Midrange affinity fluorescent Zn(II) sensors of the Zinpyr family: syntheses, characterization, and biological imaging applications

The syntheses and photophysical characterization of ZP9, 2-{2-chloro-6-hydroxy-3-oxo-5-[(2-{[pyridin-2-ylmethyl-(1H-pyrrol-2-ylmethyl)amino]methyl}phenylamino)methyl]-3H-xanthen-9-yl}benzoic acid, and ZP10, 2-{2-chloro-6-hydroxy-5-[(2-{[(1-methyl-1H-pyrrol-2-ylmethyl)pyridin-2-ylmethylamino]methyl}phenylamino)methyl]-3-oxo-3H-xanthen-9-yl}benzoic acid, two asymmetrically derivatized fluorescein-based dyes, are described. These sensors each contain an aniline-based ligand moiety functionalized with a pyridyl-amine-pyrrole group and have dissociation constants for Zn(II) in the sub-micromolar (ZP9) and low-micromolar (ZP10) range, which we define as "midrange". They give approximately 12- (ZP9) and approximately 7-fold (ZP10) fluorescence turn-on immediately following Zn(II) addition at neutral pH and exhibit improved selectivity for Zn(II) compared to the di-(2-picolyl)amine-based Zinpyr (ZP) sensors. Confocal microscopy studies indicate that such asymmetrical fluorescein-based probes are cell permeable and Zn(II) responsive in vivo.     

稀土-荧光素双重荧光响应pH探针分子的合成与荧光性质研究

将含有8-氨基喹啉与荧光素衍生物缩合的席夫碱QZ1与铕髥的TTA(噻吩基三氟乙酰丙酮)化合物作为基本原料合成了新的铕髥-荧光素化合物Eu-QZ1,并对其进行了光谱表征。化合物Eu-QZ1在470 nm光的激发下产生荧光素的特征绿色荧光(530 nm)。在从碱到酸(pH从9.0到6.0)滴定过程中该荧光的强度增强了接近5倍,其pKa值计算为7.30。在370 nm光的激发下,Eu-QZ1发射稀土铕髥离子的特征红色荧光(主峰位于612 nm)。在从酸到碱(pH从5.5到8.5)滴定过程中,该荧光的强度增强了接近10倍,其pKa值计算为7.39。这些结果表明Eu-QZ1是1个高灵敏度双荧光响应的pH探针。细胞试验表明Eu-QZ1具有良好的细胞通透性,能够在红光和绿光两个位置标记海拉细胞中的pH范围。     

Zinspy sensors with enhanced dynamic range for imaging neuronal cell zinc uptake and mobilization

Thiophene moieties were incorporated into previously described Zinspy (ZS) fluorescent Zn(II) sensor motifs (Nolan, E. M.; Lippard, S. J. Inorg. Chem. 2004, 43, 8310-8317) to provide enhanced fluorescence properties, low-micromolar dissociation constants for Zn(II), and improved Zn(II) selectivity. Halogenation of the xanthenone and benzoate moieties of the fluorescein platform systematically modulates the excitation and emission profiles, pH-dependent fluorescence, Zn(II) affinity, and Zn(II) complexation rates, offering a general strategy for tuning multiple properties of xanthenone-based metal ion sensors. Extensive biological studies in cultured cells and primary neuronal cultures demonstrate 2-{6-hydroxy-3-oxo-4,5-bis[(pyridin-2-ylmethylthiophen-2-ylmethylamino)methyl]-3H-xanthen-9-yl}benzoic acid (ZS5) to be a versatile imaging tool for detecting Zn(II) in vivo. ZS5 localizes to the mitochondria of HeLa cells and allows visualization of glutamate-mediated Zn(II) uptake in dendrites and Zn(II) release resulting from nitrosative stress in neurons.     

Rhenium tricarbonyl core complexes of thymidine and uridine derivatives

Thymidine and uridine were modified at the C2' and C5' ribose positions to form amine analogues of the nucleosides (1 and 4). Direct amination with NaBH(OAc)3 in DCE with the appropriate aldehydes yielded 1-{5-[(bis(pyridin-2-ylmethyl)amino)methyl]-4-hydroxytetrahydrofuran-2-yl}-5-methyl-1H-pyrimidine-2,4-dione (L1), 1-{5-[(bis(quinolin-2-ylmethyl)amino)methyl]-4-hydroxytetrahydrofuran-2-yl}-5-methyl-1H-pyrimidine-2,4-dione (L2), and 1-[3-(bis(pyridin-2-ylmethyl)amino)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl]-1H-pyrimidine-2,4-dione (L5), while standard coupling procedures of 1 and 4 with 5-(bis(pyridin-2-ylmethyl)amino)pentanoic acid (2) and 5-(bis(quinolin-2-ylmethyl)amino)pentanoic acid (3) in the presence of HOBT-EDCI in DMF provided a second novel series of bifunctional chelators: 5-(bis(pyridin-2-ylmethyl)amino)pentanoic acid [(3-hydroxy-5-(5-methyl-4-oxo-3,4-dihydro-2H-pyrimidin-1-yl)tetrahydrofuran-2-yl)methyl] amide (L3), 5-(bis(quinolin-2-ylmethyl)amino)pentanoic acid [(3-hydroxy-5-(5-methyl-4-oxo-3,4-dihydro-2H-pyrimidin-1-yl)tetrahydrofuran-2-yl)methyl] amide (L4), 5-(bis(pyridin-2-ylmethyl)amino)pentanoic acid [2-(2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-3-yl] amide (L6), and 5-(bis(quinolin-2-ylmethyl)amino)pentanoic acid [2-(2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-3-yl] amide (L7). The rhenium tricarbonyl complexes of L1-L4, L6, and L7, [Re(CO)3(LX)]Br (X=1-4, 6, 7: compounds 5-10, respectively), have been prepared by reacting the appropriate ligand with [NEt4][Re(CO)3Br3] in methanol. The ligands and their rhenium complexes were obtained in good yields and characterized by common spectroscopic techniques including 1D and 2D NMR, HRMS, IR, cyclic voltammetry, UV, and luminescence spectroscopy and X-ray crystallography. The crystal structure of complex 6.0.5NaPF6 displays a facial geometry of the carbonyl ligands. The nitrogen donors of the tridentate ligand complete the distorted octahedral spheres of the complex. Crystal data: monoclinic, C2, a = 24.618(3) A, b = 11.4787(11) A, c = 15.5902(15) A, beta = 112.422(4) degrees , Z = 4, D(calc) = 1.562 g/cm3.     

Synthesis and antimicrobial activity of some derivatives of (7-hydroxy-2-oxo-2H-chromen-4-yl)-acetic acid hydrazide

(7-Hydroxy-2-oxo-2H-chromen-4-yl)-acetic acid hydrazide (2) was prepared from (7-hydroxy-2-oxo-2H-chromen-4-yl)-acetic acid ethyl ester (1) and 100% hydrazine hydrate. Compound 2, is the key intermediate for the synthesis of several series of new compounds such as Schiff's bases 3a-l, formic acid N'-[2-(7-hydroxy-2-oxo-2H- chromen-4-yl)acetyl] hydrazide (4), acetic acid N'-[2-(7-hydroxy-2-oxo-2H-chromen-4- yl)-acetyl] hydrazide (5), (7-hydroxy-2-oxo-2H-chromen-4-yl)-acetic acid N'-[2-(4- hydroxy-2-oxo-2H-chromen-3-yl)-2-oxoethyl] hydrazide (6), 4-phenyl-1-(7-hydroxy-2- oxo-2H-chromen- 4-acetyl) thiosemicarbazide (7), ethyl 3-{2-[2-(7-hydroxy-2-oxo-2H- chromen-4-yl)-acetyl]hydrazono}butanoate (8), (7-hydroxy-2-oxo-2H-chromen-4-yl)- acetic acid N'-[(4-trifluoromethylphenylimino)methyl] hydrazide (9) and (7-hydroxy-2- oxo-2H-chromen-4-yl)acetic acid N'-[(2,3,4-trifluorophenylimino)-methyl] hydrazide (10). Cyclo- condensation of compound 2 with pentane-2,4-dione gave 4-[2-(3,5- dimethyl-1H-pyrazol-1-yl)-2-oxoethyl]-7-hydroxy-2H-chromen-2-one (11), while with carbon disulfide it afforded 7-hydroxy-4-[(5-mercapto-1,3,4-oxadiazol-2-yl)methyl]-2H- chromen-2-one (12) and with potassium isothiocyanate it gave 7-hydroxy-4-[(5- mercapto-4H-1,2,4-triazol-3-yl)methyl]-2H-chromen-2-one (14). Compound 7 was cyclized to afford 2-(7-hydroxy-2-oxo-2H-chromen-4-yl)-N -(4-oxo-2-phenylimino- thiazolidin-3-yl) acetamide (15).     

Synthesis,structure elucidation and plants growth promoting effects of novel quinolinyl chalcones

The readily synthesized 3-(4-Hydroxy-1-methyl-1,2-dihydro-2-oxoquinolin-3-yl)-1-phenyl-1H-pyrazole-4-carbaldehyd (5) and 3-(2-Oxo-2H-chromen-3-yl)-1-phenyl-1H-pyrazole-4-carbaldehyde (6) were utilized as a convenient starting precursor materials for synthesis of novel enone system 4-hydroxy-1-methyl-3-(4-(2H-2-oxo-chromen-3-yl)prop-2-enoyl)-1-phenyl-1H-pyrazol-4-yl)quinolin-2(1H)-one (7) and4-hydroxy-1-methyl-3-(2E)-3-(3-(2-oxo-2H-chromen-3-yl)-1-phenyl-1H-pyrazol-4-yl)acryloyl)quinolin-2(1H)-one (8). Simple homonuclear NOE experiment (NOESY 1D) method was performed for structure elucidation of the novel quinolinyl chalcones. The synthesized compounds have been estimated for their effect of growth on some selective crop of plants (Hibiscus, Mint and Basil).     

6-methylpyridyl for pyridyl substitution tunes the properties of fluorescent zinc sensors of the Zinpyr family

To prepare fluorescent zinc sensors with binding affinities lower than that of the parent 9-(o-carboxyphenyl)-2,7-dichloro-4,5-bis(bis(2-pyridylmethyl)methylaminomethyl)-6-hydroxy-3-xanthenone (ZP1), dimethylated and tetramethylated derivatives were synthesized having either two or four of the pyridyl subunits methylated at the 6-position. Like the parent ZP1, both Me(2)ZP1 and Me(4)ZP1 exhibit increased fluorescence in the presence of Zn(2+). The integrated emission of Me(2)ZP1 increases 4-fold in the presence of excess zinc, whereas Me(4)ZP1 displays 2.5-fold enhanced fluorescence for Zn(2+). Methylating the 6-positions of the pyridyl rings raises the dissociation constant of the sensors and lowers the pK(a) values associated with the tertiary amine ligands in a systematic manner. The properties of the dimethylated Me(2)ZP1 dye resemble those of ZP1, but the tetramethylated Me(4)ZP1 differs greatly from ZP1 in terms of its brightness, affinity toward Zn(2+), exchange kinetics, and metal sensitivity. Both Me(2)ZP1 and Me(4)ZP1 can enter HeLa cells and signal the presence of Zn(2+). Staining caused by both dyes is punctate, with localization patterns resembling that observed for ZP1.     

Syntheses and Crystal Structures of a Series of Coordination Complexes Derived from the Fluorescein Ligand

Solvothermal reaction of lanthanide(Ⅲ) salts with fluorescein (2-(6-hydroxy3-oxo-3H-xanthen-9-yl)benzoic acid) led to a series of new coordination polymers {[Ln(C 20 H 11 O 5)(C 20 H 10 O 5)(H 2 O)]·DMF} n (Ln=Er,Eu,Gd,Tb,Tm,Yb).The PXRD patterns of the complexes indicate they are isomorphous.The structure of complex {[Er(C 20 H 11 O 5)(C 20 H 10 O 5)(H 2 O)]·DMF} n has been determined by single-crystal X-ray diffraction,revealing a 2D framework in which DMF molecules were filled between the layers.The crystal structure belongs to the triclinic system,space group P1,with a=12.107(4),b=12.232(4),c=13.273(4),α=68.005(7),β=88.024(11),γ=77.451(8)°,V=1776.7(9) 3,Z=2,D c=1.720 g/cm 3,μ=2.434 mm-1,F(000)=918,R int=0.0584,T=293(2) K,the final R=0.0621 and wR=0.1501.     

4-Hydroxy-2-quinolones. 108. N-R-amides of 9-fluoro-1-hydroxy-5-methyl-3-oxo-6,7-dihydro-3H,5H-pyrido[3,2,1-ij]quinoline-2-carboxylic acid and their antitubercular activity

The reaction of 6-fluoro-2-methyl-1,2,3,4-tetrahydroquinoline with triethylmethanetricarboxylate gives di(9-fluoro-1-hydroxy-5-methyl-3-oxo-6,7-dihydro-3H,5H-pyrido[3,2,1-ij]quinolin-2-yl)methane and ethyl 9-fluoro-1-hydroxy-5-methyl-3-oxo-6,7-dihydro-3H,5H-pyrido[3,2,1-ij]quinoline-2-carboxylate from which alkyl-, dialkylaminoalkyl-, and hetarylamides as well as hydrazides were prepared. The structure and antitubercular properties of the compounds synthesized are discussed. __________ Translated from Khimiya Geterotsiklicheskikh Soedinenii, No. 9, pp. 1391–1407, September, 2006.     

Synthesis of a fluorescent xanthenic derivative useful for labeling amine residues

The 2,5-dioxopyrrolidin-1-yl-4-(3-hydroxy-6-oxo-6H-xanthen-9-yl)-3-methylbenzoate has been synthesized as an amine-reactive derivative able to yield stable covalently labeled biopolymers. The new derivative has been used to label polyribocytidilic acid (5'), poly(C), amine residues. TG-II-poly(C) exhibits monoexponential decay at the physiological pH range. In addition, both steady-state fluorescence intensity and fluorescence decay are also sensitive to solution pH. The large decrease in steady-state fluorescence upon hybridization allows it to be used as a nucleic acid probe in a homogeneous assay format. In summary, we report an efficient synthesis to obtain labeled RNA from commercially available materials in excellent yields.     

新型吡咯并三嗪酮类PARP-1抑制剂的设计、合成及生物活性

分别以5-溴-2-氟苯甲腈(1a)和3-溴苯甲腈(1b)为原料,经Sonogashira偶联,脱三甲基硅基保护基,三分子偶联及水解等5步反应制得中间体2-氟-5-［(4-氧代-3,4-二氢吡咯［1,2-d］［1,2,4］三嗪-1-基)甲基］苯甲酸（6a)和3-[(4-氧代-3,4-二氢吡咯［1,2-d］［1,2,4］三嗪-1-基)甲基］苯甲酸（6b）。环烷基甲酸经酰氯化,缩合和脱Boc保护基3步反应制得环烷基哌嗪-1-基甲酮(7a~7c)。 6a与NCS（1 eq.）反应制得5-［(6-氯-4氧代-3,4二氢吡咯［1,2-d］［1,2,4］三嗪-1-基)甲基］-2氟 苯甲酸(6c); 6a与NCS(2 eq.)反应制得5-［(6,7-二氯-4氧代-3,4二氢吡咯［1,2-d］［1,2,4］三嗪-1-基)甲基］-2氟-苯甲酸(6d)。 6a~6d, 6a~6c分别与7a~7c和1-（2-嘧啶基）哌嗪在TBTU（缩合剂）,DIPEA（碱）的作用下合成了13个新型吡咯并三嗪酮类PARP-1抑制剂(8a~8m),其结构经¹HNMR和MS(ESI)表征。采用Alarm blue法研究了8a~8m对肿瘤细胞MDA-MB-436的抑制活性(IC₅₀)。结果表明：8f, 8g, 8i和8j对MDA-MB-436有较强的抑制活性（IC₅₀=30.5~69.3 nmol·L^-1）。     

1H and 13C NMR assignments for two new xanthones from the endophytic fungus Xylaria sp. FRR 5657

The complete 1H and 13C NMR assignments are reported for the new natural products, 7-hydroxy-3-(hydroxymethyl)-1-methoxy-9H-xanthen-9-one (1) and 2,5-dihydroxy-8-methoxy-6-methyl-9-oxo-9H-xanthene -1-carboxylic acid (2). Both of these secondary metabolites were isolated from the fermentation culture of the endophytic fungus Xylaria sp. FRR 5657. 1D and 2D NMR experiments that included 1H, gCOSY, gHSQC and gHMBC were used for the determination of the structure and assignment of these xanthones.     

Structure,synthesis, and voltammetric investigation of 1-(quinolin-2-yl)-2-(pyran-2-yl)ethane-1,2-dione derivatives

By the XRD analysis the structure was established of 1-(7,8-dimethyl-4-chloroquinolin-2-yl)-2-[3,5-di(tert-butyl)-6-oxo-6H-pyran-2-yl]ethane-1,2-dione formed as a result of the oxidation of 3,5-di(tert-butyl)-6-[(Z)-2-(quinolin-2-yl)-1-hydroxyethen-1-yl]pyran-2-ones. By the cyclic voltammetry the oxidation of 1-(quinolin-2-yl)-2-(pyran-2-yl)ethane-1,2-dione derivatives was shown to proceed in two stages.     

New platinum(II) and palladium(II) quinoline-imine-pyridine, quinoline-imine-thiazole and quinoline-imine-imidazole complexes by metal-assisted condensation reactions

Pt(II) and Pd(II) methyl- and chloro-complexes with the tridentate N-donor ligands ((pyridin-2-yl)methylene)quinolin-8-amine (NNPy), ((pyridin-2-yl)ethylidene)quinolin-8-yl-amine (NNMePy), (phenyl(pyridin-2-yl)methylene)quinolin-8-yl-amine (NNPhPy), ((thiazol-2-yl)methylene)quinolin-8-amine (NNTh) and ((imidazol-4-yl)methylene)quinolin-8-amine (NNImH) were prepared by metal-assisted condensation of 8-aminoquinoline and an ortho-substituted aldehydo- or keto- N-heterocycle. Preliminary reactivity studies involving the coordinated tridentate N-donors, the chloro-ligand and the M-CH₃ bond were carried out, leading to the synthesis of several new complexes. During these studies, the formation of a novel five-coordinate Pt(II) carbonyl-complex was observed.     

Enantioselective synthesis of both enantiomers of the neuroexcitant 2-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl) propanoic acid (ATPA)

The preparation of both enantiomers of 2-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl) propanoic acid (ATPA), 1, an analogue of the neuroexcitant 2-amino-3-(3-hydroxy-5-methyl-4-yl) propanoic acid (AMPA) is described. The enantiomerically pure glycine derivative tert-butoxycarbonyl-2-(tert-butyl)-3-methyl-4-oxo-1-imidazolidinecarboxylate (BOC-BMI) was coupled with 4-bromomethyl-2-methoxymethyl-5-tert-butylisoxazolin-3-one 6 to give the intermediates (2R,5R)-8 and (2S,5S)-8. These alkylated products were hydrolyzed under mild conditions to give enantiopure (R)-1 and (S)-1 with e.e.'s in excess of 99% in 33% overall yield.     

Synthesis of 8-demethyl-8-hydroxy-5-deazariboflavins

1-Deoxy 1-(3,4-dihydro-8-hydroxy-2,4-dioxopyrimido[4,5-b]quinolin-10-(2H)-yl)-D-ribitol (7,8-didemethyl-8-hydroxy-5-deazariboflavin), the flavin moiety of Methanobacterium coenzyme F₄₂₀, and its 7-methyl analog were prepared by acid-catalyzed reaction of appropriately substituted 6-(N-D-ribitylanilino)uracils with trimethyl or triethyl orthoformate followed by deprotection.     

4-Substituted-3-alkyl-3,4-dihydro-2H-1,3-benzoxazin-2-ones II (1,2). Solvolytic transformations of urea and thiourea derivatives into 1-alkyldihydro-6-(2-hydroxyaryl)-1,3,5-triazine-2,4-(1H,3H) diones

A series of 3,4-Dihydro-2-oxo-(3-substituted-2H-1,3-benzoxazin-4-yl)ureas (II, Table I) and 3,4-dihydro-2-oxo-(3-substituted-2H-1,3-benzoxazin-4-yl)thioureas (Table II) was prepared by treating 3,4-dihydro-4-hydroxy-3-substituted-2H-1,3-benzoxazin-2-ones with ureas and thioureas, respectively. In the presence of alcoholic alkali these compounds underwent transacylation to dihydro-6-(2-hydroxyaryl)-1,3,5-triazine-2,4-(1H,2H)diones (Table III) and their 4-thio analogues (Table IV).     

A highly selective fluorescent sensor for distinguishing cadmium from zinc ions based on a quinoline platform

A fluorescent sensor, N-(quinolin-8-yl)-2-(quinolin-8-yloxy)acetamide (HL), based on 8-aminoquinoline and 8-hydroxyquinoline platforms has been synthesized. This sensor displays high selectivity and sensitive fluorescence enhancement to Cd(2+) in ethanol. Moreover, sensor HL can distinguish Cd(2+) from Zn(2+) via two different sensing mechanisms (photoinduced electron transfer for Cd(2+); internal charge transfer for Zn(2+)). The composition of the complex Cd(2+)/HL or Zn(2+)/L(-) has been found to be 1:1, based on the fluorescence/absorption titration and further confirmed by X-ray crystallography.     

Condensation of β,δ-triketone derived from dehydroacetic acid with aliphatic amines and copper(II) complexes of the schiff bases

The condensation of ethyl-2-hydroxy-4-oxo-4-(4-hydroxy-6-methyl-2-pyrone-3-yl)-2-butenoate with primary aliphatic monoamines occurs at 4-hydroxy group of the pyrone ring according to ¹HNMR and mass spectrometric data. The ketamine tautomeric form of the prepared Schiff bases is predominantly present in chloroform solutions. Isolated crude copper(II) complexes of the condensation products are shown to be mononuclear with the 1:2 metal to ligand ratio.     

Rational design of fluorescein-based fluorescence probes. Mechanism-based design of a maximum fluorescence probe for singlet oxygen.

Fluorescein is one of the best available fluorophores for biological applications, but the factors that control its fluorescence properties are not fully established. Thus, we initiated a study aimed at providing a strategy for rational design of functional fluorescence probes bearing fluorescein structure. We have synthesized various kinds of fluorescein derivatives and examined the relationship between their fluorescence properties and the highest occupied molecular orbital (HOMO) levels of their benzoic acid moieties obtained by semiempirical PM3 calculations. It was concluded that the fluorescence properties of fluorescein derivatives are controlled by a photoinduced electron transfer (PET) process from the benzoic acid moiety to the xanthene ring and that the threshold of fluorescence OFF/ON switching lies around -8.9 eV for the HOMO level of the benzoic acid moiety. This information provides the basis for a practical strategy for rational design of functional fluorescence probes to detect certain biomolecules. We used this approach to design and synthesize 9-[2-(3-carboxy-9,10-dimethyl)anthryl]-6-hydroxy-3H-xanthen-3-one (DMAX) as a singlet oxygen probe and confirmed that it is the most sensitive probe currently known for (1)O(2). This novel fluorescence probe has a 9,10-dimethylanthracene moiety as an extremely fast chemical trap of (1)O(2). As was expected from PM3 calculations, DMAX scarcely fluoresces, while DMAX endoperoxide (DMAX-EP) is strongly fluorescent. Further, DMAX reacts with (1)O(2) more rapidly, and its sensitivity is 53-fold higher than that of 9-[2-(3-carboxy-9,10-diphenyl)anthryl]-6-hydroxy-3H-xanthen-3-ones (DPAXs), which are a series of fluorescence probes for singlet oxygen that we recently developed. DMAX should be useful as a fluorescence probe for detecting (1)O(2) in a variety of biological systems.