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In recent years, red‐fleshed apple has attracted a lot of attention due to its pleasant appearance, taste, and being a valuable source of anthocyanins. Generally, different types of this apple are known as ornamental apple plants that are rarely found in various regions around the world like New Zealand, Kazakhstan, Kyrgyzstan, Australia, China, and Turkey. This study focused on the accumulation and changes of the bioactive and aroma‐related volatile components at different production stages of a red‐fleshed apple vinegar. The aim of this study is to produce an anthocyanin‐ and vitamin C‐rich vinegar (NAV) from red‐fleshed Niedzwetzky's apple for the first time. According to the results, in this vinegar, it was determined high concentrations of total phenolic content (904.8 mg‐GAE/mL), total flavonoid content (0.25 mg‐EGCE/mL), vitamin C content (3024.3 mg‐AAE/mL), and 1,1‐diphenyl‐2‐picrylhydrazyl (8.33 mmol‐TE/mL), as well as FRAP (13.57 mmol‐ISE/mL), respectively. Moreover, total anthocyanin content value was found to be 6.46 mg/g. One of the components that provide the specified functional effect in this vinegar is chlorogenic acid, which constitutes the main phenolic compound, and the other one is cyanidin‐3‐glucoside, which is the major anthocyanin in this vinegar. The main volatile components associated with the aroma of vinegar are ‐acetic acid, ‐phenylacetic acid (5067.7 µg/100 mL), ‐phenethyl alcohol (3096.1 µg/100 mL), and ‐nonanoic acid (2939.0 µg/100 mL) compounds. Consequently, it is recommended to expand the range of Niedzwetzky's apples in the production and consumption of food products such as functional vinegar. Thus, a new vinegar with high functional properties will be introduced to the worldwide food industry. Practical Application: In this study, vinegar was produced from the small and dark‐red‐flesh‐colored fruits of Niedzwetzky's apple tree, which is known in a small part of the world and is generally used as an ornamental plant due to its red‐pink flowers. It has been determined that the produced vinegar has high bioavailability due to the apples with red inner flesh and high anthocyanin content and has a desirable aromatic content. Thus, a food product with high functional quality and desirable aroma, accessible to consumers all over the world, has been produced from a little‐known fruit. [ABSTRACT FROM AUTHOR]

MYB transcription factors have been linked to anthocyanin synthesis and various color phenotypes in plants. In apple, MYB10 confers a red‐flesh phenotype due to a minisatellite insertion in its R6 promoter, but R6:MYB10 genotypes exhibit various degrees of red pigmentation in the flesh, suggesting the involvement of other genetic factors. Here, it is shown that MdWRKY10, a transcription factor identified via DNA pull‐down trapping, binds to the promoter of MdMYB10 and activates its transcription. MdWRKY10 specifically interacts with the WDR protein MdTTG1 to join the apple MYB‐bHLH‐WDR (MBW) complex, which significantly enhances its transcriptional activation activity. A 163‐bp InDel detected in the promoter region of the alleles of MdWRKY10 in a hybrid population of identical heterozygous genotypes regarding R6 by structural variation analysis, contains a typical W‐box element that MdWRKY10 binds to for transactivation. This leads to increased transcript levels of MdWRKY10 and MdMYB10 and enhanced anthocyanin synthesis in the flesh, largely accounting for the various degrees of flesh red pigmentation in the R6 background. These findings reveal a novel regulatory role of the WRKY‐containing protein complex in the formation of red flesh apple phenotypes and provide broader insights into the molecular mechanism governing anthocyanin synthesis in plants. [ABSTRACT FROM AUTHOR]

Abstract MYB transcription factors have been linked to anthocyanin synthesis and various color phenotypes in plants. In apple, MYB10 confers a red‐flesh phenotype due to a minisatellite insertion in its R6 promoter, but R6:MYB10 genotypes exhibit various degrees of red pigmentation in the flesh, suggesting the involvement of other genetic factors. Here, it is shown that MdWRKY10, a transcription factor identified via DNA pull‐down trapping, binds to the promoter of MdMYB10 and activates its transcription. MdWRKY10 specifically interacts with the WDR protein MdTTG1 to join the apple MYB‐bHLH‐WDR (MBW) complex, which significantly enhances its transcriptional activation activity. A 163‐bp InDel detected in the promoter region of the alleles of MdWRKY10 in a hybrid population of identical heterozygous genotypes regarding R6 by structural variation analysis, contains a typical W‐box element that MdWRKY10 binds to for transactivation. This leads to increased transcript levels of MdWRKY10 and MdMYB10 and enhanced anthocyanin synthesis in the flesh, largely accounting for the various degrees of flesh red pigmentation in the R6 background. These findings reveal a novel regulatory role of the WRKY‐containing protein complex in the formation of red flesh apple phenotypes and provide broader insights into the molecular mechanism governing anthocyanin synthesis in plants.

Cork spot-like physiological disorder (CSPD) is a newly identified issue in 'Kurenainoyume' apples, yet its mechanism remains unclear. To investigate CSPD, we conducted morphological observations on 'Kurenainoyume' apples with and without pre-harvest fruit-bagging treatment using light-impermeable paper bags. Non-bagged fruit developed CSPD in mid-August, while no CSPD symptoms were observed in bagged fruit. The bagging treatment significantly reduced the proportion of opened lenticels, with only 17.9% in bagged fruit compared to 52.0% in non-bagged fruits. In non-bagged fruit, CSPD spots tended to increase from the lenticels, growing in size during fruit development. The cuticular thickness and cross-sectional area of fresh cells in CSPD spots were approximately 16 µm and 1600 µm², respectively. Healthy non-bagged fruit reached these values around 100 to 115 days after full bloom from mid- to late August. Microscopic and computerized tomography scanning observations revealed that many CSPD spots developed at the tips of vascular bundles. Therefore, CSPD initiation between opened lenticels and vascular bundle tips may be influenced by water stress, which is potentially caused by water loss, leading to cell death and the formation of CSPD spots. [ABSTRACT FROM AUTHOR]

Coloration, a major factor contributing to apple quality, attracts consumer attraction, thereby increasing the sale of fruits in the market. Among the fruit pigments, anthocyanins and carotenoids are the most important pigments that impart coloration to apples. Pigment accumulation, which is regulated by transcriptional factors in the anthocyanin and carotenoid biosynthesis pathways, varies according to genetics, transcription, and developmental stage. To understand the regulation of color development in apples, we evaluated the expression of the genes in the anthocyanin and carotenoid biosynthetic pathways and analyzed the accumulation of pigment, including anthocyanin and carotenoid in the skin and flesh of the red-fleshed apple 'Enbu' cultivar at different fruit development stages (30, 60, 90, 120, and 150 days after full bloom, and ripe). Color development increased according to the developmental stage, with considerable variation in both the transcript levels and pigment concentrations observed in tissues, skin, and flesh. Moreover, we identified key transcription factors among the anthocyanin and carotenoid biosynthesis genes that regulated pigment accumulation. Pearson's correlation analysis showed a strong correlation between the coloration patterns and the expression levels of anthocyanin biosynthesis-related genes (MdPAL, MdCHI, and MdF3H) and carotenoid biosynthesis-related genes (MdGGPPS, MdPSY, MdZDS, MdCRTISO, MdCRHβ, and MdZEP). This study provides insight into the molecular mechanisms underlying pigment biosynthesis for breeding high-quality red-fleshed apple varieties to cater to consumer attention and preference. [ABSTRACT FROM AUTHOR]

In this research, we analyzed the protective effects of red-fleshed apple flavonoid extracts (RAFEs) on male reproductive injury induced by busulfan, using both in vitro and in vivo models. In the cell-based experiments, RAFEs significantly improved cell viability and proliferation rates compared to control groups. Similarly, in vivo testing with male mice showed that RAFEs and whole apple flavonoid extracts (WAFEs) enhanced various biochemical and liver function-related indicators in the testes; however, RAFEs demonstrated superior efficacy in mitigating testicular damage. Through immunohistochemistry, qRT-PCR, and Western blotting, we found that RAFEs notably enhanced the expression of spermatogenesis-related genes. Moreover, RAFEs increased the expression of oxidative stress- and apoptosis-related genes, thereby effectively reducing oxidative damage in the testes. These findings highlight the potential of RAFEs as natural agents for the prevention and treatment of male reproductive injury, paving the way for future research and potential therapeutic applications. [ABSTRACT FROM AUTHOR]

SUMMARY: Anthocyanins are important secondary metabolites in plants. They are important for human health because of their antioxidant activities and because their dietary intake reduces the incidence of cardiovascular and cerebrovascular diseases and tumors. The biosynthesis of anthocyanins and its regulation in fruits and vegetables is a global research hotspot. Compared with cultivated apples, the red‐fleshed apple is a relatively new and popular commodity in the market. Previous studies on red‐fleshed apples have focused on the basis for the high anthocyanin content and the transcriptional regulation of anthocyanin synthesis. In the present study, we focused on the mechanism of microRNA‐mediated post‐transcriptional regulation of anthocyanin synthesis in red‐fleshed apples. We identified a microRNA (miRNA), designated mdm‐miR858, that is specifically expressed in the flesh of apple fruit. The expression level of miR858 was significantly lower in red‐fleshed apples than in white‐fleshed apples. The overexpression of mdm‐miR858 significantly inhibited anthocyanin accumulation, whereas the silencing of mdm‐miR858 promoted anthocyanin synthesis in STTM858 transgenic apple calli. Further analyses showed that mdm‐miR858 targets the transcription factor genes MdMYB9 and MdMYBPA1 to participate anthocyanin accumulation in apple. Our results also show that MdHY5, a transcription factor in the light signaling pathway, can bind to the promoter of mdm‐miR858 to inhibit its transcription, thereby regulating anthocyanin synthesis. Based on our results, we describe a novel HY5‐miR858‐MYB loop involved in the modulation of anthocyanin biosynthesis. These findings provide new information about how plant miRNAs regulate anthocyanin anabolism and provide a basis for breeding new anthocyanin‐rich, red‐fleshed apple varieties. Significance Statement: Compared with cultivated apples, red‐fleshed apples have higher anthocyanin contents. The miRNA mdm‐miR858 is specifically expressed in the flesh of apples and targets two transcription factor genes, MdMYB9 and MdMYBPA1, to participate anthocyanin accumulation; further, a novel HY5‐miR858‐MYB loop participates in regulating anthocyanin synthesis in apple in response to light. [ABSTRACT FROM AUTHOR]

SUMMARY: Sugar and anthocyanin are important indicators of fruit quality, and understanding the mechanism underlying their accumulation is essential for breeding high‐quality fruit. We identified an R2R3‐MYB transcription factor MdMYB305 in the red‐fleshed apple progeny, which was positively correlated with fruit sugar content but negatively correlated with anthocyanin content. Transient injection, stable expression [overexpressing and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9)], and heterologous transformation of tomato confirmed that MdMYB305 promotes the accumulation of sugar and inhibits the synthesis of anthocyanin. A series of molecular experiments (such as electrophoretic mobility shift and luciferase assays) confirmed that MdMYB305 combines with sugar‐related genes (MdCWI1/MdVGT3/MdTMT2) and anthocyanin‐related genes (MdF3H/MdDFR/MdUFGT), promoting and inhibiting their activities, and finally regulating the sugar and anthocyanin content of fruits. In addition, the study also found that MdMYB305 competes with MdMYB10 for the MdbHLH33 binding site to balance sugar and anthocyanin accumulation in the fruits, which provides a reference value for exploring more functions of the MYB‐bHLH‐MYB complex and the balance relationship between sugar and anthocyanin in the future. Significance Statement: We found and confirmed that the R2R3‐MYB transcription factor MdMYB305 was positively correlated with sugar content and negatively correlated with anthocyanin content. In addition, we found that MdMYB305 competes with MdMYB10 for the MdbHLH33 binding site to balance sugar and anthocyanin accumulation in the fruits. [ABSTRACT FROM AUTHOR]

The genetic link between apple red flesh (RF) coloration and the internal flesh browning disorder (FBD) is a major challenge when breeding high fruit quality RF apple cultivars. A genome-wide association study (GWAS) was conducted in a population of about 900 red-leaved seedlings to identify genomic regions and putative candidate genes using whole genome sequencing of the pools of extreme phenotypes (XP) for the RF colour coverage (using the weighted cortex index (WCI)) and FBD. This study identified novel genomic regions contributing to WCI and FBD variation in the red-leaved seedlings. The FBD-associated regions were enriched for genes regulating senescence, heat shock proteins, cytochrome P450, ascorbate metabolism and pectin methyl esterases. Although there were no significant regions in common for WCI and FBD, there were several genes (e.g. MYB85, MYB66, ethylene insensitive 3, DNAJ heat shock protein, WRKY7, and NAC42) enriched commonly between the genomic regions associated with these traits, potentially underpinning the genetic connection between WCI and FBD. Some of the differentially expressed genes between the R6:MdMYB10 and white-fleshed ‘control’ apples resided within the GWAS hotspot for WCI (e.g. chalcone synthase, UDP-Glycosyl transferase) and FBD (e.g. Rho GTPase activating protein, lipoxygenase 1, phytoene synthase) – validating the XP-GWAS findings. Paralogs of several genes resided in the trait-associated genomic regions, suggesting that whole genome duplication plays an important role in the regulation of these traits. Adverse genetic correlations between WCI and sensory traits were observed, and strategies to develop FBD-free high fruit quality RF cultivars are discussed.

In recent years, the global incidence of liver damage has increased. Despite the many known health benefits of red-fleshed apple flavonoids, their potential liver-protective effects have not yet been investigated. In this study, we analyzed the composition of red-fleshed apple flavonoid extract (RAFE) by high-performance liquid chromatography (HPLC). We then induced liver damage in mice with carbon tetrachloride (CCl4) and performed interventions with RAFE to analyze its effect on liver damage, using bifendate as a positive control. The results showed that catechin was the most abundant flavonoid in ‘XJ4’ RAFE (49.346 mg/100 g). In liver-injured mice, the liver coefficients converged to normal levels following RAFE intervention. Moreover, RAFE significantly reduced the enzymatic activity levels of glutamic oxaloacetic transaminase (ALT), glutamic alanine transaminase (AST), and alkaline phosphatase (ALP) in mouse serum. Furthermore, RAFE significantly increased the content or enzyme activity level of total glutathione, total antioxidant capacity, and superoxide dismutase, and significantly decreased the content of malondialdehyde in the liver of mice. In parallel, we performed histopathological observations of mouse livers for each group. The results showed that RAFE restored the pathological changes caused by CCl4 around the central hepatic vein in mice and resulted in tightly bound hepatocytes. The recovery effect of RAFE was dose-dependent in the liver tissue. Regarding intestinal microorganisms, we found that RAFE restored the microbial diversity in liver-injured mice, with a similar microbial composition in the RAFE intervention group and normal group. RAFE reduced the ratio of Firmicutes to Bacteroidetes, increased the levels of probiotic bacteria, such as Lactobacillus acidophilus, and Clostridium, and reduced the levels of harmful bacteria, such as Erysipelothrix Rosenbach. Therefore, RAFE ameliorated CCl4-induced liver damage by modulating the abundance and composition of intestinal microorganisms in mice. In conclusion, RAFE alleviated CCl4-induced liver damage in mice, with H-RAFE (5 mg kg–1) significantly improving liver damage in mice but M-RAFE (1 mg kg–1) significantly improving the imbalance of intestinal microorganisms in mice. Our research suggests that RAFE could be employed for the adjuvant treatment and prevention of liver damage, and may have important applications in food and medicine.

Coloration, a major factor contributing to apple quality, attracts consumer attraction, thereby increasing the sale of fruits in the market. Among the fruit pigments, anthocyanins and carotenoids are the most important pigments that impart coloration to apples. Pigment accumulation, which is regulated by transcriptional factors in the anthocyanin and carotenoid biosynthesis pathways, varies according to genetics, transcription, and developmental stage. To understand the regulation of color development in apples, we evaluated the expression of the genes in the anthocyanin and carotenoid biosynthetic pathways and analyzed the accumulation of pigment, including anthocyanin and carotenoid in the skin and flesh of the red-fleshed apple ‘Enbu’ cultivar at different fruit development stages (30, 60, 90, 120, and 150 days after full bloom, and ripe). Color development increased according to the developmental stage, with considerable variation in both the transcript levels and pigment concentrations observed in tissues, skin, and flesh. Moreover, we identified key transcription factors among the anthocyanin and carotenoid biosynthesis genes that regulated pigment accumulation. Pearson’s correlation analysis showed a strong correlation between the coloration patterns and the expression levels of anthocyanin biosynthesis-related genes (MdPAL, MdCHI, and MdF3H) and carotenoid biosynthesis-related genes (MdGGPPS, MdPSY, MdZDS, MdCRTISO, MdCRHβ, and MdZEP). This study provides insight into the molecular mechanisms underlying pigment biosynthesis for breeding high-quality red-fleshed apple varieties to cater to consumer attention and preference.

The mitogen-activated protein kinase (MAPK) signaling cascade is a widely existing signal transduction system in eukaryotes, and plays an important role in the signal transduction processes of plant cells in response to environmental stress. In this study, we screened MdMKK9, a gene in the MAPK family. This gene is directly related to changes in anthocyanin synthesis in the 'Daihong' variety of red-fleshed apple (Malus sieversii f neidzwetzkyana (Dieck) Langenf). MdMKK9 expression was up-regulated in 'Daihong' tissue culture seedlings cultured at low levels of nitrogen. This change in gene expression up-regulated the expression of genes related to anthocyanin synthesis and nitrogen transport, thus promoting anthocyanin synthesis and causing the tissue culture seedlings to appear red in color. To elucidate the function of MdMKK9, we used the CRISPR/Cas9 system to construct a gene editing vector for MdMKK9 and successfully introduced it into the calli of the 'Orin' apple. The MdMKK9 deletion mutants (MUT) calli could not respond to the low level of nitrogen signal, the expression level of anthocyanin synthesis-related genes was down-regulated, and the anthocyanin content was lower than that of the wild type (WT). In contrast, the MdMKK9-overexpressed calli up-regulated the expression level of anthocyanin synthesis-related genes and increased anthocyanin content, and appeared red in conditions of low level of nitrogen or nitrogen deficiency. These results show that MdMKK9 plays a role in the adaptation of red-fleshed apple to low levels of nitrogen by regulating the nitrogen status and anthocyanin accumulation. [ABSTRACT FROM AUTHOR]

[Display omitted] • Young, red-fleshed apples (RFA) are rich in bioactive polyphenolic compounds. • Natural deep eutectic solvents (NDES) efficiently extract polyphenolic compounds. • Optimised NDES conditions enhance the yield of phenolics from young RFA. • Phenolic content and bioactivities of NDES and methanol extracts are similar. Red-fleshed apple cultivars with an enhanced content of polyphenolic compounds have attracted increasing interest due to their promising health benefits. Here, we have analysed the polyphenolic content of young, red-fleshed apples (RFA) and optimised extraction conditions of phenolics by utilising natural deep eutectic solvents (NDES). We also compare the antioxidant, neuroprotective and antimicrobial activities of NDES- and methanol-extracted phenolics from young RFA. High-performance liquid chromatography coupled to high-resolution mass spectrometry (HPLC-HRMS) was used for phenolics identification and quantification. Besides young RFA, ripe red-fleshed, young and ripe white-fleshed apples were analysed, revealing that young RFA possess the highest phenolic content (2078.4 ± 4.0 mg gallic acid equivalent/100 g), and that ripe white-fleshed apples contain the least amount of phenolics (545.0 ± 32.0 mg gallic acid equivalent/100 g). The NDES choline chloride-glycerol containing 40 % w/w H 2 O gave similar yields at 40 °C as methanol. In addition, the polyphenolics profile, and bioactivities of the NDES extract from young RFA were comparable that of methanol extracts. Altogether, our data show that NDES extracts of young RFA are a promising source of bioactive polyphenolics with potential applications in diverse sectors, e.g., for functional food production, smart material engineering and natural therapies. [ABSTRACT FROM AUTHOR]

Anthocyanin and proanthocyanidin play important roles in plant secondary metabolism. Although previous studies identified many transcription factors involved in anthocyanin and proanthocyanidin synthesis, the effects of MADS-box transcription factors are unclear in apple. Brassinosteroids (BRs) are steroid hormones that affect plant flavonoid biosynthesis, but the underlying regulatory mechanism is not yet well established. In this study, we identified a MADS-box transcription factor, MdJa2, which contained a highly conserved MADS-box domain and belonged to the STMADS11 subfamily. Additionally, MdJa2 was responsive to BR signal, and the overexpression of MdJa2 inhibited the synthesis of anthocyanin and proanthocyanidin. The silencing of MdJa2 in "Orin" calli promoted anthocyanin and proanthocyanidin accumulations. Moreover, MdJa2 interacted with MdBZR1. MdJa2 was revealed to independently regulate anthocyanin and proanthocyanidin synthesis pathways. The MdJa2–MdBZR1 complex enhanced the binding of MdJa2 to the promoters of downstream target genes. Our research provides new insights into how MADS-box transcription factors in the BR signaling pathway control the accumulations of anthocyanin and proanthocyanidin in red-fleshed apple. [ABSTRACT FROM AUTHOR]

Anthocyanin and proanthocyanidin play important roles in plant secondary metabolism. Although previous studies identified many transcription factors involved in anthocyanin and proanthocyanidin synthesis, the effects of MADS-box transcription factors are unclear in apple. Brassinosteroids (BRs) are steroid hormones that affect plant flavonoid biosynthesis, but the underlying regulatory mechanism is not yet well established. In this study, we identified a MADS-box transcription factor, MdJa2, which contained a highly conserved MADS-box domain and belonged to the STMADS11 subfamily. Additionally, MdJa2 was responsive to BR signal, and the overexpression of MdJa2 inhibited the synthesis of anthocyanin and proanthocyanidin. The silencing of MdJa2 in “Orin” calli promoted anthocyanin and proanthocyanidin accumulations. Moreover, MdJa2 interacted with MdBZR1. MdJa2 was revealed to independently regulate anthocyanin and proanthocyanidin synthesis pathways. The MdJa2–MdBZR1 complex enhanced the binding of MdJa2 to the promoters of downstream target genes. Our research provides new insights into how MADS-box transcription factors in the BR signaling pathway control the accumulations of anthocyanin and proanthocyanidin in red-fleshed apple.

Busulfan is currently an indispensable anti-cancer drug, but the side effects on male reproductive system are so serious. Meanwhile, red-fleshed apples are natural products with high anthocyanin content. In this research, we analyzed the effect of red-fleshed apple anthocyanin extract (RAAE) on busulfan-treated mice. Compared with the busulfan group, main plasma biochemical indicators were significantly improved after RAAE treatment. Compared with BA0 (busulfan without RAAE) group, total antioxidant capacity(T-AOC) and the activity of superoxide dismutase (SOD) and glutathione catalase (GSH-Px) in RAAE treatment groups were obviously increased, while the activity of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were significantly decreased. Malondialdehyde (MDA) was significantly decreased in the RAAE groups. In addition, we found RAAE alleviated busulfan-disrupted spermatogenesis through improving genes expression which are important for spermatogenesis, such as DDX4, PGK2, and TP1. Furthermore, we found that RAAE increased beneficial bacteria Akkermansia and Lactobacillaceae, and significantly depleted harmful bacteria Erysipelotrichia. The correlation studies indicated that RAAE ameliorated busulfan-induced rise in LysoPC levels through regulating gut microbial community and their associated metabolites. In conclusion, this study extends our understanding of the alleviated effect of RAAE on busulfan-induced male reproductive dysfunction through regulating the relationships between gut microbiota and metabolites.

Red‐fleshedapples are preferredbecause of their high content of phenolics and antioxidants in peel and pulp. Herein, we evaluated the mechanisms of apple peel polyphenolic extracts (APP) and apple flesh polyphenolic extracts (AFP) from the new red‐fleshed apple in inhibiting cell proliferation and inducing apoptosis on human breast cancer MDA‐MB‐231 cells. The antiproliferative activities were determined by the CCK8 assay. The expression of proteins was determined using Western blot. We found that the content of polyphenols and flavonoids in APP was significantly higher than that in AFP, and 14 main phenolic compounds in APP and AFP were quantified using UPLC–MS/MS techniques. Besides, the significant inhibition effects of APP and AFP were achieved through Akt pathway by inducing apoptosis (significantly upregulating reactive oxygen species [ROS] levels, and downregulating expression of pAkt, pBad, Bcl‐2, promoting Cytochrome c release, activating Cle‐Caspase 9, and inducing expressions of Cle‐Caspase 3 and Cle‐PARP), and inducing G0/G1 cell cycle arrest (increased expressions of p‐p53 and p21 and decreased expressions of PCNA and Cyclin D1). And the inhibition effect of APP was stronger than that of AFP. These results suggest that AFP and APP may be excellent sources of natural chemicals for treating triple‐negative breast cancer MDA‐MB‐231 cells. Practical Application: The effects of antiproliferation of phenolic extracts from red‐fleshed apple peels and flesh on human breast cancer MDA‐MB‐231 cells were evaluated. The data may clarify the functional parts of red‐fleshed apple and provide some basis for scientific researchers and consumers to recognize and exploit red‐fleshed apple. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate comprehensively the metabolic pathways and human bioavailability of anthocyanins and other (poly)phenols in an apple matrix, and to elucidate potential intake biomarkers. After the acute intake of a red-fleshed apple freeze-dried snack, plasma and urine were collected and analyzed by UPLC-MS/MS. A total of 37 phase-II and microbial phenolic metabolites were detected in plasma and urine. Among these, phloretin glucuronide, cyanidin-3-O-galactoside (plasma and urine) and peonidin-3-O-galactoside (urine) were the only metabolites detected in all the volunteers and not detected at basal conditions. The maximum urine excretion was detected at 2–4 h, and the main increase in plasma of phloretin glucuronide and cyanidin-3-O-galactoside was observed at 2 h post-intake (61.0 ± 6.82 and 10.3 ± 1.50 nM, respectively). These metabolites could be selected as the best intake biomarkers of red-fleshed apple and might be useful in human intervention studies when studying its health effects.

The present study was conducted to evaluate the ameliorating effect of 5-aminolevulinic acid (ALA) on physiological and biochemical changes of induced NaCl-salinity stress on in vitro shoot and callus cultured of red-fleshed apple. Shoot and callus segments of the red-flesh apple of Local Iranian genotype were cultured on MS medium containing different levels of NaCl (0, 30, 60 and 90 mM) and ALA (0, 2.5, 5, 10 and 20 µM). Physiological and biochemical variations of treated explants with or without NaCl and ALA treatments were recorded. In both explants, salinity reduced chlorophyll and carotenoids contents, but the activities of antioxidant enzymes and accumulation of total phenol and anthocyanin increased with increasing salinity level (90mM). Exogenous ALA in 5 and particularly in 10 µM was effective in enhancing chlorophyll and carotenoids contents, increasing the activities of superoxide dismutase, ascorbate peroxidase and accumulating of total phenol and anthocyanin. These results indicate that ALA has a powerful salinity-ameliorating potential on in vitro cultured shoot and callus of Iranian red-fleshed apple.

Apple (Malus domestica Borkh.) is an important fruit tree species worldwide. Apple fruits are favored by consumers because of their antioxidative, anti‐inflammatory, and antitumor effects as well as their protective effects against cardiovascular diseases and other health benefits. There is considerable interest in red‐fleshed apple fruits among breeders because of their high flavonoid and anthocyanin contents. However, the flavonoids extracted from red‐fleshed apple fruits must still be functionally characterized, especially regarding their protective effects against certain pathologies. In this study, the flavonoid components and contents in the extracts prepared from red‐fleshed apple cultivar "Meihong" were determined. Additionally, the in vitro antioxidant activities and protective effects of the extracts against CCl4‐induced acute liver injury were investigated. The red‐fleshed apple flesh flavonoid extract (RAFF) exhibited strong in vitro antioxidant activities. Compared with the model control mice treated with CCl4, the mice pretreated with high (800 mg/kg·bw), middle (400 mg/kg·bw), and low (200 mg/kg·bw) RAFF doses had significantly lower CCl4‐induced serum aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase activities. Moreover, the RAFF pretreatment also significantly decreased the liver malondialdehyde activity and prevented the CCl4‐induced decrease in liver superoxide dismutase, glutathione peroxidase, catalase, and reduced glutathione levels. Furthermore, a histopathological examination revealed that RAFF inhibited the inflammatory cell infiltration and cell boundary loss caused by CCl4 in the liver. Thus, RAFF is a natural antioxidant with significant antioxidative activities and liver protective effects. The results of this study may be relevant for enhancing the application of the red‐fleshed apple fruit extract as a food additive. Practical Application: We took the self‐selected red‐fleshed apple cultivar "Meihong" as the unique research material, and the active ingredients of its flavonoid extract, in vitro antioxidant activity and hepatoprotective effect were analyzed. It is of great significance to promote the development of the red‐fleshed apple industry, and also provides an important reference for the development of natural antioxidants. [ABSTRACT FROM AUTHOR]

The analysis of selected phenolic compounds in dried apple pulps with pentafluorophenyl (PFP) reversed-phase HPLC-UV-FL is presented. Three new apple cultivars named Nicoter Kanzi®, Shinano Gold yello®, and the red-fleshed one Luresweet Redlove® grown in one site in South Tyrol (Italy) have been investigated and compared with cv. Golden Delicious. Phenolics belonging to different classes (hydroxybenzoic acids, flavan-3-ols, caffeoylquinic derivatives, flavonols, and dihydrochalcones) have been analyzed. Extraction with aqueous methanolic mixture yielded good analytical recovery. Luresweet Redlove® showed the highest content of quercetin-3-O-glucoside, gallic acid, and chlorogenic acid and the lowest content of catechins and procyanidins; on the other hand, Nicoter Kanzi® showed the lowest concentration of quercetin-3-O-glucoside. The analytical procedure described is an easy and affordable methodology to investigate other apple varieties from local cultivations, to study their phytochemical composition, and to determine their potential nutraceutical values. The content of phenolic compounds in the three apple varieties Nicoter Kanzi®, Shinano Gold yello®, and Luresweet Redlove® was reported for the first time. [ABSTRACT FROM AUTHOR]

Anthocyanin pigmentation is an important consumption trait of apple (Malus domestica Borkh.). In this study, we focused on the identification of NAC (NAM, ATAF1/2 and CUC2) proteins involved in the regulation of anthocyanin accumulation in apple flesh. A group of MdNACs was selected for comparison of expression patterns between the white-fleshed cultivar 'Granny Smith' and red-fleshed 'Redlove'. Among them, MdNAC42 was screened, which exhibited a higher expression level in red-fleshed than in white-fleshed fruit, and has a positive correlation with anthocyanin content as fruits ripened. Moreover, overexpression of MdNAC42 in apple calli resulted in the up-regulation of flavonoid pathway genes, including MdCHS , MdCHI , MdF3H , MdDFR , MdANS and MdUFGT , thereby increasing the accumulation of anthocyanins, which confirmed the roles of MdNAC42 in anthocyanin biosynthesis. Notably, MdNAC42 was demonstrated to have an obvious interaction with MdMYB10 either in vitro or in vivo by yeast two-hybrid combined with bimolecular fluorescence complementation, further suggesting that MdNAC42 is an important part of the regulatory network controlling the anthocyanin pigmentation of red-fleshed apples. To the best of our knowledge, this is the first report identifying the MdNAC gene as related to anthocyanin accumulation in red-fleshed apples. This study provides valuable information for improving the regulatory model of anthocyanin biosynthesis in apple fruit. [ABSTRACT FROM AUTHOR]

Furan is a volatile and carcinogenic heterocyclic chemical compound that occurs in a wide range of thermally processed food. It can be induced during food-preparing processes by high temperatures and UV-C light. In the present study, the degradation of furan content in ground coffee, Maillard model system, and not-from-concentrate (NFC) apple juice by red-fleshed apple anthocyanin extract (RAAE) was studied. The results demonstrated that RAAEs had different degrees of degradation of furan content in coffee powder, and the RAAE from ‘XJ3’ had the most significant effect, with a reduction rate of up to 20%. Moreover, by adding RAAE to the Maillard model system, we found the amounts of furan were significantly reduced. At the same time, RAAE from ‘XJ3’ could observably reduce the content of furan in pasteurized NFC juice, with ‘Fuli’ NFC juice furan content decreasing the most, which was 68%. Taken together, our study demonstrated that the use of RAAE could be a feasible way to reduce furan content in ground coffee, Maillard model system, and NFC apple juice.

BACKGROUND Red‐fleshed apples are a great source of natural colorants and functional food ingredients because of their high anthocyanin content. Generally, anthocyanins are highly unstable after extraction, which limits their wide applications in the food and pharmaceutical industries. This study was aimed at investigating the effects of combining copigmentation with encapsulation on the stability of anthocyanins from red‐fleshed apples. In this study, red‐fleshed apple anthocyanins were copigmented with caffeic acid, and then the copigmented complexes were encapsulated using gum arabic and maltodextrin using spray drying and freeze drying. RESULTS: All anthocyanin microcapsules had high encapsulation efficiencies ranging from 93.84 to 96.85% with mean hydrodynamic diameter smaller than 350 nm. After heating at 80 °C for 2 h, the dispersions of microencapsulated anthocyanins with copigments exhibited the highest absorbance values at λmax (515 nm) (P < 0.05). Light stability experiments demonstrated that the half‐life of the red‐fleshed apple anthocyanins increased from 5 to 12 days after being treated with copigmentation and encapsulation. The drying methods (spray/freeze drying) did not significantly influence the stability of the microencapsulated anthocyanins. CONCLUSIONS: Applying copigmentation and spray‐drying encapsulation in tandem has great potential for enhancing the stability of red‐fleshed apple anthocyanins. Thus, such anthocyanins with enhanced stability may be increasingly used in the food and pharmaceutical industries as value‐added natural food pigments. © 2018 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Red-flesh apples as a novel anthocyanin rich food. • Complete metabolic pathways for phenolics from red-flesh apples. • Phloretin glucuronide as specific apple intake biomarker. • Cyanidin galactoside and peonidin galactoside as red-flesh apple intake biomarkers. Abstract The aim of this study was to investigate comprehensively the metabolic pathways and human bioavailability of anthocyanins and other (poly)phenols in an apple matrix, and to elucidate potential intake biomarkers. After the acute intake of a red-fleshed apple freeze-dried snack, plasma and urine were collected and analyzed by UPLC-MS/MS. A total of 37 phase-II and microbial phenolic metabolites were detected in plasma and urine. Among these, phloretin glucuronide, cyanidin-3- O -galactoside (plasma and urine) and peonidin-3- O -galactoside (urine) were the only metabolites detected in all the volunteers and not detected at basal conditions. The maximum urine excretion was detected at 2–4 h, and the main increase in plasma of phloretin glucuronide and cyanidin-3- O -galactoside was observed at 2 h post-intake (61.0 ± 6.82 and 10.3 ± 1.50 nM, respectively). These metabolites could be selected as the best intake biomarkers of red-fleshed apple and might be useful in human intervention studies when studying its health effects. [ABSTRACT FROM AUTHOR]

The aim of this study was to determine the effect of different drying methods: convective (at 50, 60, 70 °C), vacuum-microwave (at 120, 240, 360, 480 W and 360 W with reduction to 120 W) and hybrid (convective pre-drying at 50, 60, 70 °C followed by vacuum-microwave drying at 120 W) on the quality parameters of novel red-fleshed apple fruit snacks (RFAs), such as phenolics, on-line antioxidant capacity, water activity and color. Drying kinetics, including a temperature profile of dried material, and modified Page model were determined. Freeze-drying was used as a control method. The highest content of bioactive compounds in the samples was retained following freeze-drying, then hybrid, vacuum-microwave and finally convection drying. The antioxidant capacity measured by on-line 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), identified anthocyanins, flavan-3-ols and phenolic acid as the main compounds responsible for this activity. Unfavorable changes in color, formation of hydroxymethylfurfural (HMF) and degradation of polyphenolics were noted along with increasing drying temperature and magnetron power. The red-fleshed apple snacks are a promising high-quality dehydrated food product belonging to functional foods category.

In this research, we compared the phenotypical characters, total anthocyanins content, total phenols content, and antioxidant activity of red-fleshed apple cultivars ‘XJ4’, ‘QN-5’, ‘DH’ and ‘HX1’ at three fruit developmental stages. A further flavonoids metabolites study was conducted in ‘XJ4’ and ‘DH’. We found broader variation of total anthocyanins content in the peel of the four cultivars, which might result in larger differences of free radicals scavenging rate. The most significant difference in fruit phenotype, anthocyanins content, and DPPH scavenging rate was observed between ‘XJ4’ and ‘DH’ at mature stage. Therefore, the flavonoids metabolites of ‘XJ4’ and ‘DH’ at mature stage were compared to unveil the details of anthocyanins compounds. The unique compounds pelargonidin 3-O-β-d-glucoside and cyanidin-3-O-malonylhexoside were detected only in peel and flesh of ‘XJ4’ but not in ‘DH’, which might contribute to the purple peel and dark-red flesh color of ‘XJ4’. Significantly decreased upstream metabolites in the early biosynthetic genes regulated domain were found only in ‘XJ4’ peel but not in the flesh. This might explain why the anthocyanins content in ‘XJ4’ peel was decreased largely at the mature stage. Taken together, our findings will give some insight into the metabolites study in flavonoid biosynthetic pathway of red-fleshed apple.

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The present study was conducted to evaluate the ameliorating effect of 5-aminolevulinic acid (ALA) on physiological and biochemical changes of induced NaCl-salinity stress on in vitro shoot and callus cultured of red-fleshed apple. Shoot and callus segments of the red-flesh apple of Local Iranian genotype were cultured on MS medium containing different levels of NaCl (0, 30, 60 and 90 mM) and ALA (0, 2.5, 5, 10 and 20 µM). Physiological and biochemical variations of treated explants with or without NaCl and ALA treatments were recorded. In both explants, salinity reduced chlorophyll and carotenoids contents, but the activities of antioxidant enzymes and accumulation of total phenol and anthocyanin increased with increasing salinity level (90mM). Exogenous ALA in 5 and particularly in 10 µM was effective in enhancing chlorophyll and carotenoids contents, increasing the activities of superoxide dismutase, ascorbate peroxidase and accumulating of total phenol and anthocyanin. These results indicate that ALA has a powerful salinity-ameliorating potential on in vitro cultured shoot and callus of Iranian red-fleshed apple. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate consumer preference for the novel red-fleshed apple genotypes. The information presented provides us with insight into consumer propensity to accept a new variety with radical visible innovative attributes and societal demand for a more sustainable production. The survey was carried out on the population of a province in central Italy, where there is consumer demand for organic and high-quality foods. The consumers’ opinions contrasted greatly and can be divided into two groups. Over one quarter of the respondents declared that they did not like the innovative fruit. The cluster analysis and CHAID analyses were performed on the consumers’ responses to determine sample groupings. Three clusters were identified among the consumers with positive opinions, enthusiastic, health-conscious, and unmotivated illustrating how the hierarchical level of preferences of the potential consumers of the red-fleshed apple fruit are triggered by various motivations. The surveys also pointed out that most consumers of the sampled population were not biased against research activities producing innovative food products. A large number of consumers, especially women, declared that they were willing to pay a higher price considering the increased nutritional value of red-fleshed apple fruits. By examining the broad context of this study, decision-making researchers and operators of the fruit industry will be able to make informed and realistic predictions concerning possible future scenarios and to determine the possible effects and desirability. [ABSTRACT FROM AUTHOR]

Red-flesh apples are known as functional fruits because of their rich anthocyanin. The anthocyanin content of the red flesh apple cultivar ′QN-5′ we bred can reach 361 mg·kg−1 (FW), and showed higher scavenging capacity to DPPH radicals, hydroxyl radicals, and superoxide anion radicals, with scavenging rates of 80.0%, 54.0%, and 43.3%, respectively. We used this particular anthocyanin-rich ′QN-5′ apple as material to examine how nanocrystallization affects the antixodiant effect of anthocyanin. The anthocyanin extract was encapsulated with biocompatible zein to form zein-anthocyanin nanoparticles (ZANPs). Transmission electron microscopy (TEM) scanning showed that ZANPs had a regular spherical shape with an average diameter size of 50–60nm. When the ratio of the zein and the anthocyanin was 1:0.5, the results suggested that the encapsulation efficiency (EE) of the ZANPs could reach as high as 92.8%, and that scavenging rate for DPPH radicals was increased from 87.1% to 97.2% compared to the non-nanocrystallized anthocyanin extract. Interestingly, treatment under alkaline conditions (pH 9.0), high temperature (90 °C), and a storage time of 7 days could decrease the scavenging capacity of the ZANPs for DPPH radicals, but this scavenging capacity loss for ZANPs was significantly lower than that observed in the non-nanocrystallized anthocyanin, suggesting the higher stability of ZANPs is caused by encapsulation. These results would provide a theoretical basis for the application of the anthocyanin in scavenging free radicals under stress conditions.

Coloration in apple (Malus×domestica) flesh is mainly caused by the accumulation of anthocyanin. Anthocyanin is biosynthesized through the ?avonoid pathway and regulated by MYB, bHLH, and WD40 transcription factors (TFs). Here, we report that the HD-Zip I TF MdHB1 was also involved in the regulation of anthocyanin accumulation. MdHB1 silencing caused the accumulation of anthocyanin in 'Granny Smith' flesh, whereas its overexpression reduced the flesh content of anthocyanin in 'Ballerina' (red-fleshed apple). Moreover, flowers of transgenic tobacco (Nicotiana tabacum 'NC89') overexpressing MdHB1 showed a remarkable reduction in pigmentation. Transient promoter activation assays and yeast one-hybrid results indicated that MdHB1 indirectly inhibited expression of the anthocyanin biosynthetic genes encoding dihydro avonol-4-reductase (DFR) and UDP-glucose:?avonoid 3-O-glycosyltransferase (UFGT). Yeast two-hybrid and bimolecular fluorescence complementation determined that MdHB1 acted as a homodimer and could interact with MYB, bHLH, and WD40 in the cytoplasm, consistent with its cytoplasmic localization by green fluorescent protein fluorescence observations. Together, these results suggest that MdHB1 constrains MdMYB10, MdbHLH3, and MdTTG1 to the cytoplasm, and then represses the transcription of MdDFR and MdUFGT indirectly. When MdHB1 is silenced, these TFs are released to activate the expression of MdDFR and MdUFGT and also anthocyanin biosynthesis, resulting in red flesh in 'Granny Smith'. [ABSTRACT FROM AUTHOR]

This study presents a new rapid and non-destructive technique for detecting the degree of red coloration in the flesh of a red-fleshed apple cultivar ‘Kurenai no Yume’. Using a UV–vis-NIR interactance device, interactance data ranging from 190 to 1800 nm was collected non-destructively from the surface of 180 apple fruits. Levels of anthocyanins in the flesh of each sample were determined with a spectrophotometer. Partial least squares (PLS) regression, multiple linear regression (MLR) and artificial neural networks (ANN) modeling analysis were performed to develop models for estimating anthocyanin content from interactance data. Results showed that all the developed models achieved reasonable predictive accuracy. Particularly, the ANN model based on 11 key wavelengths, which achieved consistent performance in predictions with calibration ( R 2 = 0.5798) and test ( R 2 = 0.5324) datasets, were the most promising from the perspective of practical applications. This study strongly suggests that the interactance technique combined with the developed models might provide a new rapid and non-destructive approach for assessing the quality of red-fleshed apples. [ABSTRACT FROM AUTHOR]

• Red-fleshed apple is a rich source of polyphenols and biofortified in anthocyanins. • A complete elucidation of the phenolic metabolism of red-fleshed apple is presented. • Acute and sustained interventions were combined to evaluate phenolic bioavailability. • We report a strong influence of the food matrix on anthocyanins bioavailability. • A selection of the most discriminatory phenolic intake biomarkers was performed. The present study aimed to evaluate the metabolism and bioavailability of anthocyanins (ACN) and other phenolics from red-fleshed apple (RFA) and to define the intake biomarkers compared to common white-fleshed apple (WFA). Acute and sustained (6-week) interventions were combined in a randomized, controlled and parallel study with 121 hypercholesterolemic subjects. Another arm consuming ACN-rich infusion from aronia fruit (ARO) provided matched content and profile of ACN. Plasma, urine and faeces samples were analyzed using UPLC-MS/MS. Results showed higher bioavailability of ACN after ARO compared to RFA, showing a clear apple matrix effect. The dihydrochalcone phloretin-2′- O -glucuronide was the most discriminant intake biomarker of both apples. The urinary peonidin-3- O -galactoside was a good biomarker after both ARO and RFA intakes, whereas peonidin- O -arabinoside was reported to be specific from ARO. The elucidation of the phenolic metabolism and the selection of intake biomarkers is a promising approach to relate phenolic compounds and human health. [ABSTRACT FROM AUTHOR]

We have succeeded in isolating an MdMYB110a_JP gene responsible for a red-fleshed trait from a fruit of apple cultivar 'JPP35' ('Jonathan' × 'Pink Pearl'). The isolated MdMYB110a_JP gene was located on chromosome (ch.) 17, which was different from the location of known MdMYB1/ 10 gene of ch.9, and 'JPP35' and 'Pink Pearl' did not contain the known R :MdMYB10 allele responsible for the red-skin and red-fleshed trait. The MdMYB110a_JP was expressed strongly and weakly in the cortex and core of 'JPP35' fruit, respectively, at the time of coloring start in flesh, and also weakly in flower buds. Following the MdMYB110a_JP expression, the expression of the genes, MdCHS and MdLDOX, that encode the enzymes of the flavonoid pathway, was induced in flesh of 'JPP35' in accordance with anthocyanin accumulation. In contrast, the MdMYB110a_JP gene was not expressed in any tissues in red-skin and white-fleshed 'Fuji', and in red-skin and red-fleshed 'Maypole'. Instead, MdMYB1- 1 allele responsible for red-skin trait was expressed in red-skin of 'Fuji' and 'JPP35', and R :MdMYB10 allele responsible for red-skin and red-flesh trait was expressed in red-core and red-cortex in 'Maypole' as expected. Moreover, 35S: MdMYB110a_JP transgenic apple 'JM2' showed a red-foliage phenotype depending on the MdMYB110a_JP expression level. From the results, it was strongly suggested that the red-fleshed phenotype of 'JPP35' fruit was caused by up-regulation of the genes of anthocyanin pathway induced by the MdMYB110a_JP gene. [ABSTRACT FROM AUTHOR]

• Total aroma OAVs was lowest in expansion period and highest in ripen period. • The concentration of bound form aromas was higher than that of free form. • Esters, aldehydes and terpenes were the main bound form aromas in 'redlove 1'. • 'Fruity' and 'green' were the main odor notes of 'redlove 1' apples. Red-fleshed apple is attracting more and more attentions from both consumers and researchers thanks to its pleasant appearance and flavor. Accumulation and development of aromas of apple fruit are important factors to understand the fruit quality of a new cultivated variety. However, knowledge on the key aroma components development and accumulation of red-fleshed apple (Malus sieversii f. neidzwetzkyana (Dieck) Langenf) in Xinjiang through the fruit developmental periods is insufficient. In this paper, free and bound form aromas of a red-fleshed apple since young fruit period through expansion period and conversion period to ripen period were comparatively studied. We classified aroma substances according to the compound category (esters, aldehydes, alcohols, terpenes, and aromatics), and to the odor types. The concentration of bound form aroma compounds were higher than that of free form throughout the 4 periods and showed different evolution trends. Among the 4 periods, the bound aroma was the most in the expansion period; and terpenoids showed the greatest potential to release free form aromas. Total aromas were found the lowest concentration in expansion period and the highest concentration in ripen period. The observations in this study will provide theoretical support for the research of typical aroma formation regularity of red-fleshed apple, and guide for the aroma enhancement regulation. [ABSTRACT FROM AUTHOR]

Furan is a volatile and carcinogenic heterocyclic chemical compound that occurs in a wide range of thermally processed food. It can be induced during food-preparing processes by high temperatures and UV-C light. In the present study, the degradation of furan content in ground coffee, Maillard model system, and not-from-concentrate (NFC) apple juice by red-fleshed apple anthocyanin extract (RAAE) was studied. The results demonstrated that RAAEs had different degrees of degradation of furan content in coffee powder, and the RAAE from 'XJ3' had the most significant effect, with a reduction rate of up to 20%. Moreover, by adding RAAE to the Maillard model system, we found the amounts of furan were significantly reduced. At the same time, RAAE from 'XJ3' could observably reduce the content of furan in pasteurized NFC juice, with 'Fuli' NFC juice furan content decreasing the most, which was 68%. Taken together, our study demonstrated that the use of RAAE could be a feasible way to reduce furan content in ground coffee, Maillard model system, and NFC apple juice. [ABSTRACT FROM AUTHOR]

• WRKY, AP2 and MYB transcription factor with large differences were screened out. • PPO plays a major role during the early browning stage. • Membrane lipids degradated; produce free fatty acids. Enzymatic browning affects the quality of fresh-cut produce. To elucidate the mechanism regulating the browning of red-fleshed apple, we analyzed the transcriptome and metabolome of freshly cut Meihong apple samples collected at 0, 0.5, and 4 h after cutting. The main differentially abundant metabolites during the browning process were phenols, amino acids and fatty acids. Phenols are substrates for enzymatic browning and most of them are negatively correlated with browning. Amino acids and their derivatives are associated with non-enzymatic browning and most of them are positively correlated with browning. A transmission electron microscopy analysis revealed the degradation of the cell membrane of cut fruit samples, which significantly affected the fatty acid content. The differentially expressed genes were mainly related to plant–pathogen interactions. Polyphenol oxidase (PPO) plays a major role during the early browning stage, but PPO gene expression was not higher than WRKY, AP2 and MYB transcription factor. [ABSTRACT FROM AUTHOR]

The aim of this study was to determine the effect of different drying methods: convective (at 50, 60, 70 °C), vacuum-microwave (at 120, 240, 360, 480 W and 360 W with reduction to 120 W) and hybrid (convective pre-drying at 50, 60, 70 °C followed by vacuum-microwave drying at 120 W) on the quality parameters of novel red-fleshed apple fruit snacks (RFAs), such as phenolics, on-line antioxidant capacity, water activity and color. Drying kinetics, including a temperature profile of dried material, and modified Page model were determined. Freeze-drying was used as a control method. The highest content of bioactive compounds in the samples was retained following freeze-drying, then hybrid, vacuum-microwave and finally convection drying. The antioxidant capacity measured by on-line 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), identified anthocyanins, flavan-3-ols and phenolic acid as the main compounds responsible for this activity. Unfavorable changes in color, formation of hydroxymethylfurfural (HMF) and degradation of polyphenolics were noted along with increasing drying temperature and magnetron power. The red-fleshed apple snacks are a promising high-quality dehydrated food product belonging to functional foods category. [ABSTRACT FROM AUTHOR]

In this research, we compared the phenotypical characters, total anthocyanins content, total phenols content, and antioxidant activity of red-fleshed apple cultivars 'XJ4', 'QN-5', 'DH' and 'HX1' at three fruit developmental stages. A further flavonoids metabolites study was conducted in 'XJ4' and 'DH'. We found broader variation of total anthocyanins content in the peel of the four cultivars, which might result in larger differences of free radicals scavenging rate. The most significant difference in fruit phenotype, anthocyanins content, and DPPH scavenging rate was observed between 'XJ4' and 'DH' at mature stage. Therefore, the flavonoids metabolites of 'XJ4' and 'DH' at mature stage were compared to unveil the details of anthocyanins compounds. The unique compounds pelargonidin 3-O-β-d-glucoside and cyanidin-3-O-malonylhexoside were detected only in peel and flesh of 'XJ4' but not in 'DH', which might contribute to the purple peel and dark-red flesh color of 'XJ4'. Significantly decreased upstream metabolites in the early biosynthetic genes regulated domain were found only in 'XJ4' peel but not in the flesh. This might explain why the anthocyanins content in 'XJ4' peel was decreased largely at the mature stage. Taken together, our findings will give some insight into the metabolites study in flavonoid biosynthetic pathway of red-fleshed apple. [ABSTRACT FROM AUTHOR]

Red-flesh apples are known as functional fruits because of their rich anthocyanin. The anthocyanin content of the red flesh apple cultivar ′QN-5′ we bred can reach 361 mg·kg−1 (FW), and showed higher scavenging capacity to DPPH radicals, hydroxyl radicals, and superoxide anion radicals, with scavenging rates of 80.0%, 54.0%, and 43.3%, respectively. We used this particular anthocyanin-rich ′QN-5′ apple as material to examine how nanocrystallization affects the antixodiant effect of anthocyanin. The anthocyanin extract was encapsulated with biocompatible zein to form zein-anthocyanin nanoparticles (ZANPs). Transmission electron microscopy (TEM) scanning showed that ZANPs had a regular spherical shape with an average diameter size of 50–60nm. When the ratio of the zein and the anthocyanin was 1:0.5, the results suggested that the encapsulation efficiency (EE) of the ZANPs could reach as high as 92.8%, and that scavenging rate for DPPH radicals was increased from 87.1% to 97.2% compared to the non-nanocrystallized anthocyanin extract. Interestingly, treatment under alkaline conditions (pH 9.0), high temperature (90 °C), and a storage time of 7 days could decrease the scavenging capacity of the ZANPs for DPPH radicals, but this scavenging capacity loss for ZANPs was significantly lower than that observed in the non-nanocrystallized anthocyanin, suggesting the higher stability of ZANPs is caused by encapsulation. These results would provide a theoretical basis for the application of the anthocyanin in scavenging free radicals under stress conditions. [ABSTRACT FROM AUTHOR]