Study of the fluorescence and interaction between cyclodextrins and neochlorogenic acid, in comparison with chlorogenic acid

www.nature.com/scientificreports OPEN Study of the fluorescence and interaction between cyclodextrins and neochlorogenic acid, in comparison with chlorogenic acid Silvia Navarro‑Orcajada1, Adrián Matencio2, Cristina Vicente‑Herrero1, Francisco García‑Carmona1 & José Manuel López‑Nicolás1* Neochlorogenic acid, a less-studied isomer of chlorogenic acid, has been seen to posses antioxidant, antifungal, anti-inflammatory and anticarcinogenic effects, which makes it an interesting candidate for incorporation in functional foods. However, its poor solubility in water and susceptibility to oxidation make such a task difficult. To overcome that, its encapsulation in cyclodextrins (CDs) is proposed. The fluorescence of neochlorogenic acid in different pH conditions was analyzed, and caffeic acid was proved to be the fluorescent moiety in the molecule. An encapsulation model whereby the ligand poses two potential complexation sites (caffeic and D-(-)-quinic moieties), showed that α-CD and HP-β-CD formed the best inclusion complexes with neochlorogenic acid, followed by M-β-CD, β-CD and γ-CD. Molecular docking with the two best CDs gave better scores for α-CD, despite HP-β-CD providing stabilization through H-bonds. The encapsulation of chlorogenic acid led to a similar CD order and scores, although constants were higher for α-CD, β-CD and M-β-CD, lower for HP-β-CD, and negligible for γ-CD. The protonation state affected these results leading to a different order of CD preference. The solubility and the susceptibility to oxidation of neochlorogenic acid improved after complexation with α-CD and HP-β-CD, while the antioxidant activity of both isomers was maintained. Neochlorogenic acid (3-O-caffeoylquinic acid) is an isomer of chlorogenic acid (5-O-caffeoylquinic acid) formed by ester binding between caffeic acid and D-(-)-quinic acid (Fig. 1). The acid can be found in several foods, such as peaches, prunes, plums, coffee beans, apricots, rosemary leaves and cherries, and was proved to be accumulated throughout thirty days of postharvest drying at room t emperature1–3. Although chlorogenic acid is well characterised, its mechanism of action well studied4,5 and it is known for its biological properties as an antioxidant, anti-inflammatory, hepatoprotective, antimicrobial, cardioprotective, anticarcinogenic, neuroprotective, anti-obesity, anti-diabetes (among others) agent6,7, research about the physicochemical properties and biological effects of its isomer, neochlorogenic acid, is scarce. Despite this, some studies assert that neochlorogenic acid also has antioxidant, antifungal, anti-inflammatory and anticarcinogenic bioactivities8–13, and might therefore be an interesting candidate for incorporating in functional foods or nutraceuticals as a bioactive compound. However, in this respect, the molecule presents problems, including its low solubility in water and susceptibility to oxidation by the enzyme polyphenol oxidase due to its o-diphenol s tructure14,15, making it necessary to find new strategies. Several studies have demonstrated that the encapsulation of bioactive compounds in cyclodextrins (CDs) is a suitable way to overcome problems of this nature16,17. CDs are torus-shaped oligosaccharides made up of α-(1,4) linked glucose units. The most common are natural CDs, α, β and γ-CD, which contain six, seven and 1 Departamento de Bioquímica y Biología Molecular‑A, Facultad de Biología, Universidad de Murcia, Regional Campus of International Excellence “Campus Mare Nostrum”, 30100 Murcia, Spain. 2Dipartimento Di Chimica, Università di Torino, via P. Giuria 7, 10125 Turin, Italy. *email: Scientific Reports | (2021) 11:3275 | https://doi.org/10.1038/s41598-021-82915-9 1 Vol.:(0123456789) www.nature.com/scientificreports/ Figure 1.  Structure of (A) neochlorogenic acid, (B) chlorogenic acid and (C) caffeic acid; including pKa values determined for the carboxyl group and first phenolic g roup45. eight glucose units, r espectively18 and are included in the European list of additives approved for alimentary use with the corresponding E-numbers: E-457, E-459 and E-458, respectively. Apart from this, α-CD could be considered the most interesting CD for use in functional foods as it is on the register of EU health claims from the European Food and Safety Authority (EFSA). This claim asserts that the consumption of α-CD as a part of a starch-containing meal reduces the rise in blood glucose levels after that m eal19. This makes α-CD a suitable ingredient in foodstuffs intended for diabetics. CDs have a hydrophobic inner cavity due to the orientation of their hydrogen atoms, unlike their mainly hydrophilic outer surface in which the primary and secondary hydroxyl groups are exposed to the solvent, making the whole molecule fairly polar which enables its solubility in aqueous solutions. This fact means that poorly water-soluble compounds and hydrophobic moieties interact non-covalently with the CD inner cavity to form inclusion complexes, which can be more water-soluble than the free form depending on the type of CD used20. The use of these encapsulating agents in the food, pharmaceutical and cosmetic industries is rising rapidly due to their ability to increase the bioavailability of different compounds and to protect molecules against the action of external a gents21,22. In recent years, our research group has published several works concerning the ability of CDs to encapsulate different molecules of the stilbene family, such as resveratrol, oxyresveratrol, pterostilbene or piceatannol20,23–26, lipids27,28 and other bioactive c ompounds29. We observed that CDs were able to improve the solubility of phenolic compounds leading to an increase of its a ctivity24. Besides that, there are some studies available on the encapsulation of chlorogenic acid by these agents, mainly with β-CD30–33, and only a few of them evaluate the impact that this process could have on the activity of the molecule34,35. Alvarez-Parilla et al.30 proposed a competitive 1:1 model in which a CD molecule could capture chlorogenic acid either by the caffeic acid moiety or the D-()-quinic acid moiety, and when the complex is formed another CD cannot be incorporated into the system, excluding a 1:2 model with two molecules of CDs. Shao et al. (2014) and Zhao et al. (2010) demonstrated by H-NMR spectroscopy that cyclodextrins could encapsulate chlorogenic acid by these moieties. Since neochlorogenic acid has the same moieties as chlorogenic acid, this type of complexation could be considered to its isomer. Still, there is no research on the encapsulation of neochlorogenic acid or on its fluorescence properties, which extremely limits the knowledge of this molecule and, therefore, its potential incorporation as an ingredient in foods, cosmetics or drugs. Indeed, this is the first work to make an exhaustive study of the interaction between neochlorogenic acid and several natural and modified CDs, using fluorimetri (...truncated)