In this thesis, the design, synthesis, photophysical, and biological properties
of four classes of phosphorescent iridium(III) polypyridine complexes are
reported. The potential applications of these complexes as intracellular
nitric oxide (NO) sensors, cellular sialic acid (Sia) sensors, cellular metabolic
indicators, and breast cancer cell imaging reagents are described.
The reactive, ubiquitous, and paramagnetic NO plays a key role in the
signaling processes of immune, cardiovascular, and nervous systems.
Importantly, unregulated NO production could lead to various diseases such
as cancer, septic shock, Alzheimer's disease, Parkinson's disease, and
Huntington's disease. Thus, the development of sensors for intracellular NO
is of paramount importance. In Chapter 2, the synthesis and
characterization of four 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) (1a), 2-phenylpyridine (Hppy)
(2a), 2-phenylquinoline (Hpq) (3a), 2-phenylcinchoninic acid methyl ester
(Hpqe) (4a)) are reported. 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 (1b), Hppy (2b), Hpq (3b), Hpqe (4b)) were also prepared.
The electrochemical and photophysical properties of the complexes were
studied. Upon photoexcitation, the diamine complexes displayed weak
green to red triplet metal-to-ligand charge-transfer (3MLCT) (dπ(Ir) →
π*(bpy-DA/pqe)) or triplet intraligand charge-transfer (3IL) (π → π*) (pq)
emission in fluid solutions at 298 K. The emission lifetimes of these
complexes were found to be concentration-dependent and their emission
quantum yields were considerably lower than those of their triazole
counterparts, indicative of the quenching properties of the appended
o-phenylenediamine unit. Upon protonation, the diamine complexes
displayed increased emission intensity but the emission properties of their
triazole counterparts were independent on the pH of the solution.
Additionally, the diamine complexes reacted readily with NO under aerated
conditions, resulting in substantial emission enhancement of the solution.
The reactions were found to be highly selective toward NO over other
reactive oxygen and nitrogen species (RONS). The lipophilicity and cellular
uptake efficiency of the diamine complexes were examined and correlated to
their molecular structures. These complexes were basically noncytotoxic
toward HeLa cells, as revealed by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-
diphenyltetrazolium bromide (MTT) assay. The cellular internalization and
intracellular NO-sensing properties of complex 2a were investigated by
laser-scanning confocal microscopy. The results showed that this complex
was localized in the cytoplasm and enriched in mitochondria. Furthermore,
it was capable of reacting with exogenous and endogenous NO in HeLa cells
and RAW 264.7 murine macrophages, respectively.
Sialic acids (Sia) are a collection of monosaccharides that are
composed of a nine-carbon backbone. These sugars are mainly found on
the cell surfaces as terminal units of glycoproteins and glycolipids, and are
involved in many biological processes. An imbalance in the expression of
Sia has been regarded as a marker of tumorigenic and metastatic processes
The targeting of Sia has relied on the use of lectins and antibodies, whereas
the use of transition metal complexes with phosphorescence properties is
extremely rare. In Chapter 3, the synthesis and characterization of three
phosphorescent cyclometalated iridium(III) polypyridine complexes bearing a
phenylboronic acid (PBA) moiety [Ir(N^C)2(bpy-TEG-PBA)](PF6).nHCl (n = 0
or 1) (bpy-TEG-PBA = 4-(N-(13-(3-boronobenzylamino)-4,7,10-trioxatridecanyl)
aminocarbonyl-oxy-methyl)-4'-methyl-2,2'-bipyridine; HN^C =
Hppy (1a), Hpq (2a), 2-phenyl-4-quinolinecarboxylic acid (Hpqa) (3a)) are
reported. Their boronic acid-free counterparts
[Ir(N^C)2(bpy-TEG-Bn)](PF6).nHCl (n = 0 or 1) (bpy-TEG-Bn =
4-(N-(13-benzylamino-(4,7,10-trioxa-tridecanyl))-aminocarbonyl)-oxy-methyl)
-4'-methyl-2,2'-bipyridine; HN^C = Hppy (1b), Hpq (2b), Hpqa (3b)) were also
prepared for comparison studies. The electrochemical and photophysical
properties of the complexes were studied. Upon photoexcitation, these
complexes showed intense and long-lived yellow to orange-red emission in
fluid solutions at 298 K, which originated from a 3MLCT (dπ(Ir) →
π*(N^N/N^C)) or an admixture of 3MLCT and 3IL (π → π*) (N^C) excited states.
Spectrophotometric titrations revealed that the PBA complexes were capable
of interacting with Sia and the binding affinity was stronger than other
saccharides such as D-glucose, D-galactose, and D-mannose, which are
commonly found on glycoproteins. The emission intensities of the pqa
complexes 3a and 3b were found to be pH-dependent in aqueous solution.
The apparent pKa values of these complexes were determined to be 4.57 and
4.37, respectively. The lipophilicity and cellular uptake efficiency of the
complexes were examined and correlated to their molecular structures.
Additionally, the cytotoxic activity of the complexes toward HepG2 cells was
evaluated by the MTT assay. Live-cell confocal images and ICP-MS
analyses showed that the ppy and pq complexes were efficiently taken up by
the cells whereas the pqa complexes were hardly internalized. The uptake
mechanism of the ppy and pq complexes was further investigated using
different metabolic inhibitors. Furthermore, the Sia-recognition properties of
the PBA complexes were examined by laser-scanning confocal microscopy
and ICP-MS.
Glucose is a crucial source of energy for mammalian cells.
Fast-growing cancer cells usually exhibit enhanced metabolic rates as
reflected by their faster glucose uptake rates compared with normal cells.
Thus, the development of glucose-based metabolic indicators for cancer cells
has attracted much attention. In Chapter 4, the synthesis and
characterization of a series of phosphorescent cyclometalated iridium(III)
polypyridine complexes appended with a β-D-glucose moiety
[Ir(N^C)2(bpy-TEG-ONCH3-β-D-glc)](PF6) (bpy-TEG-ONCH3-β-D-glc = 4-(10-
N-methyl-N-(β-D-glucopyranosyl)-amino-oxy-2,5,8-trioxa-dec-1-yl)-4'-methyl-
2,2'-bipyridine; HN^C = 2-phenylbenzothiazole (Hbt) (1a), Hppy (2a), Hpq)
(3a), 7,8-benzoquinoline (Hbzq) (4a)) are reported. The D-galactose
analogues [Ir(N^C)2(bpy-TEG-ONCH3-β-D-gal)](PF6) (bpy-TEGONCH3-
β-D-gal = 4-(10-N-methyl-N-(β-D-galactopyranosyl)-aminooxy-
2,5,8-trioxa-dec-1-yl)-4'-methyl-2,2'-bipyridine; HN^C = Hbt (1b), Hppy
(2b), Hpq (3b), Hbzq (4b)) and a sugar-free bt complex
[Ir(bt)2(bpy-TEG-OMe)](PF6) (bpy-TEG-OMe = 4-(2,5,8,11-tetraoxa-dodec-1-
yl)-4'-methyl-2,2'-bipyridine) (1c) were also prepared. Upon photoexcitation,
these complexes displayed intense and long-lived a 3MLCT (dπ(Ir) → π*(N^N))
or 3IL (π → π*) (N^C) emission in fluid solutions at 298 K and in alcohol glass
at 77 K. The lipophilicity of all the complexes was determined by
reversed-phase HPLC. Additionally, the cellular uptake efficiency and
cytotoxicity of the complexes toward HeLa cells were examined by ICP-MS
and the MTT assay, respectively. The results showed that the cytotoxicity of
the complexes was generally correlated to their lipophilicity and cellular
uptake efficiency. Temperature dependence and chemical inhibition
experiments indicated that the transport of bt−glucose complex 1a across the
cell membrane occurred through an energy-requiring process such as
endocytosis, in additional to a pathway that was mediated by glucose
transporters (GLUTs). Additionally, the cellular uptake efficiency of this
complex was found to be strongly dependent on hormonal stimulation and
inhibition. Furthermore, laser-scanning confocal microscopy revealed that
the complex was localized in the mitochondria and was highly resistant to
photobleaching compared to a fluorescent organic glucose derivative
2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxy-D-glucose (2-NBDG).
The substrates of GLUTs are not limited to glucose; fructose has been
identified to be selectively facilitated transported by GLUT5, which is
significantly expressed in breast cancer tissues compared with other cancer
cells and normal breast tissues. Importantly, luminescent fructose
derivatives that exhibit photocytotoxic activity and high specificity toward
breast cancer cells are useful in the development of photoindcued cytotoxic
reagents for breast cancer cells. In Chapter 5, the synthesis and
characterization of two novel phosphorescent cyclometalated iridium(III)
polypyridine complexes with a fructose pendant [Ir(N^C)2(bpy-fru)](PF6)
(bpy-fru = 4-(1-deoxy-D-fructose-1-yl)aminoacarbonyl-4'-methyl-2,2'-
bipyridine; HN^C = Hppy (1a), Hpq (2a)) are reported. The fructose-free
analogues [Ir(N^C)2(bpy-Bu)](PF6) (bpy-Bu = 4-N-(n-butylaminocarbonyl)-
4'-methyl-2,2'-bipyridine; HN^C = Hppy (1b), Hpq (2b)) were also prepared
for comparison studies. Upon photoexcitation, all the complexes exhibited
yellow to orange emission originating from 3MLCT (dπ(Ir) → π*(N^N)) or an
admixture of 3MLCT and 3IL (π → π*) (N^C) excited states in fluid solutions at
298 K and in alcohol glass at 77 K. The lipophilicity and cellular uptake of
the complexes were examined and correlated to their molecular structures.
The fructose complexes 1a and 2a were much less cytotoxic toward MCF-7
cells compared to their fructose-free counterparts 1b and 2b, as revealed by
the MTT assay. Additionally, the fructose complexes were found to exhibit
photocytotoxic activity upon irradiation. Experiments involving
1,5-dihydroxynaphthalene (DHN) showed that this activity was due to the
generation of singlet oxygen. Laser-scanning confocal microscopy images
indicated that the internalization of the fructose complex 2a toward MCF-7
cells involved an energy-requiring process, and the complex was enriched in
the mitochondria of the cells. The uptake of this complex was also mediated
by fructose transporters, as revealed by the confocal images and ICP-MS
analyses involving chemical inhibition experiments. Furthermore, cell
line-dependence and coculture experiments showed that the accumulation of
the complex was selective toward MCF-7 and MDA-MB-231 breast cancer
cells.
| Date of Award | 15 Jul 2015 |
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| Original language | English |
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| Awarding Institution | - City University of Hong Kong
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| Supervisor | Kam Wing Kenneth LO (Supervisor) |
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