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To evaluate sensory intensities of tastes, astringency, or pungency with ease and high reproducibility, the present study proposes a method using a fluorescent substance as a sensor. Unlike conventional taste sensors that each require a specialized system, this approach has the advantage in that it can be conducted using common analytical equipment such as a fluorescence spectrometer. In this report, a method to detect catechins and its application toward astringent evaluation is described. The binding of a sensor molecule with eight catechins revealed that the sensor molecule has higher affinity for catechins with stronger astringency. The complexation with the more astringent catechins showed larger fluorescence quenching. The change in the fluorescence intensity of a sample solution with the sensor molecule based on a reference solution was defined as the astringent intensity. Evaluation tests of green tea astringency demonstrated that the outputs of the sensor molecule correlate highly with the results by the human sense.

The addition of ent-gallocatechin-3- O-gallate ( ent-GCg) or epigallocatechin-3- O-gallate (EGCg) to an aqueous solution of 4 G -α-glucopyranosylrutin (monoglucosylrutin, MGR) causes the color of the solution to weaken due to complexation between MGR and these flavan-3-ols. Copigmentation is a well-known color change phenomenon resulting from the complexation of flavonoids that deepens and strengthens the color of the solution, whereas MGR/catechin complexation results in the opposite change in color (i.e., weakening). In order to gain insight into the mechanism underlying the rare changes in the color of solutions of complexes between flavonoids, the structures of the MGR monomer and the complexes in aqueous solutions and their photochemical properties were investigated by computational methods. Molecular dynamics simulations and subsequent density functional theory (DFT) calculations revealed that the complex structures are stabilized through aromatic/aromatic, CH/π, and OH/O interactions as direct intermolecular forces and that many solvent water networks would contribute to the complexations. Time-dependent DFT calculations showed that the change in the color of an MGR/ ent-GCg solution is due only to a decrease in absorbance, whereas that of an MGR/EGCg solution is due to both a decrease in absorbance and a hypsochromic shift.

Associations between catechin molecules were investigated by (1)H NMR titration experiments. Eight green tea catechins formed self-assembled dimers in water, and gallate-type catechins had a greater tendency to self-associate than non-gallate-type catechins. All eight catechins also associated as 1:1 heterodimer complexes. Investigation of complex formation of epigallocatechin-3-O-gallate (EGCg) and epigallocatechin (EGC) with the other catechins showed that the affinity between EGCg and 2,3-trans-gallate-type catechins was remarkably high, and the binding affinity of EGCg for ECg was also rather strong. In contrast, the non-gallate-type catechin EGC exhibited generally low binding affinity for other catechins. Structural analyses of the complexes by ROESY experiments and density functional theory calculations demonstrated that the higher binding abilities of gallate-type catechins are due to providing multiple intermolecular interactions that remain effective in an aqueous environment, such as aromatic/aromatic or CH/π interactions.

To develop a methodology for creating a sensor with a receptor for specific taste substances, we focused on constructing a sensing system for the bitter-astringent taste intensity of green tea catechins: (-)-epigallocatechin-3-O-gallate (EGCg), (-)-epicatechin-3-O-gallate (ECg), (-)-epigallocatechin (EGC), and (-)-epicatechin (EC). (1)H NMR titration experiments revealed that beta-cyclodextrin was an adequate receptor for sensing the bitter-astringent taste intensity of catechins. A surface plasmon resonance (SPR) system immobilized beta-cyclodextrin indicated larger responses for the gallate-type catechins in comparison to the non-gallate-type catechins. These responses corresponded to the tendency of the bitter-astringent taste intensity of the catechins felt by humans. Furthermore, the SPR system detected the larger stability of the complex between the gallate-type catechins and beta-cyclodextrin, which was interpreted as the aftertaste produced in humans by the gallate-type catechins. These results demonstrate that the beta-cyclodextrin/SPR system can sense the bitter-astringent taste intensity of the green tea catechins similar to human gustation. The methodology presented in this study can be used as a basic strategy for developing taste sensors with specific receptor functions.

Eight catechins reduced the color of a monoglucosyl rutin solution, the gallate-type catechins having a greater effect than the non-gallate-type catechins. Each catechin formed a 1:1 complex with monoglucosyl rutin. The binding constants of the gallate-type catechins were larger than those of the non-gallate type. These results indicate that the degree of color change in the monoglucosyl rutin solution depended on the ability of each catechin to form a complex with monoglucosyl rutin.

The reductive effect of pectin on tea catechin astringency was investigated by using a taste sensor system and (1)H-NMR spectroscopy. The sensor analysis revealed that the astringency of gallate-type catechins (EGCg and ECg) was reduced by the addition of pectin, whereas that of non-gallate-type catechins (EGC and EC) hardly changed. Changes in the (1)H-NMR chemical shifts of the catechins and pectin in mixed solutions showed that the gallate-type catechins formed complexes with pectin more closely than the non-gallate-type catechins. These results demonstrate that complexation between the gallate-type catechins and pectin is a factor for reducing catechin astringency.

Evaluating the binding energy of a catechin/caffeine complex in water is important in order to elucidate the ability for molecular recognition of tea catechins. The results of this study revealed that the stoichiometric ratio of the complexation between tea chatechins (EGCg, ECg, EGC, and EC) and caffeine was 1:1 at least up to a concentration of 5.0 mM. The free energy (-DeltaG) values for binding in water at 301 K were evaluated to be 2.7, 2.6, 2.2, and 2.0 kcal/mol for EGCg, ECg, EGC, and EC, respectively, by the titration method with (1)H-NMR. An investigation of the (1)H-NMR chemical shift change and NOESY spectra in the catechin/caffeine solutions showed the participation of the A-rings of the catechins in complexation, as well as that of the galloyl groups or B-rings.