Hydrogen-Bonded Thiol Undergoes Unconventional Excited-State Intramolecular Proton-Transfer Reactions.

The chapter on the thiol-related hydrogen bond (H-bond) and its excited-state intramolecular proton-transfer (ESIPT) reaction was recently opened where compound 4'-diethylamino-3-mercaptoflavone (3NTF) undergoes ESIPT in both cyclohexane solution and solid, giving a 710 nm tautomer emission with an anomalously large Stokes shift of 12,230 cm-1. Considering the thiol H-bond to be unconventional compared to the conventional Pauling-type -OH or -NH H-bond, it is thus essential and timely to probe its fundamental difference between their ESIPT. However, thiol-associated ESIPT tends to be nonemissive due to the dominant nπ* character of the tautomeric lowest excited state. Herein, based on the 3-mercaptoflavone scaffold and π-elongation concept, a new series of 4'-substituted-7-diethylamino-3-mercaptoflavones, NTFs, was designed and synthesized with varied H-bond strength and 690-720 nm tautomeric emission upon ultraviolet (UV) excitation in cyclohexane. The order of their H-bonding strength was experimentally determined to be N-NTF < O-NTF < H-NTF < F-NTF, while the rate of -SH ESIPT measured by fluorescence upconversion was F-NTF (398 fs)-1 < H-NTF (232 fs)-1 < O-NTF (123 fs)-1 < N-NTF (101 fs)-1 in toluene. Unexpectedly, the strongest H-bonded F-NTF gives the slowest ESIPT, which does not conform to the traditional ESIPT model. The results are rationalized by the trend of carbonyl oxygen basicity rather than -SH acidity. Namely, the thiol acidity relevant to the H-bond strength plays a minor role in the driving force of ESIPT. Instead, the proton-accepting strength governs ESIPT. That is to say, the noncanonical thiol H-bonding system undergoes an unconventional type of ESIPT.

Tuning intramolecular charge transfer and spin-orbit coupling of AIE-active type-I photosensitizers for photodynamic therapy.

Development of photosensitizers (PSs) featuring type-I reactive oxygen species (ROS) with aggregation-induced emission (AIE) properties is a judicious approach to overcome the deficit of conventional photodynamic therapy (PDT). However, it remains a challenge to design AIE-active type-I ROS PSs using a simple theranostic scaffold paired with a delicate balance between intramolecular charge transfer (ICT) and large spin-orbit coupling (SOC) features to facilitate intersystem crossing (ISC) and hence to intensify triplet excitons for type-I ROS generation as well as to improve optical properties for the desired biomedical applications. In this work, a rationally designed series of PSs based on C-6-substituted tetraphenylethylene-fused benzothiazole-coumarin scaffolds, named TPE-nCUMs, were synthesized via a fused-ring-electron-acceptor (FREA) strategy, endowed with AIE properties in aqueous solution and thus self-monitoring type-I ROS generation under white-light irradiation to study the effects of diverse ICT and SOC potentials on their photochemical and optical properties. Both experimental and theoretical results revealed that the concomitantly increasing strengths of both ICT and SOC features promote type-I ROS generation by TPE-nCUMs. The key role of the SOC-promoting carbonyl group towards the ROS generation ability of TPE-nCUMs was then examined. Among TPE-nCUMs, gem-2OMe-TPE-2CUM displayed highly efficient type-I ROS generation. Importantly, gem-OMe-TPE-1CUM acts as a fluorescent indicator in HeLa cells (in vitro), revealing its excellent diffusion capability in both lysosomal and mitochondrial organelles with low dark toxicity, high cytotoxicity under white-light and remarkable PDT efficiency. Our study has thus elucidated a rationally designed strategy at the molecular level to fine-tune ICT and SOC features for the advance of AIE-active type-I ROS PSs, opening a new avenue for cancer treatment and image-guided therapy.

Tailoring C-6-Substituted Coumarin Scaffolds for Novel Photophysical Properties and Stimuli-Responsive Chromism.

A judicious strategy was utilized to envision the substantial regio-positional effects of substituents on the photophysical properties of the 2H-chromen-2-one-3-benzothiazole scaffold-based push-pull framework, named 6-X-CUMs. Among them, 6-NEt2-CUM reveals prominent excited-state intramolecular charge transfer with a large change of dipole moment (Δµ ∼ 18.23 D), hence displaying remarkable emission solvatochromism from the green (536 nm in cyclohexane) to far-red region (714 nm in dimethyl sulfoxide) and a high-temperature sensitivity (-0.23 nm °C-1). These, together with unique basicity and acido-/vaporchromism upon acidification elucidated by NMR and photospectroscopic studies, show stark contrast to the conventional 7-NEt2-CUM. The new series of these tailored 6-X-CUMs represents a new dimension in tailoring the photophysical properties for the development of a promising class of multistimuli-responsive materials.

Chapter Open for the Excited-State Intramolecular Thiol Proton Transfer in the Room-Temperature Solution.

We report here, for the first time, the experimental observation on the excited-state intramolecular proton transfer (ESIPT) reaction of the thiol proton in room-temperature solution. This phenomenon is demonstrated by a derivative of 3-thiolflavone (3TF), namely, 2-(4-(diethylamino)phenyl)-3-mercapto-4H-chromen-4-one (3NTF), which possesses an -S-H···O═ intramolecular H-bond (denoted by the dashed line) and has an S1 absorption at 383 nm. Upon photoexcitation, 3NTF exhibits a distinctly red emission maximized at 710 nm in cyclohexane with an anomalously large Stokes shift of 12 230 cm-1. Upon methylation on the thiol group, 3MeNTF, lacking the thiol proton, exhibits a normal Stokes-shifted emission at 472 nm. These, in combination with the computational approaches, lead to the conclusion of thiol-type ESIPT unambiguously. Further time-resolved study renders an unresolvable (<180 fs) ESIPT rate for 3NTF, followed by a tautomer emission lifetime of 120 ps. In sharp contrast to 3NTF, both 3TF and 3-mercapto-2-(4-(trifluoromethyl)phenyl)-4H-chromen-4-one (3FTF) are non-emissive. Detailed computational approaches indicate that all studied thiols undergo thermally favorable ESIPT. However, once forming the proton-transferred tautomer, the lone-pair electrons on the sulfur atom brings non-negligible nπ* contribution to the S1' state (prime indicates the proton-transferred tautomer), for which the relaxation is dominated by the non-radiative deactivation. For 3NTF, the extension of π-electron delocalization by the diethylamino electron-donating group endows the S1' state primarily in the ππ* configuration, exhibiting the prominent tautomer emission. The results open a new chapter in the field of ESIPT, covering the non-canonical sulfur intramolecular H-bond and its associated ESIPT at ambient temperature.

Simple Molecular-Engineering Approach for Enhancing Orientation and Outcoupling Efficiency of Thermally Activated Delayed Fluorescent Emitters without Red-Shifting Emission.

The inclusion of a tetraphenylbenzene (4Ph) unit in thermally activated delayed fluorescence emitters is demonstrated as a novel strategy for greatly enhancing the horizontally oriented alignment of the emitters without shifting the emission spectrum to longer wavelengths. Doping of blue-emitting 4PhOXDDMAC or greenish-blue-emitting 4PhOXDPXZ into o-DiCbzBz host layers yielded much higher degrees of horizontally oriented alignment for the emitter (up to 92%) compared to those when the 4Ph unit was excluded (69 and 75%, respectively). The enhanced alignment results in high outcoupling efficiencies of 24 and 35% in organic light-emitting diodes based on 4PhOXDDMAC and 4PhOXDPXZ, respectively, and boosts the external quantum efficiencies to values (8.8 and 29.2%, respectively) that are higher than what would be expected for randomly oriented emitters (outcoupling efficiency of 20%). These enhancements are achieved while avoiding the redshift that often occurs using the common strategy of increasing molecular length and, thereby, conjugation, to increase orientation.

SimCP3-An Advanced Homologue of SimCP2 as a Solution-Processed Small Molecular Host Material for Blue Phosphorescence Organic Light-Emitting Diodes.

We have overcome the synthetic difficulty of 9,9',9'',9''',9'''',9'''''-((phenylsilanetriyl)tris(benzene-5,3,1-triyl))hexakis(9H-carbazole) (SimCP3) an advanced homologue of previously known SimCP2 as a solution-processed, high triplet gap energy host material for a blue phosphorescence dopant. A series of organic light-emitting diodes based on blue phosphorescence dopant iridium (III) bis(4,6-difluorophenylpyridinato)picolate, FIrpic, were fabricated and tested to demonstrate the validity of solution-processed SimCP3 in the device fabrication.

Delineation of G-Quadruplex Alkylation Sites Mediated by 3,6-Bis(1-methyl-4-vinylpyridinium iodide)carbazole-Aniline Mustard Conjugates.

A new G-quadruplex (G-4)-directing alkylating agent BMVC-C3M was designed and synthesized to integrate 3,6-bis(1-methyl-4-vinylpyridinium iodide)carbazole (BMVC) with aniline mustard. Various telomeric G-4 structures (hybrid-2 type and antiparallel) and an oncogene promoter, c-MYC (parallel), were constructed to react with BMVC-C3M, yielding 35 % alkylation yield toward G-4 DNA over other DNA categories (<6 %) and high specificity under competition conditions. Analysis of the intact alkylation adducts by electrospray ionization mass spectroscopy (ESI-MS) revealed the stepwise DNA alkylation mechanism of aniline mustard for the first time. Furthermore, the monoalkylation sites and intrastrand cross-linking sites were determined and found to be dependent on G-4 topology based on the results of footprinting analysis in combination with mass spectroscopic techniques and in silico modeling. The results indicated that BMVC-C3M preferentially alkylated at A15 (H26), G12 (H24), and G2 (c-MYC), respectively, as monoalkylated adducts and formed A15-C3M-A21 (H26), G12-C3M-G4 (H24), and G2-C3M-G4/G17 (c-MYC), respectively, as cross-linked dialkylated adducts. Collectively, the stability and site-selective cross-linking capacity of BMVC-C3M provides a credible tool for the structural and functional characterization of G-4 DNAs in biological systems.

Synthesis and characterization of heteroatom-bridged bis-spirobifluorenes for the application of organic light-emitting diodes.

Pure 2-iodo-9,9'-spirobifluorene was synthesized by an efficient method without troublesome iodination of 9,9-spirobifluorene (SP) or the Sandmeyer reaction of 2-amino-9,9'-spirobifluorene. A series of main group element-bridged bis-9,9'-spirobifluorene derivatives were synthesized via coupling reactions of 2-iodo-9,9'-spirobifluorene and main group element-containing precursors. These heteroatom-bridged bis-spirobifluorenes show large triplet state energy gaps, high glass transition temperatures, and varied charge-transporting properties advantageous to the host materials for blue phosphorescence organic light-emitting diodes.

Reaction-based and single fluorescent emitter decorated ratiometric nanoprobe to detect hydrogen peroxide.

A novel reaction-based cross-linked polymeric nanoprobe with a self-calibrating ratiometric fluorescence readout to selectively detect H2O2 is reported. The polymeric nanoprobe is fabricated by using hydrophobic H2O2-reactive boronic ester groups, crosslinker units, and environmentally sensitive 3-hydroxyflavone fluorophores through a miniemulsion polymerization. On treatment with H2O2, the boronic esters in the polymer are cleaved to form hydrophilic alcohols and subsequently lead to a hydrophobic-hydrophilic transition. Covalently linked 3-hydroxyflavones manifest the change in polarity as a ratiometric transition from green to blue, accompanied by a 500-fold increase in volume. Furthermore, this nanoprobe has been used for ratiometric sensing of glucose by monitoring the H2O2 generated during the oxidation of glucose by glucose oxidase, and thus successfully distinguished between normal and pathological levels of glucose.

Facile fabrication of dextran-based fluorescent nanogels as potential glucose sensors.

Glucose-responsive nanogels based on dextran and poly(3-acrylamidophenylboronic acid) (PAAPBA) were fabricated by a facile self-assembly assisted (SAA) strategy. Further introduction of the fluorescent agent 2-[4-(3-hydroxy-4-oxo-4H-chromen-2-yl)phenoxy]ethylacrylamide (3HF-AM) allowed visualization of this glucose sensitivity, thus ensuring their potential use as glucose sensors.

Pregnenolone activates CLIP-170 to promote microtubule growth and cell migration.

Pregnenolone (P5) is a neurosteroid that improves memory and neurological recovery. It is also required for zebrafish embryonic development. However, its mode of action is unclear. Here we show that P5 promotes cell migration and microtubule polymerization by binding a microtubule plus end-tracking protein, cytoplasmic linker protein 1 (CLIP-170). We captured CLIP-170 from zebrafish embryonic extract using a P5 photoaffinity probe conjugated to diaminobenzophenone. P5 interacted with CLIP-170 at its coiled-coil domain and changed it into an extended conformation. This increased CLIP-170 interaction with microtubules, dynactin subunit p150(Glued) and LIS1; it also promoted CLIP-170-dependent microtubule polymerization. CLIP-170 was essential for P5 to promote microtubule abundance and zebrafish epiboly cell migration during embryogenesis, and overexpression of the P5-binding region of CLIP-170 delayed this migration. P5 also sustained migration directionality of cultured mammalian cells. Our results show that P5 activates CLIP-170 to promote microtubule polymerization and cell migration.

A new strategy for intracellular delivery of enzyme using mesoporous silica nanoparticles: superoxide dismutase.

We developed mesoporous silica nanoparticle (MSN) as a multifunctional vehicle for enzyme delivery. Enhanced transmembrane delivery of a superoxide dismutase (SOD) enzyme embedded in MSN was demonstrated. Conjugation of the cell-penetrating peptide derived from the human immunodeficiency virus 1 (HIV) transactivator protein (TAT) to mesoporous silica nanoparticle is shown to be an effective way to enhance transmembrane delivery of nanoparticles for intracellular and molecular therapy. Cu,Zn-superoxide dismutase (SOD) is a key antioxidant enzyme that detoxifies intracellular reactive oxygen species, ROS, thereby protecting cells from oxidative damage. In this study, we fused a human Cu,Zn-SOD gene with TAT in a bacterial expression vector to produce a genetic in-frame His-tagged TAT-SOD fusion protein. The His-tagged TAT-SOD fusion protein was expressed in E. coli using IPTG induction and purified using FMSN-Ni-NTA. The purified TAT-SOD was conjugated to FITC-MSN forming FMSN-TAT-SOD. The effectiveness of FMSN-TAT-SOD as an agent against ROS was investigated, which included the level of ROS and apoptosis after free radicals induction and functional recovery after ROS damage. Confocal microscopy on live unfixed cells and flow cytometry analysis showed characteristic nonendosomal distribution of FMSN-TAT-SOD. Results suggested that FMSN-TAT-SOD may provide a strategy for the therapeutic delivery of antioxidant enzymes that protect cells from ROS damage.

Alkaline earth metal ion induced coil-helix-coil transition of lysine-coumarin-azacrown hybrid foldamers with OFF-OFF-ON fluorescence switching.

A new metal-ion-responsive and fluorescent foldamer, OPLM(8), composed of eight lysine-coumarin-azacrown units, has been designed and synthesized. The flexible OPLM(8) can be forced into a well-defined helix structure only upon the addition of alkaline earth metal ions. The structural change is based on the crown ether moieties being positioned in the requisite arrangement along the peptide chain, that is, at i, i+4 spacing, such that the alkaline earth metal ions can mediate the formation of four sandwich complexes between them. Moreover, varying the chelator-to-metal-ion ratio from 2:1 to 1:1 resulted in disassembly of the sandwich complexes leading to collapse of the helical structure to a random coil. These metal-ion-induced structural transitions could not only be monitored by the CD amplitude change but also easily probed by unique "OFF-OFF-ON" fluorescence intensity changes from 0.7-fold to 14-fold as the structure changed from the folded helix to a random coil. To further verify that the helix formation was indeed induced by metal-ion complexation, two kinds of control octamers with only four metal-ion chelators on the side chains were studied. One, which was capable of forming two sandwich complexes between the i and i+4 residues, displayed a negative Cotton couplet with the magnitude of its A value close to half that of OPLM(8), and the second had four metal-ion chelators positioned in the same turn, and hence was incapable of forming intramolecular metal complexes and showed different induced CD signals. Collectively, the photospectroscopic data and the results of the control studies suggest that alkaline earth metal ions can efficiently promote the flexible octamer OPLM(8) into a well-organized helix by the formation of sandwich complexes between substituents at an i, i+4 spacing.

Rare solvent annealing effective benzo(1,2-b:4,5-b')dithiophene-based low band-gap polymer for bulk heterojunction organic photovoltaics.

A newly synthesized benzo(1,2-b:4,5-b')dithiophene-based low band-gap copolymer pBCN is amenable to solvent annealing in the fabrication of organic photovoltaics, of which power conversion efficiency is greatly improved to 4.2% with PC(61)BM or 4.9% with PC(71)BM.

Specifically and reversibly immobilizing proteins/enzymes to nitriolotriacetic-acid-modified mesoporous silicas through histidine tags for purification or catalysis.

Six nitriolotriacetic-acid-modified ordered mesoporous silicas (NTA-OMPSs) with different pore sizes and surface features for specific and reversible protein immobilization were fabricated and characterized. Specific immobilization of a genetically engineered undecaprenyl pyrophosphate synthase (UPPs) from cell lysate and a chemically modified His-tagged horseradish peroxidase (HRP) in these Ni-NTA-OMPSs through histidine coordination to the nickelated NTA was demonstrated and confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and sodium dodecyl sulfate polyacrylamide gel electrophoresis. Negligible leakage of these enzymes over a wide range of acidic conditions was observed. Moreover, histidine tags with different lengths (His6, His4, His3, and His2) applied to HRP were evaluated to find the minimum length for effective complexation. Enzymatic assessment studies indicated that the pore size of the OMPSs has minimal influence on the enzymatic activity, whereas chemical entities such as unreacted mercapto groups tailored on the interior surfaces of the OMPSs played certain roles in inhibiting the enzymatic activity and stability. On MCF-S-NTA, SBA-S-NTA, and film-S-NTA, which contained unreacted mercaptopropyl groups on the interior surface, immobilized His-tagged HRP showed lower catalytic activity and stability than on MCF-NTA, film-NTA, and SBA-NTA. Selective hydroxylation of optically pure L-tyrosine to (S)-2-amino-3-(3,4-dihydroxyphenyl)propanoic acid (L-DOPA) by the immobilized HRP was also demonstrated.

Highly efficient control of thrombin activity by multivalent nanoparticles.

We have demonstrated that the incorporation of sulfated galactose acid (sulf-Gal) into thrombin-binding-aptamer (TBA)-conjugated gold nanoparticles (TBA-AuNPs) enables highly effective inhibition of thrombin activity toward fibrinogen. AuNP bioconjugates (TBA(15)/TBA(29)/sulf-Gal-AuNPs) were prepared from 13 nm AuNPs, 15-mer thrombin-binding aptamer (TBA(15)), 29-mer thrombin-binding aptamer (TBA(29)), and sulf-Gal. The numbers of TBA and sulf-Gal molecules per AuNP proved to have a strong impact on inhibitory potency. The best results were observed for 15-TBA(15)/TBA(29)/sulf-Gal-AuNPs (with 15 TBA(15) and 15 TBA(29) molecules per AuNP), which, because of their particularly flexible conformation and multivalency, exhibited ultrahigh binding affinity toward thrombin (K(d)=3.4×10(-12) M) and thus extremely high anticoagulant (inhibitory) potency. Compared to the case without inhibitors (the "normal" value), their measured thrombin clotting time (TCT) was 91 times longer, whereas for TBA(15) alone it was only 7.2 times longer. Their anticoagulant activity was suppressed by TBA-complementary-sequence (cTBA)-modified AuNPs (cTBA(15)/cTBA(29)-AuNPs) at a rate that was 20 times faster than that of free cTBA(15)/cTBA(29). Thus, easily prepared, low-cost, multivalent AuNPs show great potential for biomedical control of blood clotting.

Preparation of highly luminescent mannose-gold nanodots for detection and inhibition of growth of Escherichia coli.

In this paper, we describe a novel, simple, and convenient method for preparing water-soluble biofunctional gold nanodots (Au NDs) for the sensitive and selective detection of Escherichia coli (E. coli) and the inhibition of its growth. We obtained luminescent mannose-capped Au NDs (Man-Au NDs) from as-prepared 2.9-nm Au nanoparticles (Au NPs) and 29,29'-dithio bis(3',6',9',12',15',18'-hexaoxa-nonacosyl α-D-mannopyranoside) (Man-RSSR-Man). To obtain improved quantum yield (>20%), luminescent Man-Au NDs (1.8 nm) were prepared from Au NPs (0.47 µM) and Man-RSSR-Man (2.5 mM) in the presence of sodium borohydride (NaBH(4); 1.0 mM). The highly luminescent properties of Man-Au NDs prepared by the NaBH(4)-assisted method were characterized by UV-vis absorption, photoluminescence, and X-ray photoelectron spectroscopies. The results supported the high-density coverage of the NDs surface by Man-RS ligands. Multivalent interactions between Man-Au NDs and FimH proteins located on the bacterial pili of E. coli resulted in the formation of aggregated cell clusters. After concentrating this agglutinative E. coli from a large-volume cell solution (5 mL), Man-Au NDs were displaced by mannose (100 mM) and stabilized by Man-RSSR-Man (5 mM). Monitoring the luminescence of Man-Au NDs allowed the detection of E. coli at levels as low as 150 CFU/mL. Man-Au NDs were also found to be efficient antibacterial agents, selectively inhibiting the growth of E. coli through Man-Au ND-induced agglutination. Our small-diameter Man-Au NDs, which provided an ultra high ligand density (local concentration) of mannose units for multivalent interactions with E. coli, have great potential for use as an antibacterial agent in other applications.

Fluorescent substrate analog for monitoring chain elongation by undecaprenyl pyrophosphate synthase in real time.

Farnesyl pyrophosphate (FPP) is a common substrate for a variety of prenyltransferases for synthesizing isoprenoid compounds. In this study, (2E,6E)-8-O-(N-methyl-2-aminobenzoyl)-3,7-dimethyl-2,6-octandien-1-pyrophosphate (MANT-O-GPP), a fluorescent analog of FPP, was synthesized and demonstrated as a satisfactory substrate for Escherichia coli undecaprenyl pyrophosphate synthase (UPPS) with a K(m) of 1.5 µM and a k(cat) of 1.2s(-1) based on [(14)C]IPP consumption. Interesting, we found that its emission fluorescence intensity at 420 nm increased remarkably during chain elongation, thereby useful for real-time monitoring kinetics of UPPS to yield a K(m) of 1.1 µM and a k(cat) of 1.0 s(-1), consistent with those measured using radiolabeled substrate. Using this assay, the IC(50) of a known UPPS inhibitor farnesyl thiopyrophosphate (FsPP) was confirmed. Our studies provide a convenient and environmentally friendly alternative for kinetics and inhibition studies on UPPS drug target.

En route to white-light generation utilizing nanocomposites composed of ultrasmall CdSe nanodots and excited-state intramolecular proton transfer dyes.

One single material that emits white light is of paramount interest for the development of white light-emitting diodes (WLEDs). Here we report a novel nanocomposite, in which a new type of excited-sate intramolecular proton transfer (ESIPT) molecule, namely 5-(1,2-dithiolan-3-yl)-N-(2-{[4-(3-hydroxy-4-oxo-4H-chromen-2-yl)phenyl](methyl)amino}ethyl)pentanamide (HF-N-LA), is anchored onto the surface of ultrasmall CdSe quantum dots through dithiol functionality. Authentic white light with a CIE coordinate of (0.33, 0.33) could then be generated by confluence of 440 nm emission from CdSe and 570 nm proton-transfer tautomer emission from HF-N-LA. Moreover, linear color tunability could be achieved simply by altering relative amount of the two species, i.e., number of HF-N-LA onto CdSe, in one single nanocomposite, thus opening an innovative route toward applying nanocrystals in the field of WLEDs.

Generation and spectroscopic profiles of stable multiarylaminium radical cations bridged by fluorenes.

A series of arylaminofluorene derivatives (DTFA-1, TTFA-2, TAFB-3, and TAFA-4) were synthesized, and the generation of their corresponding arylaminium cation radicals was readily achieved by Cu(ClO(4))(2) in CH(3)CN. Moreover, the cation radicals were stable at ambient temperature with substantially long life times and exhibited distinct colors. The oxidation mechanism and spectroscopic features of the resulting cation radicals were probed by UV-vis-NIR spectroscopy and electron spin resonance experiments.