A three-shell supramolecular complex enables the symmetry-mismatched chemo- and regioselective bis-functionalization of C60

Molecular Russian dolls (matryoshkas) have proven useful for testing the limits of preparative supramolecular chemistry but applications of these architectures to problems in other fields are elusive. Here we report a three-shell, matryoshka-like complex—in which C60 sits inside a cycloparaphenylene nanohoop, which in turn is encapsulated inside a self-assembled nanocapsule—that can be used to address a long-standing challenge in fullerene chemistry, namely the selective formation of a particular fullerene bis-adduct. Spectroscopic evidence indicates that the ternary complex is sufficiently stable in solution for the two outer shells to affect the addition chemistry of the fullerene guest. When the complex is subjected to Bingel cyclopropanation conditions, the exclusive formation of a single trans-3 fullerene bis-adduct was observed in a reaction that typically yields more than a dozen products. The selectivity facilitated by this matryoshka-like approach appears to be a general phenomenon and could be useful for applications where regioisomerically pure C60 bis-adducts have been shown to have superior properties compared with isomer mixtures.This work was supported by grants from MINECO-Spain (CTQ2016-77989-P and PID2019-104498GB-I00 to X.R., RTI2018-095622-B-100 to D.M. and I.I., and EUR2019-103824 to F.G.), Generalitat de Catalunya (2017SGR264 and a PhD grant to C.F.-E.) and the Severo Ochoa Center of Excellence Program (Catalan Institute of Nanoscience and Nanotechnology, grant SEV-2017-0706). X.R. is also grateful for ICREA-Acadèmia awards. M.v.D. is grateful for financial support from the Deutsche Forschungsgemeinschaft (project number 182849149-SFB953 ‘Synthetic Carbon Allotropes’), the Fonds der Chemischen Industrie (FCI), the University of Ulm and the Deutscher Akademischer Austauschdienst (PhD fellowship to O.B.). E.U. thanks Universitat de Girona for a PhD grant and we thank Serveis Tècnics de Recerca, Universitat de Girona for technical support.Peer reviewe

Label-free quantitative ¹H NMR spectroscopy to study low-affinity ligand–protein interactions in solution: A contribution to the mechanism of polyphenol-mediated astringency - Fig 1

<p><b>Molecular structures of catechins and structurally related polyphenolic compounds.</b> (-)-epicatechin (a), (-)-epigallocatechin (b), (-)-epicatechin-3-gallate (c), (-)-epigallocatechin-3-gallate (d), methyl gallate (e), and quercetin-3-O-rutinoside (f).</p

Unbound (-)-epigallocatechin gallate (EGCG) in pure buffer (reference), 10% (v/v) human whole saliva, 1% (w/v) carboxymethyl cellulose (CMC), and a combination of both.

<p>Unbound EGCG was quantified by single qHNMR measurements and is expressed as mole fractions. Error bars denote the standard deviation as obtained from integrating all the distinct, non-overlaid proton signals of EGCG. Data points behind means are provided in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184487#pone.0184487.s003" target="_blank">S3 Table</a>.</p

Label-free quantitative ¹H NMR spectroscopy to study low-affinity ligand–protein interactions in solution: A contribution to the mechanism of polyphenol-mediated astringency

<div><p>Nuclear magnetic resonance (NMR) spectroscopy is well-established in assessing the binding affinity between low molecular weight ligands and proteins. However, conventional NMR-based binding assays are often limited to small proteins of high purity and may require elaborate isotopic labeling of one of the potential binding partners. As protein–polyphenol complexation is assumed to be a key event in polyphenol-mediated oral astringency, here we introduce a label-free, ligand-focused ¹H NMR titration assay to estimate binding affinities and characterize soluble complex formation between proteins and low molecular weight polyphenols. The method makes use of the effects of NMR line broadening due to protein–ligand interactions and quantitation of the non-bound ligand at varying protein concentrations by quantitative ¹H NMR spectroscopy (qHNMR) using electronic reference to access in vivo concentration (ERETIC 2). This technique is applied to assess the interaction kinetics of selected astringent tasting polyphenols and purified mucin, a major lubricating glycoprotein of human saliva, as well as human whole saliva. The protein affinity values (<i>BC</i>_<i>50</i>) obtained are subsequently correlated with the intrinsic mouth-puckering, astringent oral sensation imparted by these compounds. The quantitative NMR method is further exploited to study the effect of carboxymethyl cellulose, a candidate “anti-astringent” protein binding antagonist, on the polyphenol–protein interaction. Consequently, the NMR approach presented here proves to be a versatile tool to study the interactions between proteins and low-affinity ligands in solution and may find promising applications in the discovery of bioactives.</p></div

Pairwise comparison matrix to assess the relative astringency of polyphenols.

<p>Pairwise comparison matrix to assess the relative astringency of polyphenols.</p

Effect of Astringent Stimuli on Salivary Protein Interactions Elucidated by Complementary Proteomics Approaches

The interaction of astringent substances with salivary proteins, which results in protein precipitation, is considered a key event in the molecular mechanism underlying the oral sensation of puckering astringency. As the chemical nature of orally active astringents is diverse and the knowledge of their interactions with salivary proteins rather fragmentary, human whole saliva samples were incubated with suprathreshold and isointensity solutions of the astringent polyphenol (−)-epigallocatechin gallate, the multivalent metal salt iron­(III) sulfate, the amino-functionalized polysaccharide chitosan, and the basic protein lysozyme. After separation of the precipitated proteins, the proteins affected by the astringents were identified and relatively quantified for the first time by complementary bottom-up and top-down mass spectrometry-based proteomics approaches. Major salivary target proteins, which may be involved in astringency perception, are reported here for each astringent stimulus

Effect of Tannic Acid on the Protective Properties of the in situ Formed Pellicle

Objectives: In the present in situ/ex vivo study the impact of tannic acid on the erosion-protective properties of the enamel pellicle was tested. Additionally, the antiadherent and antibacterial effects of tannic acid were evaluated. Methods: The pellicle was formed in situ on bovine enamel samples fixed on individual splints worn by 6 subjects. Following 1 min of pellicle formation the volunteers rinsed for 10 min with tannic acid. After further oral exposure for 19 min, 109 min, and 8 h overnight, respectively, slabs were incubated in HCl ex vivo (pH 2.0, 2.3, 3.0) over 120 s. Subsequently, kinetics of calcium and phosphate release were measured photometrically. Samples after a 1-min fluoride mouth rinse as well as enamel samples with and without a 30-min in situ pellicle served as controls. Antiadherent effects were evaluated after a 1-min rinse with tannic acid and oral exposure of the slabs overnight. DAPI (4 ′ ,6-diamidino2-phenylindole) combined with concanavalin A staining and live/dead staining was used for fluorescence microscopic visualization and quantification of adherent bacteria and glucans. Modification of the pellicle’s ultrastructure by tannic acid was evaluated by transmission electron microscopy (TEM). Results: Tannic acid significantly improved the erosion-protective properties of the pellicle in a pH-dependent manner. Bacterial adherence and glucan formation on enamel were significantly reduced after rinses with tannic acid as investigated by fluorescence microscopy. TEM imaging indicated that rinsing with tannic acid yielded a sustainable modification of the pellicle; it was distinctly more electron dense. Conclusion: Tannic acid offers an effective and sustainable approach for the prevention of caries and erosion