. Polymerization of trichome exudate appears to be driven by a soluble polyphenol oxidase (PPO). PPO constitutes up to 70% of the protein in individually collected trichomes and reaches a concentration approaching 200 μm in these organs. Trichome PPO has been purified and shown to be a monomeric copper metalloprotein with an isoelectric point of 5.5, possessing only o-diphenol oxygen oxido-reductase activity, and is larger than most other reported PPOs, with relative molecular weight of 59,000. Chlorogenic and caffeic acid were the most readily oxidized of 14 phenolic substrates tested. Polyclonal antibodies raised against the relative molecular weight 59,000 S. berthaultii trichome PPO were used to show that S. tuberosum L. trichomes express low levels of a cross-reactive protein that lacks detectable PPO activity.     

Changes in o-Diphenol Oxidase During Fibre Development in Cotton

Quantitative and electrophoretic changes in o-diphenol oxidase(o-diphenol: O₂ oxidoreductase, E.C. 1 10,3.1) were studiedduring the entire period of cotton (Gossypium arboreum L. cv.Sanjay) fibre development. A significant increase in o-diphenoloxidase activity was recorded during the fibre initiation phaseand it is suggested that a shift in redox balance towards oxidationmay play an important role in fibre initiation. Low o-diphenoloxidase activity during elongation and its high activity duringthe phase of secondary thickening, together with isoenzyme patterns,suggest an important role of this enzyme in cotton fibre development.The role of o-diphenol oxidase in relation to auxin turnoverand redox balance is discussed. Gossypium arboreum, cotton, fibre development, o-diphenol oxidase, redox balance, auxin turnover     

Properties of copper-dependent o-diphenol oxidase activity in the potato aphid Macrosiphum euphorbiae (thomas)

Potato aphid Macrosiphum euphorbiae (Thomas) was found to contain high amounts of o-diphenol oxidase activity. Enzyme activity was largely distributed into the postmitochondrial supernatant from Brij-35 extracted aphids and occurs in a latent form that was activated up to 45-fold by pretreatment with isopropanol. The aphid enzyme has a broad pH optimum near 6, and utilized L-dopa (K_m = 1.4 mM, V_max = 348 nmol/min-mg protein), dopamine, and 4-methylcatechol the best out of the twelve substrates tested. In addition, this activity is a typical copper-dependent oxidase in that it is potently inhibited by phenylthiourea (50% inhibition at 30nM) and other copper chelators, including salicylhydroxamic acid. The above properties are common to most insect tyrosinases. However, the aphid enzyme lacked the o-hydroxylase and laccase components and the optimal activity at higher temperatures that are typical of cuticular tyrosinases of other insects. The high levels of o-diphenol oxidase in aphids compared to other insects is surprising, since the major function associated with these enzymes, that of melanization and sclerotization of cuticle, is of much less importance to aphids. The possibility that aphids use this enzyme to metabolize dietary phenolics is discussed.     

Chemical Characterization and Multidirectional Biological Effects of Different Solvent Extracts of Arum elongatum: in Vitro and in Silico Approaches

Arum elongatum (Araceae) is widely used traditionally for the treatment of abdominal pain, arterial hypertension, diabetes mellitus, rheumatism and hemorrhoids. This study investigated the antioxidant properties, individual phenolic compounds, total phenolic and total flavonoid contents (HPLC/MS analysis), reducing power and metal chelating effects of four extracts obtained from A. elongatum (ethyl acetate (EA), methanol (MeOH), methanol/water (MeOH/water) and infusion). The inhibitory activity of the extracts were also determined against acetylcholinesterase, butyrylcholinesterase, tyrosinase, amylase and glucosidase enzymes. The MeOH/water extracts contained the highest amount of phenolic contents (28.85 mg GAE/g) while the highest total flavonoid content was obtained with MeOH extract (36.77 mg RE/g). MeOH/water demonstrated highest antioxidant activity against DPPH⋅ radical at 38.90 mg Trolox equivalent per gram. The infusion extract was the most active against ABTS⁺⋅ (133.08 mg TE/g). MeOH/water extract showed the highest reducing abilities with the CUPRAC value of 102.22 mg TE/g and the FRAP value of 68.50 mg TE/g. A strong metal chelating effect was observed with MeOH/water extract (35.72 mg EDTAE/g). The PBD values of the extracts ranged from 1.01 to 2.17 mmol TE/g. EA extract displayed the highest inhibitory activity against AChE (2.32 mg GALAE/g), BChE (3.80 mg GALAE/g), α-amylase (0.56 mmol ACAE/g) and α-glucosidase (9.16 mmol ACAE/g) enzymes. Infusion extract was the most active against tyrosinase enzyme with a value of 83.33 mg KAE/g. A total of 28 compounds were identified from the different extracts. The compounds present in the highest concentration were chlorogenic acids, 4-hydroxybenzoic acid, caffeic acid, p-coumaric acid, ferulic acid, isoquercitrin, delphindin 3,5-diglucoside, kaempferol-3-glucoside and hyperoside. The biological activities of A. elongatum extracts could be due to the presence of compounds such as gallic acid, chlorogenic acids, ellagic acid, epicatechin, catechin, kaempferol, 4-hydroxybenzoic acid, caffeic acid, p-coumaric acid, ferulic acid, quercetin, isoquercitrin, and hyperoside. Extracts of A. elongatum showed promising biological activities which warrants further investigations in an endeavor to develop biopharmaceuticals.     

o-Diphenol: Oxygen oxidoreductase from leaves of sugar cane

A non-particulate o-diphenol: O₂ oxidoreductase (phenolase) has been isolated from leaves of sugar cane. Gel filtration produced two fractions MW 32000 and 130000. The preferred substrate was chlorogenic acid. Other o-diphenols (caffeic acid, catechol, pyrogallol, dihydroxyphenylalanine) all of which were slowly oxidized when tested alone, increased the rates of O₂ consumption obtained with catalytic amounts of chlorogenic acid. Both enzyme fractions were inhibited by thiols; thioglycollate, which acted in a non-competitive manner, was most effective.     

Fractional changes in phenolic acids composition in root nodules of Arachis hypogaea L.

Phenolic acids are active antimicrobial compounds and root signaling molecules that play important roles in plant defense responses. They are generally present in plants as glycosides or esters. A range of soluble and bound phenolic acids were detected in roots and root nodules of Arachis hypogaea L., among which five were identified by high performance liquid chromatography (HPLC) coupled with UV–Vis diode array detector (DAD), viz., p-coumaric acid (p-com), p-hydroxybenzaldehyde (HBAld), p-hydroxybenzoic acid (HBA), caffeic acid (CA) and protocatechuic acid (PA). Para-coumaric acid was constitutively present in all fractions whereas HBA was present in the soluble form only in young nodules. CA and PA were mostly present in the wall bound fraction. The root nodules contain higher concentration of phenolic acids than non-nodulated roots and presence of peroxidase and polyphenol oxidase indicate the metabolism of phenolic acids in roots and root nodules. These results indicate that phenolic acids (p-com and CA) in bound-glycosidic or ester forms were major components in cell wall fortification which provide protection to the root nodule from pathogen attack.     

Purification and Some Properties of Chlorogenic Acid Oxidase from Apple (Malus pumila)

Chlorogenic acid oxidase was extensively purified to homogeneity from apple flesh (Malus pumila cv. Fuji). The enzyme was purified 470-fold, with a total yield close to 70% from the plastid fraction by ammonium sulfate precipitation, gel filtration and ion-exchange chromatography. The molecular weight was determined to be 65,000 by both SDS-PAGE and gel filtration chromatography. The optimum pH for the enzyme activity was around 4.0, and the enzyme was stable in the range of pH 6–8. The pI obtained by isoelectrofocusing was 5.4, and the N-terminal amino acid sequence was N-Asp-Pro-Leu-Ala-Pro-Pro-. The reaction rate of the purified enzyme was much larger for chlorogenic acid than for other o-diphenols such as (+)-catechin, (?)-epicatechin and 4-methylcatechol, and the enzyme lacked both cresolase activity and p-diphenol oxidase activity. The K_m value for the enzyme was found to be 122μM toward chlorogenic acid. The purified enzyme had far less thermal stability than the enzyme of the plastid fraction. Diethyl-dithiocarbamate, sodium azide, o-phenanthroline and sodium fluoride markedly inhibited the enzyme activity.     

Biocatalytic properties of a peroxidase-active cell-free extract from onion solid wastes: caffeic acid oxidation

The exploitation of food residual sources consists of a major factor in reducing the polluting load of food industry wastes and developing novel added-value products. Plant food residues including trimmings and peels might contain a range of enzymes capable of transforming bio-organic molecules with potential phytotoxicity, including hydrolases, peroxidases and polyphenoloxidases. Although the use of bacterial and fungal enzymes has gained interest in studies pertaining to bioremediation applications, plant enzymes have been given less attention or even disregarded. In this view, this study aimed at the investigating the use of a crude peroxidase preparation from onion solid by-products for oxidising caffeic acid, a widespread o-diphenol, whose various derivatives may occur in food industry wastes, such as olive mill waste waters. Increased enzyme activity was observed at a pH value of 5, but considerable activity was also retained for pH up to 7. Favourable temperatures for increased activity varied between 20°C and 40°C, 30°C being the optimal. Liquid chromatography-mass spectrometry analysis of a homogenate/H₂O₂-treated caffeic acid solution revealed the existence of a tetramer as major oxidation product. Based on the data generated, a putative pathway for the formation of the peroxidase-mediated caffeic acid tetramer was proposed.     

Identification of a potent xanthine oxidase inhibitor from oxidation of caffeic acid

Inhibitory activity of Fe-ion-catalyzed radical oxidation products from 22 types of phenolic compounds toward xanthine oxidase (XO) was investigated. Phenols are readily oxidizable compounds in nature and, thus, showed potent antioxidant activities. Among the phenols screened in this study, noticeable activity was observed in the oxidation product of caffeic acid, whereas almost no XO-inhibitory activity of caffeic acid was observed. Assay-guided purification of the oxidation product of caffeic acid afforded a highly potent XO inhibitor, with an IC₅₀ value that was calculated to be 60 nmol L⁻¹, which indicated XO-inhibitory activity much stronger than that of allopurinol (IC₅₀ = 1 μmol L⁻¹), a potent XO inhibitor and excellent medicine for the treatment of gout. The chemical structure of this new XO inhibitor was investigated by one- and two-dimensional NMR and HR–ESI–MS analyses, and the unique tetracyclic structure was confirmed by synthesis starting from commercially available 1,2,4-trimethoxybenzene and 3,4-dimethoxylbenzoyl chloride.     

Particulate and soluble forms of o-diphenol oxidase from potato tubers

A soluble and two different particulate forms of o-diphenol oxidase have been obtained from aged or fresh potato slices by differential and density gradient centrifugation. The particulate enzymes were shown to sediment with microsomes and peroxisomes, respectively. Over half the enzyme activity of aged slices was found to be particle bound, with approximately twice as much enzyme in the microsomes as in the peroxisomal fraction. Very similar distribution patterns have been obtained with fresh potatoes, which have an o-diphenol oxidase activity approximately one-third that of aged slices.     

Antistaphylococcal and biofilm inhibitory activities of gallic,caffeic, and chlorogenic acids

Staphylococcus aureus is a Gram-positive pathogen which is able to form biofilms, exhibiting a more pronounced resistance to antibiotics and disinfectants. The?hurdles posed in eradicating biofilms?have driven the search for new compounds able to fight these structures. Phenolic compounds constitute one of the most numerous and ubiquitous group of plant secondary metabolites with many biological activities. The aim of the present work was to study the potential antimicrobial and antibiofilm properties of gallic, caffeic, and chlorogenic acids against S. aureus as well to elucidate its mechanism of action. It was concluded that the phenolic acids studied in this work?have antistaphylococcal properties. For instance, gallic acid is able to influence the adhesion properties of S. aureus. The phenolic acids tested were also able to inhibit the production of α-hemolysin by this microorganism, with the exception of chlorogenic acid. Regarding its mechanism of action, caffeic acid interferes with the stability of the cell membrane and with the metabolic activity of the cells of S. aureus.     

Wine phenolic compounds influence the production of volatile phenols by wine-related lactic acid bacteria

Aims: To evaluate the effect of wine phenolic compounds on the production of volatile phenols (4‐vinylphenol [4VP] and 4‐ethylphenol [4EP]) from the metabolism of p‐coumaric acid by lactic acid bacteria (LAB). Methods and Results: Lactobacillus plantarum, Lactobacillus collinoides and Pediococcus pentosaceus were grown in MRS medium supplemented with p‐coumaric acid, in the presence of different phenolic compounds: nonflavonoids (hydroxycinnamic and benzoic acids) and flavonoids (flavonols and flavanols). The inducibility of the enzymes involved in the p‐coumaric acid metabolism was studied in resting cells. The hydroxycinnamic acids tested stimulated the capacity of LAB to synthesize volatile phenols. Growth in the presence of hydroxycinnamic acids, especially caffeic acid, induced the production of 4VP by resting cells. The hydroxybenzoic acids did not significantly affect the behaviour of the studied strains. Some of the flavonoids showed an effect on the production of volatile phenols, although strongly dependent on the bacterial species. Relatively high concentrations (1 g l^?1) of tannins inhibited the synthesis of 4VP by Lact. plantarum. Conclusions: Hydroxycinnamic acids were the main compounds stimulating the production of volatile phenols by LAB. The results suggest that caffeic and ferulic acids induce the synthesis of the cinnamate decarboxylase involved in the metabolism of p‐coumaric acid. On the other hand, tannins exert an inhibitory effect. Significance and Impact of the Study: This study highlights the capacity of LAB to produce volatile phenols and that this activity is markedly influenced by the phenolic composition of the medium.     

Lack of involvement of polyphenol oxidase in ortho-hydroxylation of phenolic compounds in mung bean seedlings

Complete elimination of polyphenol oxidase activity in hypocotyls and leaves of developing mung bean ( Vigna radiata L. Wilczek cv. Berkin) seedlings by tentoxin had no effect on the content of the ortho-hydroxylated flavonoids delphinidin and rutin. Tentoxin completely eliminated polyphenol oxidase-mediated ortho-hydroxylation of p -coumaric acid to caffeic acid. Despite this, tentoxin had no effect on caffeic acid derivative contents in the seedlings. High performance liquid chromatography profiles indicated that elimination of polyphenol oxidase had no effect on either the quality or the quantity of soluble phenolic compounds, These data strongly indicate that polyphenol oxidase is not involved in metabolism of phenolic compounds in developing plant tissues.     

Antioxidant defense mechanisms of cereal aphids based on ascorbate and ascorbate peroxidase

Effects of plant o-dihydroxyphenols on ascorbate (ASA) content and ascorbate peroxidase (APOX) activity in the tissues of the grain aphid Sitobion avenae and the bird cherry-oat aphid Rhopalosiphum padi were studied. Among the aphid morphs, the highest ASA content and APOX activity were noted for larvae and the lowest for wingless apterae. When exposed to o-dihydroxyphenols, aphids of both species contained significantly lower concentrations of ASA and higher APOX activity than the controls. Among the studied compounds, caffeic acid had the strongest effect on ASA-based antioxidant responses in that caffeic acid caused a 5-fold decrease of ASA in aphid tissues. The influences of the plant o-dihydroxyphenols on antioxidant defense mechanisms within the cereal aphid species are discussed.