The antioxidant effect of tannic acid on the in vitro copper-mediated formation of free radicals

Tannic acid (TA) has well-described antimutagenic and antioxidant activities. The antioxidant activity of TA has been previously attributed to its capacity to form a complex with iron ions, interfering with the Fenton reaction [Biochim. Biophys. Acta 1472, 1999, 142]. In this work, we observed that TA inhibits, in the micromolar range, in vitro Cu(II) plus ascorbate-mediated hydroxyl radical (*OH) formation (determined as 2-deoxyribose degradation) and oxygen uptake, as well as copper-mediated ascorbate oxidation and ascorbate radical formation (quantified in EPR studies). The effect of TA against 2-deoxyribose degradation was three orders of magnitude higher than classic *OH scavengers, but was similar to several other metal chelators. Moreover, the inhibitory effectiveness of TA, by the four techniques used herein, was inversely proportional to the Cu(II) concentration in the media. These results and the observation of copper-induced changes in the UV spectra of TA are indications that the antioxidant activity of TA relates to its copper chelating ability. Thus, copper ions complexed to TA are less capable of inducing ascorbate oxidation, inhibiting the sequence of reactions that lead to 2-deoxyribose degradation. On the other hand, the efficiency of TA against 2-deoxyribose degradation declined considerably with increasing concentrations of the *OH detector molecule, 2-deoxyribose, suggesting that the copper-TA complex also possesses an *OH trapping activity.     

不同条件下荔枝多酚氧化酶活性的分析(英)

经硫酸铵盐析和ＳｅｐｈａｄｅｓＧ－１００,Ｑ－Ｓｅｐｈａｒｏｓｅ和ＰｈｅｎｙｌＳｅｐｈａｒｏｓｅ柱层析分别纯化后,得到了电泳均一的荔枝多酚氧化酶,荔枝多酚氧化酶在ＳＤＳ和ＣＡＴＢ存在下测得的酶活性略有降低,但在ＴｒｉｔｏｎＸ－１００下活性有所增加,丁酸等短链的脂肪酸对酶活性起抑制作用,而亚油酸等长链的不饱和脂肪酸则起促进作用,另外,荔枝多酚氧化酶活性还明显受ＦｅＳＯ４和ＳｎＣｌ２所抑制,但能补Ｃａ     

Synthesis and evaluation of antioxidant phenolic diaryl hydrazones as potent antiangiogenic agents in atherosclerosis

A series of bis-hydrazones derived from diaryl and diaryl ether hydroxybenzaldehyde frames 1 and 2 have been synthesized as potential antioxidant and antiangiogenic agents, two properties required to limit atherogenesis and cardiovascular events. These compounds were evaluated for their ability to neutralize free radical formation, to block endothelial cell-induced low-density lipoprotein oxidation (monitored by the formation of TBARS), an essential step in atherogenesis, and subsequent toxicity, to prevent angiogenesis evoked by low oxidized LDL concentration (monitored by the formation of capillary tubes on Matrigel) and to inhibit intracellular ROS increase involved in the angiogenic signaling. A structure/activity study has been carried out and finally allowed to select the phenolic diaryl ether hydralazine derivative 2a, sharing all these protective properties, as a promising hit for further development.     

Extraction, structural characterisation and evaluation of hydroxycinnamate esters of orchard grass (Dactylis glomerata) as substrates for polyphenol oxidase

Polyphenol oxidase (PPO) activity has been reported in orchard grass (Dactylis glomerata); however, to date, no endogenous substrates have been identified. In the present study, we report the isolation and structural elucidation of PPO substrates in this species. The free phenol fraction was extracted, separated by reverse-phase chromatography and six potential substrates, including two hydroxycinnamate esters, were identified by UV spectrometry, electrospray ionisation-tandem mass spectrometry (LC-ESI-MSⁿ) and 1D and 2D NMR analyses (¹H NMR, ¹³C NMR, DEPT, COSY, HMQC and HMBC). Furthermore, three caffeoylquinic acids (3-CQA, 4-CQA and 5-CQA) were identified by comparison of their spectral data (ESI-MS) with those of known compounds and literature data. Five of these compounds were demonstrated to be substrates for orchard grass PPO.     

Biotransformation of tea catechins into theaflavins with immobilized polyphenol oxidase

Theaflavins, an active antioxidant, a natural pigment and pharmacologically active molecule obtained from black tea were bioprocessed on an immobilized tea polyphenol oxidase (PPO) system by the conversion of tea catechins extracted from green tea leaves with an overall conversion efficiency of 85% about 14-fold increase over maximum achievable in normal black teas. The immobilized enzyme (IE) system consists of activated cellulose matrix on to which the freshly extracted tea leaf polyphenol oxidase was covalently linked. Cellulose as a matrix of choice was considered primarily for its non-toxic nature, natural origin, low cost and easy availability. The kinetic parameters of the IE system were; protein loading capacity 11.8 mg/g; pH optimum 5.9; temperature optimum 37.5 °C; K_m 4.76 ± 0.08 mM; V_max 20 ± 1.80 nmol/min; enzyme activity retention (83.58%) and number of batches per turnover without loss of activity was 14. The product from IE system was identified by HPLC and ESI-QTOF-MS spectrometry.     

Tannins in plant-herbivore interactions

Tannins are the most abundant secondary metabolites made by plants, commonly ranging from 5% to 10% dry weight of tree leaves. Tannins can defend leaves against insect herbivores by deterrence and/or toxicity. Contrary to early theories, tannins have no effect on protein digestion in insect herbivores. By contrast, in vertebrate herbivores tannins can decrease protein digestion. Tannins are especially prone to oxidize in insects with high pH guts, forming semiquinone radicals and quinones, as well as other reactive oxygen species. Tannin toxicity in insects is thought to result from the production of high levels of reactive oxygen species. Tannin structure has an important effect on biochemical activity. Ellagitannins oxidize much more readily than do gallotannins, which are more oxidatively active than most condensed tannins. The ability of insects to tolerate ingested tannins comes from a variety of biochemical and physical defenses in their guts, including surfactants, high pH, antioxidants, and a protective peritrophic envelope that lines the midgut. Most work on the ecological roles of tannins has been correlative, e.g., searching for negative associations between tannins and insect performance. A greater emphasis on manipulative experiments that control tannin levels is required to make further progress on the defensive functions of tannins. Recent advances in the use of molecular methods has permitted the production of tannin-overproducing transgenic plants and a better understanding of tannin biosynthetic pathways. Many research areas remain in need of further work, including the effects of different tannin types on different types of insects (e.g., caterpillars, grasshoppers, sap-sucking insects).     

Naturally occurring polyphenolic inhibitors of amyloid beta aggregation

Alzheimer’s disease is the most common neurodegenerative disease and is one of the main causes of death in developed countries. Consumption of foods rich in polyphenolics is strongly correlated with reduced incidence of Alzheimer’s disease. Our study has investigated the biological activity of previously untested polyphenolic compounds in preventing amyloid β aggregation. The anti-aggregatory potential of these compounds was assessed using the Thioflavin-T assay, transmission electron microscopy, dynamic light scattering and size exclusion chromatography. Two structurally related compounds, luteolin and transilitin were identified as potent inhibitors of Aβ fibril formation. Computational docking studies with an X-ray derived oligomeric structure offer a rationale for the inhibitory activity observed and may facilitate development of improved inhibitors of Aβ aggregation and toxicity.     

Cloning,expression, and characterization of miR058 and its target PPO during the development of grapevine berry stone

Polyphenol oxidases catalyzing the oxygen-dependent oxidation of phenols to quinones are ubiquitous among angiosperms. They are key enzymes playing a significant role during the synthesis of lignin. The inhibition of the synthesis of lignin in grapevine can cause seedless grapevine berry development. In this study, grapevine PPO (Vv-PPO) was predicted as the target gene of Vv-miR058 by bioinformatics analysis, and it was further cloned and its homologous conservation in various plants was analyzed. The expression profiles of miR058 and its target Vv-PPO were detected by qRT-PCR in peel, pulp and seeds of three grapevine cultivars and Vv-PPO was expressed in an opposite variation way with Vv-miR058 where both of them could be detected, suggesting that Vv-miR058 can play an important role by regulating the expression of Vv-PPO. In addition, the potential target gene Vv-PPO for Vv-miR058 was verified by RLM-RACE. This result would be helpful in theoretical basis for further research and seedless grapevine berry production.     

Stage-specific induction of systemic acquired resistance by fruit extracts of Azadirachta indica

In the present study, the efficacy of aqueous fruit extract of Azadirachta indica A. Juss. (Neem) on induction of systemic acquired resistance (SAR) in tomato against Pseudomonas syringae pv. tomato through enhancement in the activity of polyphenol oxidase (PPO) at different ages was studied. Plants at 6, 8, 10 and 12?weeks of age were selected. A single leaf at the third node from the base of each plant was treated either singly or with different combinations of the pathogen and Neem fruit extract. Samples were collected from the non-treated leaves at an interval of 24?h up to five?days for enzyme assay and after two?weeks for disease development from both treated and untreated plants. The results demonstrate that the neem fruit extract could induce additional PPO isoforms both locally as well as systemically. The PPO activity was observed to be elevated in both the treated and non-treated leaves leading to induction of SAR. The induction of SAR enhances with the increase in the age of the plant.     

Immunoaffinity Chromatography of Human β-Hexosaminidase A

A highly specific method for the purification of human 8-hexos-aminidase A employing immunoaffinity chromatography is described. Using an antiserum against the unique antigenic determinant, a, of 8-hexosaminidase A, and elution with 8.0M urea, a 283-or 1+17-fold purification of the enzyme was obtained in a single step from crude human placental homogenate.