Novel polyoxometalate-templated, 3-D supramolecular networks based on lanthanide dimers: Synthesis, structure, and fluorescent properties of [Ln(2)(DNBA)(4)(DMF)(8)][Mo6O19] (DNBA=3,5-dinitrobenzoate)

The spherical Lindquist type polyoxometalate, Mo6O192-, has been used as a noncoordinating anionic template for the construction of novel three-dimensional lanthanide-aromatic monocarboxylate dimer supramolecular networks [Ln(2)(DNBA)(4)(DMF)(8)][Mo6O19] (Ln = La 1, Ce 2, and Eu 3, DNBA = 3,5-dinitrobenzoate, DMF = dimethylformamide). The title compounds are characterized by elemental analyses, IR, and single-crystal X-ray diffractions. X-ray diffraction experiments reveal that two Ln(III) ions are bridged by four 3,5-dinitrobenzoate anions as asymmetrically bridging ligands, leading to dimeric cores, [Ln(2)(DNBA)(4)(DMF)(8)](2+); [Ln(2)(DNBA)(4)(DMF)(8)](2+) groups are joined together by pi-pi stacking interactions between the aromatic groups to form a two-dimensional grid-like network; the 2-D supramolecular layers are further extended into 3-D supramolecular networks with 1-D box-like channels by hydrogen-bonding interactions, in which hexamolybdate polyanions reside. The compounds represent the first examples of 3-D carboxylate-bridged lanthanide dimer supramolecular "host" networks formed by pi-pi stacking and hydrogen-bonding interactions encapsulating noncoordinating "guest" polyoxoanion species. The fluorescent activity of compound 3 is reported.

Water-soluble fluorescent nanospheres as fluorosensor for detection of Cu2+

A novel water-soluble fluorescent nanosphere as fluorosensor was prepared by emulsifier-free emulsion copolymerization of styrene with naphthalimide derivative (A). The fluorosensor was high sensitive for detection of Cu2+. Comparied with other fluorosensors based on organic fluorophore, it has two advantages. First, there is no pollution to environment in the use of it. Second, it can be used repeatedly.

Solvent effects of a fluorescent polyquinoline chemosensor for metal ions

Red shift and quenching effects of the fluorecence spectra of polyquinoline (PQH) solutions in different solvents are observed upon the addition of metal ions, which demonstrates the extraordinary solvent effects on metal-ion recognition and sensing ability (including sensitivity and selectivity) of PQH.

Studies on the interaction of porphyrin with antibody with fluorescent spectrum and UV-Vis differential spectra

After meso-tetra (alpha, alpha, alpha, alpha-O-phenylacetyl benzene)porphyrin combined with McAb 1F2, there was a significant hyperchromic effect, indicating that the combination of porphyrin and antibody is rigid and compact, aromatic amino acids exist at the combining sites of antigen in antibody. These aromatic amino acids are Trys and Trps, but the numbers of Trp are more than that found for Trys. The stochiometric ratio of porphyrin to 1F2 is 1:1, the disassociation constant was determined as(2.084+/-0.216) x 10(-10) mol/L by a method of fluorescence quenching, showing that both have a high affinity.

Bright blue electroluminescent devices utilizing poly (N-vinylcarbazole) doped with fluorescent dye

Bright blue electroluminescent devices have been fabricated using poly (N-vinylcarbazole) (PVK) doped with perylene as the emissive layer, poly(p-phenylenevinylene) (PPV) as the hole-transporting layer, 2-(4-biphenylyl)-5-(4-tert-butylphenyl)-1,3,4-oxadiazole (PBD), tris(8-hydroxyquinoline)aluminum (Alq(3)) as the electron-transporting layer, and Al as the cathode. A luminance of 700 cd/m(2) and a luminescent efficiency of 0.8% are achieved at a drive voltage of 36 V. In the experiment, it is found that the introduction of electron-transporting layer PBD has a great effect on the emissive color of the electroluminescent devices prepared by PVK doped with perylene. Yellow-green emission is observed from the device structure of glass substrate/indium-tin-oxide/PVK:perylene/Al. The possible emissive mechanisms are given. The effect of the transporting layer on the electroluminescence is also discussed. (C) 1997 Elsevier Science S.A.

A potent anti-oxidant property: fluorescent recombinant alpha-phycocyanin of Spirulina

To express and product a fluorescent antioxidant holo-alpha-phycocyanin (PC) of Spirulina platensis (Sp) with His-tag (rHHPC; recombinant holo-alpha-phycocyaninof Spirulina platensis with His-tag) in 5-l bench scale. A vector harbouring two cassettes was constructed: cpcA along with cpcE-cpcF in one cassette; ho1-pcyA in the other cassette. Lyases CpcE/F of Synechocystis sp. PCC6803 (S6) could catalyse the 82 site Cys in apo-alpha-PC of Sp linking with bilin chromophores, and rHHPC was biosynthesized in Escherichia coli BL21. The constant feeding mode was adopted, and transformant reached the biomass of rHHPC up to 0.55 g l(-1) broth in 5-litre bench scale. rHHPC was purified by Ni2+ affinity column conveniently. The absorbance and the fluorescence emission spectra of rHHPC had lambda(max) at 621 and 650 nm, respectively. The IC50 values of rHHPC were 277.5 +/- 25.8 mu g ml(-1) against hydroxyl radicals and 20.8 +/- 2.2 mu g ml(-1) against peroxyl radicals. Combinational biosynthesis of rHHPC was feasible, and the constant feeding mode was adopted to produce good yields of rHHPC. Fluorescent rHHPC with several unique qualitative and quantitative features was effective on scavenging hydroxyl and peroxyl radicals. A potent antioxidant rHHPC was co-expressed, produced and characterized for nutritional and pharmacological values, which would help to develop phycobiliproteins' applications in their fluorescent and biological activities.

Biosynthesis of fluorescent allophycocyanin alpha-subunits by autocatalysis in Escherichia coli

APC (allophycocyanin) is widely used for fluorescence tagging and may be a promising antioxidant agent for use within the food and pharmaceutical industries. Chromophore attachment to apo-ApcA (apo-APC alpha-subunit without chromophore) can be auto-catalysed both in vitro and in vivo. In the present study, a plasmid containing genes of apo-ApcA and chromophore synthetases (HOI (ferredoxin-dependent haem oxygenase) and PcyA (phycocyanobilin:ferredoxin oxidoreductase)] was constructed and expressed in Escherichia coli. The results show that holo-ApcA (APC alpha-subunit with chromophore) can be synthesized by autocatalysis in E. coli. Recombinant holo-ApcA showed the same spectral and fluorescent properties as PC (phycocyanin) and could serve as a good substitute for native PC for fluorescent tagging. Moreover, recombinant ApcA can inhibit hydroxyl and peroxyl radicals more strongly than holo-ApcA and native APC. The EC50 values were 296.4 +/- 22.4 mu g/ml against hydroxyl radicals and 38.5 +/- 2.6 mu g/ml against peroxyl radicals.

Combinational biosynthesis of a fluorescent cyanobacterial holo-alpha-allophycocyanin in Escherichia coli

Allophycocyanin ( APC) is a phycobiliprotein with various biological and pharmacological properties. An expression vector containing five essential genes in charge of biosynthesis of cyanobacterial APC holo-alpha subunit ( holo- ApcA) was constructed, resulting in over- expression of a fluorescent holo- ApcA in E. coli. After being cultured for 16 h, the dry cell density reached 22.5 gl(-1), and the expression of holo- HT- ApcA was up to 1 gl(-1) broth. The recombinant protein showed similar spectral features to native APC.

Biosynthesis of fluorescent cyanobacterial allophycocyanin trimer in Escherichia coli

Allophycocyanin (APC), a cyanobacterial photosynthetic phycobiliprotein, functions in energy transfer as a light-harvesting protein. One of the prominent spectroscopic characteristics of APC is a strong red-shift in the absorption and emission maxima when monomers are assembled into a trimer. Previously, holo-APC alpha and beta subunits (holo-ApcA and ApcB) were successfully synthesized in Escherichia coli. In this study, both holo-subunits from Synechocystis sp. PCC 6803 were co-expressed in E. coli, and found to self-assemble into trimers. The recombinant APC trimer was purified by metal affinity and size-exclusion chromatography, and had a native structure identical to native APC, as determined by characteristic spectroscopic measurements, fluorescence quantum yield, tryptic digestion analysis, and molecular weight measurements. Combined with results from a study in which only the monomer was formed, our results indicate that bilin synthesis and the subsequent attachment to apo-subunits are important for the successful assembly of APC trimers. This is the first study to report on the assembly of recombinant ApcA and ApcB into a trimer with native structure. Our study provides a promising method for producing better fluorescent tags, as well as a method to facilitate the genetic analysis of APC trimer assembly and biological function.

Development of a sensitive fluorescent derivatization reagent 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC) and its application for determination of amino acids from seeds and bryophyte plants using high-performance liquid chromatography with fluorescence detection and identification with electrospray ionization mass spectrometry

A pre-column derivatization method for the sensitive determination of amino acids and peptides using the tagging reagent 1,2-benzo-3,4dihydrocarbazole-9-ethyl chloroformate (BCEOC) followed by high-performance liquid chromatography with fluorescence detection has been developed. Identification of derivatives was carried out by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS/MS). The chromophore of 2-(9-carbazole)-ethyl chloroformate (CEOC) reagent was replaced by 1,2-benzo-3,4-dihydrocarbazole functional group, which resulted in a sensitive fluorescence derivatizing reagent BCEOC. BCEOC can easily and quickly label peptides and amino acids. Derivatives are stable enough to be efficiently analyzed by high-performance liquid chromatography. The derivatives showed an intense protonated molecular ion corresponding m/z (M + H)(+) under electrospray ionization (ESI) positive-ion mode with an exception being Tyr detected at negative mode. The collision-induced dissociation of protonated molecular ion formed a product at m/z 246.2 corresponding to the cleavage of C-O bond of BCEOC molecule. Studies on derivatization demonstrate excellent derivative yields over the pH 9.0-10.0. Maximal yields close to 100% are observed with a 3-4-fold molar reagent excess. Derivatives exhibit strong fluorescence and extracted detzvatization solution with n-hexane/ethyl acetate (10:1, v/v) allows for the direct injection with no significant interference from the major fluorescent reagent degradation by-products, such as 1,2-benzo-3,4-dihydrocarbazole-9-ethanol (BDC-OH) (a major by-product), mono- 1,2-benzo-3,4-dihydrocarbazole-9-ethyl carbonate (BCEOC-OH) and bis-(1,2-benzo-3,4-dihydrocarbazole-9-ethyl) carbonate (BCEOC)(2). In addition, the detection responses for BCEOC derivatives are compared to those obtained with previously synthesized 2-(9-carbazole)-ethyl chloroformate (CEOC) in our laboratory. The ratios AC(BCEOC)/AC(CEOC) = 2.05-6.51 for fluorescence responses are observed (here, AC is relative fluorescence response). Separation of the derivatized peptides and amino acids had been optimized on Hypersil BDS C-18 column. Detection limits were calculated from 1.0 pmol injection at a signal-to-noise ratio of 3, and were 6.3 (Lys)-177.6 (His) fmol. The mean interday accuracy ranged from 92 to 106% for fluorescence detection with mean %CV < 7.5. The mean interday precision for all standards was < 10% of the expected concentration. Excellent linear responses were observed with coefficients of > 0.9999. Good compositional data could be obtained from the analysis of derivatized protein hydrolysates containing as little as 50.5 ng of sample. Therefore, the facile BCEOC derivatization coupled with mass spectrometry allowed the development of a highly sensitive and specific method for the quantitative analysis of trace levels of amino acids and peptides from biological and natural environmental samples. (c) 2005 Elsevier B.V. All rights reserved.

Synthesis of a terbium fluorescent chelate and its application to time-resolved fluoroimmunoassay

A new nonadentate ligand, N, N, N-1, N-1-[2,6-bis(3'-aminomethyl-1 1'-pyrazolyl)-4-phenylpyridine]tetrakis(acetic acid) (BPTA) for a Tb3+ fluorescent complex was synthesized. The Tb3+ complex is strongly fluorescent, having a large fluorescence quantum yield of 1.00 and very long fluorescence lifetime of 2.681 ms in 0.05 M berate buffer of pH 9.1. Streptavidin (SA) was labeled with SPTA by using its succinimidyl monoester, and the BPTA-Tb3+-labeled SA was used in sandwich-type time-resolved fluoroimmunoassay (TR-FIA) of alpha -fetoprotein (AFP) and carcinoembryonic antigen (CEA) in human sera. The Tb3+-labeled SA was also used in competitive type TR-FIA of bensulfuron- methyl (BSM) in water. The detection limits of these assays are 42 pg/mL for AFP, 70 pg/mL for CEA, and 0.4 ng/mL for BSM. In addition, a new simultaneous measurement method for AFP and CEA in a human serum sample was developed by using 4,4'-bis(1 " ,1 " ,1 " ,2 " ,2 " ,3 " ,3 " -heptafluoro-4 " ,6 " -hexanedion-6 " -yl)chlorosulfo-o-terphenyl ((BHHCT)-Eu3+-labeled anti-AFP antibody, biotinylated anti-CEA antibody, and BPTA-Tb3+-labeled SA. The concentrations of AFP and CEA in 39 human serum samples were determined, and the results were compared with those of the independently determined AFP and CEA by TR-FIA with a single-label method. A good correlation was obtained with the correlation coefficients of 0.991 for AFP and 0.994 for CEA.

Preparation and a time-resolved fluoroimmunoassay application of new europium fluorescent nanoparticles

New silica-based europium fluorescent nanoparticles having surface amino groups were prepared by a covalent binding-copolymerization technique. In the nanoparticles, the fluorescent Eu3+ chelate molecules were covalently bound to silicon atoms to protect the nanoparticles from dye leaking in bio-applications. The amino groups on the surface of nanoparticles made the surface modification and bioconjugation of nanoparticles easier. The nanoparticles were characterized and developed as a new type of fluorescence probe for a highly sensitive time-resolved fluoroimmunoassay (TR-FIA) of human hepatitis B surface antigen (HBsAg).