Sheltering Mono-P-Ligated Metal Complexes in Porous Polystyrene Monolith: Effect of Aryl Pendant Stabilizers on Catalytic Durability.

Metal centers that can generate coordinatively unsaturated metals in accessible and stable states have been developed using synthetic polymers with sophisticated ligand and scaffold designs, which required synthetic efforts. Herein, we report a simple and direct strategy for producing polymer-supported phosphine-metal complexes, which stabilizes mono-P-ligated metals by modulating the electronic properties of the aryl pendant groups in the polymer platform. A three-fold vinylated PPh3 was copolymerized with a styrene derivative and a cross-linker to produce a porous polystyrene-phosphine hybrid monolith. Based on the Hammett substituent constants, the electronic properties of styrene derivatives were modulated and incorporated into the polystyrene backbone to stabilize the mono-P-ligated Pd complex via Pd-arene interactions. Through NMR, TEM, and comparative catalytic studies, the polystyrene-phosphine hybrid, which induces selective mono-P-ligation and moderate Pd-arene interactions, demonstrated high catalytic durability for the cross-coupling of chloroarenes under continuous-flow conditions.

Effects of three infusion fluids with different sodium chloride contents on steady-state serum concentrations of bromide in dogs.

Potassium bromide overdose (bromism) in the management of canine epilepsy has been known. However, a protocol to reduce bromide concentrations rapidly has not been previously established. The effects of three infusion fluids with different chloride contents on the steady-state serum concentrations of bromide in beagles were determined. After stabilization of the serum bromide concentrations, seven dogs were infused with saline (Na+ 154 mmol/L; Cl- 154 mmol/L), lactated Ringer's (Na+ 131 mmol/L; Cl- 110 mmol/L), or maintenance solutions (Na+ 35 mmol/L; Cl- 35 mmol/L) at a rate of 2 or 10 ml kg-1  hr-1 for 5 hr. Serum and urine were collected hourly, and the bromide concentrations were measured. When saline and lactated Ringer's solutions were infused at a rate of 10 ml kg-1  hr-1 for 5 hr, serum bromide concentrations were decreased by 14.24% and urine bromide concentrations by 17.63%, respectively. Of all compositions of infusion fluids, only sodium and chloride contents were associated with the decreased serum concentrations and the increased renal clearance of bromide. In summary, saline and lactated Ringer's solutions reduced serum bromide concentrations in a sodium chloride-dependent manner in dogs were found when infused at 10 ml kg-1  hr-1 for 5 hr.

Diversity in the carotenoid profiles and the expression of genes related to carotenoid accumulation among citrus genotypes.

Carotenoids are not only important to the plants themselves but also are beneficial to human health. Since citrus fruit is a good source of carotenoids for the human diet, it is important to study carotenoid profiles and the accumulation mechanism in citrus fruit. Thus, in the present paper, we describe the diversity in the carotenoid profiles of fruit among citrus genotypes. In regard to carotenoids, such as ß-cryptoxanthin, violaxanthin, lycopene, and ß-citraurin, the relationship between the carotenoid profile and the expression of carotenoid-biosynthetic genes is discussed. Finally, recent results of quantitative trait locus (QTL) analyses of carotenoid contents and expression levels of carotenoid-biosynthetic genes in citrus fruit are shown.

Continuous-Flow Catalysis Using Phosphine-Metal Complexes on Porous Polymers: Designing Ligands, Pores, and Reactors.

Continuous-flow syntheses using immobilized catalysts can offer efficient chemical processes with easy separation and purification. Porous polymers have gained significant interests for their applications to catalytic systems in the field of organic chemistry. The porous polymers are recognized for their large surface area, high chemical stability, facile modulation of surface chemistry, and cost-effectiveness. It is crucial to immobilize transition-metal catalysts due to their difficult separation and high toxicity. Supported phosphine ligands represent a noteworthy system for the effective immobilization of metal catalysts and modulation of catalytic properties. Researchers have been actively pursuing strategies involving phosphine-metal complexes supported on porous polymers, aiming for high activities, durabilities, selectivities, and applicability to continuous-flow systems. This review provides a concise overview of phosphine-metal complexes supported on porous polymers for continuous-flow catalytic reactions. Polymer catalysts are categorized based on pore sizes, including micro-, meso-, and macroporous polymers. The characteristics of these porous polymers are explored concerning their efficiency in immobilized catalysis and continuous-flow systems.

Effects of Cyclic Glycopolymers Molecular Mobility on their Interactions with Lectins.

Cyclic polymers, which are found in the field of biopolymers, exhibit unique physical properties such as suppressed molecular mobility. Considering thermodynamics, the suppressed molecular mobility of cyclic polymers is expected to prevent unfavorable entropy loss in molecular interactions. In this study, we synthesized cyclic glycopolymers carrying galactose units and investigated the effects of their molecular mobility on the interactions with a lectin (peanut agglutinin). The synthesized cyclic glycopolymers exhibited delayed elution time on size exclusion chromatography and a short spin-spin relaxation time, indicating typical characteristics of cyclic polymers, including smaller hydrodynamic size and suppressed molecular mobility. The hemagglutination inhibition assay revealed that the cyclic glycopolymers exhibited weakened interactions with PNA compared to the linear counterparts, attributable to the suppressed molecular mobility. Although the results are contrary to our expectations, the impact of polymer topology on molecular recognition remains intriguing, particularly in the context of protein repellent activity in the biomedical field.

Bayesian optimization of glycopolymer structures for the interaction with cholera toxin B subunit.

The optimal structure of synthetic glycopolymers for GM1 mimetics was determined through Bayesian optimization. The interactions of glycopolymers carrying galactose and neuraminic acid units in different compositions with cholera toxin B subunit (CTB) were assessed by an enzyme-linked immunosorbent assay (ELISA). Gaussian process regression, using the ELISA results, predicted the composition of glycopolymers that would exhibit stronger interactions with CTB. Following five cycles of optimization, the glycopolymers carrying 60 mol% galactose and 25 mol% neuraminic acid demonstrated an IC50 value of 75 µM for CTB, representing the lowest value among the synthesized glycopolymers.

Design of Glycopolymers for Controlling the Interactions with Lectins.

Carbohydrates are involved in life activities through the interactions with their corresponding proteins (lectins). Pathogen infection and the regulation of cell activity are controlled by the binding between lectins and glycoconjugates on cell surfaces. A deeper understanding of the interactions of glycoconjugates has led to the development of therapeutic and preventive methods for infectious diseases. Glycopolymer is one of the classes of the materials present multiple carbohydrates. The properties of glycopolymers can be tuned through the molecular design of the polymer structures. This review focuses on research over the past decade on the design of glycopolymers with the aim of developing inhibitors against pathogens and manipulator of cellular functions.

Optimized immobilization of single chain variable fragment antibody onto non-toxic fluorescent nanoparticles for efficient preparation of a bioprobe.

Single-chain variable fragment antibody (scFv) is a small molecular weight antibody that can be used for both therapeutic and diagnostic purposes. To visualize the interaction with the target biomolecules, scFv must be labeled with fluorescent molecules. In this study, to achieve the efficient labeling of scFv, we developed scFv-fluorescent nanoparticle conjugates to utilize scFv as bioprobes. As fluorescent carriers, cadmium-free ZnS-AgInS2/ZnS core/shell nanoparticles were used, and scFv was immobilized onto the nanoparticles via the interaction of nickel ions on nitrilotriacetic acid and hexahistidine (His-tag) fused with scFv. UV-Vis, fluorescence spectra, NMR, and dynamic laser scattering were used to characterize the scFv immobilized fluorescent nanoparticles (scFv-FNPs). The amounts of scFv on FNPs were controlled by the concentration of scFv. The scFv-FNPs that were prepared were non-toxic and selectively bound to cancer cells. The scFv-FNPs could be used as bioanalytical tools, and the immobilization method described here is a promising method for labeling biomolecules with the His-tag.

Expression and functional analysis of two lycopene ß-cyclases from citrus fruits.

In the present study, two LCYb genes (CitLCYb1 and CitLCYb2) were isolated from Satsuma mandarin (Citrus unshiu Marc.), Valencia orange (Citrus sinensis Osbeck) and Lisbon lemon (Citrus limon Burm.f.) and their functions were analyzed by the color complementation assay in lycopene-accumulating E. coli cells. The results showed that CitLCYb1 and CitLCYb2 shared high identity at the amino acid level among the three citrus varieties. The N-terminal region of the two proteins encoded by CitLCYb1 and CitLCYb2 was predicted to contain a 51-residue chloroplastic transit peptide, which shared low similarity. In Satsuma mandarin, the secondary structures of the CitLCYb1 and CitLCYb2 encoding proteins without the transit peptide were quite similar. Moreover, functional analysis showed that both enzymes of CitLCYb1 and CitLCYb2 participated in the formation of ß-carotene, and when they were co-expressed with CitLCYe, α-carotene could be produced from lycopene in E. coli cells. However, although CitLCYb2 could convert lycopene to α-carotene in E. coli cells, its extremely low level of expression indicated that CitLCYb2 did not participate in the formation of α-carotene during the green stage in the flavedo. In addition, the high expression levels of CitLCYb1 and CitLCYb2 during the orange stage played an important role in the accumulation of ß,ß-xanthophylls in citrus fruits. The results presented in this study might contribute to elucidate the mechanism of carotenoid accumulation in citrus fruits.

Regulation of carotenoid accumulation and the expression of carotenoid metabolic genes in citrus juice sacs in vitro.

In the present study, to investigate the mechanisms regulating carotenoid accumulation in citrus, a culture system was set up in vitro with juice sacs of three citrus varieties, Satsuma mandarin (Citrus unshiu Marc.), Valencia orange (Citrus sinensis Osbeck), and Lisbon lemon (Citrus limon Burm.f.). The juice sacs of all the three varieties enlarged gradually with carotenoid accumulation. The changing patterns of carotenoid content and the expression of carotenoid metabolic genes in juice sacs in vitro were similar to those ripening on trees in the three varieties. Using this system, the changes in the carotenoid content and the expression of carotenoid metabolic genes in response to environmental stimuli were investigated. The results showed that carotenoid accumulation was induced by blue light treatment, but was not affected by red light treatment in the three varieties. Different regulation of CitPSY expression, which was up-regulated by blue light while unaffected by red light, led to different changes in carotenoid content in response to these two treatments in Satsuma mandarin and Valencia orange. In all three varieties, increases in carotenoid content were observed with sucrose and mannitol treatments. However, the accumulation of carotenoid in the two treatments was regulated by distinct mechanisms at the transcriptional level. With abscisic acid (ABA) treatment, the expression of the genes investigated in this study was up-regulated in Satsuma mandarin and Lisbon lemon, indicating that ABA induced its own biosynthesis at the transcriptional level. This feedback regulation of ABA led to decreases in carotenoid content. With gibberellin (GA) treatment, carotenoid content was significantly decreased in the three varieties. Changes in the expression of genes related to carotenoid metabolism varied among the three varieties in response to GA treatment. These results provided insights into improving carotenoid content and composition in citrus during fruit maturation.

4-Amino-TEMPO-Immobilized Polymer Monolith: Preparations, and Recycling Performance of Catalyst for Alcohol Oxidation.

Continuous flow reactors with immobilized catalysts are in great demand in various industries, to achieve easy separation, regeneration, and recycling of catalysts from products. Oxidation of alcohols with 4-amino-TEMPO-immobilized monolith catalyst was investigated in batch and continuous flow systems. The polymer monoliths were prepared by polymerization-induced phase separation using styrene derivatives, and 4-amino-TEMPO was immobilized on the polymer monolith with a flow reaction. The prepared 4-amino-TEMPO-immobilized monoliths showed high permeability, due to their high porosity. In batch oxidation, the reaction rate of 4-amino-TEMPO-immobilized monolith varied with stirring. In flow oxidation, the eluent permeated without clogging, and efficient flow oxidation was possible with residence times of 2-8 min. In the recycling test of the flow oxidation reaction, the catalyst could be used at least six times without catalyst deactivation.

Crucial role of the small GTPase Rac1 in insulin-stimulated translocation of glucose transporter 4 to the mouse skeletal muscle sarcolemma.

The Rho family GTPase Rac1 has been implicated in the regulation of glucose uptake in myoblast cell lines. However, no evidence for the role of Rac1 has been provided by a mouse model. The purpose of this study is to test the involvement of Rac1 in insulin action in mouse skeletal muscle. Intravenous administration of insulin indeed elicited Rac1 activation in gastrocnemius muscle, suggesting the involvement of Rac1 in this signaling pathway. We then examined whether insulin-stimulated translocation of the facilitative glucose transporter GLUT4 from its storage sites to the skeletal muscle sarcolemma depends on Rac1. We show that ectopic expression of constitutively activated Rac1, as well as intravenous administration of insulin, caused translocation of GLUT4 to the gastrocnemius muscle sarcolemma, as revealed by immunofluorescent staining of a transiently expressed exofacial epitope-tagged GLUT4 reporter. Of particular note, insulin-dependent, but not constitutively activated Rac1-induced, GLUT4 translocation was markedly suppressed in skeletal muscle-specific rac1-knockout mice compared to control mice. Immunogold electron microscopic analysis of endogenous GLUT4 gave similar results. Collectively, we propose a critical role of Rac1 in insulin-dependent GLUT4 translocation to the skeletal muscle sarcolemma, which has heretofore been predicted solely by cell culture studies.

Exogenous Application of ABA and NAA Alleviates the Delayed Coloring Caused by Puffing Inhibitor in Citrus Fruit.

Combined spraying of gibberellin (GA) and prohydrojasmon (PDJ) was an effective method to reduce peel puffing in Satsuma mandarins. However, in the GA-and-PDJ combined treatment, fruit color development was delayed during the ripening process. In the present study, to improve the coloration of the GA and PDJ-treated fruit, the effects of exogenous application of 1-naphthaleneacetic acid (NAA) and abscisic acid (ABA) on chlorophyll and carotenoid accumulation were investigated. The results showed that both ABA and NAA treatments accelerated the color changes from green to orange in the GA and PDJ-treated fruit during the ripening process. With the NAA and ABA treatments, chlorophylls contents were decreased rapidly, and the contents of ß,ß-xanthophylls were significantly enhanced in the GA and PDJ-treated fruit. In addition, gene expression results showed that the changes of the chlorophyll and carotenoid metabolisms in the NAA and ABA treatments were highly regulated at the transcriptional level. The results presented in this study suggested that the application of NAA and ABA could potentially be used for improving the coloration of the GA and PDJ-treated fruit.

Synergistic interaction of cigarette smoking and alcohol drinking with serum carotenoid concentrations: findings from a middle-aged Japanese population.

Previous studies have indicated low serum carotenoid concentrations among cigarette smokers and/or alcohol drinkers, but little is known about the interaction of smoking and drinking with serum carotenoids. We tested the hypothesis that smoking and drinking reduce serum carotenoid concentrations synergistically. A total of 1073 subjects (357 male and 716 female) who had received health examinations in the town of Mikkabi, Shizuoka Prefecture, Japan, participated in the study. The subjects were divided into six groups according to alcohol intake (non-drinkers, < 1 g/d; light drinkers, > or = 1, < 25 g/d; moderate-to-heavy drinkers, > or = 25 g/d) and smoking status (non-smokers and current smokers). The dietary intakes and serum concentrations of six carotenoids (lycopene, alpha-carotene, beta-carotene, lutein, beta-cryptoxanthin and zeaxanthin) within each group were evaluated cross-sectionally. The dietary intakes of all carotenoids did not differ in the six groups after adjusting for age and sex. The multivariate-adjusted means of the serum carotenoid concentrations in non-drinkers did not differ between non-smokers and current smokers. In contrast, the adjusted means of serum alpha-carotene, beta-carotene and beta-cryptoxanthin were significantly lower than those with increased alcohol intake, and these lower serum carotenoids among alcohol drinkers were more evident in current smokers than in non-smokers. Serum lycopene of moderate-to-heavy drinkers was significantly lower than that of non-drinkers, but it was not influenced by smoking. Neither smoking nor drinking was associated with the serum concentrations of lutein and zeaxanthin. These results suggest that smoking and drinking may reduce the serum alpha-carotene, beta-carotene and beta-cryptoxanthin concentrations in a synergistic manner.

Novel biodegradation system for bisphenol A using laccase-immobilized hollow fiber membranes.

Radiation-induced graft polymerization was applied to prepare membranes for multilayer immobilization of laccase, which has biodegradation ability for bisphenol A (BPA). Glycidyl methacrylate (GMA) was grafted onto porous polyethylene membranes as the monomer of polymer brushes, and aminoethanol (AE) was introduced to the grafted GMA membrane, creating unfolded polymer brushes that serve as a good support for multilayer immobilization of laccase. The objectives of this study were as follows: adjustment of space velocity (SV) for optimum performance; enhancement of stability in organic media through moisture retention; biodegradation of BPA at continuous operation; and investigation of the effects of redox mediators. Laccase and membrane activities were increased at higher SVs as a result of stronger substrate transport. The 1.85% moisture retention as a result of high-density AE containing polymer brushes demonstrated the improved stability of immobilized laccase over free laccase in methanol-containing solutions. BPA was removed with an activity of 0.11 mol/h/kg-membrane. The effects of three major laccase mediators on BPA oxidation was studied, and only 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) was shown to increase the oxidation of BPA to 100% at low SVs. Improved stability of laccase and high removal rates in the continuous biodegradation of BPA were achieved by the presented method.

Associations of serum carotenoid concentrations with the metabolic syndrome: interaction with smoking.

Recent epidemiological studies show the associations of serum antioxidant status with the metabolic syndrome. Oxidative stress may play an important role in the pathogenesis of diabetes and CVD. Actually, smoking is a potent oxidative stressor in man, but little is known about the interaction of serum carotenoids and the metabolic syndrome with smoking status. In this study, the associations of the serum carotenoids with the metabolic syndrome stratified by smoking habit were evaluated cross-sectionally. A total of 1073 subjects (357 male and 716 female) who had received health examinations in the town of Mikkabi, Shizuoka Prefecture, Japan, participated in the study. Among total subjects, the OR for the metabolic syndrome in the highest tertile of serum beta-carotene was 0.41 (95 % CI 0.18, 0.92) after adjusting confounders. In current smokers, significantly lower OR were observed in the middle (OR 0.10; 95 % CI 0.01, 0.72) and highest (OR 0.06; 95 % CI 0.01, 0.73) tertiles of serum beta-carotene. Furthermore, lower OR were observed in accordance with tertiles of serum alpha-carotene and beta-cryptoxanthin in current smokers (P for trend 0.042 and 0.036, respectively). In contrast, in non-smokers, a significantly lower OR was observed in the highest tertile of serum beta-carotene (OR 0.30; 95 % CI 0.10, 0.89) after multiple adjustment. Inverse associations of serum carotenoids with the metabolic syndrome were more evident among current smokers than non-smokers. These results support that antioxidant carotenoids may have a protective effect against development of the metabolic syndrome, especially in current smokers who are exposed to a potent oxidative stress.

Abscisic acid affects expression of citrus FT homologs upon floral induction by low temperature in Satsuma mandarin (Citrus unshiu Marc.).

After a long juvenile period, citrus trees undergo seasonal flowering cycles. Under natural conditions, citrus flowering is regulated mainly by low ambient temperatures around 15-20 °C and water deficit stress. Recent studies have revealed that fluctuations in the expression of citrus homologs of FLOWERING LOCUS T (FT, encoding a flowering integrator) are correlated with their presumed role as flower-promoting signals. Previous ectopic expression analyses have demonstrated the flower-promoting function of citrus FT homologs. In this study, we examined whether abscisic acid (ABA) affects the expression of FT homologs and the flowering induced by low ambient temperatures. Application of exogenous ABA to potted Satsuma mandarin (Citrus unshiu Marc.) trees resulted in transient accumulation of citrus FT homolog transcripts. The promoter of one citrus FT homolog, CiFT3, was active in transgenic A. thaliana (Arabidopsis thaliana) and responded to exogenous and endogenous ABA. CiFT3 is preferentially expressed in shoots, and its expression was affected by flower-inductive treatments. Endogenous ABA accumulated in mandarin shoots during the floral induction period at 15 °C and under field conditions. The accumulation of ABA was correlated with the accumulation of FT homolog transcripts and flowering intensity. It was consistent with changes in the expression of genes related to ABA metabolism. The abundance of carotenoid precursors that serve as substrates for ABA biosynthesis decreased in leaves during the accumulation of ABA. Our data indicate that ABA and carotenoid precursors in leaves influence the flowering of mandarin trees induced by low temperature.

Quantification of carotenoids in citrus fruit by LC-MS and comparison of patterns of seasonal changes for carotenoids among citrus varieties.

To quantify the 18 carotenoids on the basic routes of the carotenoid biosynthesis in plants simultaneously, a method for liquid chromatography-mass spectrometry (LC-MS) using atmospheric pressure chemical ionization was developed. With this method, the seasonal changes of carotenoids in the flavedo and juice sacs of 39 citrus varieties were analyzed. On the basis of the patterns of seasonal changes of carotenoids in both flavedo and juice sacs, 39 citrus varieties were classified. In flavedo, 39 varieties were classified into 5 clusters, in which the carotenoid profiles were carotenoid-poor, phytoene-abundant, violaxanthin-abundant, violaxanthin- and beta-cryptoxanthin-abundant, and phytoene-, violaxanthin-, and beta-cryptoxanthin-abundant, respectively. In juice sacs, they were classified into 4 clusters, in which the carotenoid profiles were carotenoid-poor, violaxanthin-abundant, violaxanthin- and phytoene-abundant, and violaxanthin-, phytoene-, and beta-cryptoxanthin-abundant, respectively. In flavedo, many citrus varieties, except for the carotenoid-poor and phytoene-abundant varieties, massively accumulated beta,epsilon-carotenoids (e.g., lutein), beta,beta-carotenoids (e.g., beta-cryptoxanthin and violaxanthin), and phytoene, in that order. In juice sacs, the accumulation order among beta,beta-carotenoids was observed. Violaxanthin accumulation preceded beta-cryptoxanthin accumulation in violaxanthin-, phytoene-, and beta-cryptoxanthin-abundant varieties. In each variety, the carotenoid profiles of the flavedo and juice sacs on the basis of the concentration in violaxanthin and beta-cryptoxanthin were similar, with the exception of a few varieties.

Macroporous Gel with a Permeable Reaction Platform for Catalytic Flow Synthesis.

We mimic a living system wherein target molecules permeate through capillary and cells for chemical transformation. A monolithic porous gel (MPG) was easily prepared by copolymerization of gel matrix, tertiary amine, and cross-linking monomer in one-step synthesis. Interconnected capillaries existed in the MPG, enabling flow application with high permeability. Because the capillaries were constituted of polymer gel, Pd(0)-loaded MPG provided another permeable pathway to substrates in a gel network, contributing to its much high turnover number after 30 days of use, compared with that of Pd(0)-loaded inorganic supports. Interestingly, the gel network size of the MPG influenced the catalytic frequency. Diffusivities of the substrates and product in the gel networks increased with increasing network sizes in relation to catalytic activities. The MPG strategy provides a universal reactor design in conjunction with a practical process and precisely controlled reaction platform.

Glycopolymer monoliths for affinity bioseparation of proteins in a continuous-flow system: glycomonoliths.

In this study, macroporous materials, called glycomonoliths, were produced from saccharide-containing monomers, and used for affinity bioseparation of proteins in a continuous-flow system. The porous structure formation of the glycomonoliths involved polymerization-induced phase separation of the polyacrylamide unit. The pore size could be controlled between several hundred nanometers and several micrometers by changing the alcohol used as the porogenic solvent during the preparation of the monolith. The glycomonolith pores allowed for the permeation of solutions through the monoliths, which meant that they could be used in a continuous-flow system. The adsorption capacities of the glycomonoliths for the saccharide-binding protein (concanavalin A) were larger than that of a glycopolymer-grafted material because of the higher saccharide densities in the monoliths than the grafted material. When concanavalin A was eluted from the glycomonolith, the concentration of concanavalin A in the effluent was up to 11 times higher than that in the feed solution. The adsorption of concanavalin A to the glycomonolith was specific, even in the presence of other proteins.