Development of responsive lanthanide-based magnetic resonance imaging and luminescent probes for biological applications

Lanthanide complexes have unique chemical characteristics compared with typical organic complexes, and have recently attracted much interest because of the expanding need for new bioanalytical sensors. For example, magnetic resonance imaging (MRI) permits noninvasive three-dimensional imaging inside opaque organisms, and gadolinium ion (Gd(3+)) complexes have become important tools as MRI contrast agents. However, most of them are nonspecific, and report solely on anatomy. Therefore, responsive MRI contrast agents, so-called "smart" MRI contrast agents whose ability to relax water protons is greatly enhanced by recognition of a particular biomolecule, have great potential for elucidating biological phenomena. On the other hand, lanthanide complexes such as europium (Eu(3+)) and terbium (Tb(3+)) complexes have excellent luminescence properties for biological applications, i.e., long luminescence lifetime of the order of milliseconds and a large Stoke's shift of >200 nm. Their long-lived luminescence is especially suitable for time-resolved measurements, because the interference from short-lived background fluorescence and scattered light rapidly decays to a negligible level after a pulse of excitation light is applied, and the emitted light can be collected after an appropriate delay time. These luminescent lanthanide complexes have already found commercial use as highly sensitive luminescent probes in heterogeneous and homogeneous assays. This paper reviews our research on the design and synthesis of responsive lanthanide-based MRI and luminescent probes for advanced bioimaging.     

Development of a zinc ion-selective luminescent lanthanide chemosensor for biological applications

Detection of chelatable zinc (Zn(2+)) in biological studies has attracted much attention recently, because chelatable Zn(2+) plays important roles in many biological systems. Lanthanide complexes (Eu(3+), Tb(3+), etc.) have excellent spectroscopic properties for biological applications, such as long luminescence lifetimes of the order of milliseconds, a large Stoke's shift of >200 nm, and high water solubility. Herein, we present the design and synthesis of a novel lanthanide sensor molecule, [Eu-7], for detecting Zn(2+). This europium (Eu(3+)) complex employs a quinolyl ligand as both a chromophore and an acceptor for Zn(2+). Upon addition of Zn(2+) to a solution of [Eu-7], the luminescence of Eu(3+) is strongly enhanced, with high selectivity for Zn(2+) over other biologically relevant metal cations. One of the important advantages of [Eu-7] is that this complex can be excited with longer excitation wavelengths (around 340 nm) as compared with previously reported Zn(2+)-sensitive luminescent lamthanide sensors, whose excitation wavelength is at too high an energy level for biological applications. The usefulness of [Eu-7] for monitoring Zn(2+) changes in living HeLa cells was confirmed. This novel Zn(2+)-selective luminescent lanthanide chemosensor [Eu-7]should be an excellent lead compound for the development of a range of novel luminescent lanthanide chemosensors for biological applications.     

Preparation,characterization and luminescence of transparent thin film of ionogels

The sol–gel fabrication of luminescent and transparent thin film of ionogels containing trivalent lanthanide complexes have been obtained from the silylated ionic liquid in the presence of lanthanide salts (Ln³⁺, Ln=Tb and Eu) and organic ligands. The resulting thin films were investigated by FT-IR spectroscopy, scanning electron microscopy and luminescence spectroscopy. FT-IR spectra reveal the hydrolysis and condensation of the silylated ionic liquid as well as the formation of luminescent lanthanide complexes in the thin films. Scanning electronic microscope images show the homogeneous characteristic of the thin films.     

A Cl- anion-responsive luminescent Eu3+ complex with a chiral tripod: ligand substituent effects on ternary complex stoichiometry and anion sensing selectivity

A new series of N3,O-mixed donor tripods was prepared for luminescent Eu3+ complexes, in which the soft quinoline nitrogen, tertiary amine nitrogen, and hard amide oxygen donors were cooperatively involved. The mixed donor tripods formed more stable 1 : 1 complexes with Eu(NO3)3, La(NO3)3 and Tb(NO3)3 than the corresponding N4 donor tripods, and their Eu3+ complexes particularly exhibited anion-responsive luminescence properties. NMR, UV, and luminescence spectroscopic characterizations revealed that -CH3 substitution on the tripod skeleton remarkably altered the preferred stoichiometry of the "tripod-Eu3+-anion" ternary complex and gave anion-dependent europium luminescence. Although the disubstituted tripod preferred to form non-luminescent 2 : 1 (tripod : Eu3+) complexes with Eu(NO3)3 and other salts, it formed a luminescent 1 : 1 complex with EuCl3. Thus, this type of tripod offered Cl- anion-selective luminescence enhancement that was easily observed by the naked eye.     

Encapsulation of labile trivalent lanthanides into a homobimetallic chromium(III)-containing triple-stranded helicate. Synthesis, characterization, and divergent intramolecular energy transfers

The segmental bidentate-tridentate-bidentate ligand L2 reacts with M(II) (M = Cr, Zn) and Ln(III) (Ln = La, Eu, Gd, Tb, Lu) to give the heterotrimetallic triple-stranded helicates [MLnM(L2)3]7+. For M = Zn(II), the isolated complexes [ZnLnZn(L2)3](CF3SO3)7 (Ln = Eu, Tb) display only lanthanide-centred luminescence arising from the pseudo-tricapped trigonal prismatic LnN9 coordination site. For M = Cr(II), rapid air oxidation provides Cr(III) and leads to the isolation of inert [CrLnCr(L2)3](CF3SO3)9 (Ln = Eu, Tb) complexes, in which divergent intramolecular Ln --> Cr energy transfers can be evidenced. Taking [ZnEuZn(L2)3]7+ as a luminescent standard for Eu-centred emission, a quantitative treatment of the energy migration processes indicates that the rate constant characterizing the Eu --> Cr energy transfer is more efficient in the trimetallic system, than in the analogous simple bimetallic edifice. Particular attention is focused on potential control of directional energy transfer processes in Cr-Ln pairs.     

[（n—C4H9）4N][EuxM1—x（TTA）4]（M=La、Sm、Gd、Tb）的光致发光

合成了一类组成为[(n-C4H9)4N][EuxM1-x(TTA)4](M=La、Sm、Gd、Tb）的固体配合物，通过测定其红外光谱，X射线粉末衍射谱和荧光光谱，研究了配合物结构和发光性质随Eu^3 浓度变化的规律。红外光谱和XRD谱的分析结果表明，在体系中没有新化合物生成，而荧光光谱分析结果表明发光强度与Eu^3 浓度不成线性关系，不发光的基质配合物组分对发光有不同大小和不同类型的影响，提出一种可能的发光机制解释这一共发光现象。     

Highly resolved luminescence measurements for traces of 1,3-dibetanato europium(III) and terbium(III) complexes using a microscope laser Raman spectrometer toward the forensic discrimination.

The emission spectra of luminescent trivalent europium (Eu3+) and terbium (Tb3+) complexes were measured using a microscope laser Raman spectrometer with a doubled Nd:YAG laser (532 nm) and an Ar laser (488 nm). Excitation at 532 and 488 nm corresponded to wavelengths of the 7F1 --> 5D1 band of Eu3+ and the 7F6 --> 5D4 band of Tb3+, respectively. The Eu3+ and Tb3+ complexes were discriminated by high-resolution emission spectra more distinctly and sensitively than by fluorescence spectrometry, the usual analytical method.     

A disymmetric terpyridine based ligand for the formation of luminescent di-aquo lanthanide complexes

The synthesis of ligand H3 based on a disymmetrically substituted terpyridine core functionalised by a carboxylic acid in the 6-position and a bis(carboxymethyl)aminomethyl function in the 6'-position is described. The coordination behaviour of this heptadentate (4N/3O) ligand with lanthanide cations (Ln=Eu, Gd and Tb) was studied in solution showing the formation of complexes with [Ln] stoichiometry. Complexes with general formula [Ln(H2O)2] were isolated from neutral water solutions containing equimolar amounts of cations and ligands, and the complexes were characterized in the solid state (elemental analysis, IR) and in solution (mass spectrometry). The photo-physical properties of the luminescent complexes of Eu and Tb were studied in water solution by means of absorption, steady state and time-resolved emission spectroscopies. Evolution of the luminescence lifetimes of the Eu and Tb complexes in H2O and D2O reveals the presence of two water molecules coordinated in the first coordination sphere of the cations. Despite this important hydration number, the overall luminescence quantum yields of the complexes remained elevated, especially in the case of Tb (Phi=22.0 and 6.5% respectively for Tb and Eu). Upon crystallisation the Gd complex formed dimeric species in which two gadolinium atoms are each heptacoordinated by one ligand, the coordination sphere being completed by a single water molecule and a bridging carboxylate function, pointing to different behaviours in the solid and liquid states.     

β-二酮、邻菲罗啉分别与铕和铽三元配合物的合成及发光性能研究

为了寻找新的发光材料并研究β-二酮对稀土配合物发光性能的影响, 我们合成了一个新的β-二酮配体: 1-苯 基-3-(对苯乙炔苯基)-1,3-丙二酮(HPPP), 并用HPPP、邻菲罗啉(phen)分别与Eu(III)和Tb(III)反应, 合成了两个新的三元稀土配合物: Eu(PPP)3phen和Tb(PPP)3phen, 通过红外光谱、化学分析、元素分析对三元稀土配合物的组成和结构进行了表征. 研究了配合物的荧光性质, 发现β-二酮配体对配合物的发光有较大影响, 通过量子化学计算对实验结果进行了解释.     

Ag@SiO2@Y2O3:RE3+（RE=Eu,Tb）核壳结构发光材料的制备与性能

采用均相沉淀法制备了Ag@SiO2@(Y,RE)(OH)CO3.H2O(RE=Eu,Tb)核壳结构微球,经过700℃焙烧后成功制备出Ag@SiO2@Y2O3:RE3+(RE=Eu,Tb)核壳结构发光材料。XRD谱图表明Ag核具有结晶良好的面心立方结构;SiO2层为无定型;Y2O3层为立方晶系。FTIR谱图表明核壳之间以化学键相结合。TEM照片表明合成了核壳结构的表面光滑的复合微球,分散良好,大小均匀,Ag核的粒径分布为50±20 nm;SiO2层的厚度为20～30 nm;Y2O3:RE3+(RE=Eu,Tb)层厚度约为125 nm。电子衍射图像表明Ag@SiO2@Y2O3:RE3+(RE=Eu,Tb)为多晶结构。UV-Vis光谱表明表面包覆使Ag离子的等离子体共振吸收峰发生了红移。荧光光谱表明Ag@SiO2@Y2O3:Eu3+具有Eu3+的特征红光发射,Ag@SiO2@Y2O3:Tb3+具有Tb3+的特征绿光发射,但是发光强度均比纯的Y2O3:RE3+有所减弱,说明贵金属的引入对稀土Y2O3:RE3+(RE=Eu,Tb)的发光起到了荧光猝灭的作用。     

Towards libraries of luminescent lanthanide complexes and labels from generic synthons

A synthetic approach is developed to obtain families of luminescent lanthanide complexes and markers from a generic family of precursors built from nonadentate coordination sites. The syntheses of the precursors, based on a directed regioselective nucleophilic aromatic substitution on polyfluoropyridines, are described. Functionalisation of the synthons on the aromatic moieties allowed the introduction of labelling functions and/or the extension of the electronic delocalisation, with concomitant changes in the spectroscopic properties. The synthesis of two such families of ligands and of some of their complexes of Eu(III) and Tb(III) are described, and the photo-physical properties of the complexes were measured, revealing excellent luminescence quantum yields reaching unity in some cases. For some of these complexes, the emphasis was further put on the preparation of an N-hydroxylsuccinimide (NHS) ester as activated function for labelling. The Tb and La complexes in the NHS activated form were synthesized and fully characterized. The labelling was first demonstrated on amino functionalized polymer beads and characterized by time-resolved luminescence microscopy. In a second step, the activated Tb complex was used for the labelling of GFR44 monoclonal antibody, and was applied to the detection of carcinoembryonic antigene (CEA) within the frame of a time-resolved fluoroimmunoassay. Comparison with a commercially available kit based on a europium cryptate as energy donor confirms the efficiency of Tb to act as an energy donor with an unoptimised 35% increase of the detection efficiency.     

Synthesis and characterization of red-luminescent graphene oxide functionalized with silica-coated Eu3+ complex nanoparticles

Traditional dye-doped fluorescent graphene oxide (GO) reveals a low quantum yield and a short life expectancy. Herein, red-luminescent silica-coated Eu(3+) complex nanoparticles were synthesized and covalently coupled to GO nanosheets by means of a carbodiimide-mediated amidation process. SEM and TEM studies demonstrated successful attachment of the silica-coated Eu(3+) complex nanoparticles onto the GO surface. Spectroscopic studies showed that the GO-nanoparticle conjugates exhibit strong luminescence, long lifetimes, as well as good photostability, which suggests that this new type of luminescent nanomaterial has the potential for highly sensitive time-resolved fluorescence cyto- and histochemistry imaging.     

Bimodal system (luminophore and paramagnetic contrastophore) derived from Ln(III) complexes based on a bipyridine-containing macrocyclic ligand

The synthesis of a new 15-membered polyaza-macrocyclic ligand L3H3, which is based on a 2,2'-bipyridine moiety and a diethylenetriaminetriacetic acid core, is reported. The lanthanide chelates of this octadentate ligand were programmed for bimodal probes, luminescent agents (Sm, Eu, Tb, Dy), and magnetic resonance imaging agents (Gd3+). The neutral 1:1 complexes with these Ln3+ ions were prepared and studied in aqueous solution by luminescence and NMR techniques. The main photophysical characteristics of these complexes (i.e., the absorption and luminescence spectra, the metal-centered lifetimes, and the overall luminescence yields, Phi) were measured. In addition, the role played by nonradiative pathways (vibrational energy transfer involving coordinated water molecules, involvement of ligand-to-metal charge-transfer excited states, or metal --> ligand back transfer) is discussed. The L3.Eu and L3.Tb complexes show very bright luminescence when photoexcited from the lowest-energy absorption band of the bipyridine chromophore. The luminescence quantum yields in an air-equilibrated water solution at room temperature are 0.10 and 0.21, respectively, despite the presence of one water molecule in the first coordination sphere of the metal ion. NMR data show that L3.Gd contains also one H2O molecule in the inner sphere. The proton longitudinal relaxivity, r1, of this complex is 3.4 s(-1) mM(-1) (0.47 T, 310 K) and the rotational correlation time, tau(R), is 57 ps (310 K). These values are comparable to those of the clinically used Gd-DTPA. Interestingly, the water exchange rate between the coordination site and the bulk solvent is slow (tau(M) = 3.5 micros at 310 K). The presence of water molecules in the second sphere and in rapid exchange with the solvent is discussed. Finally, it was found by luminescence and NMR experiments that these lanthanide complexes are stable versus transmetalation by several cations (especially Ca2+ and Zn2+) at physiological pH and have no interaction with blood proteins.     

Modulation of luminescence intensity of lanthanide complexes by photoinduced electron transfer and its application to a long-lived protease probe

Luminescent lanthanide complexes (Tb(3+), Eu(3+), etc.) have excellent properties for biological applications, including extraordinarily long lifetimes and large Stokes shifts. However, there have been few reports of lanthanide-based functional probes, because of the difficulty in designing suitable complexes with a luminescent on/off switch. Here, we have synthesized a series of complexes which consist of three moieties: a lanthanide chelate, an antenna, and a luminescence off/on switch. The antenna is an aromatic ring which absorbs light and transmits its energy to the metal, and the switch is a benzene derivative with a different HOMO level. If the HOMO level is higher than a certain threshold, the complex emits no luminescence at all, which indicates that the lanthanide luminescence can be modulated by photoinduced electron transfer (PeT) from the switch to the sensitizer. This approach to control lanthanide luminescence makes possible the rational design of functional lanthanide complexes, in which the luminescence property is altered by a biological reaction. To exemplify the utility of our approach to the design of lanthanide complexes with a switch, we have developed a novel protease probe, which undergoes a significant change in luminescence intensity upon enzymatic cleavage of the substrate peptide. This probe, combined with time-resolved measurements, was confirmed in model experiments to be useful for the screening of inhibitors, as well as for clinical diagnosis.     

Study of lanthanide complexes with salicylic acid by photoacoustic and fluorescence spectroscopy

Solid complexes Ln(Sal)3.H2O (Sal: salicylic acid; Ln: La3+, Nd3+, Eu3+, Tb3+) are synthesized, and their photoacoustic (PA) spectra in the UV-Vis region have been recorded. PA intensities of central lanthanide ions are interpreted in terms of the probability of nonradiative transitions. It is found that PA intensity of the ligand increases in the order of Tb(Sal)3.H2O < La(Sal3).H2O < Eu(Sal)3.H2O < Nd(Sal)3.H2O. Different PA intensities of the ligand are interpreted by comparison with the fluorescence spectra. Ternary complexes Eu(Sal)3Phen and Tb(Sal)3Phen (Phen: 1,10-phenanthroline) are synthesized. Compared with their binary complexes, PA intensity of the ligand Sal decreases for Eu(Sal)3Phen, while the reverse is true for that of Tb(Sal)3Phen. The luminescence of Eu3+ increases remarkably when Phen is introduced, and luminescence of Tb3+ decreases greatly when Phen is added. The intramolecular energy transfer and relaxation processes in the complexes are discussed from two aspects: radiative and nonradiative relaxations.     

铕(Ⅲ)、铽(Ⅲ) β-二酮配合物的制备及表征

醋酸铕、醋酸铽分别与乙酰丙酮在60°C水浴下加热回流,生成两种稀土β-二酮配合物,即乙酰丙酮铕(Eu(acac)3?3H2O)和乙酰丙酮铽(Tb(acac)3?3H2O),在紫外灯下观察配合物发光现象,乙酰丙酮铕在紫外灯下显暗红色,乙酰丙酮铽在紫外灯下显亮绿色。红外分析结果与理论相吻合,荧光分析表明它们是较好的荧光材料,热分析结果说明它们的热稳定性相对较高。     

A new luminescence method for determining dysprosium in the presence of terbium

The new luminescence method for determining dysprosium in the presence of terbium is developed based on the difference in the lifetimes of dysprosium and terbium in complexes with 3-(6-benzodioxanyl)pyrazole-5-carboxylic acid (τ = 6 and 910 μs, respectively). The possibility of determining the short-lived and weakly luminescent dysprosium(III) ion in the presence of long-living and intensively luminescent terbium(III) was demonstrated for the first time using time-resolved luminescence. The developed method was used to determine dysprosium in luminescent materials doped with both lanthanides.     

Prototype protein assembly as scaffold for time-resolved fluoroimmuno assays

Turnip yellow mosaic virus (TYMV) is an icosahedral plant virus with an average diameter of 28 nm and can be isolated in gram quantities from turnip or Chinese cabbage inexpensively. In this study, it was selected as a prototype bionanoparticle for time-resolved fluoroimmuno assay (TRFIA). Two types of reactive amino acid residues were employed to anchor luminescent terbium complexes and biotin groups based on orthogonal chemical reactions. While terbium complexes were used as luminescent signaling groups, biotin motifs acted as a model ligand for protein binding. The bioconjugation results were confirmed by MS and Western blot analysis. Steady-state and time-resolved luminescence study of the dual-modified viruses demonstrated that the spectroscopic properties of the Tb complex are unperturbed by the labeling procedure. The dual-modified particle was probed by fluorescence resonance energy transfer (FRET) experiments using avidin labeled with an Alexa488 fluorophore, which bound to the biotin on the surface of the particle, as an energy acceptor, and terbium complexes as an energy donor. The emission and excitation spectra of the dual-labeled TYMV particle displayed residual virus fluorescence and Tb luminescence upon ligand-centered excitation. The Tb luminescence lifetime was 1.62 ms and could be effectively fitted with a single-exponential behavior. In the TRFIA, an efficient transfer of 66% was observed, and the calculation using the F?rster radius of 41 A allowed for an estimation of the average donor-acceptor distance of 36 A. Our studies show that the two reactive sites can communicate with each other on the surface of a nanoscale biological assembly. In particular, the ligand-receptor binding (biotin and avidin in this paper) was not interfered with when anchored to the surface of TYMV. Therefore, as a prototype of polyvalent bionanoparticles, TYMV can be used as scaffold for sensor development with TRFIA.     

洛美沙星的铕离子荧光探针时间分辨荧光法测定

采用时间分辨荧光法,以铕离子为荧光探针,依据Eu3+-La3+-LMX-SDS稀土共发光体系,建立了测定洛美沙星(LMX)含量的新方法。研究了配合物的紫外及荧光光谱,分析了配合物的发光机理,考察了溶液pH值、缓冲溶液种类及反应试剂加入量对配合物荧光强度的影响。在优化的实验条件下,配合物荧光强度和洛美沙星浓度呈线性关系,相关系数为0.999 0,线性范围为5.0×10-7~1.0×10-5mol/L。方法的检出限为6.8×10-8mol/L,对5.0×10-6mol/L的洛美沙星溶液测定11次,RSD为0.6%。将该方法用于尿液中洛美沙星含量的测定,回收率为94%~104%。     

Brilliant Sm, Eu, Tb, and Dy chiral lanthanide complexes with strong circularly polarized luminescence

The synthesis, characterization, and luminescent behavior of trivalent Sm, Eu, Dy, and Tb complexes of two enantiomeric, octadentate, chiral, 2-hydroxyisophthalamide ligands are reported. These complexes are highly luminescent in solution. Functionalization of the achiral parent ligand with a chiral 1-phenylethylamine substituent on the open face of the complex in close proximity to the metal center yields complexes with strong circularly polarized luminescence (CPL) activity. This appears to be the first example of a system utilizing the same ligand architecture to sensitize four different lanthanide cations and display CPL activity. The luminescence dissymmetry factor, g(lum), recorded for the Eu(III) complex is one of the highest values reported, and this is the first time the CPL effect has been demonstrated for a Sm(III) complex with a chiral ligand. The combination of high luminescence intensity with CPL activity should enable new bioanalytical applications of macromolecules in chiral environments.