Investigation of Invigorating Qi and Activating Blood Circulation Prescriptions in Treating Qi Deficiency and Blood Stasis Syndrome of Ischemic Stroke Patients: Study Protocol for a Randomized Controlled Trial.

Ischemic stroke (IS) is characterized by high morbidity and high mortality. The integration of Traditional Chinese medicine (TCM) and western medicine has shown promising benefits in relieving symptoms, promoting neurological recovery, and improving the quality of life of patients with IS. In TCM, Qi-deficiency along with blood-stasis (QDBS) syndrome is one of the common types of IS that is treated by invigorating Qi and activating blood circulation. In TCM theory, improving the corresponding degree of prescription-syndrome correlation (PSC) is helpful to improve clinical efficacy. In this study, we intend to use similar prescriptions that invigorate Qi and activate blood circulation: Buyang Huanwu granules (BHG), Naoxintong capsules (NXTC), and Yangyin Tongnao granules (YTG). The goal is to evaluate their level of PSC inpatients with IS with QDBS syndrome and find relevant biomarkers to provide an objective basis for precise treatment of TCM and improve the clinical therapeutic effects. A multicenter, randomized, double-blinded, and placebo-controlled intervention trial will be conducted in IS patients with QDBS syndrome, followed by an add-on of Chinese patent medicine. A total of 160 subjects will be randomly assigned to the BHG, NXTC, YTG, and placebo groups in a 1:2:1:1 allocation ratio. All subjects will undergo 28 days of treatment and then followed for another 180 days. The primary outcome is the changes in the National Institutes of Health Stroke Scale score after 28 days of medication. The secondary outcomes include the modified Rankin scale score, activity of daily living scale score, and TCM symptom score. Data will be analyzed in accordance with a predefined statistical analysis plan. Ethical approval of this trial has been granted by the Research Ethics Committee of the First Affiliated Hospital of Zhejiang Chinese Medical University (ID: 2017-Y-004-02). Written informed consent of patients will be required. This trial is registered in the Chinese Clinical Trial Registry (ChiCTR1800015189), and the results will be disseminated to the public through peer-reviewed journals and academic conferences.

Scavenging activity and mechanism study of ferulic acid against reactive carbonyl species acrolein.

Acrolein, known as one of the most common reactive carbonyl species, is a toxic small molecule affecting human health in daily life. This study is focused on the scavenging abilities and mechanism of ferulic acid and some other phenolic acids against acrolein. Among the 13 phenolic compounds investigated, ferulic acid was found to have the highest efficiency in scavenging acrolein under physiological conditions. Ferulic acid remained at (3.04±1.89)% and acrolein remained at (29.51±4.44)% after being incubated with each other for 24 h. The molecular mechanism of the detoxifying process was also studied. Detoxifying products, namely 2-methoxy-4-vinylphenol (product 21) and 5-(4-hydroxy-3-methoxyphenyl)pent-4-enal (product 22), were identified though nuclear magnetic resonance (NMR) and gas chromatography-mass spectrometry (GC-MS), after the scavenging process. Ferulic acid showed significant activity in scavenging acrolein under physiological conditions. This study indicates a new method for inhibiting damage from acrolein.

Assay of dried blood spot from finger prick for sodium valproate via ink auxiliary headspace gas chromatography mass spectrometry.

Sodium valproate is the most commonly used antiepileptic drug that patients need to keep taking over a long period of time or on a permanent basis. Its blood concentration should be accurately detected to avoid toxicity or side-effects, especially for children and the aged. Dried blood spot (DBS) sampling from finger prick is a minimally invasive and patient-friendly procedure for blood collection. However, there are few studies about rapid detection of sodium valproate in DBS samples in current literatures. In this work, we developed an ink auxiliary headspace gas chromatography mass spectrometry (GC-MS) strategy for direct detection of sodium valproate in DBS from epilepsy patients, which does not need extra solvent extraction or elution. It was discovered that carbon black ink could provide better capacity of heat absorption and dissociation, and higher quality of headspace sampling. The detection sensitivity has been improved with reported headspace GC-MS methods, and the limit of quantitation could reach to 200 ng/mL. Finally, this strategy was practically applied to quantify sodium valproate in DBS samples from 29 epilepsy patients. The result showed higher accuracy with lower relative errors by comparing with the clinical immunoassay results. In conclusion, we developed a direct detection method for DBS samples that is suitable for high-throughput clinical test with great potential for clinical application.

Hydrolytic cleavage of both CS2 carbon-sulfur bonds by multinuclear Pd(II) complexes at room temperature.

Developing homogeneous catalysts that convert CS2 and COS pollutants into environmentally benign products is important for both fundamental catalytic research and applied environmental science. Here we report a series of air-stable dimeric Pd complexes that mediate the facile hydrolytic cleavage of both CS2 carbon-sulfur bonds at 25 °C to produce CO2 and trimeric Pd complexes. Oxidation of the trimeric complexes with HNO3 regenerates the dimeric starting complexes with the release of SO2 and NO2. Isotopic labelling confirms that the carbon and oxygen atoms of CO2 originate from CS2 and H2O, respectively, and reaction intermediates were observed by gas-phase and electrospray ionization mass spectrometry, as well as by Fourier transform infrared spectroscopy. We also propose a plausible mechanistic scenario based on the experimentally observed intermediates. The mechanism involves intramolecular attack by a nucleophilic Pd-OH moiety on the carbon atom of coordinated µ-OCS2, which on deprotonation cleaves one C-S bond and simultaneously forms a C-O bond. Coupled C-S cleavage and CO2 release to yield [(bpy)3Pd3(µ3-S)2](NO3)2 (bpy, 2,2'-bipyridine) provides the thermodynamic driving force for the reaction.

XTH31, encoding an in vitro XEH/XET-active enzyme, regulates aluminum sensitivity by modulating in vivo XET action, cell wall xyloglucan content, and aluminum binding capacity in Arabidopsis.

Xyloglucan endohydrolase (XEH) and xyloglucan endotransglucosylase (XET) activities, encoded by xyloglucan endotransglucosylase-hydrolase (XTH) genes, are involved in cell wall extension by cutting or cutting and rejoining xyloglucan chains, respectively. However, the physiological significance of this biochemical activity remains incompletely understood. Here, we find that an XTH31 T-DNA insertion mutant, xth31, is more Al resistant than the wild type. XTH31 is bound to the plasma membrane and the encoding gene is expressed in the root elongation zone and in nascent leaves, suggesting a role in cell expansion. XTH31 transcript accumulation is strongly downregulated by Al treatment. XTH31 expression in yeast yields a protein with an in vitro XEH:XET activity ratio of >5000:1. xth31 accumulates significantly less Al in the root apex and cell wall, shows remarkably lower in vivo XET action and extractable XET activity, has a lower xyloglucan content, and exhibits slower elongation. An exogenous supply of xyloglucan significantly ameliorates Al toxicity by reducing Al accumulation in the roots, owing to the formation of an Al-xyloglucan complex in the medium, as verified by an obvious change in chemical shift of (27)Al-NMR. Taken together, the data indicate that XTH31 affects Al sensitivity by modulating cell wall xyloglucan content and Al binding capacity.

LC/MS guided isolation of alkaloids from lotus leaves by pH-zone-refining counter-current chromatography.

The traditional methods used in natural product separation primarily target the major components and the minor components may thus be lost during the separation procedure. Consequently, it's necessary to develop efficient methods for the preparative separation and purification of relatively minor bioactive components. In this paper, a LC/MS method was applied to guide the separation of crude extract of lotus (Nelumbo nucifera Gaertn.) leaves whereby a minor component was identified in the LC/MS analysis. Afterwards, an optimized pH-zone-refining CCC method was performed to isolate this product, identified as N-demethylarmepavine. The separation procedure was carried out with a biphasic solvent system composed of hexane-ethyl acetate-methyl alcohol-water (1:6:1:6, v/v) with triethylamine (10 mM) added to the upper organic phase as a retainer and hydrochloric acid (5 mM) to the aqueous mobile phase eluent. Two structurally similar compounds--nuciferine and roemerine--were also obtained from the crude lotus leaves extract. In total 500 mg of crude extract furnished 7.4 mg of N-demethylarmepavine, 45.3 mg of nuciferine and 26.6 mg of roemerine with purities of 90%, 92% and 96%, respectively. Their structures were further identified by HPLC/ESI-MSn, FTICR/MS and the comparison with reference compounds.

Highly efficient and catalyst-free synthesis of substituted thioureas in water.

A highly efficient and catalyst-free method for the synthesis of substituted thiourea derivatives is presented. The synthesis is performed through the reaction of thiazolidine-2-thiones with various amines in water. A sound number of thioureas with both unsymmetrical and symmetrical architectures have been obtained in good to excellent yields.

Self-assembly and host-guest interaction of metallomacrocycles using fluorescent dipyrazole linker with dimetallic clips.

By employing diimine ligands coordinated dimetallic clips ([(bpy)2Pd2(NO3)2](NO3)2 or [(phen)2Pd2(NO3)2](NO3)2, where bpy = 2,2'-bipyridine, phen = 1,10-phenanthroline) as the corner and anthracene-, naphthalene-, and benzene-based dipyrazolate dianions as the linker, a series of positively charged metallomacrocycles ([M4L2]4+ or [M8L4]8+) have been synthesized through a directed self-assembly method in aqueous solution. Every macrocycle has a cavity to bind solvent molecules or anions. The structures were characterized by elemental analysis, 1H and 13C NMR, electrospray ionization mass spectrometry, and single crystal X-ray diffraction analysis for compound 1 x 4 PF6(-) (1 = {[(bpy)Pd]4L(1)2}4+), 3 x 4 PF6(-) x 8 CH3CN x H2O (3 = {[(bpy)Pd]4L(2)2}4+), and 7 x 4 PF6(-) x 6 H2O (7 = {[(bpy)Pd]4L(5)2}4+). The 1:1 host-guest complexation for anthracene-based dipyrazolate-bridged macrocycles with aromatic guests was investigated via UV-vis and fluorescent titration.

Resveratrol and its oligomers from wine grapes are selective (1)O2 quenchers: mechanistic implication by high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry and theoretical calculation.

Resveratrol and its oligomers, abundantly present in wine grapes, are believed to be effective phytoalexins for the phenomenon "French paradox" partially by virtue of their powerful antiradical properties. EPR spin-trapping technique was utilized, demonstrating all polyphenols were selective (1)O2 quenchers but not effective (•)OH and O2(•¯) scavengers. On the basis of the HPLC-ESI-MS(2) analysis for the simulated reactions of polyphenols with (1)O2, the molecular weights of the resulting photochemical products were 14 or 28 Da higher than those of their substrates. No fragment C2H2O (42 Da), which was rather distinctive of the resorcinol rings in these cases, had been observed, whereas their MS/MS spectra displayed characteristic neutral fragments including carbon monoxide (CO, 28 Da) and 2-hydroxy[1,4]benzoquinone (C6H4O3, 124 Da). Finally, PM3 semiempirical calculations and HR-FTICR-MS experiments were performed, supporting the assertion that their quenching mechanism involved physical and chemical pathways. Chemical quenching underwent an endoperoxide intermediate form to generate quinones.

Self-assembly of bowl-like trinuclear metallo-macrocycles.

By employing different diimine complexes and a naphthanoimidazolate (L(1)) or benzoimidazolate (L(2)) anion ligand as linker, a series of trimetallo-macrocycles have been synthesized through a directed self-assembly approach that involves spontaneous deprotonation of the ligands in aqueous solution. Some compounds, namely, [M(3)L(3)](NO(3))(3) (M = (18-crown-6-phen)Pt, 1 or 7; (15-crown-5-phen)Pt, 2; (benzo-24-crown-8-phen)Pt, 8; (benzo-24-crown-8-phen)Pd, 9; (15-crown-5-phen)Pd, 4 or 11; (18-crown-6-phen)Pd, 3 or 10; (dmbpy)Pd, 6 or 14; (bpy)Pd, 5 or 13; (bpy)Pt, 12; (N4-Phen)Pd, 15) were synthesized. In all of these compounds, the L(1)or L(2) anion ligands are in a syn, syn, syn orientation which result in a bowl-like cavity that can serve as a host to bind the methyl of a solvent CH(3)CN within the naphthanoimidazolate or benzoimidazolate-built cavity through C-H...pi hydrogen bonds in the crystal state. The structures are characterized by elemental analysis, (1)H NMR, ESI-MS, and in the cases of 4a, 11, 13, and 14a by single-crystal X-ray diffraction analysis.

Chemo-/regioselective synthesis of 6-unsubstituted dihydropyrimidinones, 1,3-thiazines and chromones via novel variants of Biginelli reaction.

A novel and facile cascade Biginelli-like assembly employing enaminone, aldehyde and urea/thiourea has been developed, which provides a highly chemo- and regioselective synthesis of new dihydropyrimidinones, 1,3-thiazines and chromones by altering particular functional groups in the reactants.

Novel regioselectivity: three-component cascade synthesis of unsymmetrical 1,4- and 1,2-dihydropyridines.

The three-component sequential reaction of alpha,beta-unsaturated aldehydes, amines, and enaminones proceeded smoothly to give 1,3,4-trisubstituted 1,4-dihydropyridines in aqueous DMF. Moreover, the unexpected regioselective formation of 1,2-dihydropyridines has been observed for the first time in such an approach. On the basis of a systematic study, the novel regioselectivity could be assigned both to steric and electronic effects originating from the amine partner.

[Studies on chemical constituents and antibacterial activity from n-butanol extract of Sarcandra glabra].

OBJECTIVE: To study the chemical constituents and the antibacterial activity from n-butanol extract of Sarcandra glabra. METHOD: The compounds were isolated by Diaion HP-20, Sephadex LH-20, MCI CHP-20 and silica gel column chromatographic methods. Their structures were identified on the basis of physicochemical and spectroscopic analysis. The antibacterial effect of the compounds were measured against Staphylococcus aureus by filterpaper slice method, finally the antibacterial ring in each group was recorded after 24 hours. RESULT: Seven constituents were isolated and elucidated as 5, 7, 3', 4'-tetrahydroxy-6-C-beta-D-glucopyranosylflavanone (1), kaempferol-3-O-beta-D-glucuronide (2), fraxidin (3), isofraxidin (4), isofraxidin-7-O-beta-D-glucopyranoside (5), kaempferol (6), pinostrobin (7). Diameters (in mm) of antibacterial ring in the compounds 2, 5, 6 were orderly recorded as follows: 14.67 +/- 0.08, 11.14 +/- 1.06, 8.26 +/- 1.26 and the compound 4 is not effective. CONCLUSION: Compounds 1-3 and 5 were isolated from S. glabra for the first time.

Preliminary studies on the chemical characterization and antioxidant properties of acidic polysaccharides from Sargassum fusiforme.

In order to investigate the antioxidant properties of the polysaccharides from the brown alga Sargassum fusiforme, the crude polysaccharides from S. fusiforme (SFPS) were extracted in hot water, and the lipid peroxidation inhibition assay exhibited that SFPS possessed a potential antioxidant activity. Hence, two purely polymeric fractions, SFPS-1 and SFPS-2 were isolated by the column of DEAE (2-diethylaminoethanol)-Sepharose Fast Flow, with their molecular weights of 51.4 and 30.3 kDa determined by high performance gel permeation chromatography (HPGPC). They were preliminarily characterized using chemical analysis in combination of infrared (IR) and nuclear magnetic resonance (NMR) spectroscopies and found to contain large amounts of uronic acids and beta-glycosidical linkages. The antioxidant activities of these two SFPS fractions were evaluated using superoxide and hydroxyl radical-scavenging assays. The results show that the antioxidant ability of SFPS-2 was higher than that of SFPS-1, probably correlating with the molecular weight and uronic acid content.

New tyrosinase inhibitory sesquiterpenes from Chloranthus henryi.

Chemical investigation of the leaves and stems of Chloranthus henryi resulted in the isolation and characterization of two new eudesmane-type sesquiterpenes, and two new germacrane-type sesquiterpenes, together with two known compounds. The inhibitory activities against tyrosinase of all isolates were also evaluated.

[Effect of puerarin and ligustrazine on cultured hippocampal neurons injury induced by Abeta25-35].

OBJECTIVE: To investigate the changes of MDA, SOD, LDH of cultured hippocampal neurons injury induced by amyloid-beta protein (Abeta 25-35) and the protective effect of puerarin and ligustrazine. METHOD: Primary hippocampal neurons were cultured and induced by Abeta 25-35. The concentrations of MDA, SOD and LDH in cultured hippocampal neurons were measured after exposed to Abeta 25-35, puerarin and ligustrazine. RESULT: The Alzheimer disease (AD) model was successfully established in cultured hippocampal neurons. AD group has remarkably increased MDA and LDH level, and decreased SOD level, Piracetan group and combined application group of have remarkably decreased MDA and LDH level and increased SOD level, compared with AD group (P < 0.01). Ligustrazine together with puerarin group has remarkably decreased MDA and LDH level and increased SOD level, compared with ligustrazine group and puerarin group (P < 0.05). CONCLUSION: Abeta 25-35 can induce cultured hippocampal neurons injury, combined application of ligustrazine, and puerarin can alleviate the injury.

Nisin-controlled extracellular production of apidaecin in Lactococcus lactis.

Apidaecins are heat-stable, nonhelical antibacterial peptides isolated from lymph fluid of the honeybee (Apis mellifera). These peptides are active against a wide range of gram-negative bacteria and they are the most prominent components of the honeybee humoral defense against microbial invasion. In the present study, one isoform of apidaecin, apidaecin Ho, was expressed extracellularly in the food-grade bacterium Lactococcus lactis. Results showed that expression driven by the lactococcal nisA promoter and Usp45 signal peptide resulted in efficient secretion of apidaecin in L. lactis subsp. cremoris NZ9000. Recombinant apidaecin was purified by gel filtration and semipreparative RP-HPLC, and about 10 mg active recombinant apidaecin was obtained from 1,000 ml culture. This is the first report on the nisin-controlled extracellular production of active apidaecin in L. lacits. The expression and delivery of apidaecin in the food-grade L. lactis may provide a clue to facilitate the widespread application of apidaecin in the control and prevention of gram-negative bacteria infections of human and animals.

Self-assembly, structures, and photophysical properties of 4,4'-bipyrazolate-linked metallo-macrocycles with dimetal clips.

By employing functional diimine ligands coordinated dipalladium(II,II) or diplatinum(II,II) clips as corners and the coplanar 4,4'-bipyrazolate dianion (L(2-)) ligand as linker, a series of bipyrazolate-bridged metallo-macrocycles, namely, [M8L4](NO3)8 (M = Pd(dmbpy), 1; Pd(bpy), 2; Pt(bpy), 3a; Pd(phen), 4; Pt(phen), 5; Pd(15-crown-5-phen), 6; Pd(18-crown-6-phen), 8; Pd(benzo-24-crown-8-phen), 10a; Pt(15-crown-5-phen), 7a, Pt(18-crown-6-phen), 9a; Pt(benzo-24-crown-8-phen), 11a) and [M6L3](NO3)6 (M = Pt(bpy), 3b; Pt(15-crown-5-phen), 7b; Pt(18-crown-6-phen), 9b; Pd(benzo-24-crown-8-phen), 10b; Pt(benzo-24-crown-8-phen), 11b), have been synthesized through a directed self-assembly approach that involves spontaneous deprotonation of the 1H-bipyrazolyl ligands in aqueous solution. All these compounds have a crown-shaped cavity that can serve as host to solvent molecules and anions. The structures are characterized by elemental analysis, (1)H and (13)C NMR, ESI-MS, and in the cases of 1a (the BF4(-) salt of 1), 2a (the BF4(-) salt of 2), and 3b by single-crystal X-ray diffraction analysis. Photophysical properties for complexes 1 and 2 are discussed.

Functional and structural characterization of apidaecin and its N-terminal and C-terminal fragments.

Two aspects were studied to elucidate the functional and structural characterization of apidaecin and its N-terminal and C-terminal fragments: (i) Functions of the N-terminal and C-terminal fragments of apidaecin were first studied by measuring their antibacterial activity, their ability to enter Escherichia coli cells and their effects on the activities of beta-galactosidase and alkaline phosphatase. The results indicate that neither the N-terminal nor the C-terminal of apidaecin contains intracellular delivery unit or active segment. (ii) The effect of apidaecin on the ATPase activity of DnaK, and the interactions of apidaecin with E.coli lidless DnaK and DnaK D-E helix were studied. Results showed that apidaecin could interact with the E.coli lidless DnaK protein and stimulate its ATPase activity, but not with E.coli DnaK D-E helix. This indicated that the antimicrobial activity of apidaecin may be shown by stimulating the ATPase activity of DnaK by binding to its conventional substrate-binding site, to decrease its cellular concentration of DnaK by competing with natural substrates and inhibit the enzymes' activities of E. coli cells. It is the first study to suggest that the apidaecin-binding site of DnaK is the conventional substrate binging site.