SEM observation of hydrous superabsorbent polymer pretreated with room-temperature ionic liquids.

Room-temperature ionic liquid (RTIL), which is a liquid salt at or below room temperature, shows peculiar physicochemical properties such as negligible vapor pressure and relatively-high ionic conductivity. In this investigation, we used six types of RTILs as a liquid material in the pretreatment process for scanning electron microscope (SEM) observation of hydrous superabsorbent polymer (SAP) particles. Very clear SEM images of the hydrous SAP particles were obtained if the neat RTILs were used for the pretreatment process. Of them, tri-n-butylmethylphosphonium dimethylphosphate ([P(4, 4, 4, 1)][DMP]) provided the best result. On the other hand, the surface morphology of the hydrous SAP particles pretreated with 1-ethyl-3-methylimidazolium tetrafluoroborate ([C2mim][BF4]) and 1-butyl-3-methylimidazolium tetrafluoroborate ([C4mim][BF4]) was damaged. The results of SEM observation and thermogravimetry analysis of the hydrous SAP pretreated with the RTILs strongly suggested that most water in the SAP particles are replaced with RTIL during the pretreatment process.

Cadmium-free sugar-chain-immobilized fluorescent nanoparticles containing low-toxicity ZnS-AgInS2 cores for probing lectin and cells.

Sugar chains play a significant role in various biological processes through sugar chain-protein and sugar chain-sugar chain interactions. To date, various tools for analyzing sugar chains biofunctions have been developed. Fluorescent nanoparticles (FNPs) functionalized with carbohydrate, such as quantum dots (QDs), are an attractive imaging tool for analyzing carbohydrate biofunctions in vitro and in vivo. Most FNPs, however, consist of highly toxic elements such as cadmium, tellurium, selenium, and so on, causing problems in long-term bioimaging because of their cytotoxicity. In this study, we developed cadmium-free sugar-chain-immobilized fluorescent nanoparticles (SFNPs) using ZnS-AgInS2 (ZAIS) solid solution nanoparticles (NPs) of low or negligible toxicity as core components, and investigated their bioavailability and cytotoxicity. SFNPs were prepared by mixing our originally developed sugar-chain-ligand conjugates with ZAIS/ZnS core/shell NPs. In binding experiments with lectin, the obtained ZAIS/ZnS SFNPs interacted with an appropriate lectin to give specific aggregates, and their binding interaction was visually and/or spectroscopically detected. In addition, these SFNPs were successfully utilized for cytometry analysis and cellular imaging in which the cell was found to possess different sugar-binding properties. The results of the cytotoxicity assay indicated that SFNPs containing ZAIS/ZnS have much lower toxicity than those containing cadmium. These data strongly suggest that our designed SFNPs can be widely utilized in various biosensing applications involved in carbohydrates.

The effect of hydrophilic ionic liquids 1-ethyl-3-methylimidazolium lactate and choline lactate on lipid vesicle fusion.

Ionic liquids (ILs) are room-temperature molten salts that have applications in both physical sciences and more recently in the purification of proteins and lipids, gene transfection and sample preparation for electron microscopy (EM) studies. Transfection of genes into cells requires membrane fusion between the cell membrane and the transfection reagent, thus, ILs may be induce a membrane fusion event. To clarify the behavior of ILs with cell membranes the effect of ILs on model membranes, i.e., liposomes, were investigated. We used two standard ILs, 1-ethyl-3-methylimidazolium lactate ([EMI][Lac]) and choline lactate ([Ch][Lac]), and focused on whether these ILs can induce lipid vesicle fusion. Fluorescence resonance energy transfer and dynamic light scattering were employed to determine whether the ILs induced vesicle fusion. Vesicle solutions at low IL concentrations showed negligible fusion when compared with the controls in the absence of ILs. At concentrations of 30% (v/v), both types of ILs induced vesicle fusion up to 1.3 and 1.6 times the fluorescence intensity of the control in the presence of [Ch][Lac] and [EMI][Lac], respectively. This is the first demonstration that [EMI][Lac] and [Ch][Lac] induce vesicle fusion at high IL concentrations and this observation should have a significant influence on basic biophysical studies. Conversely, the ability to avoid vesicle fusion at low IL concentrations is clearly advantageous for EM studies of lipid samples and cells. This new information describing IL-lipid membrane interactions should impact EM observations examining cell morphology.

Stable sugar-chain-immobilized fluorescent nanoparticles for probing lectin and cells.

Sugar chains are important molecules in cellular recognition and signaling, and quantum dots (QDs) are a very powerful tool for in vitro and in vivo imaging. Herein, we report the preparation of stable sugar-chain-immobilized fluorescent nanoparticles (SFNPs) and their application to the analysis of sugar-chain-protein interactions and cellular imaging. SFNPs were easily prepared by mixing CdTe/CdS core/shell QDs with our previously developed sugar-chain-ligand conjugates. The obtained SFNPs were very stable and could be stored for several months. In the binding analysis, ß-galactose- and α-glucose-immobilized SFNPs were specifically interacted with Ricinus communis agglutinin I and concanavalin A, respectively, and made into aggregates. The binding interaction was detected visually, fluorescently, or both. In the experiment for cellular imaging, it was found that SFNPs were predominantly taken up by human hepatocyto carcinoma cells (HepG2), suggesting the possible usage of our designed SFNPs for various biochemical analyses of sugar chains.

Chromosome observation by scanning electron microscopy using ionic liquid.

Electron microscopy has been used to visualize chromosome since it has high resolution and magnification. However, biological samples need to be dehydrated and coated with metal or carbon before observation. Ionic liquid is a class of ionic solvent that possesses advantageous properties of current interest in a variety of interdisciplinary areas of science. By using ionic liquid, biological samples need not be dehydrated or metal-coated, because ionic liquid behaves as the electronically conducting material for electron microscopy. The authors have investigated chromosome using ionic liquid in conjunction with electron microscopy and evaluated the factors that affect chromosome visualization. Experimental conditions used in the previous studies were further optimized. As a result, prewarmed, well-mixed, and low concentration (0.5∼1.0%) ionic liquid provides well-contrasted images, especially when the more hydrophilic and the higher purity ionic liquid is used. Image contrast and resolution are enhanced by the combination of ionic liquid and platinum blue staining, the use of an indium tin oxide membrane, osmium tetroxide-coated coverslip, or aluminum foil as substrate, and the adjustment of electron acceleration voltage. The authors conclude that the ionic-liquid method is useful for the visualization of chromosome by scanning electron microscopy without dehydration or metal coating.

SEM observation of wet biological specimens pretreated with room-temperature ionic liquid.

A facile pretreatment process for SEM: The use of room temperature ionic liquids (RTILs) provides an interesting method for SEM of biological specimens. We used a novel and concise method of pretreatment, excluding fixation or Au sputtering steps. Fine and smooth-textured SEM images of a wide variety of biological specimens treated in this way were observed without artefacts.

Two conformers of 10,11-dihydro-5H-dibenzo[a,d]cycloheptene spiro-linked with homobenzoquinone epoxide.

The crystal structures of the two thermally equilibrated conformational isomers of the epoxide 1',5'-dimethylspiro[10,11-dihydro-5H-dibenzo[a,d]cycloheptene-5,8'-4'-oxatricyclo[5.1.0.0(3,5)]octane]-2',6'-dione, C23H20O3, have been determined by X-ray diffraction. In the tricyclic dione skeleton, the oxirane and cyclopropane rings adopt an anti structure with respect to the conjunct quinone frame. The spiro-linked 10,11-dihydro-5H-dibenzo[a,d]cycloheptene ring of the major isomer has a fairly twisted boat form, folding opposite to the adjoining cyclopropane methyl substituent, whereas the seven-membered ring of the minor isomer has an almost ideal twist-boat form, inversely folding to the side of the relevant methyl group. The conformational structures of these isomers have been compared with those of the corresponding isomers of the unepoxidized homobenzoquinone.

Acid-catalyzed transannular cyclization of 3aH-cyclopentene[8]annulene-1,4-(5H,9aH)-diones and some proposed mechanisms.

Bicyclic 3aH-cyclopentene[8]annulene-1,4-(5H,9aH)-diones underwent three types of acid-induced transannular reactions, Michael cyclization, [3 + 2] cycloaddition, and Friedel-Crafts ipso-alkylation, depending on the cyclopentenone ring substituent (Me or Ph) and the position of [8]annulenone substituent as well as the nature of acids (BF3, MeSO3H, CF3SO3H). The Me-substituent permitted the Michael reaction for all acids used to give tricyclic diones by the activation of cyclopentenone carbonyl group. However, the Ph-substituent inhibited the Michael reaction for BF3 and MeSO3H but allowed the [3 + 2] cycloaddition and Friedel-Crafts reaction for CF3SO3H depending on the position of annulenone substituent. These CF3SO3H reactions exhibited the following novel rearrangements, affording 2-naphthalenone and 7-acenaphthylene derivatives, respectively. The factors that control the reaction mode of these transannular cyclizations were discussed in view of the constraint twist-boat conformation of [8]annulenone ring as well as the ring substituent effects on the intramolecular cyclization. In addition, these [8]annulenone rings were found to easily undergo the intramolecular [2 + 2] photocyclization to provide the tetracyclic cage compounds which exhibited the facile cycloreversion under the influence of acid.

Site selectivity switch in Lewis acid catalysis. Mechanism and kinetic simulation of skeletal rearrangement of cyclobutene-fused homoquinones.

[reactions: see text] We investigated the site selectivity switch in BF3-catalyzed dual skeletal rearrangements of cyclobutene-fused diarylhomobenzoquinones by changing the stoichiometric amount of acid concentration. From the lower to the higher equivalency of BF3 x Et2O, the branching product ratios (path A/path B) obeyed nonlinear sigmoid curves against the equivalency of BF3 x Et2O. The observed selectivity profiles were simulated to elucidate factors that govern thermodynamic aspects (binding affinity K of each carbonyl function with acid) and kinetic aspects (rate constants k for the cyclobutene-ring cleavage).

Kinetic evidence for remote pi-aryl participation in the BF3-catalyzed rearrangement of [2 + 2] photocycloadducts of diarylhomobenzoquinones with diphenylacetylene.

[reaction: see text] The BF(3)-catalyzed rearrangement of cyclobutene-fused m- and p-substituted diarylhomobenzoquinones exclusively gave the keto-alcohols via a Wagner-Meerwein vinyl-anion migration followed by the annulation of a delta-located endo-aryl group. The Hammett treatments for the endo/exo substituent effects, as well as the kinetic solvent effects, indicated that this reaction proceeds through the concerted S(N)2-like mechanism involving a rate-determining endo-aryl-assisted transition state.

Mechanism of novel consecutive rearrangements of cyclobutene-fused diphenylhomobenzoquinones catalyzed by lewis acids.

Lewis acid catalyzed rearrangements of highly strained [2 + 2] photoadducts 1a-d of diphenylhomobenzoquinone with various acetylenes were investigated under the influence of AlCl(3), SnCl(4), BF(3), and TiCl(4). With the relief of steric strain, these tricyclo[5.2.0.0(3,5)]non-8-ene-2,6-diones underwent the three steps of consecutive skeletal transformations. The first step was the two-way cyclobutene ring-cleavage reaction with a Wagner-Meerwein vinyl migration to either Lewis acid activated carbonyl function. This process virtually occurred under the anchimeric assistance of the endo-phenyl ring to give, after proton transfer, the phenylene-bridged tetracyclic keto alcohols 2 and 3, respectively. The next step was the acid-induced cyclopropane ring cleavage of only 3 to lead to bicyclic diones 4 via a following stereoselective proton transfer. The last one involved a Michael-type intramolecular cyclization of 4 accompanied by a proton transfer to afford thermodynamically less stable tricyclic diones 5alpha which epimerized to 5beta only by TiCl(4). The factors that control the selectivity and the reactivity of these tandem reactions were addressed on the basis of the X-ray crystal analyses as well as the PM3 calculations. It was found the present Lewis acid-catalyzed rearrangements were very dependent on the substituents of 1a-d and the nature of the Lewis acids.