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To demonstrate the accurate analysis of catecholamines and amino acid using derivatization reagents, we investigated the reaction conditions for 2,4,6-triethyl-3,5-dimethyl pyrylium trifluoromethanesulfonate (Py-Tag), derivatization of the targets dopamine (DA) and γ-aminobutyric acid (GABA) on tissue sections, and constructed an optimized reaction compartment. Ten different Py-Tag reaction conditions with the targets were considered. The optimal condition for the Py-Tag reaction with the targets was identified as a 70% methanol with 5% trimethylamine (v/v) solution at 60 °C under homogenous conditions. To reproduce this reaction on tissue sections, we constructed a reaction compartment to maintain humidity levels and facilitate the derivatization reaction. Moreover, visualization of DA and GABA was archived by derivatized-imaging mass spectrometry. Brain sections of unilateral 6-OHDA lesioned Parkinson's disease model rats showed Py-Tag DA (m/z 328.3) in the unilateral striatum and Py-Tag GABA (m/z 278.3) in the cerebral cortex, striatum, hippocampus and hypothalamus. Using the Parkinson's disease model rat brain, images with left-right differences were obtained for the localization of DA and GABA. These findings indicate that it is important to consider the reaction conditions that allow high reaction efficiency between DA or GABA and Py-Tag as well as high quality imaging of sections.

The incidence of inflammatory bowel diseases (IBD) is increasing worldwide. Mesenchymal stem/stromal cells (MSCs) have immunomodulatory functions and are a promising source for cell transplantation therapy for IBD. However, owing to their heterogeneous nature, their therapeutic efficacy in colitis is controversial and depends on the delivery route and form of transplanted cells. Cluster of differentiation (CD)73 is widely expressed in MSCs and used to obtain a homogeneous MSC population. Herein, we determined the optimal method for MSC transplantation using CD73+ cells in a colitis model. mRNA sequencing analysis showed that CD73+ cells exhibit downregulation of inflammatory genes and upregulation of extracellular matrix-related genes. Furthermore, three-dimensional CD73+ cell spheroids showed enhanced engraftment at the injured site through the enteral route, facilitated extracellular matrix remodeling, and downregulated inflammatory gene expression in fibroblasts, leading to attenuation of colonic atrophy. Therefore, the interaction between intestinal fibroblasts and exogenous MSCs via tissue remodeling is one mechanism that can be exploited for colitis prevention. Our results highlight that transplantation of homogeneous cell populations with well-characterized properties is beneficial for IBD treatment.

3Z,5E-Octa-3,5-diene-1,3,4-tricarboxylic acid-3,4-anhydride (ODTAA, 1) was isolated from Paecilomyces sp. FKI-6801 for its selective IMP-1 MBL inhibitory activity. The first total synthesis of 1 from the commercially available compound was achieved in 9 steps with 28% overall yield. Introduction of catechol to the maleic anhydride of 1 improved the IC

Ivermectin (IVM), an avermectin-derivative anthelmintic, specifically binds to glutamate-gated chloride ion channels (GluCls), causing paralysis in invertebrates. IVM also exhibits other biological activities such as Wnt/β-catenin pathway inhibition in vertebrates that do not possess GluCls. This study showed that affinity purification using immobilized IVM B1a isolated TELO2, a cofactor of phosphatidylinositol 3-kinase-related kinases (PIKKs), as a specific IVM B1a-binding protein.

We compared four derivatization reagents to analyze catecholamines and amino acids by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. 2,4,6-Trimethylpyrylium tetrafluoroborate (TMPy), 2,4-diphenyl-pyranylium tetrafluoroborate (DPP-TFB), 4-(anthracen-9-yl)-2-fluoro-1-methylpyridin-1-ium iodide (FMP-10), and triphenyl pyrilium (TPP) were used as derivatization reagents that can specifically modify primary amines or hydroxy groups in target molecules. Three derivatization reagents, not including TPP, reacted with all target molecules. The derived catecholamines dopamine and L-DOPA, and the amino acids GABA and glycine, were efficiently ionized in comparison with non-derivatized targets. Comparative analysis indicated that TMPy and FMP-10 produced general increases in signal-to-noise ratios (S/N), whereas DPP and TPP produced specific increases in the S/N of GABA and DA. Notably, TMPy is a small molecule that efficiently reacts with target molecules due to the absence of high bulk and steric hinderance.

A simple and rapid analytical method was developed for determination of four biogenic amines [histamine (Him), cadaverine (Cad), tyramine (Tym), 2-phenylethylamine (Pea)] in fish and fish products. This method uses a new derivatization reagent, 2,4,6-triethyl-3,5-dimethyl pyrylium trifluoromethanesulfonate (Py-Tag). The four biogenic amines in the samples were extracted with trichloroacetic acid. The diluted extract was derivatized with Py-Tag (15 min at 50°C) and then subjected to liquid chromatography-tandem mass spectrometry (LC-MS/MS). The limits of quantification for the method were 2 mg/kg for Him, Tym, and Pea and 10 mg/kg for Cad. The matrix effects derived from the tested fish and fish products were negligible in the LC-MS/MS analysis. The impact of the sample matrices on the Py-Tag derivatization was also negligible. The trueness and repeatability of the method were assessed by performing replicate analyses (n = 5) of five samples of fish and fish products, each spiked with the four biogenic amines at three different concentration levels. Analysis of the samples found 87%-104% of the spiked concentrations and the relative standard deviations were6.1%. A reference sample and quality control canned fish samples were analyzed by the method, and the concentrations of the Him were within acceptable limits. The developed method was successfully used to determine concentrations of the four biogenic amines in 48 fish and fish products on the Japanese market. The developed method does not require cleanup using a solid-phase extraction column or similar, and the derivatization reaction time was only 15 min. The results suggested that the present method is reliable and suitable for rapid analysis of the four biogenic amines in fish and fish products.

Baculovirus-infected silkworms are promising bioreactors for producing recombinant glycoproteins, including antibodies. Previously, we developed a method for isotope labeling of glycoproteins for nuclear magnetic resonance (NMR) studies using silkworm larvae reared on an artificial diet containing 15N-labeled yeast crude protein extract. Here, we further develop this method by introducing a technique for the expression of isotope-labeled glycoproteins by silkworm pupae, which has several potential advantages relative to larvae-based techniques in terms of production yield, ease of handling, and storage. Here, we fed fifth instar larvae an artificial diet with an optimized composition containing [methyl-13C]methionine, leading to pupation. Nine-day-old pupae were then injected with recombinant Bombyx mori nucleopolyhedrovirus (BmNPV) bacmid for expression of recombinant human immunoglobulin G (IgG). From the whole-body homogenates of pupae, 0.35 mg/pupa of IgG was harvested, which is a yield that is five times higher than can be obtained from larvae. Recombinant IgG, thus prepared, exhibited mainly three kinds of pauci-mannose-type oligosaccharides and had a 13C-enrichment ratio of approximately 80%. This enabled selective observation of NMR signals originating from the methionyl methyl group of IgG, confirming its conformational integrity. These data demonstrate the utility of silkworm pupae as factories for producing recombinant glycoproteins with amino-acid-selective isotope labeling.

A computed tomography (CT) scan revealed 2 nodules in the right upper and middle lobes of the lung and swelling of an upper mediastinal lymph node (#2R) in a 77-year-old male. Positron emission tomography (PET)/CT showed abnormal uptake only in the right middle lobe nodule, so we suspected a double primary lung cancer (cT1bN0M0, stage ⅠA), and performed a right upper and middle lobectomy with ND2a-2 dissection. Pathological investigation revealed that the lung nodules were adenocarcinomas and the lymph node swelling #2R was a metastasis of thyroid cancer. After surgery, careful examination was done for thyroid but the primary lesion was not found. Careful observation for an occult thyroid cancer is continuing at the outpatient.

Site specific introduction of the polar hydrophobic trifluoromethyl-λ6-tetrafluorosulfanyl (CF3SF4) group can effectively control the secondary structure of a heptapeptide, the minimum repeat unit of an α-helix. The structural influence of CF3SF4-containing amino acid on the heptapeptide was established using NMR methods.

The quantification of amino acids, peptides and proteins are important in fundamental science to understand intravital event, as well as in applied fields such as biological, medical, and pharmaceutical sciences. Py-Tags (Py0, Py6 and Py12) are primary amine-reactive isotope-labelled tagging reagents. Py-Tags can selectively and efficiently react with the primary amino group which has more than a two-carbon chain from the alpha-carbon of the target sample by the reaction of pyrylium salts with nucleophiles under alkaline conditions, thereby increasing the mass by 175.15, 181.17, and 187.19 Da, respectively, and generating positive ions. We used Py-Tags to label not only the amino acids but also peptides with a lysine residue(s), fragment peptides with a lysine(s), digested proteins, and intact proteins. All Py-Tag-labelled peptide sets showed similar mass differences. Thus, we could easily analyze expression ratio by single measurement. Furthermore, the ratio of the measured ion intensities was representative of the relative amount of each material.

Objectives We examined the usability of theophylline extended-release dry syrups, including their powder fluidity and ingestibility using one branded and six generic formulations (formulations A, B, C, D, E, F, and G, respectively). Methods To evaluate the ease of handling by pharmacists, powder fluidity was evaluated by measuring the repose angle, whereas to determine ingestibility, the viewpoints of patients or caregivers were evaluated using a questionnaire survey. Results and discussion: The repose angles of formulations A, B, C, D, E, and F were approximately 30°–40°. These formulations had sufficient powder fluidity, indicating the ease of weighing for most pharmacists. However, the repose angle of formulation G was Conclusions The present study provides useful information for selecting branded or generic theophylline extended-release dry syrups.

Objectives In this study, we examined the taste and dissolution of theophylline extended-release dry syrups using one branded and six generic formulations. Methods Healthy volunteers underwent sensory testing to assess the syrup taste. Drug dissolution was examined using in vitro dissolution tests. Results and discussion: Syrup taste, including sweetness, bitterness, and aftertaste, was different between formulations. These differences can likely be attributed to the presence of different pharmaceutical additives between formulations. Our results indicated that syrup aftertaste was dependent on the level of sweetness and bitterness. In addition, dissolution behaviors were different between formulations, suggesting bio-inequivalence between formulations. These differences likely reflect the use of different controlled-release technologies between formulations. Conclusions This study provides useful information for selecting branded or generic theophylline extended-release dry syrups for individualized treatments.

The chemistry of the 3,3,3-trifluoroprop-1-enyl (TFPE) group has attractive characteristics in medicinal chemistry as a new fluorine motif. However, there are no reports on the properties of this group because it is difficult to construct molecules with it. For the convenient construction of the TFPE group, a new fluorination reagent, CF3CH=CHTMS (1), was developed from commercially available chemicals with easy purification processes and excellent yields. The utility of 1 as a trifluoropropenylation reagent was exhibited in several types of reaction such as the Sonogashira cross-coupling reaction. Furthermore an indometacin analogue bearing a TFPE group showed greater pharmaceutical activity than the original indometacin. This review describes the details of these research studies under three topics: 1) synthesis of 1; 2) Sonogashira cross-coupling reaction of 1 with acetylene, followed by cyclization into an indole ring; and 3) synthesis of an indometacin analogue with a TFPE group.

The angiogenic gene therapy is an attractive approach for the treatment of ischemic muscle diseases, including peripheral arterial disease and ischemic heart diseases. Although a variety of gene transfer methods have been developed, the efficiency of gene transfer is still limited. We have been developing the needleless high-energy bioinjector device, Pyro-drive Jet Injector (PJI), based on pyrotechnics using a combination of ignition powder and gunpowder, however, the utility of PJI in gene transfer into muscle tissues remains unclear. pcDNA3.1 plasmid containing Flag was injected to the thigh muscles of C57BL/6J mice using PJI or needle, as a control. Histological analysis demonstrated that the protein expression of Flag was observed in a wider range in PJI group than in needle group. To assess the validity of PJI for gene therapy, pcDNA3.1-human fibroblast growth factor 2 (FGF2), which has angiogenic activity and tissue protective properties, was injected into the ischemic thigh muscles with PJI or needle. ELISA assay revealed that the protein expression of FGF2 was increased in the thigh muscle tissues by PJI-mediated gene delivery. Significantly, histological analyses revealed that muscle fiber cross-sectional area and the number of endothelial marker CD31 (+) cells was increased in ischemic hind-limb tissues of the PJI-FGF2 group but not in those of needle-FGF2 group. To expand the applicability of the PJI-mediated gene transfer, pcDNA3.1-venus plasmid was injected into murine hearts with PJI or needle. PJI method was successful in gene transfer into murine hearts, especially into cardiomyocytes, with high efficiency when compared to needle method. Collectively, the non-needle, non-liposomal and non-viral gene transfer by PJI could be a novel therapeutic approach for muscle diseases.

We describe the preparation of a cyclic hemiacetal-lactone 6, which is a key intermediate in the total synthesis of Aurodox (1) and also a degradation product of aurodox. The bicyclic core of 6 was obtained by lactonization and hemiacetalization after stereo-selective epoxidation at late stage. The construction of olefins utilized a Wittig reaction and a Horner-Wadsworth-Emmons reaction using d -pantolactone (14).

The addition of InBr3 to the oxidative Sonogashira cross-coupling reaction of 2-ethynylaniline with (E)-trimethyl(3,3,3-trifluoroprop-1-enyl)silane led to a dramatic increase in the reactivity to afford the corresponding 1,3-enynes bearing a trifluoromethyl group on their terminal sp(2) carbon. The subsequent cyclization of these 1,3-enynes under palladium catalysis provides access to the corresponding indoles bearing a 3,3,3-trifluoroprop-1-enyl group at their 2-position.

Oxidative Sonogashira cross-coupling reactions of (E)-trimethyl(3,3,3-trifluoroprop-1-enyl)silane with arylacetylene were achieved using silver fluoride and a palladium catalyst, to afford high yields of various 1,3-enynes with a CF3 group on the terminal sp(2) carbon. Silver fluoride promoted C-Si bond dissociation and oxidation of palladium, enabling catalytic use of palladium.

The Hiyama cross-coupling reaction of (E)-trimethyl(3,3,3-trifluoroprop-1-enyl)silane (1) with 2-iodoaniline (2) proceeded without any protection of the amino group. The coordination of copper(II) fluoride to 2,2'-bipyridyl provided the fluoride source required to trigger this reaction, affording (E)-2-(3,3,3-trifluoroprop-1-enyl)aniline (3). In the presence of a stoichiometric amount of [Cu(OTf)]2·C6H6, the treatment of 3 with an aryl aldehyde at 200 °C provided the 2-aryl-3-trifluoromethylquinoline (4) via the oxidative cyclization of an in situ-generated imine substructure.

The addition of InBr3 to the oxidative Sonogashira reaction of aryl acetylenes (I) and (V) with silane (II) leads to a dramatic increase in reactivity to afford the corresponding 1,3-enynes (III) and (VI), respectively.

(E)-trimethyl-(3,3,3-trifluoroprop-1-enyl)silane (1) was synthesized as a reagent for use in Hiyama cross-coupling reactions for the production of β-trifluoromethylstyrene derivatives. Cross-coupling of 1 with electronically diverse aryl iodides was achieved by treatment with CsF in the presence of catalytic amounts of palladium to afford the desired products in moderate to good yields.

Oxidative Sonogashira cross-coupling reactions of (E)-trimethyl(3,3,3-trifluoroprop-1-enyl)silane with arylacetylene were achieved using silver fluoride and a palladium catalyst, to afford high yields of various 1,3-enynes with a CF3 group on the terminal sp2 carbon. Silver fluoride promoted C–Si bond dissociation and oxidation of palladium, enabling catalytic use of palladium.

Adsorption isotherm measurements were carried out for N2 on the basal plane of graphite held at a temperature 77 K and preplated with n-alkanes or conjugated species such as mono-substituted benzenes, anthracene, 9,10-dimethylanthracene, and phthalocyanine copper. The isotherm step arising from the fluid–solid phase transition has been found to be abnormally depressed by the conjugated species, whereas the n-alkanes have no effect on the phase behavior of N2. A similar phase transition for CO has also been found to be depressed by preadsorbed phthalocyanine copper. It is suggested that a small amount of these conjugated species dissolved in N2 and CO locally induce the 3×3 commensurate structure of N2 and CO to increase the molecular density in the fluid phase and, therefore, to depress abnormally the fluid–solid phase transition of N2 and CO.

The cyclocondensation reaction to yield quinolines (V) is only suitable for aromatic aldehydes (IV), but not aliphatic aldehydes.

We compared two ionization methods, matrix assisted laser desorption/ionization (MALDI) and nanoparticle assisted laser desorption/ionization (Nano-PALDI) mass spectrometry (MS), for the analysis of amino acids derivatized with Py-Tag™ that consists pyrylium-based compound. Py-Tag is a useful stable derivatization reagent due to wide mass differences (using 13C as the sole stable labelling isotope). For Py-Tag labelled lysine, sensitive signals that showed less noise with a ten times higher sensitivity, showed a wider mass difference by Nano-PALDI MS compared to MALDI MS.

Starting from commercially available materials, a gram-scale preparation of the key compound (II) is described in three steps with an overall yield of 74%.

The fluid–solid (FS) phase transition of N 2 adsorbed on graphite is known to be depressed abnormally by conjugated coplanar molecules such as benzene and copper phthalocyanine. This paper describes an investigation of the influence of a non-aromatic conjugated species (1,3-butadiene), a non-aromatic cation acceptor (18-crown-6), a strong electron donor (tetrathiafulvalene) and a strong electron acceptor (tetracyanoquinodimethane) on the FS phase transition of N 2 , when they were preplated on graphite. These species were, in fact, found to have no significant effect. This implies that molecular properties such as cation acceptance and electron transfer are of no importance, whereas the aromaticity of co-existing species plays a crucial role in disturbing the phase behaviour of N 2 .