Enantiomeric separation of basic compounds using heptakis(2,3-di-O-methyl-6-O-sulfo)-beta-cyclodextrin in combination with potassium camphorsulfonate in nonaqueous capillary electrophoresis: optimization by means of an experimental design

The enantiomeric separation of a series of basic pharmaceuticals (beta-blockers, local anesthetics, sympathomimetics) has been investigated in nonaqueous capillary electrophoresis (NACE) systems using heptakis(2,3-di-O-methyl-6-O-sulfo)-beta-cyclodextrin (HDMS-beta-CD) in combination with potassium camphorsulfonate (camphorSO3-). For this purpose, a face-centered central composite design with 11 experimental points was applied. The effect of the concentrations of HDMS-beta-CD and camphorSO3- on enantioresolution was statistically evaluated and depended largely on the considered analyte. The presence of camphorSO3- was found to be particularly useful for the enantioseparation of compounds with high affinity for the anionic CD. CamphorSO3- seems to act as a competitor, reducing the affinity for the CD, probably by ion-pair formation with these analytes. For compounds with lower affinity for HDMS-beta-CD, the combination of camphorSO3- and the CD appeared to have a favorable effect on enantioresolution only if the optimal CD concentration could be reached. On the other hand, for compounds characterized by a very low affinity for the anionic CD, the association of camphorSO3- and HDMS-beta-CD is always unfavorable. Finally, experimental conditions were selected by means of the multivariate approach in order to obtain the highest resolution (Rs) value for each studied compound.     

Influence of the nature of the electrolyte on the chiral separation of basic compounds in nonaqueous capillary electrophoresis using heptakis(2,3-di-O-methyl-6-O-sulfo)-beta-cyclodextrin

The influence on the enantiomeric resolution of the nature of the cationic BGE component (sodium, ammonium or potassium) and that of the anionic component (chloride, formate, methanesulfonate or camphorsulfonate) as well as the concentration of heptakis(2,3-di-O-methyl-6-O-sulfo)-beta-cyclodextrin (HDMS-beta-CD), the selected chiral selector, was studied in nonaqueous capillary electrophoresis (NACE). For this purpose, two D-optimal designs with 33 and 26 experimental points were applied. Three beta-blockers (atenolol, celiprolol and propranolol) and three local anesthetics (bupivacaine, mepivacaine and prilocaine) were selected as basic model compounds. Both cationic and anionic BGE components were found to have a deep impact on the enantiomeric resolution of the investigated analytes but it is the cationic component that has shown the strongest influence. Indeed, in some cases, the change of the latter led to a complete loss of enantioresolution. Based on the observed results, two NACE systems were recommended, namely ammonium formate and potassium camphorsulfonate in a methanolic solution containing HDMS-beta-CD and acidified with formic acid, in order to separate efficiently the enantiomers of basic drugs.     

Nonaqueous capillary electrophoresis method for the enantiomeric purity determination of S-timolol using heptakis(2,3-di-O-methyl-6-O-sulfo)-beta-cyclodextrin: validation using the accuracy profile strategy and estimation of uncertainty

Nonaqueous capillary electrophoresis (NACE) was successfully applied to the enantiomeric purity determination of S-timolol maleate using heptakis(2,3-di-O-methyl-6-O-sulfo)-beta-cyclodextrin (HDMS-beta-CD) as chiral selector. With a background electrolyte made up of a methanolic solution of 0.75 M formic acid, 30 mM potassium camphorsulfonate and containing 30 mM HDMS-beta-CD, the determination of 0.1% of R-timolol in S-timolol could be performed with an enantiomeric resolution of 8.5. Pyridoxine was selected as internal standard. The NACE method was then fully validated by applying a novel strategy using accuracy profiles. It is based on beta-expectation tolerance intervals for the total measurement error which includes trueness and intermediate precision. The uncertainty of measurements derived from beta-expectation tolerance intervals was estimated at each concentration level of the validation standards. To confirm the suitability of the developed and validated method, several real samples of S-timolol maleate containing R-timolol maleate at different concentrations were analysed and the results were compared to those obtained by liquid chromatography.     

Chiral separation of anticholinergic drug enantiomers in nonaqueous capillary electrophoresis

Nonaqueous capillary electrophoretic (NACE) method for the separation of nine structurally similar chiral anticholinergic drugs was developed. The eight drug enantiomers were separated on baseline within 18 min using 20mM phosphoric acid and 10 mM NaOH, containing 10 mM heptakis(2,3-dimethyl-6-sulfato)-4beta-cyclodextrin (HDMS-beta-CD) in methanol. The results were compared with those obtained in the high performance liquid chromatography system.     

Synergistic effects of ion-pairing in the enantiomeric separation of basic compounds with cyclodextrin derivatives in nonaqueous capillary electrophoresis

The enantiomeric separation of various kinds of basic pharmaceuticals has been investigated in nonaqueous capillary electrophoresis (NACE) systems using an ion-pairing reagent in combination with cyclodextrins (CDs). The simultaneous addition to the methanolic background electrolyte (BGE) of (+)-S-camphorsulfonate or alkanesulfonates and an anionic beta-cyclodextrin derivative, heptakis(2,3-dimethyl-6-sulfato)-beta-cyclodextrin (HDMS-beta-CD), led to partial or complete enantioresolution in most cases. In the absence of ion-pairing reagent, the enantiomeric resolution obtained with this CD derivative was most often completely lost or strongly reduced, indicating the important role of ion-pairing in the chiral recognition mechanism in these NACE systems. The influence of the nature and concentration of the counterion and the anionic CD derivative on the enantioseparation of basic compounds was studied. Synergistic effects between these two kinds of charged additives were clearly observed.     

Enantiomeric separation of omeprazole and its metabolite 5-hydroxyomeprazole using non-aqueous capillary electrophoresis

This study demonstrates the development and validation of a non-aqueous capillary electrophoresis (NACE) method for enantiomeric determination of omeprazole and its metabolite 5-hydroxyomeprazole. Heptakis-(2,3-di-O-methyl-6-O-sulfo)-beta-cyclodextrin (HDMS-beta-CD) was chosen as the chiral selector in an ammonium acetate buffer acidified with formic acid in methanol. Parameters such as CD concentration, concentration of buffer electrolyte, voltage and temperature were studied in order to optimize both the enantioresolution and migration times. An experimental design was utilized for method optimization, using software Modde 5.0. Validation of the developed method showed good linearity, which was tested over a concentration range of 2.5-500 microM. The regression coefficients for S-omeprazole, S-5-hydroxyomeprazole, R-omeprazole and R-5-hydroxyomeprazole were between 0.996 and 0.997. The limits of detection for the four enantiomers were in the range from 45 to 51microM and the limits of quantification were between 149 and 170 microM with UV detection at 301nm. Using a reduced temperature of 16 degrees C gave improved resolution values, reproducibility and also decreased the occurrence of current loss within the capillary. RSD values for peak migration time were calculated to be between 0.41 and 1.48% using an inter-day study.     

Establishment of the Model CD40 Cell Membrane Chromatography and Its Chromatographic Characteristics

A new model of atherosclerosis cell membrane chromatography has been established by using a CD40 cell membrane stationary phase (CD40 CMSP) prepared by immobilizing the CD40 cell membrane onto the surface of a silica carrier. The surface and chromatographic characteristics of CD40 CMSP were studied. The retention characteristics of anti-CD40 antibody and statins (lovastatin, simvastatin and pravastatin) were also investigated using this model. Affinities of the anti-CD40 antibody and statins toward CD40 cell membrane and receptors were based on the determination of log k′ values (the logarithm of capacity factor of a solute). There was a significant correlation between the affinity in the CD40–CMC and the effect in vitro for the pharmacological effect.     

Activation of B cells using Schneider 2 cells expressing CD40 ligand for the enhancement of antigen presentation in vitro

CD40 ligand (CD40L) expressed by activated CD4+ T cells is a family member of membrane bound TNF family ligand and its interaction with CD40 expressed in APC has been shown to contribute in enhancing immune response. Exogenous stimulation through CD40 has been performed using soluble trimeric CD40L, anti-CD40 monoclonal antibody and cells expressing CD40L. Schneider 2 (S2) cells, a cell line derived from Drosophila melanogaster, was transfected with a plasmid vector, pAc5.1/V5-HisA, for the constitutive expression of CD40L (S2-CD40L). Upon incubation of S2-CD40L with B-lymphocytes for 6 days, activated B cells were examined by counting B cell numbers and for activation markers including CD86 and HLA Class II molecules. The activated B cells were tested for its efficient APC function by mixed lymphocyte reactions (MLR) and enzyme-linked Immunospot (ELISPOT) assay. S2-CD40L was cultured for a year and maintained CD40L expression (>90%). S2-CD40L induced B cell activation as demonstrated by increment of total B cells and up-regulation of CD86 and MHC Class II molecules. Activated B cells pulsed with peptide from human cytomegalovirus pp65 antigen efficiently induced both proliferation and IFN-gamma secretion of T cells. Our result suggests that S2-CD40L can efficiently and conveniently generate B cells as a functional APC and represents a potential role for B-cell mediated cancer immunotherapy.     

C40,Nb@C40^＋,La@C40^＋的从头算研究

用量子化学从头算方法研究了Ｃ４０、Ｎｂ＠Ｃ４０＾＋，Ｌａ＾＠Ｃ４０＾＋的几何构型和电子结构，Ｃ４０最稳定构型具有Ｄ２对称性。Ｌａ和Ｎ原子内含于Ｃ４０笼中，形成金属夹心碳笼Ｎｂ＠Ｃ４０＾＋、Ｌａ＠Ｃ４０＾＋。Ｃ４０结合能大于Ｍ＠Ｃ４０＾＋（Ｍ＝Ｎｂ，Ｌａ）。     

The dynamic nature of amyloid beta (1-40) aggregation

In this paper, we characterize the dynamic nature of the full amyloid beta (1-40) (Aβ (1-40)) aggregates. We labeled the peptide with either 5-carboxytetramethylrhodamine (TAMRA) or with fluorescein-isothiocyanate (FITC). The labeled peptides were mixed after separate fibrillization, and the dynamic changes in the structure of the fibrils were imaged using confocal microscopy. Fluorescence resonance energy transfer (FRET) measurements showed that the Aβ (1-40) peptides detach from and reattach to the fibrils in a biologically relevant timescale (days). With time, the two peptides mix at the molecular level. This process is concentration dependent and occurs primarily in the external parts of the aggregates with a half time between 4 and 7 days. This study shows that the combination of confocal microscopy and FRET analysis is a facile method for studying dynamic processes in supra-molecular aggregates.     

The amphiphilic multiarm copolymers based on hyperbranched polyester and lysine: Synthesis and self-assembly

The amphiphilic multiarm copolymers were synthesized through the modification of commercially available hyperbranched polyesters(Boltorn H40) with N-ε-carbobenzoxy-L-Lysine N-carboxyanhydride(ZLys-NCA).After being condensed with N-Boc-phenylalanine(Boc-~NPhe) and deprotected the Boc-groups in trifluoroacetic acid(TFA),the original terminal hydroxyl groups were transformed into the amino groups and then initiated the ring-opening polymerization of ZLys-NCA.The hydrophilic poly(L-lysine) was grafted to the surface of Boltorn H40 successfully after the protecting benzyl groups were removed by the HBr solution in glacial acetic acid(33 wt%).The resulting multiarm copolymers were characterized by the ~1H-NMR,GPC and FTIR.The arm length calculated by NMR and GPC analysis was about 3 and 13 lysine-units for H40-Phe-PLysl and H40-Phe-PLys2 respectively.Due to the amphiphilic molecular structure,they displayed ability to self-assemble into spherical micelles in aqueous solution with the average diameter in the range from 70 nm to 250 nm.The CMC of H40-Phe-PLysl and H40-Phe-PLys2 was 0.013 mg/mL and 0.028 mg/mL,respectively, indicating that H40-Phe-PLysl with shorter arm length is easier to self-assemble than H40-Phe-PLys2 with longer arm length.     

Efficient syntheses of the keto-carotenoids canthaxanthin, astaxanthin, and astacene

Three keto-carotenoids were prepared by the oxidation of the stable C(40) trisulfone 6, which has been used as the key compound in our beta-carotene synthesis. The first allylic oxidation to the unsaturated ketone and the second oxidation to the alpha-hydroxyketone produced the C(40) trisulfones 7 and 10, respectively. The Ramberg-Backlund reaction of the oxidized C(40) trisulfone was efficiently effected by the use of a mild base, NaOMe, in the presence of CCl(4) as a halogenating agent to give the C(40) disulfones 8 and 11. Base-promoted dehydrosulfonation reaction of the disulfone compounds produced the fully conjugated polyenes of canthaxanthin (1), astaxanthin (2), and astacene (3).     

Preparation of silica-on-titania patterns with a wettability contrast

The preparation of patterned inorganic surfaces consisting of silica (SiO2) and titania (TiO2) is described. The approach is based on a combination of standard photolithography and plasma-enhanced chemical vapor deposition. Silicon wafers coated with a titania layer (40 nm) were patterned by use of a positive photoresist and then a thin silica layer (10-40 nm) was plasma-deposited. The photoresist was removed by decomposition at 800 degrees C. The inorganic patterned surfaces possessed excellent high-temperature resistance. Since the silica patches were effectively dehydroxylated during the thermal treatment, the patterns consisted of moderately hydrophobic (silica) and hydrophilic (titania) domains with a significant wettability contrast (40 degrees for water). The surface was further hydrophobized with a self-assembled monolayer of fluoroalkylsilane (FAS) and exposed to UV light. The FAS layer was locally oxidized on the TiO2 patches and the wettability contrast was maximized to 120 degrees (the highest possible value on smooth surfaces).     

In situ synthesis of oligonucleotide arrays by using surface tension

This work describes the in situ synthesis of oligonucleotide arrays on glass surfaces. These arrays are composed of features defined and separated by differential surface tension (surface tension arrays). Specifically, photolithographic methods were used to create a series of spatially addressable, circular features containing an amino-terminated organosilane coupled to the glass through a siloxane linkage. Each feature is bounded by a perfluorosilanated surface. The differences in surface energies between the features and surrounding zones allow for chemical reactions to be readily localized within a defined site. The aminosilanation process was analyzed using contact angle, X-ray photoelectron spectroscopy (XPS), and time-of-flight/secondary ion mass spectroscopy (TOF-SIMS). The efficiency of phosphoramidite-based oligonucleotide synthesis on these surface tension arrays was measured by two methods. One method, termed step-yields-by-hybridization, indicates an average synthesis efficiency for all four (A,G,C,T) bases of 99.9 +/- 1.1%. Step yields measured for the individual amidite bases showed efficiencies of 98.8% (dT), 98.0% (dA), 97.0% (dC), and 97.6% (dG). The second method for determining the amidite coupling efficiencies was by capillary electrophoresis (CE) analysis. Homopolymers of dT (40- and 60mer), dA (40mer), and dC (40mer) were synthesized on an NH(4)OH labile linkage. After cleavage, the products were analyzed by CE. Synthesis efficiencies were calculated by comparison of the full-length product peak with the failure peaks. The calculated coupling efficiencies were 98.8% (dT), 96.8% (dA), and 96.7% (dC).     

Increased Availability of Tryptophan in 5-Methyltryptophan-Tolerant Shoots of Catharanthus roseus and Their Postharvest in vivo Elicitation Induces Enhanced Vindoline Production

Ten 5-methyltryprophan (5-MT)-resistant multiple shoot culture lines in three genotypes of Catharanthus roseus were selected in vitro. The variant shoot lines displayed a differential threshold tolerance limit against the analogue stress, ranged from 20 to 70?mg/l 5-MT in the medium. The lines tolerant to 40?mg/l 5-MT stress were most stable and fast proliferating. All the selected lines in the presence of 5-MT stress recorded increased level of tryptophan in their free amino acid pool. Highest tryptophan accumulation occurred in lines P40, P30, D40, and N40 (i.e., 296.5, 241.0, 200.6, and 202.0???g/g dry wt., respectively). A concomitant increase in the total alkaloid content (2.3?C3.8?% dry wt.) under the analogue stress was also noticed in these lines when compared to 1.0?C1.58?% dry wt. in the respective wild-type shoot maintained on a stress-free medium. The HPLC analysis of the alkaloid extracts of the 5-MT-tolerant lines grown under analogue stress also revealed vindoline as a major constituent with maximum accumulation in lines N40, N30, D30, D40, and P40 (0.046, 0.032, 0.034, and 0.022?% dry wt., respectively). The rooted shoots of 5-MT-tolerant lines were successfully acclimatized under glasshouse environment wherein they grew normally and set seeds. Flowering twigs or leaves excised from 1-year-old glasshouse-grown plants of 5-MT variant lines upon postharvest in vivo elicitation with 30?mg/l 5-MT or 5.0?mg/l tryptophan registered an eight-to-tenfold increment in their vindoline content within 24?C48?h.     

含铁杂多酸盐Mossbauer谱研究 IV: 钼磷铁杂多酸盐的合成和Mossbauer谱

合成了钼磷铁杂多黄(NH4)4[PFeMo11O40H2]·9H2O和杂多蓝(NH4)4[PFeMo11O40H3]·11H2O的铵盐晶体, 用^5^7Fe同位素增丰Mossbauer谱法, 结合磁化率测定, 研究其结构与性质, 并讨论了四极分裂值Q2和Keggin结构杂多阴离子中MO6(M=Mo, Fe)八面体畸变大小的关系, 进一步证实Keggin结构杂多蓝阴离子中Mo(V)-O-Fe(III)间存在反磁交换作用。     

Modification of polysulfone membranes with polyethylene glycol and lignosulfate: electrical characterization by impedance spectroscopy measurements

Two sets of composite membranes having an asymmetric sulfonated polysulfone membrane as support layer have been obtained and electrically characterized (membranes SPS-PEG and PA-LIGS). The skin layer of the membrane SPS-PEG contains different percentages of polyethylene glycol in the casting solution (5, 25, 40, and 60 wt%), while lignosulfonate was used for manufacturing PA-LIGS membranes (5, 10, 20, and 40 wt%). Membrane electrical characterization was done by means of impedance spectroscopy (IS) measurements, which were carried out with the membranes in contact with NaCl solutions at different concentrations (10(-3) < or = c(M) < or = 5x10(-2)). Electrical resistance and equivalent capacitance of the different membrane samples were determined from IS plots by using equivalent circuits as models. Results show a clear decrease in the membrane electrical resistance as a result of both polysulfone sulfonation and the increase of the concentration of modifying substances, although a kind of limit concentration was obtained for both polyethylene glycol and lignosulfonate (40 and 20%, respectively). Results also show a decrease of around 90% in electrical resistance due to polysulfone sulfonation, while the value of the dielectric constant (hydrated state) clearly increases.     

Experimental approach to assess sorptive loss properties of volatile organic compounds in the sampling bag system

In this study, the sorptive loss patterns for volatile organic compounds were evaluated by gaseous standards containing 13 compounds (benzene, toluene, styrene, p‐xylene, methyl ethyl ketone, methyl isobutyl ketone, isobutyl alcohol, butyl acetate, acetaldehyde, propionaldehyde, butyraldehyde, isovaleraldehyde, and valeraldehyde). The gaseous standards, prepared initially at two contrasting concentration levels (40 and 4000 ppb) in a polyester aluminum bag, were measured after two consecutive transfers into empty bags. It indicates that the percent loss patterns, if assessed for all 13 target compounds, are affected most sensitively by the initial concentration levels of samples to yield 2.62 ± 2.22% (at 40 ppb) and 9.57 ± 6.74% (at 4000 ppb). Moreover, the sorptive loss patterns at high concentration samples (4000 ppb) tend to increase in relation with increasing molecular weight of target compounds, although such pattern disappears in low concentration samples (40 ppb). The observed loss patterns, if evaluated in relation to some key parameters like concentration or compound type, suggest the possibility that the sorptive loss of target compounds in storage media can occur in a predictable manner.     

Mild benzene-thermal route to GaP nanorods and nanospheres

GaP nanorods and nanospheres were synthesized from a mild benzene-thermal route at 240 and 300 degrees C, respectively, using Na, P, and GaCl(3) as the starting materials. The structure of the products was identified as zinc blende phase by X-ray powder diffraction (XRD). Transmission electron microscopy (TEM) images showed that, when the reaction temperature was 240 degrees C, the products were nanorods with widths of 20-40 nm and lengths of 200-500 nm and nanospheres with diameters of 20-40 nm. However, when the reaction temperature was increased to 300 degrees C, the products were only nanospheres, and the diameters increased to 40-60 nm. The reaction proceeded through a metallic gallium intermediate, and a solution-liquid-solid (SLS) mechanism was proposed for the one-dimensional growth. The products were also investigated by UV-vis absorption and X-ray photoelectron spectroscopy.     

Inhibitory effect of H_(3 + x)PMo_(12－x)V_xO_(40)-Ton the self-polymerization of methyl methacrylate

In this work,a series of molybdovanadophosphoric heteropoly acid quaternary ammonium salts H_(3+x)PMo_(12 -x)V_xO40-T were synthesized and employed as a reaction inhibitor in the selfpolymerization of methyl methacrylate(MMA).The polymerization inhibition effect of H_(3+x)PMoPMo_(12 -x)V_xO40-T)with different number of vanadium atoms and reaction dosages was investigated using differential scanning calorimetry(DSC).It shows that the inhibitory effect was improved with the increasing dosages of H_(3+x)PMoPMo_(12 -x)V_xO40-T),and the polymerization inhibition was also affected by the number of vanadium atoms in the H_(3+x)PMo_(12 -x)V_xO40-T .Furthermore,cyclic voltammograms(CV)was used to probe the mechanism of the inhibition reaction with H3+xPMo12xVxO40-T.The result of CV indicates that the inhibition reaction is an oxidation–reduction reaction.H_(3+x)PMo_(12 -x)V_xO40-T can react directly with the MMA monomer radicals,which eliminated the MMA monomers,and therefore the self-polymerization of the MMA can be effectively inhibited by H_(3+x)PMo_(12 -x)V_xO40-T.