Origin of the Conformational Stability of Dirhodium(II) Tetrakis[N-phthaloyl-(S)-tert-leucinate] and Its Halogenated Derivatives.

In order to elucidate the origins of the stable structures of dirhodium(II) tetrakis[N-phthaloyl-(S)-tert-leucinate] and the four derivatives with halogenated aromatic rings, the conformational stability and intramolecular interactions were investigated by DFT calculations. In all of these complexes, the conformation in which all ligands face in the same direction is the most stable. When adjacent ligands are in the same orientation, destabilization due to exchange repulsion is larger than that when they are in opposite orientations. However, this destabilizing effect is reversed by the sum of the stabilizing effects of the electronic and charge transfer interactions. The imide carbonyl group plays an important role in these stabilizing interactions. The negatively charged site and bond orbitals in the imide carbonyl group interact with the positively charged sites and bond orbitals in the aromatic ring, the carboxylate group, and the α-position of the carboxylate group in the adjacent ligands. In addition, the lone-pair orbitals of the halogen atoms contribute to conformational stabilization by interacting with the vacant orbitals in the adjacent ligands. However, the combinations of these charged sites or bond orbitals, which effectively contribute to the stabilization, are different for each complex.

Standardization of strawberry sourness and sweetness intensities using a taste sensor system and an invariant taste standard solution.

Taste is an essential factor for evaluating the quality of agricultural products. However, it is usually difficult to compare data acquired at different times or by different people because there is no invariant reference and because the evaluation methods are largely subjective. Here, we addressed these problems by developing a method for standardizing strawberry sourness and sweetness intensities using a taste sensor approach with a taste standard solution composed of sour and sweet compounds. This standard solution allows highly efficient sensor measurements because it contains the standard compounds citric acid and sucrose. In addition, we found that polyphenol destabilized the sensor response for strawberry sweetness, and its removal from the sample by appropriate treatment with polyvinylpolypyrrolidone allowed stable evaluation of the sweetness intensity. The taste sensor data obtained using this method were in good agreement with the chemical analysis values related to human sensory evaluation.

Detection of catechins using a fluorescent molecule and its application toward the evaluation of astringent intensity.

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.

Standardization of tomato juice tastes using a taste sensor approach.

To enable the taste evaluation of many food samples at a time as well as the comparison of taste evaluation data acquired at different times, a standardization method for taste intensities was developed by a combination of a taste sensor system and a standard solution prepared with taste substances. In the case of tomato juices, citric acid, sucrose, and monosodium glutamate were used as standard taste substances for sourness, sweetness, and umami taste, respectively. Each standard point of the taste intensities was determined using only one standard solution including these standard substances. The taste intensity was described as a value on a scale based on discrimination thresholds of human gustation, where intensities of sourness, sweetness, and umami taste of the tomato juices were classified into multiple levels. Organoleptic evaluation supported these results. Validation for the present standardization method revealed that this approach has enough precision for practical tomato taste evaluation.

Mechanism of Copigmentation of Monoglucosylrutin with Caffeine.

4G-α-Glucopyranosylrutin (monoglucosylrutin, MGR) is a flavonol glycoside with quercetin as an aglycone, is pale yellow in color, and engages in both copigmentation and anticopigmentation. In this study, we elucidated the mechanism underlying the copigmentation of MGR upon complexation with caffeine. Three approaches were used: binding analyses based on changes in the absorbance spectrum, NOESY experiments, and DFT and TDDFT calculations using an explicit solvation model. Our findings show that copigmentation mainly results from a bathochromic shift in the absorbance spectrum and not a from hyperchromic effect. MGR and caffeine form a complex in both 1:1 and 1:2 stoichiometric ratios. The calculated optimized 1:1 and 1:2 complex structures were supported by the NOESY spectrum and form a cluster with 13 and 11 water molecules, respectively, through hydrogen bonds. Although HOMO and LUMO contribute most to the excitation of both the MGR monomer and the complexes, these frontier molecular orbitals in the complexes are distributed more widely than those in the MGR monomer. In particular, LUMO in the complexes spreads into the copigment caffeine and the solvent water molecules. This increase in electron delocalization reduces the energy gap between the frontier molecular orbitals, resulting in copigmentation with a bathochromic shift.

Complex Structures of Monoglucosylrutin with ent-Gallocatechin-3- O-gallate and Epigallocatechin-3- O-gallate in Aqueous Solutions and the Mechanism of Color Change Induced by Complexation.

The addition of ent-gallocatechin-3- O-gallate ( ent-GCg) or epigallocatechin-3- O-gallate (EGCg) to an aqueous solution of 4G-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.

Effect of Orbital Interactions between Vicinal Bonds and between Hydroxy Groups on the Conformational Stabilities of 1,2-Ethanediol and 2,3-Butanediols.

The geometries of the two hydroxy groups in 1,2-ethanediol or 2,3-butanediols are more stable in a gauche orientation than those in an anti orientation. This has been generally explained in terms of the gauche effect, which is stabilization due to antiperiplanar electron delocalization between an antibonding orbital of the C-O bond (σCO*) and a bonding orbital of the C-H or C-C bond (σCH or σCC). However, a C-C single bond rotation simultaneously determines the geometries of the six vicinal bonds. Therefore, it is important to understand the effects on conformational stability of other interactions of the bond orbitals adjacent to the rotating C1-C2 bond. Bond model analysis revealed that antiperiplanar bond orbital interactions as a whole contribute to the higher stabilities of hydroxy/hydroxy gauche conformers, where the C-O/C-H or C-O/C-C combination including the σCO*/σCH or σCO*/σCC delocalization is not the dominant interaction stabilizing hydroxy/hydroxy gauche conformers. Rather, our results show that a large destabilization due to the antiperiplanar C-O/C-O combination in hydroxy/hydroxy anti conformers relatively increases the stabilities of hydroxy/hydroxy gauche conformers. This destabilization results mainly from the repulsion between the antiperiplanar bonding orbitals (σCO/σCO), which have a larger overlap compared to the synclinal σCO/σCO combination. The sum of the interbond energies between the vicinal bond orbitals of these 1,2-alkanediols is more advantageous for stability in gauche conformers. In addition, interactions between the gauche-oriented hydroxy groups provide large stabilization energies and the corresponding interactions in anti conformers are negligible. The relative conformational stabilities of 1,2-ethanediol and erythro-2,3-butanediol can be explained by the interactions between the antiperiplanar bond orbitals, between the vicinal bond orbitals, or between the hydroxy groups in addition to the combination of interactions between the vicinal bond orbitals and between the hydroxy groups. In contrast, in threo-2,3-butanediol, differences in the relative stabilities of the three conformers can be understood by the combination of the interactions between the vicinal bond orbitals and between the hydroxy groups.

Conformations of Flavan-3-ols in Water: Analysis Using Density Functional Theory.

To elucidate the conformations and their relative stabilities of flavan-3-ols in water, the optimized structures and energies were calculated by density functional theory, in which M06-2X, ωB97X-D, B3LYP, and CAM-B3LYP were examined as exchange-correlation functionals. The results were evaluated by comparing calculated spin-spin coupling constants of vicinal protons with the corresponding values observed in experimental 1H NMR spectra in D2O. The M06-2X and ωB97X-D results showed good agreement with the experimental NMR data and revealed that the B-ring (pseudoequatorial)/3-O (pseudoaxial) conformers were more stable [5.4-8.7 kJ/mol (1.3-2.1 kcal/mol)] than the B-ring (pseudoaxial)/3-O (pseudoequatorial) conformers in the 2,3-cis-flavan-3-ols, while the B-ring (pseudoequatorial)/3-O (pseudoequatorial) conformers and the B-ring (pseudoaxial)/3-O (pseudoaxial) conformers had similar stabilities in the 2,3-trans-flavan-3-ols.

Association of Catechin Molecules in Water: Quantitative Binding Study and Complex Structure Analysis.

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.

Construction of a high-density reference linkage map of tea (Camellia sinensis).

A few linkage maps of tea have been constructed using pseudo-testcross theory based on dominant marker systems. However, dominant markers are not suitable as landmark markers across a wide range of materials. Therefore, we developed co-dominant SSR markers from genomic DNA and ESTs and constructed a reference map using these co-dominant markers as landmarks. A population of 54 F(1) clones derived from reciprocal crosses between 'Sayamakaori' and 'Kana-Ck17' was used for the linkage analysis. Maps of both parents were constructed from the F(1) population that was taken for BC(1) population. The order of most of the dominant markers in the parental maps was consistent. We constructed a core map by merging the linkage data for markers that detected polymorphisms in both parents. The core map contains 15 linkage groups, which corresponds to the basic chromosome number of tea. The total length of the core map is 1218 cM. Here, we present the reference map as a central core map sandwiched between the parental maps for each linkage group; the combined maps contain 441 SSRs, 7 CAPS, 2 STS and 674 RAPDs. This newly constructed linkage map can be used as a basic reference linkage map of tea.

Removal of acidic or basic α-amino acids in water by poorly water soluble scandium complexes.

To recognize α-amino acids with highly polar side chains in water, poorly water soluble scandium complexes with both Lewis acidic and basic portions were synthesized as artificial receptors. A suspension of some of these receptor molecules in an α-amino acid solution could remove acidic and basic α-amino acids from the solution. The compound most efficient at preferentially removing basic α-amino acids (arginine, histidine, and lysine) was the receptor with 7,7'-[1,3-phenylenebis(carbonylimino)]bis(2-naphthalenesulfonate) as the ligand. The neutral α-amino acids were barely removed by these receptors. Removal experiments using a mixed amino acid solution generally gave results similar to those obtained using solutions containing a single amino acid. The results demonstrated that the scandium complex receptors were useful for binding acidic and basic α-amino acids.

Annual variation of natural 15N abundance in tea leaves and its practicality as an organic tea indicator.

To obtain basic knowledge about the relationship between the application of organic fertilizers and the δ15N values of leaves of organically grown tea plants, annual variations in the δ15N values of the tea leaves were investigated. Although variations did not immediately arise after the application of organic fertilizers, differences in the δ15N values between organic and conventional cultivations appeared basically after three years from the beginning of the organic cultivation except when an organic fertilizer with a low δ15N value was applied, and the variation depended on the δ15N values of the fertilizers. In addition, the effectiveness of the δ15N values as a practical indicator of organic teas was examined. The tea leaves collected from organic farms did not always have higher δ15N values than the commercially available nonorganic teas. This result demonstrates that it is not easy to discriminate organic teas from nonorganic teas simply by their δ15N values.

Development of Expressed Sequence Tag (EST)-based Cleaved Amplified Polymorphic Sequence (CAPS) markers of tea plant and their application to cultivar identification.

To develop cleaved amplified polymorphic sequence (CAPS) markers for cultivar identification of the tea leaf, 5 primer pairs designed on the basis of genes that encode proteins related to nitrogen assimilation and 26 primer pairs based on expressed sequence tag (EST) sequences of the root of tea plant were screened. From combinations of primer pair and restriction enzyme that showed polymorphism among tea plants, 16 markers were selected and applied to DNA fingerprinting of Japanese tea cultivars. Sixty-three cultivars, except for a bud sport (Kiraka) and its original cultivar (Yabukita) and a pair that was the progeny of the same crossing parent (Harumoegi and Sakimidori), were distinguished from one another. By combining the 16 markers with previously developed CAPS markers and observing the physical appearance, 67 cultivars were distinguishable. The cultivars involve approximately 95% of total tea cultivating area in Japan; therefore, about 95% of tea leaves produced in Japan can be authenticated by labeling their cultivars.

Beta-cyclodextrin/surface plasmon resonance detection system for sensing bitter-astringent taste intensity of green tea catechins.

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.

Hypochromic effect of an aqueous monoglucosyl rutin solution caused by green tea catechins.

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.

Identification of Japanese and chinese green tea cultivars by using simple sequence repeat markers to encourage proper labeling.

To identify commercial Japanese monovarietal green tea and imported green tea samples, leading Japanese cultivars were fingerprinted by using six simple sequence repeat markers analyzed by a capillary sequencer. Two well-authenticated imported Chinese monovarietal green tea samples were also fingerprinted by the same markers, one of which, was Fuyun, was a clonally propagated cultivar, and the other, Jiukengzhong, was seed-propagated. At least three markers used in this study identified 16 leading Japanese cultivars and Fuyun. Although Jiukengzhong was a mixed population with diverse genotypes, some individuals had a unique allele in one simple sequence repeat marker that was not detected in the 16 leading Japanese cultivars, an additional 39 cultivars, and Fuyun. This allele was effective as a detection marker for Jiukengzhong. These results support the use of simple sequence repeat markers for the identification of Japanese monovarietal green tea and also of imported green tea made from foreign cultivars.

Evaluation of the umami taste intensity of green tea by a taste sensor.

A method for evaluating the umami taste intensity of green tea by a taste sensor system was established. Interference in the measurement from catechins was solved by removing the catechins from sample solutions with poly(vinylpolypyrrolidone). A 5.00 mM aqueous solution of glutamic acid monosodium salt was used as the standard solution. Sensor outputs were converted into EIT uma (estimated intensity of taste concerning umami) values. One unit on the EIT uma scale was defined as the amount of the sensor output corresponding to a difference in 1.2 times the concentration of the standard substance (glutamic acid monosodium salt). The umami taste intensity of green tea was classified into six grades on the EIT uma scale. Sensory tests proved that the EIT uma value had a high correlation to the human gustatory sense.

A water-soluble acyclic phane receptor recognizing 2,3-trans-gallate-type catechins.

To recognize gallate-type catechins in aqueous solution, two water-soluble acyclic phane receptors containing three aromatic rings were synthesized. The binding of these receptors to eight catechin analogues was investigated with (1)H NMR spectroscopy. The stoichiometric ratios of the complexes between the receptors and the catechins were 1:1 in every case. The binding abilities were estimated by (1)H NMR titration. The meta-substituted receptor showed excellent binding ability for the 2,3-trans-gallate-type catechins. This study revealed that a simple acyclic phane receptor can distinguish differences in the structures of the catechin analogues.

Analysis of the bacterial community found in clay wall material used in the construction of traditional Japanese buildings.

Clay wall (also called mud wall in English and tsuchikabe in Japanese) material is used in traditional Japanese buildings. Clay wall material is manufactured by fermenting a mixture of clay, sand, and rice straw. A culture-independent study based on 16s rRNA sequences revealed that Clostridiales of Firmicutes, alpha-, gamma- and delta-proteobacteria of Proteobacteria, and Acidobacterium of Acidobacteria exist in the clay wall material. Of these, we focused on Clostridium and Geobacter and their possible roles in the degradation of the cellulose component of the straw and the reduction of ferric iron in the clay during fermentation.

Evaluation of the astringency of black tea by a taste sensor system: scope and limitation.

Grading the astringency of black tea by a taste sensor system was studied. The black tea samples manufactured in India and Sri Lanka were classified into ten steps on the basis of two standard solutions (0.65 mM and 0.26 mM EGCg aqueous solutions). An organoleptic test demonstrated that the sensor output was correlative to the human gustatory sense.