Understanding the molecular regulation of flavonoid 3'-hydroxylase in anthocyanin synthesis: insights from purple qingke.

BACKGROUND: The Flavonoid 3'-hydroxylase gene(F3'H) is an important structural gene in the anthocyanin synthesis pathway of plants, which has been proven to be involved in the color formation of organs such as leaves, flowers, and fruits in many plants. However, the mechanism and function in barley are still unclear. RESULTS: In order to explore the molecular mechanism of the grain color formation of purple qingke, we used the cultivated qingke variety Nierumzha (purple grain) and the selected qingke variety Kunlun 10 (white grain) to conduct transcriptomic sequencing at the early milk, late milk and soft dough stage. Weighted Gene Co-expression Network Analysis (WGCNA) was used to construct weighted gene co-expression network related to grain color formation, and three key modules (brown, yellow, and turquoise modules) related to purple grain of qingke were selected. F3'H (HORVU1Hr1G094880) was selected from the hub gene of the module for the yeast library, yeast two-hybrid (Y2H), subcellular localization and other studies. It was found that in purple qingke, HvnF3'H mainly distributed in the cytoplasm and cell membrane and interacted with several stress proteins such as methyltransferase protein and zinc finger protein. CONCLUSIONS: The results of this study provide reference for the regulation mechanism of anthocyanin-related genes in purple grain qingke.

LC-MS/MS-based targeted carotenoid and anthocyanidin metabolic profile of Auricularia cornea under blue and red LED light exposure.

Light exposure significantly impacted the coloration and metabolism of Auricularia cornea, although the underlying mechanisms remain unclear. This study aimed to test the apparent color and pigment metabolic profiles of A. cornea in response to red (λp = 630 nm) and blue (λp = 463 nm) visible light exposure. Colorimeter analysis showed that fruiting bodies appeared bright-white under red-light and deeper-red under blue-light, both with a yellow tinge. On the 40th day of light-exposure, bodies were collected for metabolite detection. A total of 481 metabolites were targeted analysis, resulting in 18 carotenoids and 11 anthocyanins. Under red and blue light exposure, the total carotenoids levels were 1.1652 µg/g and 1.1576 µg/g, the total anthocyanins levels were 0.0799 µg/g and 0.1286 µg/g, respectively. Four differential metabolites and three putative gene linked to the visual coloration of A. cornea were identified. This pioneering study provides new insights into the role of light in regulating A. cornea pigmentation and metabolic profile.

Preparation of a colorimetric hydrogel indicator reinforced with modified aramid nanofiber employing natural anthocyanin to monitor shrimp freshness.

The pH-responsive hydrogels have potential applications in food visualization detection, but their fragile mechanical properties limit their applicability. The excellent mechanical properties and thermal stability of aramid nanofibers (ANFs) can improve the structural stability of hydrogels. In this study, the surface properties of ANFs were enhanced through modification to improve their surface activity. The modified ANFs, designated as ANF-SN, were produced following treatment with a mixture of sulfuric acid (H2SO4) and nitric acid (HNO3), which led to increased reactivity and dispersibility of the ANFs due to the proliferation of active groups on their nanofiber surface. The preferred anthocyanin extract from purple sweet potatoes (purple sweet potato extract [PSPE]) had significant color responses to pH (2-12) and ammonia vapor. A stable dual-network colorimetric hydrogel was fabricated by combining ANF-SN, polyvinyl alcohol/sodium alginate (PVA/SA), and PSPE through a two-step method (freeze-thawing and staining). Characterization analysis showed that the strong acid modification of ANFs effectively improved their chemical reactivity. ANF-SN was better than ANF in promoting the formation of hydrogen bond networks, enhancing hydrogel network structures, and improving the viscoelasticity of hydrogels. The optimal hydrogel indicator PVA/SA/ANF-SN/PSPE had good color responsiveness and sensitivity to ammonia. It can also be used to further determine shrimp freshness value using a smartphone and RGB color-picking software.

WRKY75 regulates anthocyanin accumulation in juvenile citrus tissues.

The anthocyanin accumulation in juvenile tissues can enhance the ornamental value, attract pollinators, and help improve abiotic stress. Although transcriptional regulation studies of anthocyanin have been relatively extensive, there are few reports on the mechanism of anthocyanin accumulation in young tissues. This study reveals that many juvenile citrus tissues (flowers, leaves, and pericarp) undergo transient accumulation of anthocyanins, exhibiting a red coloration. Using weighted gene co-expression network analysis (WGCNA) identified CitWRKY75 as a candidate gene. After detecting the expression levels of CitWRKY75 in various citrus juvenile tissues, the expression trend of CitWRKY75 was highly consistent with the red exhibiting and fading. Overexpression of CitWRKY75 in tobacco significantly increased the anthocyanin content. LUC and yeast one-hybrid assay demonstrated that CitWRKY75 could bind to the promoter of CitRuby1(encoding the key transcription factor promoting anthocyanin accumulation) and promote its expression. Finally, comparing the expression levels of CitWRKY75 and CitRuby1 in the late development stage of blood orange found that CitWRKY75 was not the main regulatory factor for anthocyanin accumulation in the later stage. This study used reverse genetics to identify a transcription factor, CitWRKY75, upstream of CitRuby1, which promotes anthocyanin accumulation in citrus juvenile tissues. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-024-01490-9.

Identifying potential anthocyanin biosynthesis regulator in Chinese cherry by comprehensive genome-wide characterization of the R2R3-MYB transcription factor gene family.

BACKGROUND: Chinese cherry [Cerasus pseudocerasus (Lindl.) G.Don] (syn. Prunus pseudocerasus Lindl.) is an economically important fruiting cherry species with a diverse range of attractive colors, spanning from the lightest yellow to the darkest black purple. However, the MYB transcription factors involved in anthocyanin biosynthesis underlying fruit color variation in Chinese cherry remain unknown. RESULTS: In this study, we characterized the R2R3-MYB gene family of Chinese cherry by genome-wide identification and compared it with those of 10 Rosaceae relatives and Arabidopsis thaliana. A total of 1490 R2R3-MYBs were classified into 43 subfamilies, which included 29 subfamilies containing both Rosaceae MYBs and AtMYBs. One subfamily (S45) contained only Rosaceae MYBs, while three subfamilies (S12, S75, and S77) contained only AtMYBs. The variation in gene numbers within identical subfamilies among different species and the absence of certain subfamilies in some species indicated the species-specific expansion within MYB gene family in Chinese cherry and its relatives. Segmental and tandem duplication events primarily contributed to the expansion of Chinese cherry R2R3-CpMYBs. The duplicated gene pairs underwent purifying selection during evolution after duplication events. Phylogenetic relationships and transcript profiling revealed that CpMYB10 and CpMYB4 are involved in the regulation of anthocyanin biosynthesis in Chinese cherry fruits. Expression patterns, transient overexpression and VIGS results confirmed that CpMYB10 promotes anthocyanin accumulation in the fruit skin, while CpMYB4 acts as a repressor, inhibiting anthocyanin biosynthesis of Chinese cherry. CONCLUSIONS: This study provides a comprehensive and systematic analysis of R2R3-MYB gene family in Chinese cherry and Rosaceae relatives, and identifies two regulators, CpMYB10 and CpMYB4, involved in anthocyanin biosynthesis in Chinese cherry. These results help to develop and utilize the potential functions of anthocyanins in Chinese cherry.

Lactobacillus fermentation accelerated biotransformation of cranberry anthocyanins towards phenol-pyranoanthocyanins and their stability and antioxidant property.

Phenol-pyranoanthocyanins, a structurally modified type of anthocyanin, has higher stability than anthocyanins. However, their conversion occurs slowly. Therefore, it is crucial to improve the conversion efficiency and production of pyranoanthocyanins. In this study, cranberry anthocyanin (CRAN) was fermented using two Lactobacillus strains along with caffeic acid to form cranberry-derived pyranoanthocyanins (PY-CRAN). PY-CRAN was characterized and identified. The physicochemical properties, antioxidant activity, and tyrosinase inhibitory capacity of PY-CRAN were assessed. The results showed that phenol-pyranoanthocyanins can be rapidly produced through fermentative transformation using Lactiplantibacillus plantarum and Lacticaseibacillus paracasei. Lacticaseibacillus paracasei exhibits a higher propensity for producing phenol-pyranoanthocyanins. PY-CRAN exhibits high stability under light and various pH conditions. Moreover, they possess excellent antioxidant properties and the ability to inhibit tyrosinase. These results suggest that fermentative biotransformation conducted by Lactobacillus is an ideal method for producing cranberry pyranoanthocyanins. The resulting anthocyanins have potential as antioxidant and whitening agents, making them promising bioactive ingredients.

Unveiling the influence of high hydrostatic pressure and protein interactions on the color and chemical stability of cyanidin-3-O-glucoside.

This study explored how high hydrostatic pressure (HHP) and proteins (i.e., BSA and HSA) influence the color and chemical stability of cyanidin-3-O-glucoside (C3G) at neutral pH. HHP treatments (100-500 MPa, 0-20 min, 25 °C) did not affect C3G content in phosphate buffer (PB) and MOPS buffer. However, significant color loss of C3G occurred in PB due to pressure-induced pH reduction (e.g., from 7 to 4.8 at 500 MPa), which accelerated the hydration of C3G, converting it from colored to colorless species. Consequently, MOPS buffer was employed for subsequent stability experiments to assess the impact of protein and HHP on the thermal, storage, and UV light stability of C3G. Initially, rapid color loss occurred during heating and storage, primarily due to the reversible hydration of C3G until equilibrium with colorless species was reached, followed by slower parallel degradation. HSA increased the fraction of colored species at equilibrium but accelerated thermal degradation, while BSA had minimal effects. UV light irradiation accelerated the degradation of C3G colored species, causing direct degradation without conversion to colorless species, a process further intensified by the presence of proteins. HHP exhibited a negligible effect on C3G stability regardless of protein addition. These findings provide insights into anthocyanin stability under HHP and protein interactions, contributing to the development of future formulation and processing strategies for improved stability and broader applications.

Enzymatic acylation of cyanidin-3-O-glucoside with aromatic and aliphatic acid methyl ester: Structure-stability relationships of acylated derivatives.

Anthocyanins are water-soluble pigments, but they tend to be unstable in aqueous solutions. Modification of their molecular structure offers a viable approach to alter their intrinsic properties and enhance stability. Aromatic and aliphatic acid methyl esters were used as acyl donors in the enzymatic acylation of cyanidin-3-O-glucoside (C3G), and their analysis was conducted using ultraperformance liquid chromatography-mass spectrometry (UPLC-MS). The highest conversion rate achieved was 96.41 % for cyanidin-3-O-(6″-feruloyl) glucoside. Comparative evaluations of stability revealed that aromatic acyl group-conjugated C3G exhibited superior stability enhancement compared with aliphatic acyl group derivatives. The stability of aliphatic C3G decreased with increasing carbon chain length. The molecular geometries of different anthocyanins were optimized, and energy level calculations using density functional theory (DFT) identified their sites with antioxidant activities. Computational calculations aligned with the in vitro antioxidant assay results. This study provided theoretical support for stabilizing anthocyanins and broadened the application of acylated anthocyanins as food colorants and nutrient supplements.

Effect of glycosylation, acylation and pyranylation at cyanidin C-ring on its interaction with vitamin C in apple juice beverage matrix.

BACKGROUND: Synchronous degradation between anthocyanin and vitamin C was found in fruit and vegetable juice matrices. To investigate whether the C-ring of anthocyanin is the key site of this interaction, cyanidin with four different C-ring modifications (3-glucosylation, 3,5-diglucosylation, 6″-malonylation, pyranylation) was added to vitamin C-containing apple juice, and the changes of anthocyanin retention, vitamin C retention, color, antioxidative activity and differential metabolites were analyzed. RESULTS: The anthocyanin retention was in the order of pyranylation >6″-malonylation >3,5-diglucosylation >3-glucosylation. The vitamin C retention was in the order of 6″-malonylation > pyranylation >3,5-diglucosylation >3-glucosylation. The order of color stability was the same as that of anthocyanin retention, and the order of antioxidative activity was opposite to that of vitamin C retention. The results showed that modification at the C-ring limited the activity of anthocyanin, and suggested that the C-ring was one of the key sites for anthocyanin and vitamin C interaction. The shared differential metabolite of all apple juice matrices added with different anthocyanins was trans-hinokiresinol, which was likely generated from anthocyanin skeleton reacted with certain compounds in apple juice. CONCLUSION: This study showed that modification of the anthocyanin C-ring could affect the anthocyanin and vitamin C interaction to some extent, which provided valuable insights for the application of anthocyanin C-ring modification in shelf-life quality control of typical fruit and vegetable beverages with the coexistence of anthocyanin and vitamin C. © 2024 Society of Chemical Industry.

Phytochemical, Antimicrobial, and Antioxidant Activity of Different Extracts from Frozen, Freeze-Dried, and Oven-Dried Jostaberries Grown in Moldova.

In this paper, the qualitative and quantitative profile is evaluated of the bioactive compounds, antioxidant activity (AA), microbiostatic properties, as well as the color parameters of jostaberry extracts, obtained from frozen (FJ), freeze-dried (FDJ), and oven-dried berries (DJ). The optimal extraction conditions by ultrasound-assisted extraction (UAE) and microwave-assisted extraction (MAE) were selected after determination of the total polyphenol content (TPC), total flavonoid content (TFC), total antocyanin content (TA), AA by 2,2-diphenyl-1-picrylhydrazyl-hydrate (DPPH), and the free radical cation 2,2-azinobis-3-ethylbenzothiazoline-6-sulfonates (ABTS). Non-conventional extraction methods are less destructive to anthocyanins, while drying the berries reduced TA, regardless of the extraction method. The oven-drying process reduced the concentration of TA in DJ extracts by 99.4% and of ascorbic acid by 92.42% compared to FJ. AA was influenced by the jostaberry pretreatment methods. The DPPH and ABTS tests recorded values (mg Trolox equivalent/g dry weight) between 17.60 and 35.26 and 35.64 and 109.17 for FJ extracts, between 7.50 and 7.96 and 45.73 and 82.22 for FDJ, as well as between 6.31 and 7.40 and 34.04 and 52.20 for DJ, respectively. The jostaberry pretreatment produced significant changes in all color parameters. Mutual information analysis, applied to determine the influence of ultrasound and microwave durations on TPC, TFC, TA, AA, pH, and color parameters in jostaberry extracts, showed the greatest influence on TA (0.367 bits) and TFC (0.329 bits). The DPPH and ABTS inhibition capacity of all FJ' extracts had higher values and varied more strongly, depending on pH, heat treatment, and storage time, compared to the AA values of FDJ' and DJ' extracts. A significant antimicrobial effect was observed on all bacterial strains studied for FJP. FDJP was more active on Bacillus cereus, Staphylococcus aureus, and Escherichia coli. DJP was more active on Salmonella Abony and Pseudomonas aeruginosa. The antifungal effect of DJP was stronger compared to FDJP. Jostaberry extracts obtained under different conditions can be used in food production, offering a wide spectrum of red hues.

NAC Transcription Factor PpNAP4 Promotes Chlorophyll Degradation and Anthocyanin Synthesis in the Skin of Peach Fruit.

Chlorophyll (Chl) catabolism and anthocyanin synthesis play pivotal roles in determining the final skin color of fruits during maturation. However, in peach (Prunus persica) fruit, the regulatory mechanism governing skin color, especially the Chl catabolism, remains largely elusive. In this study, we identified ten Chl catabolic genes (CCGs), with PpSGR emerging as a key regulator in Chl degradation in peaches. Furthermore, a NAC-like, activated by AP3/P1 (NAP) transcription factor (TF), PpNAP4, was identified as a positive modulator of Chl breakdown. PpNAP4 induced the expression of PpSGR and other CCGs, including PpPPH, PpPAO, and PpTIC55-2, by directly binding to their promoters. Overexpression of PpNAP4 resulted in a heightened expression of these genes and accelerated Chl degradation. Notably, PpNAP4 also positively regulated the expression of PpANS and PpMYB10.1, one key structural gene and a core transcriptional regulator of anthocyanin synthesis, thereby contributing to fruit coloration. In summary, our findings elucidate that PpNAP4 serves as a pivotal regulator in determining the final skin color of peach by orchestrating Chl degradation and anthocyanin accumulation through direct activation of multiple CCGs and anthocyanin related genes.

Enhancing the color and astringency of red wines through white grape seeds addition: Repurposing wine production byproducts.

The clear juice fermentation technique for white wines suggests that white grape seeds, rich in flavan-3-ols and proanthocyanidins, are not effectively utilized in the winemaking process. This study incorporated 'Gewürztraminer' grape seeds into 'Cabernet Sauvignon' must before cold soak to investigate how the resultant red wines' phenolic compound profiles, color, and astringency were affected. The results showed that adding seeds primarily inhibited the leaching of flavan-3-ols from both skins and seeds. A significant increase in the levels of flavan-3-ols, tannins, and phenolic acids, as well as direct and aldehyde-bridged flavan-3-ol-anthocyanin polymers, were observed in the wines with additional seeds. This led to the improvement in the wine' red hue and its resistance to SO2 bleaching. Furthermore, the wine added with seeds exhibited stronger astringency compared to those without. The findings provide a promising winemaking strategy to improve color stability and intensify the astringency of red wines through the utilization of grape seeds.

Combined Metabolomics and Network Pharmacology Analysis Reveal the Effect of Rootstocks on Anthocyanins, Lipids, and Potential Pharmacological Ingredients of Tarroco Blood Orange (Citrus sinensis L. Osbeck).

The benefits of citrus fruits are strongly associated with their secondary metabolites. In this study, we conducted widely targeted metabolomics analyses to compare the variability of the ingredients in four scion-rootstock combinations. A total of 376 differential metabolites were obtained by a multivariate statistical analysis, and a KEGG pathway analysis showed that the enriched metabolic pathways were mainly related to the biosynthesis of flavonoids as well as lipid metabolism. The anthocyanin-targeted metabolomic features showed that cyanidin 3-O-glucoside, cyanidin 3-O-(6-O-malonyl-beta-D-glucoside), cyanidin 3-O-sophoroside, and cyanidin 3-O-xyloside were the pigments responsible for the red color of Tarocco. A lipid metabolomics analysis revealed that when Tarocco was hetero-grafted with rootstock H, there was an increase in the content of each lipid subclass, accompanied by an increase in the levels of unsaturated fatty acids, including polyunsaturated linoleic and linolenic acids, thus impacting the ratio of unsaturated fatty acids to saturated fatty acids. Additionally, we determined their antioxidant capacity ('Trifoliate orange' (Z) > 'Citrange' (ZC) > 'Hongju' (H) > 'Ziyang Xiangcheng' (X)) using in vitro assays. Finally, we utilized a network pharmacology analysis to explore the antioxidant mechanisms and potential pharmacological ingredients; we obtained 26 core targets proteins and 42 core metabolites associated with oxidative damage, providing a basis for future preventive and therapeutic applications of these metabolites.

A sandwich-type photoelectrochemical biosensor based on anthocyanin-sensitized ZnO/P5FIn heterojunction for the sensitive detection of CYFRA21-1.

. A sandwich-type photoelectrochemical (PEC) immunosensor based on a ZnO/poly(5-formylindole) (P5FIn)/anthocyanin heterostructure was developed to achieve sensitive background-free detection of the tumor marker CYFRA21-1. ZnO with good photovoltaic properties is combined with narrow bandgap P5FIn to form a p-n type heterojunction. This structure reduces the electron-hole pair recombination, thereby enhancing the photocurrent response of the composite. Anthocyanidins are environmentally friendly natural compounds with excellent antioxidant, redox properties, and remarkable electrochemical activity. After sensitization by anthocyanins, the absorption and utilization of visible light in the composites are enhanced, further improving the PEC luminescence efficiency of the materials. Additionally, boron nitride quantum dots (BN QDs) are combined with Ab2 via polydopamine (PDA) as a secondary antibody marker, enhancing its sensitivity. The biosensor exhibited a linear detection range of 0.001-100 ng mL-1 with a limit of detection (LOD) of 0.00033 ng mL-1. Furthermore, this biosensor demonstrates excellent selectivity, reproducibility, and stability, as well as successful results in analyzing actual human serum samples. This approach provides a feasible method for tumor marker detection.

Physiological and molecular responses of 'Hamlin' sweet orange trees expressing the VvmybA1 gene under cold stress conditions.

MAIN CONCLUSION: Overexpression of VvmybA1 transcription factor in 'Hamlin' citrus enhances cold tolerance by increasing anthocyanin accumulation. This results in improved ROS scavenging, altered gene expression, and stomatal regulation, highlighting anthocyanins' essential role in citrus cold acclimation. Cold stress is a significant threat to citrus cultivation, impacting tree health and productivity. Anthocyanins are known for their role as pigments and have emerged as key mediators of plant defense mechanisms against environmental stressors. This study investigated the potential of anthocyanin overexpression regulated by grape (Vitis vinifera) VvmybA1 transcription factor to enhance cold stress tolerance in citrus trees. Transgenic 'Hamlin' citrus trees overexpressing VvmybA1 were exposed to a 30-day cold stress period at 4 °C along with the control wild-type trees. Our findings reveal that anthocyanin accumulation significantly influences chlorophyll content and their fluorescence parameters, affecting leaf responses to cold stress. Additionally, we recorded enhanced ROS scavenging capacity and distinct expression patterns of key transcription factors and antioxidant-related genes in the transgenic leaves. Furthermore, VvmybA1 overexpression affected stomatal aperture regulation by moderating ABA biosynthesis, resulting in differential responses in a stomatal opening between transgenic and wild-type trees under cold stress. Transgenic trees exhibited reduced hydrogen peroxide levels, enhanced flavonoids, radical scavenging activity, and altered phytohormonal profiles. These findings highlighted the role of VvmybA1-mediated anthocyanin accumulation in enhancing cold tolerance. The current study also underlines the potential of anthocyanin overexpression as a critical regulator of the cold acclimation process by scavenging ROS in plant tissues.

A pH-sensitive film based on chitosan/gelatin and anthocyanin from Zingiber striolatum Diels for monitoring fish freshness.

As a new type of packaging method, the anthocyanin-based pH-sensitive indicator film has gained much attention owing to low cost, small size, and visually informative property. In this study, an intelligent film based on chitosan/gelatin (CG) matrix with Zingiber striolatum Diels (ZSD) anthocyanin for fish freshness monitoring was developed. The film properties, including thickness, moisture content, color, mechanical properties, UV-vis light barrier property, as well as pH and ammonia sensitivity, were evaluated. The CG-ZSD films exhibited a more compact structure when compared with the CG film. The CG-ZSD20 film showed the highest elongation at break (6.33 ± 0.62%) and lowest tensile strength (20.0 ± 0.58 MPa). FTIR spectra revealed the strong hydrogen bond interactions between ZSD and polymer matrix. Film incorporated with 15% anthocyanin extract has increased melting temperature at 118.9 °C, and a lower weight loss (13.8%) at melting temperature. In pH 1-14 buffer, the color of CG-ZSD films underwent a significant change from red to yellow-green. The CG-ZSD15 film was utilized for monitoring fish freshness and showed visible color changes from deep purple to brown. The total volatile basic nitrogen content and pH value changes of fish were closely related to the visual color changes in film. This demonstrated that the film was a highly pH-sensitive film for quantifying fish freshness in real-time.

Metabolomics Uncovers the Mechanisms of Nitrogen Response to Anthocyanins Synthesis and Grain Quality of Colored Grain Wheat (Triticum aestivum L.).

Nitrogen (N) is a key factor for plant growth and affects anthocyanin synthesis. This study aimed to clarify the potential mechanisms of N levels (LN, 0 kg·ha-1; MN, 150 kg·ha-1; HN, 225 kg·ha-1) in anthocyanin synthesis and grain quality of colored grain wheat. HN increased the yield component traits and grain morphology traits in colored grain wheat while decreasing the processing and nutrient quality traits. Most quality traits were significantly negatively correlated with the yield composition and morphological traits. Anthocyanin was more accumulated under LN conditions, but other related yield and morphological traits of colored grain wheat declined. The anthocyanin content was the highest in blue wheat, followed by that in purple wheat. Cyanidin-3-O-(6-O-malonyl-ß-d-glucoside) and cyanidin-3-O-rutinoside were the predominant anthocyanins in blue and purple wheat. The identified anthocyanin-related metabolites were associated with flavonoid biosynthesis, anthocyanin biosynthesis, and secondary metabolite biosynthesis. Therefore, the study provided information for optimizing nitrogen fertilizer management in producing high quality colored wheat and verified the close relationship between anthocyanin and low N condition.

Design of a liposome casein hydrogel as an efficient front-end homeostatic anthocyanin loading system.

Proteins have been studied and applied to improve the stability of anthocyanins (ACNs), but the changes in the pH microenvironment during the preparation of steady-state systems are often ignored, and more attention is given to the stability of the system after preparation. In this study, we propose the "anthocyanin front-end homeostasis strategy", which involves designing a system can protect anthocyanins under acidic conditions so that more anthocyanin prototypes can be loaded inside the protein. Anthocyanins are encapsulated in liposomes (Lip) at pH 3.0 and combined with casein methacrylate (CSMA) to form Anthocyanin-loaded liposomes/CSMA hydrogel (Lip@ACNs/CSMA), with good physical properties and good blood compatibility. The system increased the hydrogen peroxide scavenging capacity by 1.16 mg Vc equiv./mg ACNs and the cellular antioxidant activity by 17.55 µM quercetin/100 mg ACNs, the photo and thermal storage stability increased by 36.50 % and 30.71 %, the digestive rate increased by 17.50 %, and the biological availability increased by 0.0049 mg/mL. This study designed a liposome casein hydrogel as an efficient front-end homeostatic anthocyanin loading system and provided a new approach for improving the stability of anthocyanins.

RBOH-dependent signaling is involved in He-Ne laser-induced salt tolerance and production of rosmarinic acid and carnosol in Salvia officinalis.

BACKGROUND: In the past two decades, the impacts of Helium-Neon (He-Ne) laser on stress resistance and secondary metabolism in plants have been studied, but the signaling pathway which by laser regulates this process remains unclear. Therefore, the current study sought to explore the role of RBOH-dependent signaling in He-Ne laser-induced salt tolerance and elicitation of secondary metabolism in Salvia officinalis. Seeds were primed with He-Ne laser (6 J cm- 2) and peroxide hydrogen (H2O2, 5 mM) and 15-old-day plants were exposed to two salinity levels (0, 75 mM NaCl). RESULTS: Salt stress reduced growth parameters, chlorophyll content and relative water content (RWC) and increased malodialdehyde (MDA) and H2O2 contents in leaves of 45-old-day plants. After 48 h of salt exposure, higher transcription levels of RBOH (encoding NADPH oxidase), PAL (phenylalanine ammonia-lyase), and RAS (rosmarinic acid synthase) were recorded in leaves of plants grown from seeds primed with He-Ne laser and/or H2O2. Despite laser up-regulated RBOH gene in the early hours of exposing to salinity, H2O2 and MDA contents were lower in leaves of these plants after 30 days. Seed pretreatment with He-Ne laser and/or H2O2 augmented the accumulation of anthocyanins, total phenol, carnasol, and rosmarinic acid and increased total antioxidant capacity under non-saline and more extensively at saline conditions. Indeed, these treatments improved RWC, and K+/Na+ ratio, enhanced the activities of superoxide dismutase and ascorbate peroxidase and proline accumulation, and significantly decreased membrane injury and H2O2 content in leaves of 45-old-day plants under salt stress. However, applying diphenylene iodonium (DPI as an inhibitor of NADPH oxidase) and N, N-dimethyl thiourea (DMTU as a H2O2 scavenger) after laser priming reversed the aforementioned effects which in turn resulted in the loss of laser-induced salt tolerance and secondary metabolism. CONCLUSIONS: These findings for the first time deciphered that laser can induce a transient RBOH-dependent H2O2 burst, which might act as a downstream signal to promote secondary metabolism and salt stress alleviation in S. officinalis plants.

The introgression of BjMYB113 from Brassica juncea leads to purple leaf trait in Brassica napus.

The purple leaves of Brassica napus are abundant in anthocyanins, which are renowned for their role in conferring distinct colors, stress tolerance, and health benefits, however the genetic basis of this trait in B. napus remains largely unelucidated. Herein, the purple leaf B. napus (PL) exhibited purple pigments in the upper epidermis and a substantial increase in anthocyanin accumulation, particularly of cyanidin, compared to green leaf B. napus (GL). The genetic control of the purple leaf trait was attributed to a semi-dominant gene, pl, which was mapped to the end of chromosome A03. However, sequencing of the fragments amplified by the markers linked to pl indicated that they were all mapped to chromosome B05 from B. juncea. Within this B05 chromosomal segment, the BjMYB113 gene-specific marker showed perfect co-segregation with the purple leaf trait in the F2 population, suggesting that the BjMYB113 introgression from B. juncea was the candidate gene for the purple leaf trait in B. napus. To further verify the function of candidate gene, CRISPR/Cas9 was performed to knock out the BjMYB113 gene in PL. The three myb113 mutants exhibited evident green leaf phenotype, absence of purple pigments in the adaxial epidermis, and a significantly reduced accumulation of anthocyanin compared to PL. Additionally, the genes involved in positive regulatory (TT8), late anthocyanin biosynthesis (DFR, ANS, UFGT), as well as transport genes (TT19) were significantly suppressed in the myb113 mutants, further confirming that BjMYB113 was response for the anthocyanin accumulation in purple leaf B. napus. This study contributes to an advanced understanding of the regulation mechanism of anthocyanin accumulation in B. napus.