Hua-Wei Liu

Luminescent Cyclometalated Iridium(III) Polypyridine Di-2-picolylamine Complexes: Synthesis, Photophysics, Electrochemistry, Cation Binding, Cellular Internalization, and Cytotoxic Activity

A series of luminescent cyclometalated iridium(III) polypyridine complexes containing a di-2-picolylamine (DPA) moiety [Ir(N^C)(2)(phen-DPA)](PF(6)) (phen-DPA = 5-(di-2-picolylamino)-1,10-phenanthroline) (HN^C = 2-phenylpyridine, Hppy (1a), 2-(4-methylphenyl)pyridine, Hmppy (2a), 2-phenylquinoline, Hpq (3a), 4-(2-pyridyl)benzaldehyde, Hpba (4a)) and their DPA-free counterparts [Ir(N^C)(2)(phen-DMA)](PF(6)) (phen-DMA = 5-(dimethylamino)-1,10-phenanthroline) (HN^C = Hppy (1b), Hmppy (2b), Hpq (3b), Hpba (4b)) have been synthesized and characterized, and their photophysical and electrochemical properties investigated. Photoexcitation of the complexes in fluid solutions at 298 K and in alcohol glass at 77 K resulted in intense and long-lived luminescence. The emission of the complexes has been assigned to a triplet metal-to-ligand charge-transfer ((3)MLCT) (dπ(Ir) → π*(N^N)) or triplet intraligand ((3)IL) (π → π*) (N^C) excited state and with substantial mixing of triplet amine-to-ligand charge-transfer ((3)NLCT) (n → π*) (N^N) character, depending on the identity of the cyclometalating and diimine ligands. Electrochemical measurements revealed an irreversible amine oxidation wave at ca. +1.1 to +1.2 V vs saturated calomel electrode, a quasi-reversible iridium(IV/III) couple at ca. +1.2 to +1.6 V, and a reversible diimine reduction couple at ca. -1.4 to -1.5 V. The cation-binding properties of these complexes have been studied by emission spectroscopy. Upon binding of zinc ion, the iridium(III) DPA complexes displayed 1.2- to 5.4-fold emission enhancement, and the K(d) values determined were on the order of 10(-5) M. Jobs plot analysis confirmed that the binding stoichiometry was 1:1. Additionally, selectivity studies showed that the iridium(III) DPA complexes were more sensitive toward zinc ion among various transition metal ions examined. Furthermore, the cytotoxicity of these complexes toward human cervix epithelioid carcinoma cells have been studied by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide assay and their cellular-uptake properties by inductively coupled plasma mass spectrometry and laser-scanning confocal microscopy.

Luminescent Dendritic Cyclometalated Iridium(III) Polypyridine Complexes: Synthesis, Emission Behavior, and Biological Properties

Luminescent dendritic cyclometalated iridium(III) polypyridine complexes [{Ir(N--C)(2)}(n)(bpy-n)](PF(6))(n) (HN--C = 2-phenylpyridine, Hppy, n = 8 (ppy-8), 4 (ppy-4), 3 (ppy-3); HN--C = 2-phenylquinoline, Hpq, n = 8 (pq-8), 4 (pq-4), 3 (pq-3)) have been designed and synthesized. The properties of these dendrimers have been compared to those of their monomeric counterparts [Ir(N--C)(2)(bpy-1)](PF(6)) (HN--C = Hppy (ppy-1), Hpq (pq-1)). Cyclic voltammetric studies revealed that the iridium(IV/III) oxidation and bpy-based reduction occurred at about +1.24 to +1.29 V and -1.21 to -1.27 V versus SCE, respectively, for all the complexes. The molar absorptivity of the dendritic iridium(III) complexes is approximately proportional to the number of [Ir(N--C)(2)(N--N)] moieties in one complex molecule. However, the emission lifetimes and quantum yields are relatively independent of the number of [Ir(N--C)(2)(N--N)] units, suggesting negligible electronic communications between these units. Upon photoexcitation, the complexes displayed triplet metal-to-ligand charge-transfer ((3)MLCT) (dpi(Ir) --> pi*(bpy-n)) emission. The interaction of these complexes with plasmid DNA has been investigated by agarose gel retardation assays. The results showed that the dendritic iridium(III) complexes, unlike their monomeric counterparts, bound to the plasmid, and the interaction was electrostatic in nature. The lipophilicity of all the complexes has been determined by reversed-phase high-performance liquid chromatography (HPLC). Additionally, the cellular uptake of the complexes by the human cervix epithelioid carcinoma (HeLa) cell line has been examined by inductively coupled plasma mass spectrometry (ICP-MS), laser-scanning confocal microscopy, and flow cytometry. Upon internalization, all the complexes were localized in the perinuclear region, forming very sharp luminescent rings surrounding the nuclei. Interestingly, in addition to these rings, HeLa cells treated with the dendritic iridium(III) complexes showed specific labeled compartments, which have been identified to be the Golgi apparatus. Furthermore, the cytotoxicity of these iridium(III) complexes has been evaluated by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT) assay.

Luminescent rhenium(I) polypyridine complexes appended with an α-D-glucose moiety as novel biomolecular and cellular probes.

Glucose is the most important carbohydrate in cellular metabolism and an energy source for the growth of cells. One of the most characteristic phenotypes of rapidly growing cancer cells is their propensity to catabolize glucose at high rates, possibly due to the overexpression of glucose transporters (GLUTs). Thus, the in vitro and in vivo monitoring of glucose utilization in cancer cells has attracted much attention. Different reporting and therapeutic units such as radioactive labels, IRDye 800CW, organic fluorophores, and two-photon dyes have been conjugated to glucose or 2-deoxyglucose for the diagnosis and treatment of various tumors or cancers. Despite the development of these reagents, the possibility of using luminescent transition-metal glucose conjugates as glucoseuptake tracers and cancer cell imaging reagents has not been explored. With our ongoing interest in luminescent rhenium(I) polypyridine complexes as biological probes, we envisage that modification of these complexes with an ad-glucose pendant will generate useful luminescent probes for biomolecules and cancer cells. Herein we report three rhenium(I) polypyridine glucose complexes [Re ACHTUNGTRENNUNG(N^N)(CO)3ACHTUNGTRENNUNG(py-3-glu)] ACHTUNGTRENNUNG(PF6) (py-3-glu =3(N-(6-(N’-(4-(a-d-glucopyranosyl)phenyl)thioureidyl)hexyl)thioureidyl)pyridine, N^N=1,10-phenanthroline (phen) (1), 3,4,7,8-tetramethyl-1,10-phenanthroline (Me4-phen) (2), 4,7-diphenyl-1,10-phenanthroline (Ph2-phen) (3)) and their glucose-free counterparts [Re ACHTUNGTRENNUNG(N^N)(CO)3ACHTUNGTRENNUNG(py-3-Et)]ACHTUNGTRENNUNG(CF3SO3) (py-3-Et=3-(ethylthioureidyl)pyridine, N^N= phen (1 a), Me4-phen (2 a), Ph2-phen (3 a)) (Scheme 1). The glucose complexes were synthesized from the addition reaction of the isothiocyanate complexes [Re ACHTUNGTRENNUNG(N^N)(CO)3ACHTUNGTRENNUNG(py-3NCS)] ACHTUNGTRENNUNG(PF6)[5a] with AcO-glu-C6-NH2 in acetone, followed by deacetylation (see the Supporting Information, Schemes S1 and S2). All the complexes were characterized by H NMR spectroscopy, positive-ion ESI-MS, and IR spectroscopy and gave satisfactory elemental analyses (see the Supporting Information). Upon irradiation, the complexes exhibited intense and long-lived green-to-yellow triplet metal-to-ligand charge-transfer (MLCT; dp(Re)!p* ACHTUNGTRENNUNG(N^N)) emission (see the Supporting Information, Table S1). The structured emission band and very long lifetime of complex 2 in alcohol glass at 77 K are probably due to the involvement of IL (p!p*) (Me4-phen) character in its emissive state. Since the lectin concanavalin A (Con A) binds a-d-mannopyranoside and a-d-glucopyranoside, the possible use of the glucose complexes 1–3 as a luminescent sensor for this lectin has been investigated. Upon addition of Con A to a [a] M.-W. Louie, Dr. H.-W. Liu, M. H.-C. Lam, Dr. Y.-W. Lam, Dr. K. K.-W. Lo Department of Biology and Chemistry City University of Hong Kong, Tat Chee Avenue Kowloon, Hong Kong (P.R. China) Fax: (+852) 3442-0522 E-mail : bhkenlo@cityu.edu.hk Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/chem.201101399. Scheme 1. Structures of the rhenium(I) polypyridine complexes.

Dual-Emissive Cyclometalated Iridium(III) Polypyridine Complexes as Ratiometric Biological Probes and Organelle-Selective Bioimaging Reagents

In this Article, we present a series of cyclometalated iridium(III) polypyridine complexes of the formula [Ir(N^C)2(N^N)](PF6) that showed dual emission under ambient conditions. The structures of the cyclometalating and diimine ligands were changed systematically to investigate the effects of the substituents on the dual-emission properties of the complexes. On the basis of the photophysical data, the high-energy (HE) and low-energy (LE) emission features of the complexes were assigned to triplet intraligand ((3)IL) and triplet charge-transfer ((3)CT) excited states, respectively. Time-dependent density functional theory (TD-DFT) calculations supported these assignments and indicated that the dual emission resulted from the interruption of the communication between the higher-lying (3)IL and the lower-lying (3)CT states by a triplet amine-to-ligand charge-transfer ((3)NLCT) state. Also, the avidin-binding properties of the biotin complexes were studied by emission titrations, and the results showed that the dual-emissive complexes can be utilized as ratiometric probes for avidin. Additionally, all the complexes exhibited efficient cellular uptake by live HeLa cells. The MTT and Annexin V assays confirmed that no cell death and early apoptosis occurred during the cell imaging experiments. Interestingly, laser-scanning confocal microscopy revealed that the complexes were selectively localized on the cell membrane, mitochondria, or both, depending on the nature of the substituents of the ligands. The results of this work will contribute to the future development of dual-emissive transition metal complexes as ratiometric probes and organelle-selective bioimaging reagents.

Photophysical and cellular uptake properties of novel phosphorescent cyclometalated iridium(III) bipyridine D-fructose complexes.

We report here the synthesis, characterization and photophysical properties of two novel phosphorescent cyclometalated iridium(III) polypyridine D-fructose complexes and their fructose-free counterparts. The cellular uptake of the complexes and their cytotoxicity have also been examined.

Conferring Phosphorogenic Properties on Iridium(III)‐Based Bioorthogonal Probes through Modification with a Nitrone Unit

The use of bioorthogonal probes that display fluorogenic or phosphorogenic properties is advantageous to the labeling and imaging of biomolecules in live cells and organisms. Herein we present the design of three iridium(III) complexes containing a nitrone moiety as novel phosphorogenic bioorthogonal probes. These probes were non-emissive owing to isomerization of the C=N group but showed significant emission enhancement upon cycloaddition reaction with strained cyclooctynes. Interestingly, the connection of the nitrone ligand to the cationic iridium(III) center led to accelerated reaction kinetics. These nitrone complexes were also identified as phosphorogenic bioorthogonal labels and imaging reagents for cyclooctyne-modified proteins. These findings contribute to the development of phosphorogenic bioorthogonal probes and imaging reagents.

Rhenium(I) Polypyridine Diamine Complexes as Intracellular Phosphorogenic Sensors: Synthesis, Characterization, Emissive Behavior, Biological Properties, and Nitric Oxide Sensing

We report the development of a series of rhenium(I) polypyridine complexes appended with an electron-rich diaminoaromatic moiety as phosphorogenic sensors for nitric oxide (NO). The diamine complexes [Re(N^N)(CO)3 (py-DA)][PF6 ] (py-DA=3-(N-(2-amino-5-methoxyphenyl)aminomethyl)pyridine; N^N=1,10-phenanthroline (phen) (1 a), 3,4,7,8-tetramethyl-1,10-phenanthroline (Me4 -phen) (2 a), 4,7-diphenyl-1,10-phenanthroline (Ph2 -phen) (3 a)) have been synthesized and characterized. In contrast to common rhenium(I) diimines, these diamine complexes were very weakly emissive due to quenching of the triplet metal-to-ligand charge-transfer ((3) MLCT) emission by the diaminoaromatic moiety through photoinduced electron transfer (PET). Upon treatment with NO, the complexes were converted into the triazole derivatives [Re(N^N)(CO)3 (py-triazole)][PF6 ] (py-triazole=3-((6-methoxybenzotriazol-1-yl)methyl)pyridine; N^N=phen (1 b), Me4 -phen (2 b), Ph2 -phen (3 b)), resulting in significant emission enhancement (I/I0 ≈60). The diamine complexes exhibited high reaction selectivity to NO, and their emission intensity was found to be independent on pH. Also, these complexes were effectively internalized by HeLa cells and RAW264.7 macrophages with negligible cytotoxicity. Additionally, the use of complex 3 a as an intracellular phosphorogenic sensor for NO has been demonstrated.

Rhenium(I) polypyridine complexes functionalized with a diaminoaromatic moiety as phosphorescent sensors for nitric oxide

A series of rhenium(I) polypyridine complexes functionalized with a diaminoaromatic moiety has been developed as phosphorescent sensors for nitric oxide (NO). The diamine complexes [Re(N⁁N)(CO)3(py-diamine)](CF3SO3) [py-diamine = 3,4-diaminopyridine; N⁁N = 1,10-phenanthroline (phen) (1a), 2,9-dimethyl-1,10-phenanthroline (Me2-phen) (2a), 3,4,7,8-tetramethyl-1,10-phenanthroline (Me4-phen) (3a), 4,7-diphenyl-1,10-phenanthroline (Ph2-phen) (4a)] were synthesized and characterized. These complexes were only weakly emissive due to the diaminoaromatic moiety that quenches the 3MLCT [dπ(Re) → π*(N⁁N)] emission by photoinduced electron transfer (PET). However, in the presence of NO, these diamine complexes were converted to the triazole derivatives [Re(N⁁N)(CO)3(py-triazole)](CF3SO3) [py-triazole = 1H-1,2,3-triazolo[4,5,c]pyridine; N⁁N = phen (1b), Me2-phen (2b), Me4-phen (3b), Ph2-phen (4b)], which revealed intense emission upon excitation. The emission of the complexes was independent to pH under neutral and basic conditions (pH > 6). The cytotoxicity and cellular uptake properties of these complexes were studied by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT) assay and ICP-MS, respectively. The potential application of these complexes as intracellular NO sensors was also investigated.

Cyclometalated Iridium(III) Bipyridyl–Phenylenediamine Complexes with Multicolor Phosphorescence: Synthesis, Electrochemistry, Photophysics, and Intracellular Nitric Oxide Sensing

We present a new class of phosphorescent cyclometalated iridium(III) bipyridyl–phenylenediamine complexes [Ir(N^C)2(bpy‐DA)](PF6) (bpy‐DA=4‐(N‐(2‐amino‐5‐methoxyphenyl)aminomethyl)‐4′‐methyl‐2,2′‐bipyridine; HN^C=2‐(2,4‐difluorophenyl)pyridine (Hdfppy) (1 a), 2‐phenylpyridine (Hppy) (2 a), 2‐phenylquinoline (Hpq) (3 a), 2‐phenylcinchoninic acid methyl ester (Hpqe) (4 a)) and their triazole counterparts [Ir(N^C)2(bpy‐T)](PF6) (bpy‐T=4‐((6‐methoxybenzotriazol‐1‐yl)methyl)‐4′‐methyl‐2,2′‐bipyridine; HN^C=Hdfppy (1 b), Hppy (2 b), Hpq (3 b), Hpqe (4 b)). Upon photoexcitation, the diamine complexes exhibited fairly weak green to red phosphorescence under ambient conditions whereas the triazole derivatives emitted strongly. The photophysical properties of complexes 2 a and 2 b have been studied in more detail. Upon protonation, the diamine complex 2 a displayed increased emission intensity, but the emission properties of its triazole counterpart complex 2 b were independent on the pH value of the solution. Also, complex 2 a was found to be readily converted into complex 2 b upon reaction with NO under aerated conditions, resulting in substantial emission enhancement of the solution. The reaction was highly specific toward NO over other reactive oxygen and nitrogen species (RONS) as revealed by spectroscopic analyses. The lipophilicity and cellular uptake efficiency of the diamine complexes have been examined and correlated to their molecular structures. Also, cell‐based assays showed that these complexes were noncytotoxic toward human cervix epithelioid carcinoma (HeLa) cells (at 10 μM, 4 h, percentage survival ≈80–95 %). Additionally, the diamine complexes have been used to visualize intracellular NO generated both exogenously in HeLa cells and endogenously in RAW 264.7 murine macrophages by laser‐scanning confocal microscopy.

Rhenium(I) polypyridine dibenzocyclooctyne complexes as phosphorescent bioorthogonal probes: Synthesis, characterization, emissive behavior, and biolabeling properties.

We report the development of rhenium(I) polypyridine complexes appended with a dibenzocyclooctyne (DIBO) moiety as bioorthogonal probes for azide-modified biomolecules. Three phosphorescent rhenium(I) polypyridine DIBO complexes [Re(N^N)(CO)3(py-C6-DIBO)][CF3SO3] (py-C6-DIBO=3-(N-(6-(3,4:7,8-dibenzocyclooctyne-5-oxycarbonylamino)hexyl)aminocarbonyl)pyridine; N^N=1,10-phenanthroline (phen) (1a), 3,4,7,8-tetramethyl-1,10-phenanthroline (Me4-phen) (2a), 4,7-diphenyl-1,10-phenanthroline (Ph2-phen) (3a)) and their DIBO-free counterparts [Re(N^N)(CO)3(py-C6-BOC)][CF3SO3] (py-C6-BOC=3-(N-(6-(tert-butoxycarbonylamino)hexyl)aminocarbonyl)pyridine; N^N=phen (1b), Me4-phen (2b), Ph2-phen (3b)) were synthesized and characterized. Upon photoexcitation, all the complexes displayed intense and long-lived yellow triplet metal-to-ligand charge-transfer ((3)MLCT) (dπ(Re)→π*(N^N)) emission. The DIBO complexes underwent facile reactions with benzyl azide in methanol at 298 K with second-order rate constants (k2) in the range of 0.077 to 0.091 M(-1) s(-1). As revealed from SDS-PAGE analysis, the DIBO complexes can selectively label azide-modified proteins and the resulting bioconjugates displayed strong phosphorescence upon photoexcitation. Results of inductively coupled plasma mass spectrometry (ICP-MS) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays indicated that the DIBO complexes accumulated in Chinese Hamster Ovary (CHO) cells with considerable cytotoxic activity. Upon incubation of CHO cells with these complexes, relatively weak intracellular emission was observed. In contrast, upon pretreatment of the cells with 1,3,4,6-tetra-O-acetyl-N-azidoacetyl-D-mannosamine (Ac4ManNAz), intense emission was observed from the cell membrane and some internal compartments. The results suggest that the DIBO complexes are promising candidates for imaging azide-labeled biomolecules.