On-Site Evaluation of Constituent Content and Functionality of Perilla frutescens var. crispa Using Fluorescence Spectra.

Perilla frutescens leaves are hypothesized to possess antioxidant and amyloid-ß (Aß) aggregation inhibitory properties primarily due to their polyphenol-type compounds. While these bioactivities fluctuate daily, the traditional methods for quantifying constituent contents and functional properties are both laborious and impractical for immediate field assessments. To address this limitation, the present study introduces an expedient approach for on-site analysis, employing fluorescence spectra obtained through excitation light irradiation of perilla leaves. Standard analytical techniques were employed to evaluate various constituent contents (chlorophyl (Chl), total polyphenol content (TPC), total flavonoid content (TFC), and rosmarinic acid (RA)) and functional attributes (DPPH radical scavenging activity, ferric reducing antioxidant power (FRAP), oxygen radical absorbance capacity (ORAC), and Aß aggregation inhibitory activity). Correlations between the fluorescence spectra and these parameters were examined using normalized difference spectral index (NDSI), ratio spectral index (RSI), and difference spectral index (DSI) analyses. The resulting predictive model exhibited a high coefficient of determination, with R2 values equal to or greater than 0.57 for constituent contents and 0.49 for functional properties. This approach facilitates the convenient, simultaneous, and nondestructive monitoring of both the chemical constituents and the functional capabilities of perilla leaves, thereby simplifying the determination of optimal harvest times. The model derived from this method holds promise for real-time assessments, indicating its potential for the simultaneous evaluation of both constituents and functionalities in perilla leaves.

Chemical analysis of amyloid ß aggregation inhibitors derived from Geranium thunbergii.

Amyloid ß (Aß) aggregates in the brains of patients with Alzheimer's disease (AD) and accumulates via oligomerization and subsequent fiber elongation processes. These toxicity-induced neuronal damage and shedding processes advance AD progression. Therefore, Aß aggregation-inhibiting substances may contribute to the prevention and treatment of AD. We screened for Aß42 aggregation inhibitory activity using various plant extracts and compounds, and found high activity for a Geranium thunbergii extract (EC50 = 18 µg/mL). Therefore, we screened for Aß42 aggregation inhibitors among components of a G. thunbergii extract and investigated their chemical properties in this study. An active substance was isolated from the ethanol extract of G. thunbergii based on the Aß42 aggregation inhibitory activity as an index, and the compound was identified as geraniin (1) based on spectral data. However, although geraniin showed in vitro aggregation-inhibition activity, no binding to Aß42 was observed via saturation transfer difference-nuclear magnetic resonance (STD-NMR). In contrast, the hydrolysates gallic acid (2) and corilagin (5) showed aggregation-inhibiting activity and binding was observed via STD-NMR. Therefore, the hydrolysates produced under the conditions of the activity test may contribute to the Aß42 aggregation-inhibition activity of G. thunbergii extracts. Geraniin derivatives may help prevent and treat AD.

Simple amino silyl ether organocatalyst for asymmetric hetero Diels-Alder reaction of isatins with enones.

New two catalysts component system comprising of a primary ß-amino silyl ethers as an organocatalyst and N-protected amino acids as a co-catalyst put together worked as an efficient organocatalyst system in the hetero Diels-Alder reaction of isatins with enones affording the chiral spirooxindole-tetrahydropyranones in good chemical yields and stereoselectivities (up to 94%, up to dr 78:22., up to 85% ee).

Daurichromenic Acid from the Chinese Traditional Medicinal Plant Rhododendron dauricum Inhibits Sphingomyelin Synthase and Aß Aggregation.

Species of the genus Rhododendron have been used in traditional Chinese medicine, with the medicinal herb "Manshanfong" used as an expectorant and for the treatment of acute bronchitis. Daurichromenic acid (DCA), a constituent of Rhododendron dauricum, is a meroterpenoid with antibacterial, anti-HIV, and anti-inflammatory activities. However, the mechanisms underlying these pharmacologic activities are poorly understood. To develop new drugs based on DCA, more information is required regarding its interactions with biomolecules. The present study showed that DCA inhibits the activity of the enzyme sphingomyelin synthase, with an IC50 of 4 µM. The structure-activity relationships between DCA and sphingomyelin synthase were evaluated using derivatives and cyclized hongoquercin A. In addition, DCA was found to inhibit amyloid ß aggregation. These results may help in the design of effective drugs based on DCA.

Enantioselective Diels-Alder reaction of 1,2-dihydropyridines with aldehydes using ß-amino alcohol organocatalyst.

The enantioselective Diels-Alder reaction of 1,2-dihydropyridines with aldehydes using an easily prepared optically active ß-amino alcohol catalyst was found to provide optically active isoquinuclidines, an efficient synthetic intermediate of pharmaceutically important compounds such as oseltamivir phosphate, with a satisfactory chemical yield and enantioselectivity (up to 96%, up to 98% ee). In addition, the obtained highly optically pure isoquinuclidine was easily converted to an optically active piperidine having four successive carbon centers.

New boro amino amide organocatalysts for asymmetric cross aldol reaction of ketones with carbonyl compounds.

Distinct types of new boron fused primary amino amide organocatalysts were designed and synthesized from commercially available amino acids. Their catalytic activities were investigated in asymmetric crossed aldol reaction of ketones with aromatic aldehydes to afford the corresponding chiral anti-aldol adducts with good chemical yields, moderate diastereoselectivity and good to excellent enantioselectivities (up to 94% yields, up to 90 : 10 dr, up to 94% ee).

Cultivation Factors That Affect Amyloid-ß Aggregation Inhibitory Activity in Perilla frutescens var. crispa.

Alzheimer's disease (AD) is thought to be caused by the deposition of amyloid-ß (Aß) in the brain. Aß begins to aggregate approximately 20 years before the expression of its symptoms. Previously, we developed a microliter-scale high-throughput screening (MSHTS) system for inhibitors against Aß aggregation using quantum dot nanoprobes. Using this system, we also found that plants in the Lamiaceae, particularly Perilla frutescens var. crispa, have high activity. The cultivation environment has the potential to enhance Aß aggregation inhibitory activity in plants by changing their metabolism. Here, we report on cultivation factors that affected the activity of P. frutescens var. crispa cultivated in three fields under different cultivation conditions. The results revealed that the activity of P. frutescens var. crispa harvested just before flowering was highest. Interestingly, the activity of wind-shielded plants that were cultivated to prevent exposure to wind, was reduced to 1/5th of plants just before flowering. Furthermore, activity just before flowering increased following appropriate nitrogen fertilization and at least one week of drying from the day before harvest. In addition, we confirmed that the P. frutescens var. crispa leaf extracts suppressed Aß-induced toxicity in nerve growth factor-differentiated PC12 cells. In this study, we demonstrated that flowering, wind, soil water content, and soil nitrogen content affected Aß aggregation inhibitory activity, necessary to suppress Aß neurotoxicity, in P. frutescens var. crispa extracts. This study provides practical cultivation methods for P. frutescens var. crispa with high Aß aggregation inhibitory activity for the prevention of AD.

Thiourea fused γ-amino alcohol organocatalysts for asymmetric Mannich reaction of ß-keto active methylene compounds with imines.

Catalytic functionality of new optically active thiourea fused γ-amino alcohols was examined in the asymmetric Mannich reaction of ß-keto active methylene compounds with imines to afford chiral Mannich products, ß-amino keto compounds, with continuous chiral centers, that are versatile synthetic intermediates for deriving various biologically active compounds. In particular, the thiourea fused γ-amino alcohols showed satisfactory catalytic activity in this reaction and afforded chiral Mannich products in excellent chemical yield (up to 88%) and stereoselectivities (up to syn : anti/93 : 7 dr, up to 99% ee).

Farm use of calcium hydroxide as an effective barrier against pathogens.

Livestock farming is affected by the occurrence of infectious diseases, but outbreaks can be prevented by proper sanitary control measures. Calcium hydroxide (Ca(OH)2), commonly called slaked lime, powder is traditionally used as a disinfectant to prevent infectious diseases in livestock. Since Ca(OH)2 can inactivate a wide variety of pathogens, has a small environmental impact, does not require a disinfection tank (i.e., can be spread directly on the ground) and is produced inexpensively worldwide, it is used for the prevention of epidemics on farms worldwide. Water is essential for the strong alkalinity that underlies its disinfecting effect, but it is unknown how much water is required under field conditions. In addition, Ca(OH)2 reacts with carbon dioxide in the environment, reducing its pH, but it is unclear how long its degradation takes under actual field use. Thus, we measured the water adsorption ability of Ca(OH)2-based disinfectants and its relation to disinfectant activity, as assessed by colony counts and live/dead staining and observation. We found that 15-20% (w/w) water in Ca(OH)2 was necessary for disinfection to occur in practice. Moreover, we found that the pH of Ca(OH)2 decreased within about two weeks to one month under actual use in practical conditions and lost its ability to disinfect. We further showed that granules prepared from Ca(OH)2 and zeolite maintained high alkalinity more than twice as long as calcium powder. These findings will help to establish a suitable method of applying Ca(OH)2 to protect farms from infectious diseases.

New small γ-turn type N-primary amino terminal tripeptide organocatalyst for solvent-free asymmetric aldol reaction of various ketones with aldehydes.

New small γ-turn type N-primary amino terminal tripeptides were synthesized and their functionality as an organocatalyst was examined in the asymmetric aldol reaction of various ketones with different aromatic aldehydes under solvent-free neat conditions to afford the desired chiral anti-aldol products in good to excellent chemical yields, diastereoselectivities and enantioselectivities (up to 99%, up to syn : anti/13 : 87 dr, up to 99% ee).

Evaluation of correlation between dissolution rates of loxoprofen tablets and their surface morphology observed by scanning electron microscope and atomic force microscope.

We observed the surface morphological structures of 60 mg tablets of Loxonin, Loxot, and Lobu using scanning electron microscope (SEM) and atomic force microscope (AFM) to evaluate the dissolution rates. We found a significant difference among the initial dissolution rates of the three kinds of loxoprofen sodium tablets. Petal forms of different sizes were commonly observed on the surface of the Loxonin and Loxot tablets in which loxoprofen sodium was confirmed by measuring the energy-dispersible X-ray (EDX) spectrum of NaKalpha using SEM. However, a petal form was not observed on the surface of the Lobu tablet, indicating differences among the drug production processes. Surface area and particle size of the principal ingredient in tablets are important factors for dissolution rate. The mean size of the smallest fine particles constituting each tablet was also determined with AFM. There was a correlation between the initial dissolution rate and the mean size of the smallest particles in each tablet. Visualizing tablet surface morphology using SEM and AFM provides information on the drug production processes and initial dissolution rate, and is associated with the time course of pharmacological activities after tablet administration.

Evaluation of Amyloid ß42 Aggregation Inhibitory Activity of Commercial Dressings by A Microliter-Scale High-Throughput Screening System Using Quantum-Dot Nanoprobes.

The aggregation and accumulation of amyloid ß (Aß) in the brain is a trigger of pathogenesis for Alzheimer's disease. Previously, we developed a microliter-scale high-throughput screening (MSHTS) system for Aß42 aggregation inhibitors using quantum-dot nanoprobes. The MSHTS system is seldom influenced by contaminants in samples and is able to directly evaluate Aß42 aggregation inhibitory activity of samples containing various compounds. In this study, to elucidate whether the MSHTS system could be applied to the evaluation of processed foods, we examined Aß42 aggregation inhibitory activity of salad dressings, including soy sauces. We estimated the 50% effective concentration (EC50) from serial diluted dressings. Interestingly, all 19 commercial dressings tested showed Aß42 aggregation inhibitory activity. It was suggested that EC50 differed by as much as 100 times between the dressings with the most (0.065 ± 0.020 v/v%) and least (6.737 ± 5.054 v/v%) inhibitory activity. The highest activity sample is traditional Japanese dressing, soy sauce. It is known that soy sauce is roughly classified into a heat-treated variety and a non-heat-treated variety. We demonstrated that non-heat-treated raw soy sauce exhibited higher Aß42 aggregation inhibitory activity than heat-treated soy sauce. Herein, we propose that MSHTS system can be applied to processed foods.

Simple primary ß-amino alcohols as organocatalysts for the asymmetric Michael addition of ß-keto esters to nitroalkenes.

Simple primary ß-amino alcohols act as an efficient organocatalysts in the asymmetric Michael addition of ß-keto esters with nitroalkenes affording highly pure chiral Michael adducts. Also, both enantiomers of the adducts were obtained, depending on the specific catalyst used and reaction temperature.

Simple organocatalyst component system for asymmetric hetero Diels-Alder reaction of isatins with enones.

A simple two catalyst component system consisting of primary ß-amino alcohols as a catalyst and amino acids as a co-catalyst put together works as an efficient organocatalyst system in the hetero Diels-Alder reaction of isatins with enones to afford the chiral spirooxindole-tetrahydropyranones in good chemical yields and stereoselectivities (up to 86%, up to 85 : 15 dr., up to 95% ee).

Development of clinical application for a nutritional prescription support system for total parenteral/enteral nutrition.

One of the important roles of pharmacists as members of a nutrition support team is nutritional prescription support. We developed a nutritional prescription support system (NPSS) that facilitates prescription support and analysis and evaluated its usefulness in nutritional therapy. An NPSS for prescription support and the management of patient information was created. With this NPSS, the nutritional status was assessed, and, on the basis of the results, such variables as the total energy expenditure were calculated. This system allows prescription support for parenteral nutrition (PN) therapy, enteral nutrition (EN) therapy, and the transition period between them. This system was used for 2 representative patients and evaluated. In a malnourished patient receiving oral warfarin, EN solutions were compared by means of the NPSS, and an appropriate EN solution was selected. In addition, the prothrombin time-international normalized ratio was monitored, and favorable results were obtained regarding the adjustment of the warfarin dose and nutritional management. In a patient with aspiration pneumonia, continuous nutritional management to EN from PN therapy was straightforwardly performed with the NPSS. This NPSS allows rapid, comprehensive nutritional management during the transition period to EN from PN therapy, despite these therapies being considered separately in conventional nutritional management. The NPSS is useful for simplifying prescription support and facilitating information sharing among members of a nutrition support team.

An automated microliter-scale high-throughput screening system (MSHTS) for real-time monitoring of protein aggregation using quantum-dot nanoprobes.

Protein aggregation is the principal component of numerous protein misfolding pathologies termed proteinopathies, such as Alzheimer's disease, Parkinson's disease, prion disease, and AA amyloidosis with unmet treatment needs. Protein aggregation inhibitors have great potential for the prevention and treatment of proteinopathies. Here we report the development of an automated real-time microliter-scale high throughput screening (MSHTS) system for amyloid aggregation inhibitors using quantum-dot nanoprobes. Screening 504 crude extracts and 134 low molecular weight aromatic compounds revealed the relationship of amyloid-ß (Aß) aggregation inhibitory activities of plant extracts using a plant-based classification. Within the eudicots, rosids, Geraniales and Myrtales showed higher activity. Screening low molecular weight aromatic compounds demonstrated that the structure of tropolone endows it with potential Aß aggregation inhibitory activity. The activity of the most active tropolone derivative was higher than that of rosmarinic acid. MSHTS also identified three chaperone molecules as tau aggregation inhibitors. These results demonstrate that our automated MSHTS system is a novel and robust tool that can be adapted to a wide range of compounds and aggregation-prone polypeptides.

Electronic effects of para-substitution on acetophenones in the reaction of rat liver 3alpha-hydroxysteroid dehydrogenase.

Stereoselective reductive metabolism of various p-substituted acetophenone derivatives was studied using isolated rat liver 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD). Kinetic experiments were performed and analyzed by measuring the products by HPLC using a chiral column. The results demonstrated that the presence of an electron-withdrawing substituent on the benzene ring plays an important role in determining the reduction rate in the syntheses of various (S)-alcohols from their corresponding carbonyl compounds. A plot of log {(V(max)/K(m))X/(V(max)/K(m))H} versus the substituent parameter (pi, sigma(para), Es) shows an increasing rate mainly for electron-withdrawing substituents, with a correlation coefficient (r(2)) of 0.97 which was obtained for triplicate data that were significant at the p<0.0001 level. With this in mind, new drugs can be designed that exploit this reduction pathway by introducing an electron-withdrawing group adjacent to the reduction site when a reduction reaction is desired, or by adding an electron-donating group when minimization of the reduction pathway is desired.

Inhibitory effect of the alkyl side chain of caffeic acid analogues on lipopolysaccharide-induced nitric oxide production in RAW264.7 macrophages.

Caffeic acid esters, one of the components of propolis, are known to show a variety of biological effects such as anti-tumor, anti-oxidant, and anti-inflammatory activities. Although, the anti-inflammatory activities of caffeic acid esters have been studied by analyzing their structure, the detailed mechanisms of their activities remain unclear. Thus, in this study, we examined the function of the ester functional group and the alkyl side chain (alcoholic part) and transformed caffeic acid to several derivatives. The inhibitory effect of these derivatives on NO production in murine macrophage RAW264.7 cells was dependent on the length and size of the alkyl moiety, and undecyl caffeate was the most potent inhibitor of NO production. In addition, individual experiments using undecanol, caffeic acid, undecanol plus caffeic acid, and undecyl caffeate showed that the connection between caffeic acid and the alkyl chain is critical for activity. Amide and ketone derivatives showed that not only the ester functional group but also the amide and ketone functional groups exhibit an inhibitory effect on NO production.

New Hybrid-type Squaramide-Fused Amino Alcohol Organocatalyst for Enantioselective Domino Michael Addition/Cyclization Reaction of Oxoindolines with Cyclic 1,3-Diketones.

The new hybrid-type squaramide-fused amino alcohol containing both a Brønsted basic site and hydrogen-bonding sites in the molecule showed a high catalytic activity as an organocatalyst in the enantioselective domino Michael addition/cyclization reaction of oxoindolines with cyclic 1,3-diketones to afford the chiral spiro-conjugated oxindoles featuring 2-aminopyrans fusing with carbo-heterocyclic ring systems with excellent chemical yields (up to 98%) and enantioselectivities (up to 95% ee). The obtained chiral spiro-conjugated 2-aminopyrans bearing quaternary stereogenic carbon center could be used as synthetic precursors for several natural products that have a broad spectrum of fascinating biological activities.

Rosmarinic acid is a novel inhibitor for Hepatitis B virus replication targeting viral epsilon RNA-polymerase interaction.

Current therapeutics for hepatitis B virus (HBV) patients such as nucleoside analogs (NAs) are effective; however, new antiviral drugs against HBV are still desired. Since the interaction between the epsilon (ε) sequence of HBV pregenomic RNA and viral polymerase (Pol) is a key step in the HBV replication cycle, we aimed to identify small compounds for its inhibition, and established a pull-down assay system for the detection of ε-RNA-binding-Pol. Screening showed that 5 out of 3,965 compounds inhibited ε-Pol binding, and we identified rosmarinic acid, which exhibited specificity, as a potential antiviral agent. In order to examine the anti-HBV effects of rosmarinic acid, HBV-infected primary human hepatocytes from a humanized mouse liver were treated with rosmarinic acid. The rosmarinic acid treatment decreased HBV components including the amounts of extracellular HBV DNA with negligible cytotoxicity. We also investigated the combined effects of rosmarinic acid and the NA, lamivudine. rosmarinic acid slightly enhanced the anti-HBV activity of lamivudine, suggesting that the HBV replication step targeted by rosmarinic acid is distinct from that of NA. We analyzed an additional 25 rosmarinic acid derivatives, and found that 5 also inhibited ε-Pol. Structural comparisons between these derivatives implied that the "two phenolic hydroxyl groups at both ends" and the "caffeic acid-like structure" of rosmarinic acid are critical for the inhibition of ε-Pol binding. Collectively, our results demonstrate that rosmarinic acid inhibits HBV replication in HBV-infected cells by specifically targeting ε-Pol binding.