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Structural transformation of cyanidin-3-O-glycoside as a function of pH. The three cyanidin species are given with their expected colours as outlined. Deprotonation from the C5-OH has been omitted for simplicity.
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Choices of blue food colourants are extremely limited, with only two options in the USA, synthetic blue no. 1 and no. 2, and a third available in Europe, patent blue V. The food industry is investing heavily in finding naturally derived replacements, with limited success to date. Here, we review the complex and multifold mechanisms whereby blue pig...
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... colour therefore is the result of the ratio of the three coloured species in equilibrium at any specific pH. At very acidic pH values (around pH < 2), the formation of the red flavylium ion (AH + ) is favoured (Figure 1). This species is fully protonated and has a delocalised positive charge across the chromophore. ...
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... the pH increases above pK a1 , the first deprotonation occurs, converting flavylium ions into the neutral quinonoid base (A) and the colour changes from red to purple. At pH values above pK a2 , the quinonoid base is deprotonated further, forming the anionic quinonoid base (A − ) with a negative delocalised charge [16] (Figure 1). This deprotonation sequence is accompanied by two shifts in λ max , the first (AH + → A) typically of 20-30 nm and the second (A→ A − ) a further shift of 50-60 nm [15]. ...
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... diversity of colours observed in floral systems cannot be rationalised by differences in chemical substituent groups and the pH equilibrium alone; without interactions that stabilise the chromophore at physiological pH within the vacuole (which is usually about pH 5.5), most flowers would be devoid of their attractive hues and would instead be colourless or pale yellow (as a result of hemiketal and chalcone formation, respectively; Figure 1). Anthocyanins are large, planar compounds which lend themselves to the formation of non-covalent interactions with molecules termed co-pigments (Figure 2). ...
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... which are acylated with aromatic hydroxycinnamic (HCA) derivatives form non-covalent intramolecular associations with their covalently linked HCA moieties, which fold over the chromophore, protecting the chromophore from hydration [37] (Figure 9). These interactions can be loosely categorised into two types: type 1 where the anthocyanin forms intramolecular co-pigmentation interactions with HCAs both above and below the chromophore, and type 2 where the anthocyanin forms intramolecular interactions on only one plane, leaving one plane exposed for the formation of self-association interactions with other anthocyanins, or co-pigmentation with a non-anthocyanin co-pigment [25] (Figure 10). ...
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... acylation also increases the strength of π stacking interactions and slows irreversible colour loss by chemical degradation involving cleavage of the C-ring, although these latter processes are not well understood [39]. Consequently, aromatic acylation causes big increases in the stability of anthocyanin pigments, as illustrated in Figure 11 for extracts of di-acylated anthocyanins from the desert bluebell (Phacelia campanularia) and quadri-acylated anthocyanins from butterfly pea (Clitoria ternatea) compared to the non-acylated cyanidin 3-O-rutinoside. ...
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... observations have been interpreted in terms of reduced glycosylation favouring AVI formation, which fits well with in vitro observations that 5-O-glycosylation reduces AVI formation in vivo [44]. The role of AVI formation in patterning the flowers of lisianthus is illustrated in Figure 12. Anthocyanins from blue flowers commonly also contain aliphatic acyl groups, such as malonic acid. ...
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... best studied cases of fuzzy metal complexes are those present in Hydrangea macrophylla. These flowers range in colour from red to mauve to blue, and all contain delphinidin 3-O-glucoside, co-pigments (mainly 3-O-caffeoyl and 3-O-p-coumaroyl quinic acids), and Al 3+ in differing concentrations ( Figure 13). Figure 13. ...
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... flowers range in colour from red to mauve to blue, and all contain delphinidin 3-O-glucoside, co-pigments (mainly 3-O-caffeoyl and 3-O-p-coumaroyl quinic acids), and Al 3+ in differing concentrations ( Figure 13). Figure 13. Hydrangeas produce blue flowers due to the formation of metal chelates between delphinidin, co-pigments, and Al 3+ . ...
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... concentration of caffeoyl/pcoumaroyl 3-O-quinic acids, which serve as co-pigments, are high in several cell layers of the sepals of hydrangea. However, it is the high levels of aluminium in the subepidermal layer that confer the specificity of blue, achieved through the association of anthocyanins, co-pigments, and Al 3+ ions in the subepidermal layer of blue flowers ( Figure 13). In the comparator red-flowered variety, levels of Al 3+ were found to be much lower in the subepidermal cell layer of the sepals [26]. ...
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... formation of these complexes requires similar structural properties to those seen in the more typical metal-coordination with anthocyanins-two unsubstituted neighbouring hydroxyl groups on the B-ring. Both the anthocyanins and flavones self-associate in pairs with distances between the aromatic rings of the respective pair of approximately 3.3 Å, indicative of hydrophobic interaction [51] (Figure 14). To date, only a few metalloanthocyanins have been identified with colours ranging from purple to blue. ...
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... is an attractive force between electrical quadrupoles, overpowering the repulsion of π-electron clouds. Where aromatic systems share a similar electron density, σ-π interactions are favourable and should therefore favour an edge-to-face or off-centre parallel arrangement, disfavouring face-centred arrangements (Figure 15). The addition of electron-withdrawing substituents to an aromatic (arene) polarises π electron density away from the aromatic centre, reversing the direction of the quadrupole [23]. ...
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... addition of electron-withdrawing substituents to an aromatic (arene) polarises π electron density away from the aromatic centre, reversing the direction of the quadrupole [23]. Pairs of electron-rich and electron-deficient aromatics preferentially pair in a face-centred arrangement, referred to as "aromatic donor-acceptor interaction" (Figure 15). ...
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... molecules solvating anthocyanins and their co-pigments form a highly organised hydrogen bond network around the aromatic intramolecular region (hydrophobic cavity) [25]. As the pigment and co-pigment come together, the hydrogen bonds between water molecules in the solvation shell are broken and released from the hydrophobic cavity, increasing the system entropy and enthalpy ( Figure 16). The barrier to complex formation is further reinforced by the required decrease in the system entropy (requiring energy) as the acyl group rotates about the C6" axis to form a highly ordered structure. ...
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... barrier to complex formation is further reinforced by the required decrease in the system entropy (requiring energy) as the acyl group rotates about the C6" axis to form a highly ordered structure. Figure 16. Classical and non-classical hydrophobic forces involved in the formation of copigmentation complexes. ...
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... the most effective approach to identifying better natural blue food colourants may be to continue screening blue flowers and fruits to find the anthocyanins with the best tones over a wide range of pH values and good stability, of the order of months rather than days, under conditions used for cooking and processing. Such candidates include the ternatin extracts produced from the flowers of butterfly pea, which have remarkable stability even at high pH ( Figure 17C) and in cooked goods compared to spirulina natural blue and the red betacyanins of beetroot natural food colourants ( Figure 17F). ...
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... the most effective approach to identifying better natural blue food colourants may be to continue screening blue flowers and fruits to find the anthocyanins with the best tones over a wide range of pH values and good stability, of the order of months rather than days, under conditions used for cooking and processing. Such candidates include the ternatin extracts produced from the flowers of butterfly pea, which have remarkable stability even at high pH ( Figure 17C) and in cooked goods compared to spirulina natural blue and the red betacyanins of beetroot natural food colourants ( Figure 17F). ...
Citations
... The stability of anthocyanins enhanced by acylation depends on the acyl group and its structure, size, number, and attachment sites [24]. The stabilization of ternatins by hydroxycinnamic acids occurs through intramolecular or intermolecular co-pigmentation reactions, resulting in a reduction in polarity and steric hindrance to prevent nucleophilic attack [4]. The robust stability of anthocyanins derived from BF extracts enhances their antioxidant activities. ...
The antioxidant properties of butterfly pea flower (BF), which is rich in natural anthocyanins, have garnered significant attention. The impact of digestion and metabolism on BF extracts and evaluate their subsequent antioxidant activities in vivo were explored in the present study. After in vitro digestion, 42.03 ± 2.74% of total anthocyanins from BF extracts remained, indicating a negative influence of the digestion process on the bioaccessibility of phenolic compounds derived from BF. Furthermore, UPLC-LTQ-Orbitrap-MS2 analysis identified a total of four prototypes and twenty-seven metabolites in rat plasma or urine samples following the intake of BF extracts. The kinetics of key metabolites including delphinidin 3-glucoside (D3G), cyanidin-3-glucoside (C3G), and 4-hydroxybenzoic acid were subsequently determined in blood, and the Cmax values were 69.034 ± 8.05 nM and 51.65 ± 3.205 nM. These key metabolites derived from BF anthocyanins, including C3G and D3G, and flavonoid quercetin exhibited main antioxidant attributes that improved the plasmic and hepatic activities of various antioxidant enzymes and the total antioxidant capacity (T-AOC) and malondialdehyde (MDA) in a D-galactose-induced rat model. These findings provide insights into the bioaccessibility and bioavailability of bioactive constitutes derived from BF extracts, which are crucial for determining the actual efficacy of BF as well as developing functional foods based on BF.
... They stabilize the chromophore, making the pigments less susceptible to color loss under varying pH levels and light exposure (Bakowska-Barczak, 2005). Additionally, acyl groups act as barriers against enzymatic degradation and oxidation, which commonly affect non-acylated anthocyanins (Houghton, Appelhagen, & Martin, 2021). This finding provides valuable insights into the stability of natural colorants derived from butterfly pea extract, offering understanding of colorant changes during storage. ...
... Due to their chemical instability, free-state anthocyanins rarely exist in nature and are easily degraded during processing and storage [2][3][4][5]. There are different methods by which anthocyanins can be stabilized [5][6][7]. Copigmentation, for example, is a strategy that has a racted considerable a ention from scientists, as this process has been commonly observed in many anthocyanins acylated with hydroxycinnamic aromatic derivatives. It involves non-covalent interactions (primarily π-π stacking) between the anthocyanidin backbone and its covalently bound acylation fractions, which can protect the chromophore from hydration and result in a bathochromic change of the visible band. ...
... Due to their chemical instability, free-state anthocyanins rarely exist in nature and are easily degraded during processing and storage [2][3][4][5]. There are different methods by which anthocyanins can be stabilized [5][6][7]. Copigmentation, for example, is a strategy that has attracted considerable attention from scientists, as this process has been commonly observed Alzheimer's disease (AD) is the most common cause of dementia and an increasingly common cause of morbidity and mortality in the elderly. In many studies, it has been discovered that resveratrol (RV) has many potential health benefits, like antioxidant, cardioprotective, neurological, anti-inflammatory, antiplatelet, blood glucose-lowering, and anticancer activities. ...
Anthocyanins are colored water-soluble plant pigments. Upon consumption, anthocyanins are quickly absorbed and can penetrate the blood–brain barrier (BBB). Research based on population studies suggests that including anthocyanin-rich sources in the diet lowers the risk of neurodegenerative diseases. The copigmentation caused by copigments is considered an effective way to stabilize anthocyanins against adverse environmental conditions. This is attributed to the covalent and noncovalent interactions between colored forms of anthocyanins (flavylium ions and quinoidal bases) and colorless or pale-yellow organic molecules (copigments). The present work carried out a theoretical study of the copigmentation process between cyanidin and resveratrol (CINRES). We used three levels of density functional theory: M06-2x/6-31g+(d,p) (d3bj); ωB97X-D/6-31+(d,p); APFD/6-31+(d,p), implemented in the Gaussian16W package. In a vacuum, the CINRES was found at a copigmentation distance of 3.54 Å between cyanidin and resveratrol. In water, a binding free energy ∆G was calculated, rendering −3.31, −1.68, and −6.91 kcal/mol, at M06-2x/6-31g+(d,p) (d3bj), ωB97X-D/6-31+(d,p), and APFD/6-31+(d,p) levels of theory, respectively. A time-dependent density functional theory (TD-DFT) was used to calculate the UV spectra of the complexes and then compared to its parent molecules, resulting in a lower energy gap at forming complexes. Excited states’ properties were analyzed with the ωB97X-D functional. Finally, Shannon aromaticity indices were calculated and isosurfaces of non-covalent interactions were evaluated.
... Historically, blue food colorants were obtained from synthetic sources like indigo carmine, patent blue V, and brilliant blue (Buchweitz, 2016), raising consumer concerns about consuming blue-colored foods due to health issues associated with synthetic colorants. To address health concerns, the food industry is investing more in obtaining natural food colorants (Houghton et al., 2021). However, natural blue color sources are challenging to find in nature, posing the most significant challenge in providing a safe blue color for food. ...
... While the acylation patterns vary, mainly found as monoor di-acylated forms of p-hydroxybenzoic acid, caffeic acid and ferulic acid. Acylated anthocyanins were more stable than unacylated anthocyanins [11,12]. The very stable Figure 1. ...
... The discontinuous color and spectra for pH 7, 8, and 9 were caused by the buffer systems, with pH 1.0 to 7.0 as citrate-phosphate buffers, pH 8.0 as a phosphate buffer, and pH 9.0 to 13.0 as glycine-NaOH buffers. Increasing pH from 9 to 13 resulted in a color change from pink to blue/green and then yellow, indicating an alternation in anthocyanin structure from the red flavylium cation to the blue/green quinonoid base and then yellow chalcones as reported [11,13]. ...
... Many colorful foods are made by anthocyanin-rich fruits and vegetables, but it is difficult for them to maintain color consistency. Both internal and external factors would affect the color stability of anthocyanins [6,11,12,14]. ...
Purple sweetpotato anthocyanins (PSPA) exhibit significant potential as food colorants with associated health benefits. However, challenges related to browning and instability have hindered the application of PSPA. In this study, various pre-treatments and solvents for PSPA extraction were evaluated based on color, anthocyanin yields, antioxidant capabilities, and brown index. Browning markedly influenced the color and reduced the antioxidant capacity. Optimal results were obtained with the pre-treatment of “steaming of unpeeled whole sweetpotato” and the solvent “1% citric acid-ddH2O”. Furthermore, the color stability of purified PSPA solutions was evaluated under pH levels from 1 to 13 at 25 °C and 65 °C. The PSPA solutions showed a color spectrum from magenta, blue/green, and then to yellow across the pH range. The blue/green hues at pH 10–12 rapidly degraded, while the magenta hue at lower pH showed higher color stability. Elevated temperatures significantly accelerated the PSPA degradation. However, PSPA solutions at pH 1–2 exhibited remarkable color stability, with no spectral decay at either 65 °C for 12 h or 25 °C for 32 days. These results provide valid guidance for the extraction, preservation, and application of PSPA in the food industry.
... No malvidin, petunidin, or peonidin were detected in these tulips, which may be due to a lack of the methyltransferase required for their synthesis [30]. The absence of these enzymes could mean that the pathways for synthesizing malvidin, petunidin, and peonidin were incomplete or even absent. ...
Flower color is one of the most important ornamental traits of tulips (Tulipa gesneriana). Five typical tulip cultivars were selected to identify the flavonoid components and analyze their key gene expression in their tepals. Firstly, after preliminary determination of the pigment type, the flavonoids were identified by UPLC-Q-TOF-MS. A total of 17 anthoxanthins were detected in the five cultivars. The total anthoxanthin content in the white tulip and the red tulip showed a similar decreasing trend, while an increasing trend was observed in the black tulip. Similarly, a total of 13 anthocyanins were detected in five tulip cultivars. The black tulip contained the largest number of anthocyanins, mainly delphinidin derivatives (Dp) and cyanidin derivatives (Cy). The total anthocyanin content (TAC) in the orange, red, and black cultivars was higher than that in the white and yellow cultivars and presented an overall increase trend along with the flower development. TgCHS, TgFLS, TgF3H, TgF3′H, TgF3′5′H, and TgDFR, as key structural genes, were involved in the flavonoid synthesis pathway, and the expression patterns of these genes are basically consistent with the components and accumulation patterns of flavonoids mentioned above. Taken together, the flower color in tulips was closely related to the composition and content of anthocyanins and anthoxanthins, which were indeed regulated by certain key structural genes in the flavonoid pathway.
... It is important to highlight that anthocyanins' color is the result of the ratio of the three colored species in equilibrium at any specific pH. In this way, while at the lowest pH (<2), anthocyanins show red color; meanwhile, at a higher pH (6-7), they exhibited a purple color [101]. The extracts most often used for obtaining natural purple colorants are grape-skin extract (containing glucosides of peonidin, malvidin, delphinidin, and petunidin), and blackcurrant extract (containing cyanidin 3-rutinoside, delphinidin 3-rutinoside, cyanidin 3-glucoside, and delphinidin 3-glucoside). ...
... As has been previously explained, the color of the anthocyanins depends on the pH. In this sense, these compounds exhibit a blue color at pH 7-8 [101]. Butterfly pea (Clitoria ternatea L.) flowers are used for culinary purposes in the southern Asian region. ...
In recent years, the demand of healthier food products and products made with natural ingredients has increased overwhelmingly, led by the awareness of human beings of the influence of food on their health, as well as by the evidence of side effects generated by different ingredients such as some additives. This is the case for several artificial colorants, especially azo colorants, which have been related to the development of allergic reactions, attention deficit and hyperactivity disorder. All the above has focused the attention of researchers on obtaining colorants from natural sources that do not present a risk for consumption and, on the contrary, show biological activity. The most representative compounds that present colorant capacity found in nature are anthocyanins, anthraquinones, betalains, carotenoids and chlorophylls. Therefore, the present review summarizes research published in the last 15 years (2008–2023) in different databases (PubMed, Scopus, Web of Science and ScienceDirect) encompassing various natural sources of these colorant compounds, referring to their obtention, identification, some of the efforts made for improvements in their stability and their incorporation in different food matrices. In this way, this review evidences the promising path of development of natural colorants for the replacement of their artificial counterparts.
... Acidic solutions are stable, and alkaline and neutral solutions are unstable (Koh et al., 2020). As the pH in the environment continues to increase, the structure of anthocyanins continues to deprotonate, and its color changes from red to purple and then to blue-green (Houghton et al., 2021). The stability of anthocyanins is greatly influenced by environmental factors such as Ph value, temperature, light, and storage. ...
... Due to the acidic environment, anthocyanidins are present in the form of a positively charged flavylium cation. When the pH increases, the flavylium cation turns into a neutral (pH 6-7) and then an anionic (pH 7-8) quinonoid base structure [29]. Examining the pH dependence of anthocyanin binding, we found that the neutral and anionic structures present at nearly neutral pH bind better to gliadin in the case of the cyanidin glycoside-rich AC we examined. ...
Several types of gluten-related disorders are known, in which the common starting point is gluten-induced zonulin release. Zonulin results in varying degrees of increased permeability in certain gluten-related disorders but is largely responsible for the development of further pathogenic processes and symptoms. Therefore, it is important to know the barrier-modulating role of individual nutritional components and to what extent the antioxidant substance supports the protection of gliadin-induced membrane damage with its radical scavenging capacity. We investigated the pH dependence of the gliadin-anthocyanin interaction using UV photometry, during which a concentrationdependent interaction was observed at pH 6.8. The barrier modulatory effect of the anthocyanin-rich
sour cherry extract (AC) was analyzed on Caco-2 cell culture with pepsin-trypsin-resistant gliadin (PT-gliadin) exposure by TEER measurement, zonula occludens-1 (ZO-1), and Occludin immunohistochemistry. In addition to the TEER-reducing and TJ-rearranging effects of PT-gliadin, NF-κB
activation, an increase in cytokine (TNF-α, IFN-γ, and IL-8) release, and mitochondrial ROS levels
were observed. We confirmed the anti-inflammatory, stabilizing, and restoring roles of AC extract during gliadin treatment on the Caco-2 monolayer. The extract was able to significantly reduce cytokine and ROS levels despite the known interaction of the main components of the extract with PT-gliadin.
... Due to the acidic environment, anthocyanidins are present in the form of a positively charged flavylium cation. When the pH increases, the flavylium cation turns into a neutral (pH 6-7) and then an anionic (pH 7-8) quinonoid base structure [29]. Examining the pH dependence of anthocyanin binding, we found that the neutral and anionic structures present at nearly neutral pH bind better to gliadin in the case of the cyanidin glycoside-rich AC we examined. ...
Several types of gluten-related disorders are known, in which the common starting point is gluten-induced zonulin release. Zonulin results in varying degrees of increased permeability in certain gluten-related disorders but is largely responsible for the development of further pathogenic processes and symptoms. Therefore, it is important to know the barrier-modulating role of individual nutritional components and to what extent the antioxidant substance supports the protection of gliadin-induced membrane damage with its radical scavenging capacity. We investigated the pH dependence of the gliadin-anthocyanin interaction using UV photometry, during which a concentration-dependent interaction was observed at pH 6.8. The barrier modulatory effect of the anthocyanin-rich sour cherry extract (AC) was analyzed on Caco-2 cell culture with pepsin-trypsin-resistant gliadin (PT-gliadin) exposure by TEER measurement, zonula occludens-1 (ZO-1), and Occludin immunohistochemistry. In addition to the TEER-reducing and TJ-rearranging effects of PT-gliadin, NF-κB activation, an increase in cytokine (TNF-α, IFN-γ, and IL-8) release, and mitochondrial ROS levels were observed. We confirmed the anti-inflammatory, stabilizing, and restoring roles of AC extract during gliadin treatment on the Caco-2 monolayer. The extract was able to significantly reduce cytokine and ROS levels despite the known interaction of the main components of the extract with PT-gliadin.
