Phyllodulcin from the hexane fraction of Hydrangea macrophylla inhibits glucose-induced lipid accumulation and reactive oxygen species generation in Caenorhabditis elegans.

We confirmed that the hexane layer of Hydrangea macrophylla leaf extract (HLH) is rich in phyllodulcin (PD), an alternative sweetener, through high performance liquid chromatography (HPLC) analysis. To investigate in vivo activity of HLH and its PD, acute toxicity and growth rate of Caenorhabditis elegans were tested and there are no clinical abnormalities at 125-500 µg/mL of HLH. HLH decreased the total lipid and triglyceride contents dose-dependently in glucose-induced obese worms. Also, HLH increased survival rates under oxidative and thermal stress and decreased body reactive oxygen species (ROS) contents significantly. Such antioxidant properties of HLH were attributed to the enhanced activity of the antioxidant enzyme catalase. To determine whether the effect of HLH was due to PD, worms were treated with PD (concentration contained in HLH), and inhibitory effects on total lipids and ROS were observed. Our results suggest that HLH and its PD as a natural alternative sweetener can be used as materials to improve metabolic diseases.

Unlocking the Potential of ß-Fragmentation of Aminophosphoranyl Radicals for Sulfonyl Radical Reactions.

Exploiting ß-scission in aminophosphoranyl radicals for radical-mediated transformations has been a longstanding challenge. In this study, we investigated the untapped potential of ß-fragmentation in aminophosphoranyl radicals by leveraging the unique properties of the P-N bond and the substituents of P(III) reagents. Our approach carefully considers factors such as cone angle and electronic properties of phosphine and employs density functional theory (DFT) calculations to probe structural and molecular orbital influence. We successfully induced ß-fragmentation through N-S bond cleavage of aminophosphoranyl radicals under visible light and mild conditions, generating a range of sulfonyl radicals derived from pyridinium salts via the photochemical activity of electron donor-acceptor (EDA) complexes. This innovative synthetic strategy exhibits broad applicability, including late-stage functionalization, and paves the way for valuable sulfonyl radical-mediated reactions, such as alkene hydrosulfonylation, difunctionalization, and pyridylic C-H sulfonylation.

Visible-Light-Active Coumarin- and Quinolinone-Based Photocatalysts and Their Applications in Chemical Transformations.

Organic dyes have been actively studied as useful photocatalysts because they allow access to versatile structural flexibility and green synthetic applications. The identification of a new class of robust organic chromophores is, therefore, in high demand to increase structural diversity and variability. Although coumarins and quinolinones have long been acknowledged as organic chromophores, their ability to participate in photoinduced transformations is somewhat less familiar. Fascinated by their chromophoric features and adaptable platform, our group is interested in the identification of fluorescent bioactive molecules and in the development of new photoinduced synthetic methods using coumarins and quinolinones as photocatalysts. This account provides an overview of our recent progress in the discovery and application of light-absorbing coumarin and quinolinone derivatives in photochemistry and medicinal chemistry.

Cost-benefit analysis of the integrated pharmaceutical supply chain information service after the establishment of the Korean Pharmaceutical Information Service.

Background: The Korean Pharmaceutical Information Service (KPIS) was established in October 2007 to increase the transparency of the pharmaceutical supply chain by integrating relevant information. This study aimed to describe the KPIS program and perform a cost-benefit analysis of the KPIS. Methods: We conducted a cost-benefit analysis based on cost savings in terms of National Health Insurance (NHI). The outcome measures were the net financial benefit and benefit-cost ratio over the 12 years since the establishment of the KPIS. The cost estimate included the costs of labor and business operations, the development of an information entry system, and office maintenance. Financial benefits were defined as savings resulting from the implementation of the program based on KPIS data. Social benefits were defined as the prevention of recalled medicines from entering the supply chain and the decrease in inventory and disposal. Results: The KPIS clearly resulted in a net financial benefit, saving 37.2 million USD, which was 2.6 times higher than the cost of implementation. While the benefit-cost ratio was less than one during the first period, it exceeded 3.4 during the second period. After calculating and integrating social benefits, the net benefit increased to 571.6 million USD, and the benefit-cost ratio was 24.8. A sensitivity analysis of the annual benefit showed that the net benefit varied from a low of -1.5 million USD to a high of 24.7 million USD according to the program implementation year. Conclusion: The establishment of the KPIS and a system for collecting information on the pharmaceutical supply chain showed meaningful financial and social benefits when compared to the input cost. Since no other countries have an integrated pharmaceutical information system that incorporates all information from production to administration, the example of the KPIS can provide a precedent for other countries.

Site-Selective Pyridine C-H Alkylation with Alcohols and Thiols via Single-Electron Transfer of Frustrated Lewis Pairs.

A unified strategy for the deoxygenative or desulfurative pyridylation of various alcohols and thiols has been developed through a single-electron transfer (SET) process of frustrated Lewis pairs (FLPs) derived from pyridinium salts and PtBu3 . Mechanistic studies revealed that N-amidopyridinium salts serve as effective Lewis acids for the formation of FLPs with PtBu3 , and the generated phosphine radical cation ionically couples with the in situ generated xanthate, eventually affording the alkyl radical through facile ß-scission under photocatalyst-free conditions. The reaction efficiency was further accelerated by visible-light irradiation. This method is conceptually appealing by using encounter complexes in FLP chemistry to promote SET, which provides a previously unrecognized opportunity for the selective heteroarylation of a diverse range of alcohols and thiols with various functional groups, even in complex settings under mild reaction conditions.

N-Functionalized Pyridinium Salts: A New Chapter for Site-Selective Pyridine C-H Functionalization via Radical-Based Processes under Visible Light Irradiation.

The radical-mediated C-H functionalization of pyridines has attracted considerable attention as a powerful tool in synthetic chemistry for the direct functionalization of the C-H bonds of the pyridine scaffold. Classically, the synthetic methods for functionalized pyridines often involve radical-mediated Minisci-type reactions under strongly acidic conditions. However, the site-selective functionalization of pyridines in unbiased systems has been a long-standing challenge because the pyridine scaffold contains multiple competing reaction sites (C2 vs C4) to intercept free radicals. Therefore, prefunctionalization of the pyridine is required to avoid issues observed with the formation of a mixture of regioisomers and overalkylated side products.Recently, N-functionalized pyridinium salts have been attracting considerable attention in organic chemistry as promising radical precursors and pyridine surrogates. The notable advantage of N-functionalized pyridinium salts lies in their ability to enhance the reactivity and selectivity for synthetically useful reactions under acid-free conditions. This approach enables exquisite regiocontrol for nonclassical Minisci-type reactions at the C2 and C4 positions under mild reaction conditions, which are suitable for the late-stage functionalization of bioactive molecules with greater complexity and diversity. Over the past five years, a variety of fascinating synthetic applications have been developed using various types of pyridinium salts under visible light conditions. In addition, a new platform for alkene difunctionalization using appropriately designed N-substituted pyridinium salts as bifunctional reagents has been reported, offering an innovative assembly process for complex organic architectures. Intriguingly, strategies involving light-absorbing electron donor-acceptor (EDA) complexes between pyridinium salts and suitable electron-rich donors further open up new reactivity under photocatalyst-free conditions. Furthermore, we developed enantioselective reactions using pyridinium salts to afford enantioenriched molecules bearing pyridines through single-electron N-heterocyclic carbene (NHC) catalysis.Herein, we provide a broad overview of our recent contributions to the development of N-functionalized pyridinium salts and summarize the cornerstones of organic reactions that successfully employ these pyridinium salts under visible light conditions. The major advances in the field are systematically categorized on the basis of the pyridines' N-substituent, N-X (X = O, N, C, and SO2CF3), and its reactivity patterns. Furthermore, the identification of new activation modes and their mechanistic aspects are discussed by providing representative contributions to each paradigm. We hope that this Account will inspire broad interest in the continued innovation of N-functionalized pyridinium salts in the exploration of new transformations.

Site-Selective Pyridylic C-H Functionalization by Photocatalytic Radical Cascades.

An efficient pyridylic C(sp3 )-H functionalization has been developed through photocatalytic radical-mediated fluoroalkylation or cascade reactions. This method is enabled by the reversible formation of alkylidene dihydropyridine intermediates via the facile enolate formation of C4-alkyl N-amidopyridinium salts in the absence of an external base, thereby establishing the conditions necessary for subsequent intermolecular radical trapping. Rapid structural diversification of the pyridylic site can be achieved through photocatalytic multicomponent cascade reactions involving alkene trifluoromethylation, SO2 -reincorporation, and sulfonyl radical addition. This operationally simple method features a broad substrate scope and high chemoselectivity and offers a unique approach for the rational modification of the heterobenzylic C-H bonds of pyridines and quinolines with uniform site-selective control. Furthermore, experimental and theoretical studies were performed to elucidate the reaction mechanism.

Divergent reactivity of sulfinates with pyridinium salts based on one- versus two-electron pathways.

One of the main goals of modern synthesis is to develop distinct reaction pathways from identical starting materials for the efficient synthesis of diverse compounds. Herein, we disclose the unique divergent reactivity of the combination sets of pyridinium salts and sulfinates to achieve sulfonative pyridylation of alkenes and direct C4-sulfonylation of pyridines by controlling the one- versus two-electron reaction manifolds for the selective formation of each product. Base-catalyzed cross-coupling between sulfinates and N-amidopyridinium salts led to the direct introduction of a sulfonyl group into the C4 position of pyridines. Remarkably, the reactivity of this set of compounds is completely altered upon exposure to visible light: electron donor-acceptor complexes of N-amidopyridinium salts and sulfinates are formed to enable access to sulfonyl radicals. In this catalyst-free radical pathway, both sulfonyl and pyridyl groups could be incorporated into alkenes via a three-component reaction, which provides facile access to a variety of ß-pyridyl alkyl sulfones. These two reactions are orthogonal and complementary, achieving a broad substrate scope in a late-stage fashion under mild reaction conditions.

Rational Computational Design of Fourth-Generation EGFR Inhibitors to Combat Drug-Resistant Non-Small Cell Lung Cancer.

Although the inhibitors of singly mutated epidermal growth factor receptor (EGFR) kinase are effective for the treatment of non-small cell lung cancer (NSCLC), their clinical efficacy has been limited due to the emergence of various double and triple EGFR mutants with drug resistance. It has thus become urgent to identify potent and selective inhibitors of triple mutant EGFRs resistant to first-, second-, and third-generation EGFR inhibitors. Herein, we report the discovery of potent and highly selective inhibitors of EGFR exon 19 p.E746_A750del/EGFR exon 20 p.T790M/EGFR exon 20 p.C797S (d746-750/T790M/C797S) mutant, which were derived via two-track virtual screening and de novo design. This two-track approach was performed so as to maximize and minimize the inhibitory activity against the triple mutant and the wild type, respectively. Extensive chemical modifications of the initial hit compounds led to the identification of several low-nanomolar inhibitors of the d746-750/T790M/C797S mutant. Among them, two compounds exhibited more than 104-fold selectivity in the inhibition of EGFRd746-750/T790M/C797S over the wild type. The formations of a hydrogen bond with the mutated residue Ser797 and the van der Waals contact with the mutated residue Met790 were found to be a common feature in the interactions between EGFRd746-750/T790M/C797S and the fourth-generation inhibitors. Such an exceptionally high selectivity could also be attributed to the formation of the hydrophobic contact with a Gly loop residue or the hydrogen bond with Asp855 in the activation loop. The discovery of the potent and selective EGFRd746-750/T790M/C797S inhibitors were actually made possible by virtue of the modified protein-ligand binding free energy function involving a new hydration free energy term with enhanced accuracy. The fourth-generation EGFR inhibitors found in this work are anticipated to serve as a new starting point for the discovery of anti-NSCLC medicines to overcome the problematic drug resistance.

Visible-Light-Induced Cysteine-Specific Bioconjugation: Biocompatible Thiol-Ene Click Chemistry.

Bioconjugation methods using visible-light photocatalysis have emerged as powerful synthetic tools for the selective modification of biomolecules under mild reaction conditions. However, the number of photochemical transformations that allow successful protein bioconjugation is still limited because of the need for stringent reaction conditions. Herein, we report that a newly developed water-compatible fluorescent photosensitizer QPEG can be used for visible-light-induced cysteine-specific bioconjugation for the installation of QPEG by exploiting its intrinsic photosensitizing ability to activate the S-H bond of cysteine. The slightly modified QCAT enables the effective photocatalytic cysteine-specific conjugation of biologically relevant groups. The superior reactivity and cysteine selectivity of this methodology was further corroborated by traceless bioconjugation with a series of complex peptides and proteins under biocompatible conditions.

Borden Type I Sigmoid Sinus Dural Arteriovenous Fistula Presenting as Subarachnoid Hemorrhage from a Feeding Artery Aneurysm of the Anterior Inferior Cerebellar Artery: A Case Report.

Dural arteriovenous fistula is an acquired vascular anomaly that can cause various symptoms. Here, we report a rare case of Borden type I sigmoid sinus dural arteriovenous fistula presenting as subarachnoid hemorrhage. Bleeding occurred from a side-wall aneurysm in the lateral pontomedullary segment of the anterior inferior cerebellar artery, which was a minor pial feeder. Features on imaging modalities, including brain CT, CT angiography, MR imaging/angiography and digital subtraction angiography, are described with a literature review.

Hepatoprotective Effect of Fresh Grape Juice Prepared by a Low-Speed Masticating Juicer in db/db Mice.

This study investigated the hepatoprotective effect of fresh grape juice prepared using a low-speed masticating (LSM) juicer or a high-speed centrifugal (HSC) juicer in mice. Six-week-old db/db mice were fed on an AIN-93G diet or a diet containing 1% freeze-dried LSM or HSC grape juice for 7 weeks. Treatment with LSM grape juice significantly decreased hepatic triglycerides, serum aspartate transaminase activities, and homeostasis model assessment for insulin resistance values, whereas HSC juice did not significantly influence these parameters. The LSM grape juice showed higher antioxidant and anti-inflammatory activities than HSC juice. The benefits of LSM grape juice are probably due to a much higher proanthocyanidin content than that of HSC juice. These results suggest that LSM grape juice can exert hepatoprotective effects in db/db mice, partly through improving insulin resistance and promoting antioxidant and inflammatory activities.

Enzymatic browning reaction of apple juices prepared using a blender and a low-speed masticating household juicer.

The aim of this study was to investigate the effect of juicer type (blender or LSM household juicer) on the browning reaction of apple juice and evaluate the remaining antioxidant activity in the juice. The blender apple juice showed a darker brown color and 4.5 times higher PPO activity than LSM apple juice. This result suggested that the blender caused severer damage to plastids in cells leading to leakage of PPO into the juice than the LSM juicer. The total polyphenol and flavonoid content of LSM apple juice was approximately 2 times higher than that of blender apple juice because polyphenols and flavonoids can be used as substrates by PPO. The antioxidant activity of LSM juice was higher than that of blender juice. Together, these results suggested that the LSM juicer is superior to the blender for preparation of fresh apple juices due to the minimization of enzymatic oxidation reactions. Abbreviations: LSM: low-speed masticating; PPO: polyphenol oxidase; ABTS: 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid); DPPH: 2,2-diphenyl-1-picrylhydrazyl.

Antioxidant activities of fresh grape juices prepared using various household processing methods.

The purpose of this study was to evaluate and compare the antioxidant activities, and their contents, in grape juices prepared by various household juicers, and grape flesh (GF). The grape juices were prepared using a low-speed masticating (LSM) juicer, a high-speed centrifugal (HSC) juicer, and a blender (BLD). The total polyphenol, total flavonoid, total monomeric anthocyanin, and vitamin C contents were highest in the LSM grape juice, and decreased in the order: LSM > BLD > HSC > GF. The antioxidant activities such as DPPH radical scavenging activity, and SOD-like activity were significantly higher in the LSM juice than in other juices and grape flesh. The antioxidant activities and the quality of grape juices were significantly affected by the household juicing method used, and an LSM juicer is strongly recommended for making healthy grape juice, rich in antioxidants.

Biosynthesis of glucosyl glycerol, a compatible solute, using intermolecular transglycosylation activity of amylosucrase from Methylobacillus flagellatus KT.

A putative α-amylase gene (accession number, CP000284) of Methylobacillus flagellatus KT ATCC51484 was cloned in Escherichia coli, and its gene product was expressed and characterized. The purified recombinant enzyme (MFAS) displayed a typical amylosucrase (ASase) activity by the demonstration of multiple activities of hydrolysis, isomerization, and polymerization although it was designated as an α-amylase. The optimal reaction temperature and pH for the sucrose hydrolysis activity of MFAS were determined to be 45 °C and pH 8.5, respectively. MFAS has relatively high thermostable characteristics compared with other ASases, as demonstrated by a half-life of 19.3 min at 50 °C. MFAS also showed polymerization activity using sucrose as a sole substrate. Glycerol was transglycosylated by the intermolecular transglycosylation activity of MFAS. Two major products were observed by thin-layer chromatography and isolated by paper chromatography and recycling HPLC. Using (1)H and (13)C NMR, their chemical structures were determined to be (2S)-1-O-α-D-glucosyl-glycerol or (2R)-1-O-α-D-glucosyl-glycerol and 2-O-α-D-glucosyl-glycerol, in which a glucose molecule is linked to glycerol via an α-glycosidic linkage.

Biosynthesis of (+)-catechin glycosides using recombinant amylosucrase from Deinococcus geothermalis DSM 11300.

Amylosucrase (ASase, EC 2.4.1.4) is a glucosyltransferase that hydrolyzes sucrose into glucose and fructose and produces amylose-like glucan polymers from the released glucose. (+)-Catechin is a plant polyphenolic metabolite having skin-whitening and antioxidant activities. In this study, the ASase gene from Deinococcus geothermalis (dgas) was expressed in Escherichia coli, while the recombinant DGAS enzyme was purified using a glutathione S-transferase fusion system. The (+)-catechin glycoside derivatives were synthesized from (+)-catechin using DGAS transglycosylation activity. We confirmed the presence of two major transglycosylation products using TLC. The (+)-catechin transglycosylation products were isolated using silica gel open column chromatography and recycling-HPLC. Two (+)-catechin major transfer products were determined through (1)H and (13)C NMR to be (+)-catechin-3'-O-α-D-glucopyranoside with a glucose molecule linked to (+)-catechin and (+)-catechin-3'-O-α-D-maltoside with a maltose linked to (+)-catechin. The presence of (+)-catechin maltooligosaccharides in the DGAS reaction was also confirmed via recycling-HPLC and enzymatic analysis. The effects of various reaction conditions (temperature, enzyme concentration, and molar ratio of acceptor and donor) on the yield and type of (+)-catechin glycosides were investigated.

Structural features of the Nostoc punctiforme debranching enzyme reveal the basis of its mechanism and substrate specificity.

The debranching enzyme Nostoc punctiforme debranching enzyme (NPDE) from the cyanobacterium Nostoc punctiforme (PCC73102) hydrolyzes the alpha-1,6 glycosidic linkages of malto-oligosaccharides. Despite its high homology to cyclodextrin/pullulan (CD/PUL)-hydrolyzing enzymes from glycosyl hydrolase 13 family (GH-13), NPDE exhibits a unique catalytic preference for longer malto-oligosaccharides (>G8), performing hydrolysis without the transgylcosylation or CD-hydrolyzing activities of other GH-13 enzymes. To investigate the molecular basis for the property of NPDE, we determined the structure of NPDE at 2.37-A resolution. NPDE lacks the typical N-terminal domain of other CD/PUL-hydrolyzing enzymes and forms an elongated dimer in a head-to-head configuration. The unique orientation of residues 25-55 in NPDE yields an extended substrate binding groove from the catalytic center to the dimeric interface. The substrate binding groove with a lengthy cavity beyond the -1 subsite exhibits a suitable architecture for binding longer malto-oligosaccharides (>G8). These structural results may provide a molecular basis for the substrate specificity and catalytic function of this cyanobacterial enzyme, distinguishing it from the classical neopullulanases and CD/PUL-hydrolyzing enzymes.

Antioxidant effect of garlic and aged black garlic in animal model of type 2 diabetes mellitus.

Hyperglycemia in the diabetic state increases oxidative stress and antioxidant therapy can be strongly correlated with decreased risks for diabetic complications. The purpose of this study is to determine antioxidant effect of garlic and aged black garlic in animal model of type 2 diabetes. The antioxidant activity of garlic and aged black garlic was measured as the activity in scavenging free radicals by the trolox equivalent antioxidant capacity (TEAC) assay. Three week-old db/db mice were fed AIN-93G diet or diet containing 5% freeze-dried garlic or aged black garlic for 7 weeks after 1 week of adaptation. Hepatic levels of lipid peroxides and activities of antioxidant enzymes were measured. TEAC values of garlic and aged black garlic were 13.3 +/- 0.5 and 59.2 +/- 0.8 micromol/g wet weight, respectively. Consumption of aged black garlic significantly decreased hepatic thiobarbituric acid reactive substances (TBARS) level compared with the garlic group which showed lower TBARS level than control group (p<0.05). Activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) of garlic and aged black garlic group were significantly elevated compared to the control group. Catalase (CAT) activity of aged black garlic group was increased compared with the control group. These results show that aged black garlic exerts stronger antioxidant activity than garlic in vitro and in vivo, suggesting garlic and aged black garlic, to a greater extent, could be useful in preventing diabetic complications.

Role of maltogenic amylase and pullulanase in maltodextrin and glycogen metabolism of Bacillus subtilis 168.

The physiological functions of two amylolytic enzymes, a maltogenic amylase (MAase) encoded by yvdF and a debranching enzyme (pullulanase) encoded by amyX, in the carbohydrate metabolism of Bacillus subtilis 168 were investigated using yvdF, amyX, and yvdF amyX mutant strains. An immunolocalization study revealed that YvdF was distributed on both sides of the cytoplasmic membrane and in the periplasm during vegetative growth but in the cytoplasm of prespores. Small carbohydrates such as maltoheptaose and beta-cyclodextrin (beta-CD) taken up by wild-type B. subtilis cells via two distinct transporters, the Mdx and Cyc ABC transporters, respectively, were hydrolyzed immediately to form smaller or linear maltodextrins. On the other hand, the yvdF mutant exhibited limited degradation of the substrates, indicating that, in the wild type, maltodextrins and beta-CD were hydrolyzed by MAase while being taken up by the bacterium. With glycogen and branched beta-CDs as substrates, pullulanase showed high-level specificity for the hydrolysis of the outer side chains of glycogen with three to five glucosyl residues. To investigate the roles of MAase and pullulanase in glycogen utilization, the following glycogen-overproducing strains were constructed: a glg mutant with a wild-type background, yvdF glg and amyX glg mutants, and a glg mutant with a double mutant (DM) background. The amyX glg and glg DM strains accumulated significantly larger amounts of glycogen than the glg mutant, while the yvdF glg strain accumulated an intermediate amount. Glycogen samples from the amyX glg and glg DM strains exhibited average molecular masses two and three times larger, respectively, than that of glycogen from the glg mutant. The results suggested that glycogen breakdown may be a sequential process that involves pullulanase and MAase, whereby pullulanase hydrolyzes the alpha-1,6-glycosidic linkage at the branch point to release a linear maltooligosaccharide that is then hydrolyzed into maltose and maltotriose by MAase.

Characterization of a novel debranching enzyme from Nostoc punctiforme possessing a high specificity for long branched chains.

A novel debranching enzyme from Nostoc punctiforme PCC73102 (NPDE) exhibits hydrolysis activity toward both alpha-(1,6)- and alpha-(1,4)-glucosidic linkages. The action patterns of NPDE revealed that branched chains are released first, and the resulting maltooligosaccharides are then hydrolyzed. Analysis of the reaction with maltooligosaccharide substrates labeled with (14)C-glucose at the reducing end shows that NPDE specifically liberates glucose from the reducing end. Kinetic analyses showed that the hydrolytic activity of NPDE is greatly affected by the length of the substrate. The catalytic efficiency of NPDE increased considerably upon using substrates that can occupy at least eight glycone subsites such as maltononaose and maltooctaosyl-alpha-(1,6)-beta-cyclodextrin. These results imply that NPDE has a unique subsite structure consisting of -8 to +1 subsites. Given its unique subsite structure, side chains shorter than maltooctaose in amylopectin were resistant to hydrolysis by NPDE, and the population of longer side chains was reduced.