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Objective: To analyze the effect of the juice obtained from two varieties of sweet orange (Citrus sinensis L. Osbeck), Moro (a blood orange) and Navelina (a blond orange), on fat accumulation in mice fed a standard or a high-fat diet (HFD). Methods: Obesity was induced in male C57/Bl6 mice by feeding a HFD. Moro and Navelina juices were provided instead of water. The effect of an anthocyanin-enriched extract from Moro oranges or purified cyanidin-3-glucoside (C3G) was also analyzed. Body weight and food intake were measured regularly over a 12-week period. The adipose pads were weighted and analyzed histologically; total RNA was also isolated for microarray analysis. Results: Dietary supplementation of Moro juice, but not Navelina juice significantly reduced body weight gain and fat accumulation regardless of the increased energy intake because of sugar content. Furthermore, mice drinking Moro juice were resistant to HFD-induced obesity with no alterations in food intake. Only the anthocyanin extract, but not the purified C3G, slightly affected fat accumulation. High-throughput gene expression analysis of fat tissues confirmed that Moro juice could entirely rescue the high fat-induced transcriptional reprogramming. Conclusion: Moro juice anti-obesity effect on fat accumulation cannot be explained only by its anthocyanin content. Our findings suggest that multiple components present in the Moro orange juice might act synergistically to inhibit fat accumulation.

Some blood orange juice (BOJ) flavanones may have antioxidant, anti-inflammatory, anti-allergic, hypolipidaemic, vasoprotective and anticarcinogenic properties. The aim of the present study was to evaluate the pharmacokinetics of hesperetin and naringenin in human subjects after BOJ intake. In a cross-over study, seven healthy female volunteers consumed 150 and 300 ml BOJ corresponding to about 51–102 mg hesperetin and to 6–12 mg naringenin, respectively. Plasma samples were collected before, each hour for 8 h and 24 h after BOJ administration and analysed for their content of hesperetin and naringenin by liquid chromatography–MS/MS. The plasma concentrations of these compounds were dose dependent and the peak concentration (Cmax) was reached in 5·1 (sd 0·6) h after BOJ intake. The Cmax of hesperetin was 43·4 (sd 32·4) and 79·8 (sd 60·1) ng/ml after 150 and 300 ml intake, respectively. For naringenin the plasma peak was 16·4 (sd 11·9) and 34·0 (sd 20·6) ng/ml. Moreover, the conjugated forms of these flavanones represent more than 95 % of the plasma concentration. The results indicate that both hesperetin and naringenin are bioavailable after BOJ intake; naringenin seemingly more so than hesperetin.

Red blush is one of the key quality markers of blood orange fruit ( Citrus sinensis L. Osbeck). Therefore, the influence of pre- (1, 2, and 3 mM) and postharvest (0.5, 1, and 1.5 mM) methyl jasmonate (MeJA) and salicylic acid (SA) (1, 2, and 3 mM) treatments on redness, colouring pigments, and health-promoting compounds in cold stored ‘Tarocco Ippolito’ blood oranges was investigated. Preharvest application of 3 mM MeJA significantly increased rind citrus colour index (CCI) by 43.3% and colorimetric coordinate a* (redness) by 3.1% and decreased h ° (hue angle, 0 = red), L* (lightness), and b* (yellowness) by 13.7%, 12.6%, and 23.4%, respectively. This treatment also enhanced CCI (57.4%), reduced h ° (16.5%) and L* (36.4%), and increased total anthocyanins (88%), monomeric anthocyanins (117%), and flavonoids (77%) in the juice. Postharvest dip of 1.5 mM MeJA (5 min) improved CCI of juice (53.6%) and a* (5.5%), reduced h ° (15.9%), L* (19.8%), and b* (19.4%), and increased total anthocyanins (66.7%), monomeric anthocyanins (74%), and flavonoids (23.4%) in the juice. Preharvest application of 1 mM SA increased rind CCI (50.8%) and reduced L* (13.6%), b* (16.4%), and h ° (29.5%). All preharvest SA treatments significantly increased total phenolics in the juice. Lycopene was increased (61.7%) by 5 min postharvest dip of 3 mM SA. In conclusion, 3 mM preharvest spray application of MeJA four weeks before harvest and postharvest dip of 1.5 mM MeJA (5 min) are effective in improving red colour of rind and juice and health-promoting compounds in blood orange juice.

Anthocyanins are natural pigments that give a red, purple, and blue color to many plant, flower, fruit, and vegetable species. Their presence within the genus Citrus was first reported in 1916, and it is well-known that the red color of the flesh and rind of blood (red or pigmented) oranges (Citrus sinensis L. Osbeck) is due to the presence of anthocyanins. They are also present in the young shoots, flowers, and peel of lemon (Citrus limon (L.) Burm. f.), citron (Citrus medica L.), and other citrus species. Since then, the scientific community has expressed increasing interest in studying their profile and distribution, with many published studies focused on the quali-quantitative pattern in the different vegetative tissues belonging to the genus Citrus. Moreover, with the discovery of their relevant antioxidant activity, owing to their ability to capture free radicals, much research has been performed in the last two decades on their radical scavenging power, in vitro and in vivo biological properties, and anticarcinogenic capacity, also focusing attention on their bioavailability for humans. The present work is intended as a comprehensive review of the advances in scientific research on anthocyanin pigments naturally occurring within the genus Citrus, including their natural distribution, antioxidant capacity, bioavailability, and biological value and properties. The scientific evidence herein reported can be used to further increase the knowledge of this class of compounds and represents a valuable and comprehensive contribution to promoting anthocyanin-rich citrus fruit consumption as a healthy dietary habit.

Background Anthocyanins are the most important compounds for nutritional quality and economic values of blood orange. However, there are few reports on the pre-harvest treatment accelerating the accumulation of anthocyanins in postharvest blood orange fruit. Here, we performed a comparative transcriptome and metabolomics analysis to elucidate the underlying mechanism involved in seasonal drought (SD) treatment during the fruit expansion stage on anthocyanin accumulation in postharvest ‘Tarocco’ blood orange fruit. Results Our results showed that SD treatment slowed down the fruit enlargement and increased the sugar accumulation during the fruit development and maturation period. Obviously, under SD treatment, the accumulation of anthocyanin in blood orange fruit during postharvest storage was significantly accelerated and markedly higher than that in CK. Meanwhile, the total flavonoids and phenols content and antioxidant activity in SD treatment fruits were also sensibly increased during postharvest storage. Based on metabolome analysis, we found that substrates required for anthocyanin biosynthesis, such as amino acids and their derivatives, and phenolic acids, had significantly accumulated and were higher in SD treated mature fruits compared with that of CK. Furthermore, according to the results of the transcriptome data and weighted gene coexpression correlation network analysis (WGCNA) analysis, phenylalanine ammonia-lyase ( PAL3 ) was considered a key structural gene. The qRT-PCR analysis verified that the PAL3 was highly expressed in SD treated postharvest stored fruits, and was significantly positively correlated with the anthocyanin content. Moreover, we found that other structural genes in the anthocyanin biosynthesis pathway were also upregulated under SD treatment, as evidenced by transcriptome data and qRT-PCR analysis. Conclusions The findings suggest that SD treatment promotes the accumulation of substrates necessary for anthocyanin biosynthesis during the fruit ripening process, and activates the expression of anthocyanin biosynthesis pathway genes during the postharvest storage period. This is especially true for PAL3 , which co-contributed to the rapid accumulation of anthocyanin. The present study provides a theoretical basis for the postharvest quality control and water-saving utilization of blood orange fruit.

Background Organic acids and anthocyanins are the most important compounds for the flavor and nutritional quality of citrus fruit. However, there are few reports on the involvement of co-regulation of citrate and anthocyanin metabolism. Here, we performed a comparative transcriptome analysis to elucidate the genes and pathways involved in both citrate and anthocyanin accumulation in postharvest citrus fruit with ‘Tarocco’ blood orange (TBO; high accumulation) and ‘Bingtangcheng’ sweet orange (BTSO; low accumulation). Results A robust core set of 825 DEGs were found to be temporally associated with citrate and anthocyanin accumulation throughout the storage period through transcriptome analysis. Further according to the results of weighted gene coexpression correlation network analysis (WGCNA), the turquoise and brown module was highly positively correlated with both of the content of citrate and anthocyanin, and p-type ATPase ( PH8 ), phosphoenolpyruvate carboxylase kinase ( PEPCK ), chalcone isomerase ( CHI ), flavanone 3-hydroxylase ( F3H ), flavonoid 3’-hydroxylase ( F3’H ) and glutathione S transferase ( GST ) were considered key structural genes. Moreover, MYB family transcription factor ( PH4 ), Zinc finger PHD-type transcription factor ( CHR4 , HAC12 ), Zinc finger SWIM-type transcription factor ( FAR1 ) and Zinc finger C3H1-type transcription factor ( ATC3H64 ) were considered hub genes related to these structural genes. Further qRT-PCR analysis verified that these transcription factors were highly expressed in TBO fruit and their expression profiles were significantly positively correlated with the structural genes of citrate and anthocyanin metabolism as well as the content of citrate and anthocyanin content. Conclusions The findings suggest that the CHR4, FAR1, ATC3H64 and HAC12 may be the new transcription regulators participate in controlling the level of citrate and anthocyanin in postharvest TBO fruit in addition to PH4. These results may providing new insight into the regulation mechanism of citrate and anthocyanin accumulation in citrus fruit.

Background Transcriptome and proteome analyses on fruit pulp from the blood orange ‘Zaohong’ and the navel orange ‘twenty-first century’ were performed to study Citrus sinensis quality-related molecular changes during consecutive developmental periods, including young fruit, fruit-coloring onset and fruit delayed-harvest for two months, during which fruit remained on the trees. Results The time-course analysis for the fruit developmental periods indicated a complex, dynamic gene expression pattern, with the numbers of differentially expressed genes (DEGs) between the two cultivars being 119, 426 and 904 at the three continuous stages tested during fruit development and ripening. The continuous increase in total soluble solids over the course of fruit development was correlated with up-regulated sucrose phosphate synthase (SPS) transcription levels in both cultivars. Eleven differentially expressed genes between the two cultivars involved in the flavonoid pathway were significantly enriched at the onset of the fruit-coloring stage when anthocyanins were detected in blood orange alone. Among 5185 proteins, 65 up-regulated and 29 down-regulated proteins were co-expressed with their cognate mRNAs with significant transcription and protein expression levels when the fruits from the two cultivars were compared at the fruit delayed-harvest stage. Additionally, important genes participating in the γ-aminobutyric acid (GABA) shunt were activated in blood orange at two significant expression levels in the fruit delayed-harvest stage. Thus, organic acids in fruit continuously decreased during this stage. Conclusions This research was the first to provide a more comprehensive understanding of the differentially expressed genes involved in anthocyanin, sucrose and citrate metabolism at the transcriptome and proteome levels in C. sinensis , especially during the fruit delayed-harvest stage.

Blood oranges have high concentrations of bioactive compounds that are beneficial to health. In Europe, the cultivation of blood oranges is increasing due to their excellent nutritional properties. In Citrus crops, rootstocks play an important role in juice and can increase the content of bioactive compounds. The morphological, qualitative and nutritional parameters were analyzed in cultivars ‘Tarocco Ippolito’, ‘Tarocco Lempso’, ‘Tarocco Tapi’ and ‘Tarocco Fondaconuovo’ grafted onto Citrus macrophylla and Citrus reshni. ‘Tarocco Lempso’ grafted onto Citrus macrophylla obtained the highest values of weight (275.78 g), caliber (81.37 mm and 76.79 mm) and juice content (162.11 g). ‘Tarocco Tapi’ grafted onto Citrus reshni obtained the most interesting qualitative parameters (15.40 °Brix; 12.0 MI). ‘Tarocco Lempso’ grafted onto Citrus reshni obtained the most intense red juice (a* = 9.61). Overall, the highest concentrations of primary metabolites were in proline, aspartate, citric acid, and sucrose. The results showed that ‘Tarocco Ippolito’ juice grafted onto Citrus reshni had the highest levels of total hydroxycinnamic acids (263.33 mg L−1), total flavones (449.74 mg L−1) and total anthocyanins (650.42 mg L−1). To conclude, ‘Tarocco Lempso’ grafted onto Citrus macrophylla obtained the best values of agronomic parameters, and the cultivars grafted onto Citrus reshni obtained significantly higher concentrations in primary and secondary metabolites.

The commercial value of blood oranges ( ) is closely linked to the intensity of red pigmentation in the peel and flesh, driven by the accumulation of anthocyanins. While light is a crucial environmental factor for anthocyanin synthesis, the specific effects of different light spectra, particularly ultraviolet (UV) radiation, on peel pigmentation have not been fully elucidated. In this study, the effects of light spectra on anthocyanin biosynthesis in blood orange peel were systematically studied through three treatments of visible light (VL), UV and complete shading (CK). These treatments were combined with transcriptome, anthocyanin targeted metabolome and weighted gene coexpression network analysis (WGCNA). After 40 days, UV-treated fruit exhibited significantly higher anthocyanin content and color index (CI) than other treatments, with a significantly positive correlation between the two. Metabolomics identified four key anthocyanins, including cyanidin-3-o-glucoside and its 2 derivatives, as the primary contributors to pericarp coloration, with their levels significantly increased under UV exposure. WGCNA screened three core gene modules closely associated with anthocyanin metabolism, and further identified three glycosyltransferase genes (ugt79b1, bz1 and GT1) as hub genes involved in anthocyanin accumulation. This study demonstrates that UV light enhanced anthocyanin synthesis in blood orange peel by activating downstream glycosylation pathways, providing both a theoretical basis and technical approach for improving commercially market value of blood orange through light regulation.

Tarocco is a prevalent blood-orange variety in China, has a bud variant identified in Wenzhou City. To characterize the quality traits and molecular mechanisms underlying inhibition of anthocyanin synthesis in this variety, we collected fruits of Tarocco (WT) and the bud variant Ouya (MT) at nine developmental stages. Their anthocyanin, soluble sugar, and organic acid profiles were examined, and transcriptomes and metabolites were analyzed at three developmental stages. The results revealed that MT is a new blood orange variety with weak anthocyanins and a better sugar-acid ratio than the WT. The content of anthocyanin in MT fruits were significantly lower than those in WT fruits, especially cyanidin-like anthocyanins, while the flavone contents exhibited no remarkable variation. A total of 64 differentially expressed genes (DEGs), including five transcription factors (TFs), five methylation-related genes, and one flavonoid biosynthesis gene, were identified between WT and MT at three fruit developmental stages. The potential regulatory networks of these TFs were further constructed using weighted gene co-expression network analysis.Furthermore, in MT fruit treated with the 5-azacytidine, we observed hypomethylation of anthocyanins accumulated in the pulp and the promoters and genebodies of some anthocyanin synthesis-related genes. These results provide new insights into the influence of DNA methylation on anthocyanin accumulation in MT and also provide support for the promotion of MT as a new variety.