Syntheses of new model compounds related to an antigenic epitope from Bupleurum falcatum L. and their distributions in various ganglioside-phospholipid monolayers

6-N-[2-(Tetradecyl)hexadecanamido]hexyl beta-D-glucopyranosyluronic acid-(1-->6)-beta-D-galactopyranosyl-(1-->6)-beta-D-galactopyranoside (1) and its clustering compound (2) carrying a tetravalent sugar unit, which are new model compounds related to a major antigenic epitope from antiulcer pectic polysaccharide of Bupleurum falcatum L., were synthesized and the distributions of 1 and 2 in mixed ganglioside (GM1, GD1a or GT1b)/phospholipid (DPPC) monolayers were observed using atomic force microscopy (AFM). AFM images showed that 1 was distributed in the GM1, GD1a and GT1b region of the mixed monolayers, in which 1 was miscible with GD1a. Specific distribution of 1 was observed in the mixed GM1/DPPC monolayer. Compound 2 was miscible with GM1, while 2 formed associations with GD1a and GT1b in the mixed monolayers. The distribution mode of 1 and 2 was different among the mixed ganglioside/DPPC monolayers.     

Inhibition effects of gangliosides G(M1), G(D1a) and G(T1b) on base-catalyzed isomerization of prostaglandin A(2)

Micellar inhibition effect of gangliosides on a degradation of drug was investigated, where ganglioside G(M1) (GM1), G(D1a) (GD1a) and G(T1b) (GTlb) whose sialic acid residue is one, two and three, respectively, were used. The base-catalyzed isomerization of prostaglandin A(2) (PGA(2)) to prostaglandin B(2) (PGB(2)) was chosen as a model experiment. The rate for the isomerization of PGA(2) was determined by measuring the concentration of PGA(2) (and PGB(2)) with a high-performance liquid chromatography. Gangliosides micelles inhibited the isomerization of PGA(2). The inhibition effect of GT1b micelles was larger than that of GD1a micelles. This result would be due to the larger absolute value of surface potential of GT1b micelles, which brings about a larger electrostatic repulsion between micellar surface and OH(-). The terminal sialic acid residue of ganglioside was effective to inhibit the isomerization of PGA(2). GM1 micelles without terminal sialic acid residue but with large aggregation number exhibited a superior steric shielding effect rather than an electrostatically repulsive effect. The inhibition effect of GM1 micelles was enhanced by the mixed micellization with the other ganglioside with a terminal sialic acid residue. GM1-GD1a or GM1-GT1b mixed micelles remarkably inhibited the isomerization of PGA(2). The physiological activity of PGs in the biological membranes containing gangliosides was also discussed.     

Synthesis and characterization of an anomeric sulfur analogue of CMP-sialic acid

alpha-2,3-Sialyltransferase catalyzes the transfer of sialic acid from CMP-sialic acid (1) to a lactose acceptor. An analogue of 1 was synthesized in which the anomeric oxygen atom was replaced with a sulfur atom (1S). The key step in the synthesis of 1S was a tetrazole-promoted coupling of a cytidine-5'-phosphoramidite with a glycosyl thiol of a protected sialic acid. Compounds 1 and 1S were characterized for their activity in a sialyl transfer assay. The rate of solvolysis in aqueous buffer of analogue 1S was 50-fold slower than that of 1. Analogue 1S was found to be substrate for alpha-2,3-sialyltransferase. The K(m) of 1S was just 3-fold higher than that of 1, while the k(cat) of 1S was 2 orders of magnitude lower compared to 1.     

Ab initio study on deactivation pathways of excited 9H-guanine

The complete active space with second-order perturbation theory/complete active space self-consistent-field method was used to explore the nonradiative decay mechanism for excited 9H-guanine. On the 1pipi* (1L(a)) surface we determined a conical intersection (CI), labeled (S0pipi*)(CI), between the 1pipi* (1L(a)) excited state and the ground state, and a minimum, labeled (pipi*)min. For the 1pipi* (1L(a)) state, its probable deactivation path is to undergo a spontaneous relaxation to (pipi*)min first and then decay to the ground state through (S0pipi*)(CI), during which a small activation energy is required. On the 1n(N)pi* surface a CI between the 1n(N)pi* and 1pipi* (1L(a)) states was located, which suggests that the 1n(N)pi* excited state could transform to the 1pipi* (1L(a)) excited state first and then follow the deactivation path of the 1pipi* (1L(a)) state. This CI was also possibly involved in the nonradiative decay path of the second lowest 1pipi* (1L(b)) state. On the 1n(O)pi* surface a minimum was determined. The deactivation of the 1n(O)pi* state to the ground state was estimated to be energetically unfavorable. On the 1pisigma* surface, the dissociation of the N-H bond of the six-membered ring is difficult to occur due to a significant barrier.     

Dynamic Ring-chain Equilibrium of Nucleophilic Thiol-yne “Click” Polyaddition for Recyclable Poly(dithioacetal)s

We report a dynamic polymerization system based on the reversible nucleophilic Michael polyaddition of activated alkynes and dithiols. Four poly(dithioacetal)s(P1-P4) were prepared via the base-catalyzed thiol-yne "click" polyaddition of two dithiols(1,4-butanedithiol(4 S) and 1,5-pentanedithiol(5 S)) and two alkynones(3-butyn-2-one(Y1) and 1-phenyl-2-propyn-1-one(Y2)) at high concentrations. We systematically investigated the base-catalyzed polymerization of 4 S and Y1(for polymer P1) under dif...     

De novo design, synthesis, and function of semiartificial myoglobin conjugated with coiled-coil two-alpha-helix peptides

The introduction of a flavin chromophore on the myoglobin (Mb) surface and an effective electron-transfer (ET) reaction through the flavin were successfully achieved by utilizing the self-assembly of heterostranded coiled-coil peptides. We have prepared a semiartificial Mb, named Mb-1alphaK, in which an amphiphilic and cationic alpha-helix peptide is conjugated at the heme propionate (Heme-1alphaK). Heme-1alphaK has a covalently bound iron-protoporphyrin IX (heme) at the N terminus of a 1alphaK peptide sequence. This sequence was designed to form a heterostranded coiled-coil in the presence of a counterpart amphiphilic and anionic 1alphaE peptide sequence in a parallel orientation. Two peptides, Fla(1)-1alphaE and Fla(31)-1alphaE, both incorporating a 10-methylisoalloxazine moiety as an artificial flavin molecule, were also prepared (Fla=2-[7-(10-methyl)isoalloxazinyl]-2-oxoethyl). Heme-1alphaK was successfully inserted into apomyoglobin to give Mb-1alphaK. Mb-1alphaK recognized the flavin-modified peptides and a two-alpha-helix structure was formed. In addition, an efficient ET from reduced nicotinamide adenine dinucleotide to the heme center through the flavin unit was observed. The ET rate was faster in the presence of Fla(1)-1alphaE than in the presence of Fla(31)-1alphaE or the equivalent molecule that has no peptide chain. These results demonstrate that the introduction of a functional chromophore on the Mb surface can be achieved by using specific peptide-peptide interactions. Moreover, the dependence of the ET rate on the position of the flavin indicated that the distance between the heme active site and the flavin chromophore was regulated by the three-dimensional structure of the designed polypeptide.     

Reactivity of 1-hydro-5-carbaphosphatrane based on tautomerization between pentavalent phosphorane and trivalent cyclic phosphonite

The reaction behavior of 1-hydro-5-carbaphosphatrane (1 a) was examined. Treatment of 1 a with oxidants such as 3-chloroperoxybenzoic acid (mCPBA) and tBuOCl gave cyclic phosphonate 2 and 1-chloro-5-carbaphosphatrane (4), respectively, via cyclic phosphonite 3, a tautomer of 1 a. Compound 4 was readily hydrolyzed to afford 2. Compound 1 a was also sulfurized via 3 by elemental sulfur to afford cyclic thioxophosphonate 5, which was also obtained by reactions in the presence of bases. Treatment of 1 a with bases also proceeded through 3 to give an equilibrium mixture of the corresponding phenoxide anion 10 and the phosphoranide anion 9, which was quenched with MeI to afford a mixture of 11 and 1-methyl-5-carbaphosphatrane (1 b). Such reactivities are typical for neutral phosphoranes. Theoretical investigations of these reactivities were also performed.     

A fluorescent sensor for 2,3-bisphosphoglycerate using a europium tetra-N-oxide bis-bipyridine complex for both binding and signaling purposes

Host 1 was designed and synthesized as a fluorescent sensor for 2,3-bisphosphoglycerate (BPG, 3). The design features a tris-functionalized triethylbenzene core to preorganize binding groups. The three cationic moieties, a tetra-N-oxide bipyridine-europium complex and two ammonium groups, were included to complement the three anionic functionalities on the guest. Beyond acting as a binding site, the europium complex was used to signal binding of the guest through modification of the charge transfer emission. A 1:1 complex with BPG was determined in 50 % methanol/acetonitrile with a K(a) of 6.7 x 10(5) mol(-1) by monitoring the reduction of the fluorescence signal upon guest addition. In the titration of related glycolytic intermediates lacking a second phosphate (4-6) into host 1, 2:1 host to guest binding was observed. Similarly, control compound 2, which lacks the ammonium groups, binds BPG and 4-6 in a 2:1 fashion. Also, phenylphosphate 7 binds to host 1 in a 1:1 stoichiometry with a K(a) over three times less than 3.     

Determination of ivermectin B(1a) in animal plasma by liquid chromatography combined with electrospray ionization mass spectrometry

A novel, sensitive and specific method for the quantitative determination of ivermectin B(1a) in animal plasma using liquid chromatography combined with positive electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) is presented. Abamectin was used as the internal standard. Extraction of the samples was performed with a deproteinization step using acetonitrile. Chromatographic separation was achieved on a Nucleosil ODS 5 microm column, using gradient elution with 0.2% (v/v) acetic acid in water and 0.2% (v/v) acetic acid in acetonitrile. The method was validated according to the requirements defined by the European Community. Calibration curves using plasma fortified between 1 and 100 ng ml(-1) showed a good linear correlation (r > or = 0.9989, goodness-of-fit coefficient < or =8.1%). The trueness at 2 and 25 ng ml(-1) (n = 6) was +4.2 and -17.1%, respectively. The trueness and between-run precision for the analysis of quality control samples at 25 ng ml(-1) was -4.0 and 11.0%, respectively (n = 16). The limit of quantification of the method was 1.0 ng ml(-1), for which the trueness and precision also fell within acceptable limits. Using a signal-to-noise ratio of 3 : 1, the limit of detection was calculated to be 0.2 ng ml(-1). The specificity was demonstrated with respect to ivermectin B(1b).The method was successfully used for the quantitative determination of ivermectin B(1a) in plasma samples from treated bovines, demonstrating the usefulness of the developed method for application in the field of pharmacokinetics.     

二苯并[b,f][1,5]二氮杂芳辛四烯衍生物的合成及拆分

以邻氨基二苯甲酮为原料,经自身缩合环化合成了3种二苯并[1,5]二氮杂芳辛四烯衍生物(1a~1c);以邻苯二甲酸酐和溴苯为原料经傅-克反应制得中间体2-(4-溴苯甲酰溴)苯甲酸(M1);M1经叠氮化后自缩合制得6,12-二(4-溴苯基)二苯并[b,f][1,5]二氮杂环辛四烯(1d);以邻氨基苯甲酸甲酯为原料,经自身缩合环化制得中间体二苯并[b,f][1,5]二氮杂环辛四烯-6,12(5H,11H)-二酮(M2);M2经氯化合成6,12-二氯二苯并[b,f][1,5]二氮杂环辛四烯(1e),化合物1a~1e的结构经~1H NMR,~(13)C NMR和ESI-MS表征,其中化合物1c为新化合物。利用超临界色谱(SFC)技术对化合物1a~1e实现了手性拆分,获得5对具有高旋光度的光学活性异构体(ee99%)。     

Benzocyclobutadienyl anion: formation and energetics of an antiaromatic molecule

Benzocyclobutadienyl diazirine (2) was synthesized and reacted with hydroxide ion in a Fourier transform mass spectrometer to afford the conjugate base of benzocyclobutadiene (1a). Authentication of the ion structure was carried out by a derivatization experiment (i.e., la was converted to benzocyclobutenone enolate, which has previously been studied), and its reactivity was explored. Thermochemical data for benzocyclobutadiene (1) were obtained (deltaH(o)acid (1) = 386 +/- 3 kcal mol(-1), EA(1r) = 1.8 +/- 0.1 eV, and C-H BDE (1) = 114 +/- 4 kcal mol(-1)), compared to MP2 and B3LYP calculations, and contrasted to a series of model compounds. Cyclobutadienyl radical appears to be quite different from benzocyclobutadienyl radical (1r) and worth further exploration.     

新型光控保护基(2-重氮基-3-苯基-4H-磺酰基苯并噻喃)在氨基酸保护-光控脱保护反应中的应用

将新型磷酸基光控保护基(2-重氮基-3-苯基-4H-磺酰基苯并噻喃,PPG)应用于氨基酸［甘氨酸(1a), L 丙氨酸(1b), L 苯丙氨酸(1c)和L-脯氨酸(1d)］的保护 光控脱保护反应。产物结构经¹H NMR, ¹³C NMR和HR-FAB MS表征。研究了物料比r［n(PPG) : n(1a)］,催化剂和反应温度对2a收率的影响。结果表明：在最佳反应条件［1a 0.1 mmol, r=1.25,于室温反应65 h］下,2a收率80%。FL监测光控脱保护反应表明：反应在15 min内完成。     

生物素-亲和素体系测定雌酮

雌酮与牛血清白蛋白共价结合,合成雌酮的完全抗原。利用此抗原免疫小鼠,通过细胞融合技术制备了雌酮的单克隆抗体,经纯化表征知,抗体是IgG1型,相对分子量为164000,与固定抗的亲和常数为8.2×10^8L/mol。以生物素化的羊抗鼠免疫球蛋白及辣根过氧化物酶标记的链亲和素为标记体系,通过竞争抑制的方式测定游离的雌酮,结果表明：雌酮在10～10000pg/mL内呈线性关系。     

Isolation,Structural Elucidation,Antioxidant and Hypoglycemic Activity of Polysaccharides of Brassica rapa L.

The aim of this study was to investigate the effects of microwave ultrasonic-assisted extraction (MUAE) on the content, structure, and biological functions of Brassica rapa L. polysaccharide (BRP). Response surface methodology (RSM) was used to optimize the parameters of MUAE, and it obtained a polysaccharide with yield of 21.802%. Then, a neutral polysaccharide named BRP-1-1 with a molecular weight of 31.378 kDa was isolated and purified from BRP using DEAE-650 M and Sephadex G-100. The structures of the BRP-1-1 were elucidated through a combination of FT-IR, GC-MS, NMR, and methylation analysis. The results showed that BRP-1 consisted of mannose (Man) and glucose (Glu) in a molar ratio of 7.62:1. The backbone of BRP-1-1 mainly consisted of →6)-α-D-Glup-(1→4-β-D-Glup-(1→2)-α-D-Manp-(1→2)-α-D-Glup-(1→, the branch was [T-α-D-Manp-(1]_n→. BRP-1-1 intervention significantly inhibited α-glucosidase activity; an inhibition rate of 44.623% was achieved at a concentration of 0.5 mg/mL. The results of the in vitro biological activity showed that BRP-1-1 has good antioxidant and hypoglycemic activity, suggesting that BRP-1-1 could be developed as a functional medicine.     

1,1,3,3-四(全氟己基乙基)二锡氧烷二聚体在环氧化合物开环反应中的催化作用

采用氟碳-有机溶剂两相催化体系,考察了1,1,3,3-四(全氟己基乙基)二锡氧烷二聚体(1)在环氧化合物开环反应中的催化作用.结果表明,催化剂(1)在氟碳-有机溶剂两相体系中使环氧苯乙烯和甲醇的开环反应产率高达95%,¹³CNMR谱表明,开环反应的区域选择性为100%.在氟碳-有机溶剂两相催化体系中以一锅法制备了3-苯基丙酸2-甲氧基-2-苯乙醇酯,收率高,方法简便,催化剂几乎可以定量回收循环使用.     

Highly sensitive routine method for urinary 3-hydroxybenzo[a]pyrene quantitation using liquid chromatography-fluorescence detection and automated off-line solid phase extraction

Many workers and also the general population are exposed to polycyclic aromatic hydrocarbons (PAHs), and benzo[a]pyrene (BaP) was recently classified as carcinogenic for humans (group 1) by the International Agency for Research on Cancer. Biomonitoring of PAHs exposure is usually performed by urinary 1-hydroxypyrene (1-OHP) analysis. 1-OHP is a metabolite of pyrene, a non-carcinogenic PAH. In this work, we developed a very simple but highly sensitive analytical method of quantifying one urinary metabolite of BaP, 3-hydroxybenzo[a]pyrene (3-OHBaP), to evaluate carcinogenic PAHs exposure. After hydrolysis of 10 mL urine for two hours and concentration by automated off-line solid phase extraction, the sample was injected in a column-switching high-performance liquid chromatography fluorescence detection system. The limit of quantification was 0.2 pmol L(-1) (0.05 ng L(-1)) and the limit of detection was estimated at 0.07 pmol L(-1) (0.02 ng L(-1)). Linearity was established for 3-OHBaP concentrations ranging from 0.4 to 74.5 pmol L(-1) (0.1 to 20 ng L(-1)). Relative within-day standard deviation was less than 3% and relative between-day standard deviation was less than 4%. In non-occupationally exposed subjects, median concentrations for smokers compared with non-smokers were 3.5 times higher for 1-OHP (p<0.001) and 2 times higher for 3-OHBaP (p<0.05). The two urinary biomarkers were correlated in smokers (ρ=0.636; p<0.05; n=10) but not in non-smokers (ρ=0.09; p>0.05; n=21).     

5-羟基-3,4-二氢异喹啉-1-酮的合成

以异喹啉为起始原料,经成盐、磺化得异喹啉-5-磺酸(3),3与熔融的氢氧化钠和氢氧化钾反应得1,5-二羟基异喹啉(4),以5%Pd-C为催化剂还原4得5-羟基-3,4-二氢异喹啉-1-酮(1),总收率47.6%。1的结构经1H NMR和MS确证。     

Solid-state fermentation of agricultural wastes into food through pleurotus cultivation

The technical feasibility of using agricultural wastes (mango and date industry wastes) as a substrate for the cultivation ofPleurotus ostreatus NRRL-0366 is evaluated. When comparing the biological efficiency of mushroom production, the highest yield of fruiting bodies was obtained using a mixture of date waste and rice straw at a ratio (1:1) (11.96%), followed by a mixture 3:1 (11.16%). The lowest one was the mixture 2:1 (9.19%). FungusPleurotus ostreatus NRRL-0366 can also be cultivated on mango waste supplemented with rice straw at a different ratio. The best one was the 1:1 mixture (10.18%), whereas the lowest was a mixture 3:1 (6.4%). Comparing the results obtained favored the use of date waste as a substrate for growingPleurotus ostreatus NRRL-0366. Spawn was cultured on three different substrates as follows: Date waste alone (I); 1:1 (by wt) date waste and rice straw (II); 1:1:1 date waste, rice straw, and corncobs (III). Final dry weight and composition of the fruiting bodies are tabulated for the three sets of conditions. Date waste and rice straw mixture (II) is a good source of nonstarchy carbohydrate (67%) and protein (27.44%) containing amounts of essential amino acids, especially lysine and low RNA (3.81%). Elemental analysis were studied in the fruit bodies of the three media.     

Development of a Quantitative Method for Detection of Multiclass Veterinary Drugs in Feed Using Modified QuPPe Extraction and LC–MS/MS

A method was developed for the rapid and quantitative analysis of 30 veterinary drugs belonging to 17 classes (amphenicols (1), anthelmintics (1), cephalosporins (4), coccidiostats (1), lincosamides (1), macrolide (1), nitroimidazole (1), penicillins (3), phenylhydrazines (1), polypeptides (1), pyrethrins (1), quinolones (5), sulfonamides (3), tetracycline (3), neuroleptic agents (1), triazene trypanocidal agents (1), other. (1)) in feeds. The proposed method with a modified Quick Polar Pesticides (QuPPe) sample preparation was validated for the determination of 30 veterinary drugs in feed samples by liquid chromatography triple-quadrupole mass spectrometry (LC–MS/MS). The sample was extracted with methanol containing 1% acetic acid and purified by dispersive solid-phase extraction (d-SPE) with C18. Good linearity (r² ≥ 0.98) was observed, and the LOQ values ranged from 10 to 200 µg/kg. Average recoveries ranged from 70.8 to 118.4%, and the relative standard deviation was ≤ 18.7%. This validated method was used in the determination of 30 veterinary drugs in 142 feed samples obtained from South Korea. The results show that lincomycin was present in only one of the tested feed samples, although it was detected at a value lower than the LOQ. In conclusion, this multi-residue method can be used for screening through the detection and quantitation of residual multiclass veterinary drugs in feed samples.     

Quantitative analysis of 5 alpha-reductase inhibitors in DU145 cells using matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry and high-performance liquid chromatography/tandem mass spectrometry

N-(Dicyclohexyl)acetylpiperidine-4-benzylidene-4-carboxylic acid (1) is an excellent in vitro inhibitor of 5 alpha-reductase (5 alpha R). Compound 1 showed, however, much lower inhibition activity of 5 alpha R in vivo than in vitro, which might be caused by poor membrane permeability. The methyl ester of 1 (1a) was therefore tested as a model prodrug to see if it has better permeability properties than the corresponding acid 1. It was also monitored that this methyl ester was cleaved into the active compound 1 within the DU145 cells. Quantitative matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) and high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) methods were established with reliable linearity factors (0.996 for MALDI-TOFMS and 0.998 for HPLC/MS/MS) and reproducibility (relative standard deviation = 6.5% for MALDI-TOFMS and 2.8% for HPLC/MS/MS). The samples for MS analysis were effectively prepared from the cell homogenates using solid-phase extraction, with a high recovery of 90% on average. The intracellular amount of 1a (1.7 nmol) was much higher than that of 1 (0.032 nmol) in DU145 cells after 6 h of incubation. After incubation with the ester (1a), the cleaved acid (1) was detected within the cells. The concentration of acid 1 (0.045 nmol) in this experiment was higher than the acid content (0.032 nmol) after direct incubation with 1. Surprisingly, high amounts of the cleaved compound 1 were found outside the cells after 6 h of incubation with 1a.