Enantioselective fluorescence sensing of amino acids by modified cyclodextrins: role of the cavity and sensing mechanism

Two selectors based on modified cyclodextrins containing a metal binding site and a dansyl fluorophore-6-deoxy-6-N-(N(alpha)-[(5-dimethylamino-1-naphthalenesulfonyl)aminoethyl]phenylalanylamino-beta-cyclodextrin-containing D-Phe (3) and L-Phe (4) moieties were synthesized. The conformations of the two selectors were studied by circular dichroism, two-dimensional NMR spectroscopy and time-resolved fluorescence spectroscopy. Cyclodextrin 4 was found to have a predominant conformation in which the dansyl group is self-included in the cyclodextrin cavity, while 3 showed a larger proportion of the conformation with the dansyl group outside the cavity. As a consequence, the two cyclodextrins were found to bind copper(II) with different affinities, as revealed by fluorescence quenching in competitive binding measurements. Addition of D- or L-amino acids induced increases in fluorescence intensity, which were dependent on the amino acid used and in some cases on its absolute configuration. The cyclodextrin 4 was found to be more enantioselective than 3, suggesting that the self-inclusion in the cyclodextrin cavity strongly increases the chiral discrimination ability of the copper(II) complex. Accordingly, a linear fluorescent ligand N(alpha)-[(5-dimethylamino-1-naphthalenesulfonyl)aminoethyl]-N(1)-propyl-phenylalaninamide, which has the same binding site and absolute configuration as 4, showed very low chiral discrimination ability. The enantioselectivity in fluorescence response was found to be due to the formation of diastereomeric ternary complexes, which were detected by ESI-MS and by circular dichroism. Time-resolved fluorescence studies showed that the fluorescence of the dansyl group was completely quenched in the ternary complexes formed, and that the residual fluorescence was due to uncomplexed ligand.     

聚胸腺嘧啶单链DNA-铜纳米簇新型荧光探针检测水中硫离子

本文建立了一种快速灵敏检测水中硫离子的新方法。该方法利用聚胸腺嘧啶单链DNA保护的铜纳米簇为荧光探针。以聚胸腺嘧啶单链DNA为模板制备了具有荧光性质的铜纳米簇,当加入S2-后,铜纳米簇荧光显著猝灭。铜纳米簇荧光猝灭量与S2-浓度在0.125~8μmol/L范围内有良好的线性,检测限为22nmol/L。该方法对S2-有较好的选择性,实际样品检测结果显示回收率良好,说明该方法可以用于实际水样中S2-的检测。由于聚胸腺嘧啶单链DNA为模板制备的铜纳米簇制备过程简单快速,可在5min内完成,使得检测时间大大缩短。     

电气材料铜及铜合金中痕量杂质砷元素的氢化物发生-原子荧光光谱法测定

本文建立了氢化物发生-原子荧光光谱法测定电气材料铜及铜合金中痕量杂质元素砷的方法。通过使用L-半胱氨酸与铜基体形成络合物,抗坏血酸充分预还原砷,消除了基体干扰,并实现了铜及铜合金中痕量杂质砷的准确测定。在经过优化的实验条件下,砷的检出限为0.12 ng·m L-1,线性相关系数优于0.999,相对标准偏差小于4%。用加标回收法测定实际样品,回收率在92%到103%之间。该法操作简便,灵敏度高,无需基体预分离。     

Photoluminescent energy transfer from poly(phenyleneethynylene)s to near‐infrared emitting fluorophores

Photoluminescent energy transfer was investigated in conjugated polymer‐fluorophore blended thin films. A pentiptycene‐containing poly(phenyleneethynylene) was used as the energy donor, and 13 fluorophores were used as energy acceptors. The efficiency of energy transfer was measured by monitoring both the quenching of the polymer emission and the enhancement of the fluorophore emission. Near‐infrared emitting squaraines and terrylenes were identified as excellent energy acceptors. These results, where a new fluorescent signal occurs in the near‐infrared region on a completely dark background, offer substantial possibilities for designing highly sensitive turn‐on sensors. © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 48: 3382–3391, 2010     

氧弹分解-原子荧光法测定煤中汞的研究进展

介绍氧弹分解-原子荧光法测定煤中汞含量的研究进展。影响该方法主要因素为称样量、氧气浓度、吸收剂、还原剂、煤种以及灰分含量。分别综述了各个因素对该方法的影响,可为相关标准的制定提供参考。该方法能够简单、经济、快速地测定煤中汞的含量,但是目前学者们对于该方法的相关研究还存在分歧,需要进一步的研究验证;同时,还需要注意以下几个研究方向:吸收剂和还原剂的选择对汞回收率的影响;吸收剂在密闭氧弹中对汞的回收率及吸收时间的研究;氧弹废气中汞含量的测定;煤种和灰分含量对汞含量测定的影响。     

Interface characterization in latex blend films by fluorescence energy transfer

Immiscible polymer blend films were formed by air drying aqueous dispersions containing mixtures of a high-T_g latex, poly(methyl methacrylate), and a film-forming low-T_g latex, poly(butyl methacrylate-co-butyl acrylate). Fluorescence energy transfer experiments were used to characterize the interfaces in these films, in which one component was labeled with a donor dye and the other with an acceptor. The quantum efficiency of energy transfer (Φ_ET) between the donors and acceptors is influenced by the interfacial contact area between the two polymer phases. As the amount of soft component in the blend is increased, Φ_ET approaches an asymptotic value, consistent with complete coverage of the hard polymer surface with soft polymer. This limiting extent of energy transfer is very sensitive to the total surface area in the film, with correspondingly more energy transfer at constant volume fraction for small hard particles. Some of the details of the energy transfer are revealed through a fluorescence lifetime distribution analysis. The presence of ionic surfactant (sodium dodecyl sulfate) in the dispersion from which the latex blend film is prepared reduces the cross-boundary energy transfer by 30%, which implies that in these films the surfactant decreases the interfacial contact. After annealing the surfactant-free blends above 100°C, we observe an increase in energy transfer, consistent with a broader interface between the two polymers. © 1998 John Wiley & Sons, Inc. J Polym Sci B: Polym Phys 36: 1115–1128, 1998     

荧光共振能量转移测定牛血清白蛋白的研究

随着生物分析技术进入了后基因组时代,生命科学领域里的研究课题不断深入,DNA、RNA、蛋白质和其他生物大分子的检测技术发展十分迅速,生命科学中单分子分析技术不断揭示出生命活动的客观规律.相关的新的分析方法和仪器不断取得进展,成为生命科学的前沿领域.     

TNT及RDX蒸气对基于芘氧敏感膜荧光猝灭的研究

制备了以芘为敏感材料,聚苯乙烯(PS)以及聚氯乙烯(PVC)粉末为支持体系的两种荧光猝灭氧敏感膜.PS以及PVC氧敏感膜的最大激发波长分别为349 nm和355 nm,最大发射波长分别为399 nm和398 nm.分子氧对PS以及PVC氧敏感膜的荧光均有猝灭作用.同时,实验发现2,4,6-三硝基甲苯(TNT)和环三亚甲基三硝基胺(RDX)对这两种氧敏感膜的荧光也有一定的猝灭.     

Excitation energy transfer in poly(p-phenylphenylenevinylene) determined by polarized fluorescence spectroscopy

Polarized fluorescence spectroscopy is used to investigate the photophysical behavior of poly(p-phenylphenylenevinylene) (PPPV) in polystyrene matrix in comparison with oligomeric model compounds. For this purpose PPPV is modeled by a chain consisting of a distribution of independent oligomeric segments. Excitation energy transfer (EET) between the segments depends on the wavelength of excitation, not only for transfer along an isolated polymer chain, but also for intermolecular transfer at high concentration of PPPV. The spatial range of EET, as indicated by fluorescence depolarization, is reduced for excitation at the long wavelength edge of the absorption spectrum (“red-edge-effect”). © 1995 John Wiley & Sons, Inc.     

Homogeneous time-resolved fluoroimmunoassay of bensulfuron-methyl by using terbium fluorescence energy transfer

A sensitive homogenous time-resolved fluoroimmunoassay (TR-FIA) method for bensulfuron-methyl (BSM) based on fluorescence resonance energy transfer (FRET) from a Tb(3+) fluorescent chelate with N,N,N('),N(')-[2,6-bis(3'-aminomethyl-1'-pyrazoly)-4-phenylpyridine] tetrakis(acetic acid) (BPTA-Tb(3+)) to organic dye, Cy3 or Cy3.5 has been developed. New method combined the use of BPTA-Tb(3+) labeled streptavidin, Cy3 or Cy3.5 labeled anti-BSM monoclonal antibody and biotinylated BSM-BSA conjugate (BSA is bovine serum albumin) for competitive-type immunoassay. After BPTA-Tb(3+) labeled streptavidin was reacted with a competitive immune reaction solution containing biotinylated BSM-BSA, BSM sample and Cy3 or Cy3.5 labeled anti-BSM monoclonal antibody, the sensitized and long-lived emission of Cy3 or Cy3.5 derived from FRET was measured, and thus the concentration of BSM in sample was calculated. The present method has the advantages of rapidity, simplicity and high sensitivity since the B/F (bound reagent/free reagent) separation steps and the solid-phase carrier are not necessary. The method gives the detection limit of 2.10 ngml(-1). The coefficient variations of the method are less than 1.5% and the recoveries are in the range of 95-105% for BSM water sample measurement.     

Flexibility of the aldolase molecule measured using quenching-induced variations of the Forster distance for fluorescence energy transfer

The range of flexibility of the rabbit muscle aldolase molecule was studied using fluorescent labelled aldolase. The protein molecule was specifically labelled on the opposite sites of the enzyme subunit with the fluorescence energy donor and acceptor residues. Labelled aldolase with full enzymatic activity was used as a tool in the FRET studies between IAEDANS — donor or Cys-289 and IAF — acceptor on Cys-239. A range of Forster distance (R) was obtained by collisional quenching of the donor emission. The experiments of donor fluorescence quenching with wide range of acrylamide concentrations have shown the changes of donor-acceptor distances. In the absence of quencher the D-A distance distribution in characterized by an average value of 40.4 ?, and a half-width of 0.13 ?. A dramatic increase in half-width to 17.7? is observed after expositions of the enzyme to high acrylamide concentrations (0.13 M-0.68 M).     

Selective fluorescence recovery after bleaching of single E2GFP proteins induced by two-photon excitation.

We report the two-photon excitation and emission or a recently developed green fluorescent protein (GFP) mutant, E(2)GFP. Two main excitation bands are found at 780 and 870 nm. Blinking and irreversible and reversible bleaching were observed. Fluorescence blinking occurs in the millisecond range and has been ascribed to conversions between the neutral, anionic and dark zwitterionic states. Bleaching is observed after approximately 10 to 400 ms depending on the excitation power, and it is probably due to a conversion to a dark state. The striking feature of this GFP mutant is that the fluorescence can be recovered with very high efficiency only upon irradiation at 720 +/- 10 nm. This GFP mutant therefore seems promising as an almost permanent chromophore for two-photon excitation (TPE) microscopy or for applications in single-molecule memory arrays.     

吖啶橙-罗丹明B能量转移荧光猝灭法测定叶酸

研究了吖啶橙(AO)与罗丹明B(RB)间发生能量转移的最佳条件,在pH=6.80的Britton-Robinson(B-R)缓冲溶液,十二烷基苯磺酸钠的介质中,AO-RB间发生有效能量转移,使RB荧光大大增强,叶酸(FA)的加入使能量转移体系的RB的荧光强度降低,即发生猝灭.以此建立了利用AO-RB能量转移荧光猝灭法测...     

波长色散X射线荧光光谱法测定钴精矿中钴、铜和锰含量

采用高纯氧化物经四硼酸锂和偏硼酸锂熔融制备人工标准样品,以氧化镱为内标,制作校准曲线,建立了波长色散型X射线荧光光谱法测定钴精矿中钴、铜、锰元素的分析方法。重点研究了混合熔剂、试样稀释比、氧化剂和内标选择、方法检出限、方法准确度和精密度等,结果表明,各元素校准曲线线性范围宽,相关系数均大于0.999, 钴、铜、锰元素检出限分别达到0.002%、0.001%和0.001%,测试准确度和精密度可靠,方法满足行业检测需求。     

吖啶红-罗丹明B荧光共振能量转移法测定汞

研究了吖啶红(供体)和罗丹明B(受体)之间荧光共振能量转移的最佳条件,建立了荧光共振能量转移猝灭法测定污水和废旧电池中痕量汞的方法。室温中,采用十六烷基三甲基溴化胺(CTMAB),在pH=7.0条件下,吖啶红与罗丹明B之间能产生有效的共振能量转移,汞离子的加入能使体系中罗丹明B荧光峰强猝灭从而测定汞的含量。汞离子浓度在0.05～2.5μg/mL范围内与罗丹明B荧光强度变化ΔF呈现良好线性关系(r=0.9997),检出限(3σ/K)为0.95ng/mL,加标回收率98.0%～104.5%。该方法可用于污水和废旧电池中痕量汞的测定。     

Oxidation of aminothiols by molecular oxygen catalyzed by copper ions. Stoichiometry of the reaction

Catalysis of oxidation of aminothiols by copper ions was studied depending on the structure of aminothiols and pH of the medium. The catalytic reaction proceeds in the inner coordination sphere of Cu⁺. At pH 7—9, oxidation of bidentate aminothiols involves reduction of O₂ to H₂O₂. At pH 9—13, oxidation of chelating aminothiols is accompanied by reduction of O₂ to H₂O, whereas oxidation of weak-chelating aminothiols still proceeds by the former mechanism. In this process, the thiolate anions coordinated to the Cu⁺ ions lose one electron each and are oxidized to amino disulfides, which go from the inner sphere of the Cu⁺ complex into a solution. Procedures developed for the determination of amino disulfides, the chemiluminescence determination of H₂O₂ in the presence of aminothiols as luminescence quenchers, and a modified polarographic procedure for the determination of O₂ allowed us to establish that oxidation of aminothiols is not accompanied by catalytic decomposition of H₂O₂ that formed.     

Study on molecular interactions between proteins on live cell membranes using quantum dot-based fluorescence resonance energy transfer

Mouse anti-human CD71 monoclonal antibody (anti-CD71) was conjugated with red quantum dots (QDs; 5.3 nm, emission wavelength λ _em = 614 nm) and used to label HeLa cells successfully. Then green QD-labeled goat anti-mouse immunoglobulin G (IgG; the size of the green QDs was 2.2 nm; λ _em = 544 nm) was added to bind the red-QD-conjugated anti-CD71 on the cell surface by immunoreactions. Such interaction between anti-CD71 and IgG lasted 4 min and was observed from the fluorescence spectra: the fluorescence intensity of the “red” peak at 614 nm increased by 32%; meanwhile that of the “green” one at 544 nm decreased by 55%. The ratio of the fluorescence intensities (I _544 nm/I _614 nm) decreased from 0.5 to 0.2. The fluorescence spectra as well as cell imaging showed that fluorescence resonance energy transfer took place between these two kinds of QDs on the HeLa cells through interactions between the primary antibody and the secondary antibody.