Polyphenol metabolism of developing apple skin of a scab resistant and a susceptible apple cultivar

During fruit development, the concentration of main polyphenols (flavonols, flavanols, dihydrochalcones, hydroxycinnamic acids, anthocyanins) and the activities of related enzymes (phenylalanine ammonia lyase, chalcone synthase/chalcone isomerase, flavanone 3-hydroxylase, dihydroflavonol 4-reductase, flavonol synthase, peroxidase) were monitored in apple (Malus domestica Borkh.). The seasonal survey was performed at five different sampling dates and included the healthy peel of the resistant cultivar ‘Florina’ and healthy peel, scab symptomatic spot and the tissue around the infected spot of the susceptible cultivar ‘Golden Delicious’. From all enzymes tested, chalcone synthase/chalcone isomerase had the highest activity in both cultivars, while phenylalanine ammonia lyase had the lowest. The healthy peels of the susceptible and the resistant cultivar did not show differences in the accumulation of the main polyphenol groups present in the apple skin. However, in the resistant cultivar ‘Florina’, an increase of polyphenol enzyme activities could be observed in late stages of fruit development, which seems to be related to the anthocyanin accumulation in ripe fruits. Significant differences in the polyphenol metabolism were observed in the three different tissues of the susceptible cultivar ‘Golden Delicious’. Increased concentrations of hydroxycinnamic acids, dihydrochalcones and flavan-3-ols were found in the scab symptomatic spots and surrounding tissues. Phenylalanine ammonia-lyase, dihydroflavonol 4-reductase, flavanone 3-hydroxylase and peroxidase showed higher activities in the scab symptomatic spot compared to other analysed tissues, whereas the activities of other enzymes remained unchanged. Highest induction of polyphenol accumulation after scab infection was observed in early developmental stages, whereas enzyme activities were increased in later stages.     

Phenolic compounds in apple leaves after infection with apple scab

Leaves of the scab-susceptible apple (Malus domestica) cultivar Golden Delicious were harvested from May to August 2008 and 2009. Some leaves were healthy and some infected with fungus Venturia inaequalis. The phenolic compounds were analysed in healthy leaves, infected leaves and in the scab spot tissue. In comparison to healthy leaves, the infected leaves showed higher contents of hydroxycinnamic acid, flavanols and phloridzin, while lower contents on procyanidins, quercetins and phloretin. The total amount of phenolic compounds in the infected tissue was 10 to 20 % higher than in the healthy leaves. Accumulation of phenolic compounds is a post-infection response, and probably their further transformation is a prerequisite for plant resistance.     

Individual phenolic response and peroxidase activity in peel of differently sun-exposed apples in the period favorable for sunburn occurrence

Extreme weather events like high solar radiation can cause stress in apple fruits (Malus domestica Borkh.). The aim of the study was to make a screening of individual phenols and peroxidase activity in apple peel as a response to sunburn and different sun-exposures in the period when weather conditions are suitable for sunburn occurrence. Apple fruits of ‘Golden Delicious’ and ‘Braeburn’ were sampled. Fruit temperature and color were measured prior HPLC–MS² and peroxidase activity analyses. Sunburned peel was darker and more yellow-red in comparison to healthy peel, which appeared yellow-green. Fruit temperature, total as well as individual flavonols and dihydrochalcones, total hydroxycinnamics and perixodase activity were highest in sunburned peel in comparison with healthy sun-exposed peel, furthermore both were different than shaded sides of both fruits and peel of apples inside the tree crown; moreover in sunburned peel dihydrochalcones were determined for the first time. Chlorogenic acid was up to 2.5 times higher, 3-hydroxy-phloretin-2′-O-xyloglucoside was up to 10 times higher and quercetin-3-galactoside was up to 33 times higher in sunburned peel, comparing to shaded sided peels. Flavanols did not show a distinct pattern. A deeper insight in phenolic response against environmental stress caused by high solar radiation and high air temperatures has been made.     

Apple juice inhibits human low density lipoprotein oxidation.

Dietary phenolic compounds, ubiquitous in vegetables and fruits and their juices possess antioxidant activity that may have beneficial effects on human health. The phenolic composition of six commercial apple juices, and of the peel (RP), flesh (RF) and whole fresh Red Delicious apples (RW), was determined by high performance liquid chromatography (HPLC), and total phenols were determined by the Folin-Ciocalteau method. HPLC analysis identified and quantified several classes of phenolic compounds: cinnamates, anthocyanins, flavan-3-ols and flavonols. Phloridzin and hydroxy methyl furfural were also identified. The profile of phenolic compounds varied among the juices. The range of concentrations as a percentage of total phenolic concentration was: hydroxy methyl furfural, 4-30%; phloridzin, 22-36%; cinnamates, 25-36%; anthocyanins, n.d.; flavan-3-ols, 8-27%; flavonols, 2-10%. The phenolic profile of the Red Delicious apple extracts differed from those of the juices. The range of concentrations of phenolic classes in fresh apple extracts was: hydroxy methyl furfural, n.d.; phloridzin, 11-17%; cinnamates, 3-27%; anthocyanins, n.d.-42%; flavan-3-ols, 31-54%; flavonols, 1-10%. The ability of compounds in apple juices and extracts from fresh apple to protect LDL was assessed using an in vitro copper catalyzed human LDL oxidation system. The extent of LDL oxidation was determined as hexanal production using static headspace gas chromatography. The apple juices and extracts, tested at 5 microM gallic acid equivalents (GAE), all inhibited LDL oxidation. The inhibition by the juices ranged from 9 to 34%, and inhibition by RF, RW and RP was 21, 34 and 38%, respectively. Regression analyses revealed no significant correlation between antioxidant activity and either total phenolic concentration or any specific class of phenolics. Although the specific components in the apple juices and extracts that contributed to antioxidant activity have yet to be identified, this study found that both fresh apple and commercial apple juices inhibited copper-catalyzed LDL oxidation. The in vitro antioxidant activity of apples support the inclusion of this fruit and its juice in a healthy human diet.     

Purification and properties of polyphenoloxidase of mango peel (Mangifera indica)

Polyphenoloxidase from mango(Mangifera indica) peel was purified to homogeneity by ammonium sulphate fractionation, chromatography on DEAE-Sephadex and gel filtration of Sephadex G-200. The enzyme had an apparent molecular weight of 136,000. Its pH and temperature optimum were 5.4 and 50‡C, respectively. The enzyme possessed catecholase activity and was specific too-dihydroxy phenols. The enzyme also exhibited peroxidase activity. Some non-oxidizable phenolic compounds inhibited the enzyme competitively. High inhibitory effects were also shown by some metal chelators and reducing agents, Mango peel polyphenol oxidase when immobilized onto DEAE Sephadex showed slightly higher Km for catechol and lower pH and temperature optima.     

Phenolic acid content in potato peel determines natural infection of common scab caused by <Emphasis Type="Italic">Streptomyces</Emphasis> spp.

Common scab of potato caused by the actinomycete Streptomyces scabies is a common pathogen in almost all the potato growing areas of the world. Twenty cultivars of potato were screened in naturally scab infested farmers fields at two locations Tikari and Bachhawan, Varanasi, in two successive crop seasons (2006–2007 and 2007–2008). Among the cultivars, five cultivars were recorded to be least susceptible and the others ranged from medium susceptible to very highly susceptible. Most of the cultivars showed a stable resistance reaction in both the years. Qualitative as well as quantitative estimation of phenolic acids present in peels of the potato cultivars showed their possible role in protection of the potato cultivars against common scab. All the red skinned potato cultivars that were least susceptible to common scab infection were usually found to be rich in phenolic acid contents in their peels. This showed a positive correlation between cultivar resistance to common scab and phenolic acid content in the peel.     

Phenolic Compounds of Apple and their Relationship to Scab Resistance

The contents of 15 different phenolic compounds in apple leaves and fruit skin from various orchards were determined by HPLC. High concentrations of flavan-3-ols were found in those orchards which were not infected by Venturia inaequalis . This observation corroborates earlier findings on different cultivars. The hypothesis that these compounds could be involved in the resistance of apple to scab was tested by inhibiting the key enzyme of the phenol biosynthesis, the phenylalanine-ammonia-lyase, in young shoots of the resistant genotype 'Sir Prize'. After inoculation of the inhibitor-treated leaves, severe symptoms of the disease occurred. The fungal infection was confirmed by histological studies. No flavonal-accumulation could be observed at the infection site which was the case in the non-inhibited but inoculated controls.     

Antifungal phenolic acids in apple fruits after infection with Sclerotinia fructigena

Several compounds were separated from the antifungal materials produced in Edward VII apple fruits attacked by the brown rot organism Sclerotinia fructigena. Six phenolic compounds were isolated in the crude state by column chromatography and their fungicidal properties examined. Two of the phenolic acids present were purified and identified as 4-hydroxybenzoic acid and 4-hydroxy-3-methoxybenzoic acid. These phenolic compounds were shown to arise from the action of the pathogen on the juice of the fruit and not from the peel or the juice-free pulp.     

Variation and genetic parameters of fruit colour and polyphenol composition in an apple seedling population segregating for red leaf

Red flesh colour is a relatively new target for apple breeding programmes and understanding genetic relationships between this trait and other fruit characters, including polyphenol compounds, is important for both breeders and marketers of new red flesh cultivars. In this study, fruit peel and flesh colours and concentrations of up to 20 individual fruit polyphenols within each tissue were examined in fruit harvested from a 14-family apple seedling population segregating for red and green leaf. Red leaf seedlings always produced red flesh fruit that varied from pale red to complete dark red cortical tissue (type 1 red flesh). Some (20 %) of green leaf seedlings also produced fruit with red flesh, albeit at low intensity (type 2 red flesh). Cyanidin 3-O-galactoside was the dominant anthocyanin in both fruit tissues, with concentrations being 1,900 times higher in the flesh and 2.5 times higher in the peel of fruit from red than from green leaf seedlings. Red leaf seedlings also had 59 % more flesh epicatechin and 17 % less total peel flavonols, but other polyphenols were not associated with leaf colour. Heritability estimates for red flesh colour, flesh cyanidin 3-O-galactoside, flesh and peel catechins were high in red leaf and low in green leaf seedlings. Conversely, estimates for red peel coverage and two peel anthocyanins were higher in green compared to those from red leaf seedlings. Other than these, heritability estimates were high only for dihydrochalcones and hydroxycinnamic acids from each tissue for both leaf colours but low for all other flesh and peel flavan-3-ols, procyanidins and most peel flavonols irrespective of leaf colour. Genetic correlations between polyphenol compounds varied considerably, but were broadly similar for red and green leaf seedlings. Genetic correlations were mostly moderate to high between compounds of the same metabolic group, but low between compounds from different groups. These results are discussed in relation to the genetic control of flesh colour and polyphenol accumulation in apple, as well as to implications for breeding red flesh apples with altered polyphenol composition.     

Bioactive Compounds of Fruit Parts of Three Eugenia uniflora Biotypes in Four Ripening Stages

Variability of secondary metabolites in edible (peel and pulp) and inedible (seeds) parts of three pitanga varieties, red, red-orange and purple, was investigated during the maturation process. Hydrolysable tannins, anthocyanins, and flavonoids were quantified by HPLC/DAD and carotenoids by absorbance. Peel/pulp showed greater complexity of constituents (carotenoids, anthocyanins, flavonoids, and hydrolysable tannins), while only tannins were identified in seeds, but in quantities of 10 to 100 times greater. The red-orange variety showed the highest levels of phenolic compounds in seeds and peel/pulp, except anthocyanins. The analysis of the principal response curves showed that the pitanga biotype has greater influence on metabolite variation than ripening stages. During peel/pulp maturation, a reduction in the levels of flavonoids and tannins contrasted with an increase in carotenoids and cyanidin-3-O-glucoside in all varieties, whereas in the seeds oenothein B, the major tannin, increased up to 1.32 g/100 g fresh weight. Such marked differences between fruit parts demonstrate that the seeds in stages E3 and E4 are a source of hydrolysable tannins, compounds known for their antitumor activity, while peel/pulp of all varieties in the ripe stage provide natural antioxidants, such as carotenoids and flavonoids. Lastly, the purple biotype can be a rich source of the cyanidin-3-O-glucoside pigment a potent bioactive compound.     

Synergism and phytotoxicity: The effects of tank-mix additives on the biological efficacy of Cu2+ against Venturia inaequalis and Podosphaera leucotricha

The wetting behaviour of the spray and the biological efficacy of Cu²⁺ active ingredients in agrochemical formulations may be enhanced by tank-mix additives. We investigated how three BREAK-THRU® additives (BT301: biodegradable, BT133 and BT420: bio-based and biodegradable) tank-mixed with commercial copper preparations influenced the spray distribution, leaf uptake and biological efficacy of copper additive mixtures against apple scab and apple powdery mildew under controlled conditions. We quantified the synergetic effects of these additives in foliar applications. In addition, we determined the phytotoxic potential and evaluated their impacts on photosynthetic activity, non-photochemical quenching and ROS activity. The additives BT301 and BT420 strongly reduced surface tension and contact angle of copper treatments. The fluorescence observations revealed that BT301 achieved better spreading of copper formulation with more complete coverage of the leaf surface than BT420 and BT133, whereas ‘coffee-ring’ spreading was observed with BT133. The additive BT301 showed an increase in relative fluorescence area, indicating higher ROS production as a signal of intra-cellular tissue activity. The photochemical efficiency of photosystem II (Fv/Fm) was not negatively influenced by copper or additive treatment. Thus, we observed no phytotoxic effects of copper-additive mixtures on apple leaves at treatment doses of 4 g Cu²⁺ L⁻¹. All copper treatments reduced apple scab infections significantly, by 53%–76%. Interestingly, addition of BT301 to copper preparations showed the strongest biological efficacy (83% reduction) against Venturia inaequalis, whereas addition of BT420 showed the strongest effect against Podosphaera leucotricha (89% infection reduction). The synergetic effects of additives on the biological efficacy without phytotoxic effects on plants may have potential for reducing copper loads in horticultural production systems.     

Effects of nitrogen supply on growth, contents of phenolic compounds and pathogen (scab) resistance of apple trees

The effect of long-term N-supply on growth, scab resistance and phenolic compounds in the leaves of two apple cultivars was studied. The different pools of phenylpropanoids (hydroxycinnamic acids, dihydrochalcones) and flavonoids (flavonols, catechins, procyanidins) were quanitfied by HPLC from non-infected and inoculated leaves representing different ontogenetic stages. Scab incidence was also evaluated. Strictly following the carbon-nutrient-balance hypothesis, apple trees responded to high N-supply with increased shoot growth and with a reduced accumulation of total phenolic compounds in their leaves. This was shown for the cultivar 'Golden Delicious', which is susceptible to the scab disease caused by Venturia inaequalis , and for the resistant cultivar 'Rewena'. Whereas high N-fertilization increased the susceptibility of 'Golden Delicious', it did not decrease the resistance of 'Rewena' despite of the pronounced reduction of phenolic concentrations. Thus, a simple C trade off between growth-related metabolism and secondary metabolism cannot solely explain changes in defensive potential.     

Quantification of polyphenolic compounds and relative gene expression studies of phenylpropanoid pathway in apple (Malus domestica Borkh) in response to Venturia inaequalis infection

Apple (Malus domestica Borkh) is rich in phenolic compounds, which may enhance resistance to scab disease caused by Venturia inaequalis. In present study, apple cv. Golden Delicious was used for estimation of total phenols, flavonoids and quantification of six individual phenolic compounds. between control vs inoculated samples at different inoculation stages. The relative gene expression of phenylalanine ammonia lyase, chalcone synthase and flavanone 3 hydroxylase increased and polyphenolic compounds were constitutively upregulated at different post-inoculation stages. Data suggest that synthesis and accumulation of polyphenols is closely related with disease resistance against Venturia inaequalis. This study may play a vital role in understanding and finding out the governing mechanisms of scab resistance.     

Eradication of the perithecial stage of apple scab with surfactants

Clone M3 apple leaves infected with scab (Venturia inaequalis) were dipped in one of a series of surface-active compounds or mixtures tested at a concentration of 5% a.i. Seven of these reduced the concentration of scab ascopores released in the spring by more than 99%.     

Molecular selection in apple for resistance to scab caused by Venturia inaequalis

Large-scale marker-assisted selection requires highly reproducible, consistent and simple markers. The use of genetic markers is important in woody plant breeding in general, and in apple in particular, because of the high level of heterozygosity present in Malus species. We present here the transformation of two RAPD markers, which we found previously to be linked to the major scab resistance gene Vf, into more reliable and reproducible markers that can be applied directly to apple breeding. We give an example of how the use of such markers can speed up selection for the introduction of scab resistance genes into the same plant, reducing labour and avoiding time-consuming test crosses. We discuss the nature and relationship of the scab resistance gene Vf to the one present in Nova Easygro, thought to be Vr.     

Acibenzolar-S-methyl-Induced Systemic Resistance against Anthracnose and Powdery Mildew Diseases on Cucumber Plants without Accumulation of Phytoalexins

In a study to elucidate the possible involvement of phytoalexins in acibenzolar-S-methyl (ASM)-induced systemic resistance in cucumber plants ( Cucumis sativus L.), the phenolic compounds were extracted from ASM-treated and inoculated plants and compared with those from Milsana^®-treated plants previously reported to accumulate phytoalexins in cucumber. The glycoside-linked phenolic compounds from cucumber leaves were tested for their antifungal activity to the growth of pathogens which were most effective against the cucumber anthracnose fungus Colletotrichum orbiculare , followed by the scab fungus Cladosporium cucumerinum but ineffective against the Corynespora leaf spot fungus Corynespora cassiicola . Nevertheless, the accumulations of active compounds appeared to increase with the growth stages of cucumber plants irrespective of ASM treatment. In ASM-pretreated cucumber plants either inoculated with anthracnose or the powdery mildew fungus, there was no increase in phytoalexin-like phenolics.     

High-pressure processing of apple juice: kinetics of pectin methyl esterase inactivation

High-pressure (HP) inactivation kinetics of pectin methyl esterase (PME) in apple juice were evaluated. Commercial PME was dispensed in clarified apple juice, sealed in dual peel sterilizable plastic bags, and subjected to different high-pressure processing conditions (200-400 MPa, 0-180 min). Residual enzyme activity was determined by a titration method estimating the rate of free carboxyl group released by the enzyme acting on pectin substrate at pH 7.5 (30 degrees C). The effects of pressure level and pressure holding time on enzyme inactivation were significant (p < 0.05). PME from the microbial source was found to be more resistant (p < 0.05) to pressure inactivation than PME from the orange peel. Almost a full decimal reduction in the activity of commercial PME was achieved by HP treatment at 400 MPa for 25 min. Inactivation kinetics were evaluated on the basis of a dual effect model involving a pressure pulse effect and a first-order rate model, and the pressure sensitivity of rate constants was modeled by using the z-value concept.     

<Emphasis Type="Italic">Vr</Emphasis><Subscript><Emphasis Type="Italic">2</Emphasis></Subscript>: a new apple scab resistance gene

Reports from several European countries of the breakdown of the Vf resistance, the most frequently used source of resistance in breeding programs against apple scab, emphasize the urgency of diversifying the basis of apple scab resistance and pyramiding different apple scab resistances with the use of their associated molecular markers. GMAL 2473 is an apple scab resistant selection thought to carry the resistance gene Vr. We report the identification by BSA of three AFLP markers and one RAPD marker associated with the GMAL 2473 resistance gene. SSRs associated with the resistance gene were found by (1) identifying the linkage group carrying the apple scab resistance and (2) testing the SSRs previously mapped in the same region. One such SSR, CH02c02a, mapped on linkage group 2, co-segregates with the resistance gene. GMAL 2473 was tested with molecular markers associated with other apple scab resistance genes, and accessions carrying known apple scab resistance genes were tested with the SSR linked to the resistance gene found in GMAL 2473. The results indicate that GMAL 2473 does not carry Vr, and that a new apple scab resistance gene, named Vr ₂, has been identified.     

Potential of phenolic compounds from pomegranate (Punica granatum L.) by-product with significant antioxidant and therapeutic effects: A narrative review

The wealth of epidemiological evidence in the scientific world underscores the possibility that a plant-based diet can reduce the prevalence of common diseases such as diabetes, cardiovascular disease, cancer, and stroke. The therapeutic effects of plant sources are partly explained by phenolic secondary metabolites or polyphenolic compounds. Therefore, polyphenolic compounds, which are widely distributed in plants, are of great interest for the development of effective specific drugs with antioxidant and anti-inflammatory effects. Moreover, polyphenol compounds have no harmful effects due to their natural biocompatibility and safety. Numerous studies have highlighted the potential of some industrial food wastes from plant material processing, including apple peels and mashed potatoes, grape skins, tomato and carrot peels, pomegranate peels and seeds, and many others. These byproducts are considered low-cost sources of natural biological compounds, including antioxidants, which have beneficial effects on human health.The polyphenol complex of pomegranate peel (Punica granatum L.), which makes up half of the pomegranate fruit, has more pronounced antioxidant and anti-inflammatory properties than other parts. And the most important active components of pomegranate peel, which are found only in this plant, are punicalagin, followed by ellagic acid and gallic acid. It is known that these polyphenolic compounds of pomegranate peel have the most pronounced therapeutic effect. Several studies have shown the protective effect of ellagic acid, punicalagin, against oxidative stress damage caused by free radicals. The potential of pomegranate peel as an antioxidant and therapeutic component in various biological systems is high, according to scientific sources.However, despite extensive research in recent years, a review of sources has shown that there is insufficient evidence to support the therapeutic effects of polyphenolic compounds from pomegranate peels. The role of pomegranate peel polyphenolic compounds, including flavonoids, as antioxidants in various biological systems also requires further research. Of particular importance are the mechanisms by which antioxidants influence the cellular response against oxidative stress. The purpose of this review was to report our current knowledge of plant polyphenolic compounds and their classification, and to evaluate the potential of phenolic compounds from pomegranate peels with significant antioxidant and therapeutic effects.     

Genome mapping of an apple scab,a powdery mildew and a woolly apple aphid resistance gene from open-pollinated Mildew Immune Selection

Apple is host to a wide range of pests and diseases, with several of these, such as apple scab, powdery mildew and woolly apple aphid, being major causes of damage in most areas around the world. Resistance breeding is an effective way of controlling pests and diseases, provided that the resistance is durable. As the gene pyramiding strategy for increasing durability requires a sufficient supply of resistance genes with different modes of action, the identification and mapping of new resistance genes is an ongoing process in breeding. In this paper, we describe the mapping of an apple scab, a powdery mildew and a woolly apple aphid gene from progeny of open-pollinated mildew immune selection. The scab resistance gene Rvi16 was identified in progeny 93.051 G07-098 and mapped to linkage group 3 of apple. The mildew and woolly aphid genes were identified in accession 93.051 G02-054. The woolly aphid resistance gene Er4 mapped to linkage group 7 to a region close to where previously the genes Sd1 and Sd2, for resistance to the rosy apple leaf-curling aphid, had been mapped. The mildew resistance gene Pl-m mapped to the same region on linkage group 11 where Pl2 had been mapped previously. Flanking markers useful for marker-assisted selection have been identified for each gene.