Threoninol as a scaffold of dyes (threoninol-nucleotide) and their stable interstrand clustering in duplexes

Functional molecules such as dyes (Methyl Red, azobenzene, and Naphthyl Red) were tethered on D-threoninol as base surrogates (threoninol-nucleotide), which were consecutively incorporated at the center of natural oligodeoxyribonucleotides (ODNs). Hybridization of two ODNs involving threoninol-nucleotides allowed interstrand clustering of the dyes on D-threoninol and greatly stabilized the duplex. When two complementary ODNs, both of which had tethered Methyl Reds on consecutive D-threoninols, were hybridized, the melting temperature increased proportionally to the number of Methyl Reds, due to stacking interactions. Clustering of Methyl Reds induced both hypsochromicity and narrowing of the band, demonstrating that Methyl Reds were axially stacked relative to each other (H-aggregation). Since hybridization lowered the intensity of circular dichroism peaks at the pi-pi* transition region of Methyl Red (300-500 nm), clustered Methyl Reds were scarcely wound in the duplex. Alternate hetero dye clusters could also be prepared only by hybridization of two ODNs with different threoninol-nucleotides, such as Methyl Red-azobenzene and Methyl Red-Naphthyl Red combinations. A combination of Methyl Red and azobenzene induced bathochromic shift and broadening of the band at the Methyl Red region due to the disturbance of exciton interaction among Methyl Reds. But interestingly, the Methyl Red and Naphthyl Red combination induced merging of each absorption band to give a single sharp band, indicating that exciton interaction occurred among the different dyes. Thus, D-threoninol can be a versatile scaffold for introducing functional molecules into DNA for their ordered clustering.     

Covalent incorporation of methyl red dyes into double-stranded DNA for their ordered clustering

An ordered dye cluster of Methyl Reds was formed in double-stranded DNA by hybridizing two complementary DNA-dye conjugates, each involving a Methyl Red moiety on a threoninol linker and a 1,3-propanediol spacer arranged alternately in the middle of the DNA sequence. In the duplex, Methyl Reds from each strand were axially stacked antiparallel to each other, as determined from NMR analysis. This clustering of Methyl Reds induced distinct changes in both UV/Vis and CD spectra. Single-stranded DNA-Methyl Red conjugates on D-threoninol linkers and (1,3-propanediol) spacers exhibited broad absorption spectra with lambda(max) at around 480 nm, and almost no CD was observed at around the absorption maximum of Methyl Red. However, as Methyl Reds were clustered by hybridization, lambda(max) shifted towards shorter wavelengths with respect to its monomeric transition. This hypsochromic shift increased as the number of Methyl Red molecules increased. Furthermore, a positive couplet was also strongly induced here. These dye clusters are H-aggregates, in which molecular excitons are coupled. The positive couplet demonstrates that the clusters on D-threoninol form a right-handed helix. In contrast, the induced CD became much weaker with Methyl Red on L-threoninol, which intrinsically prefers counterclockwise winding. Thus, mutual orientation of the stacked dye molecules was controlled by the chirality of the linker.     

基于1,8-萘酰亚胺与罗丹明B间荧光共振能量转移的高选择性Hg2+比率荧光探针

以罗丹明B与1,8-萘二甲酰亚胺反应合成了1个高选择性Hg²⁺比率荧光探针(RN). 在甲醇/乙腈/4-羟基哌嗪乙磺酸缓冲溶液(pH=7.2, 体积比8:1:1)中, RN对Hg²⁺具有比色和比率荧光双重响应. 加入Hg²⁺后, RN的紫外-可见光谱在约556 nm处产生强吸收, 溶液由浅绿色变为橙色, 其它金属离子对RN的紫外-可见光谱几乎无影响. 无Hg²⁺存在时, RN的荧光光谱在540 nm处出现萘二甲酰亚胺荧光团的特征峰; 加入Hg²⁺后, 540 nm处的发射带逐渐消失, 同时在580 nm附近产生强荧光, 荧光颜色从绿色变为橙色. 这归因于从萘酰亚胺到开环罗丹明B的荧光共振能量转移(FRET), 探针RN对Hg²⁺的比率荧光响应具有高选择性, 不受其它共存金属离子的干扰.     

Reversed Assembly of Dyes in an RNA Duplex Compared with Those in DNA

We prepared reversed dye clusters by hybridizing two RNA oligomers, each of which tethered dyes (Methyl Red, 4′‐methylthioazobenzene, and thiazole orange) on D ‐threoninols (threoninol nucleotides) at the center of their strands. NMR spectroscopic analyses revealed that two dyes from each strand were axially stacked in an antiparallel manner to each other in the duplex, and were located adjacent to the 3′‐side of a natural nucleobase. Interestingly, this positional relationship of the dyes was completely the opposite of that assembled in DNA that we reported previously: dyes in DNA were located adjacent to the 5′‐side of a natural nucleobase. This observation was also consistent with the circular dichroism of dimerized dyes in which the Cotton effect of the dyes (i.e., the winding properties of two dyes) was inverted in RNA relative to that in DNA. Further spectroscopic analyses revealed that clustering of the dyes on RNA duplexes induced distinct hypsochromicity and narrowing of the band, thus demonstrating that the dyes were axially stacked (i.e., H‐aggregates) even on an A‐type helix. On the basis of these results, we also prepared heterodimers of a fluorophore (thiazole orange) and quencher (Methyl Red) in an RNA duplex. Fluorescence from thiazole orange was found to be strongly quenched by Methyl Red due to the excitonic interaction, so that the ratio of fluorescent intensities of the RNA–thiazole orange conjugate with and without its complementary strand carrying a quencher became as high as 27. We believe that these RNA–dye conjugates are potentially useful probes for real‐time monitoring of RNA interference (RNAi) mechanisms.     

DNA-dye conjugates for controllable H aggregation(1)

Methyl Red H aggregate of predetermined size is successfully synthesized from the DNA conjugate involving multiple Methyl Red moieties in sequence. In the single stranded state, hypsochromicity monotonically increases with the number of incorporated dyes: the peak maximum of the conjugate involving six Methyl Reds appears at 415 nm, and the shift is as great as 69 nm (3435 cm(-)(1)) with respect to the monomeric transition. This large hypsochromicity accompanied by the narrowing of the band clearly demonstrates that H aggregate is formed in the single strand. H aggregation is further promoted at higher ionic strength. Upon addition of complementary DNA below the T(m), however, this H band disappears and a new peak appears at 448 nm, indicating that aggregated structure is changed by the duplex formation. This spectral change is completely reversible so that the H band at 415 nm appears again above T(m). Thus, aggregated structure can be reversibly controlled by the formation and dissociation of the DNA duplex.     

Excitonic interaction: another photophysical process for fluorescence‐controlled nucleic acid sensing

An excitonic interaction caused by the H‐aggregation of fluorescent dyes is a new type of useful photophysical process for fluorescence‐controlled nucleic acid sensing. We designed a fluorescence‐labeled nucleotide in which two thiazole orange dyes were linked covalently. A DNA strand containing this fluorescence‐labeled nucleotide showed absorption at 480 nm before hybridization, whereas an absorption band at 510 nm became predominant when the DNA was hybridized with the complementary strand. The shift in the absorption bands shows the existence of an excitonic interaction between dyes in the nucleotide, and as a result, emission from the doubly thiazole orange‐labeled DNA was well controlled. This clear change in fluorescence intensity depending on hybridization is applicable to multicolor RNA imaging in living cells. © 2010 The Japan Chemical Journal Forum and Wiley Periodicals, Inc. Chem Rec 10: 188–196; 2010: Published online in Wiley InterScience ( www.interscience.wiley.com ) DOI 10.1002/tcr.201000003     

A coumarin-based fluorescent probe for selective detection of Cu2+ in water

Fluorescent Red GK, a commercially available coumarin-based dye, was developed as a “turn-off” fluorescent probe for detection of Cu²⁺ in aqueous solution. It exhibited high selectivity and sensitivity at room temperature. Upon addition of Cu²⁺, the strong fluorescence of Fluorescent Red GK was severely quenched and its color changed from orange to colorless under illumination with a UV lamp; the color of the solution also changed from pink to colorless. So, it can be used as a specific colorimetric and fluorescent probe for Cu²⁺ with a detection limit as low as 0.0634?μM.     

Spectrophotometric determination of nitrite

A new reagent for the absorptiometric determination of nitrite ion is used. Nitrite is reacted with 1-amino-4-naphthalenesulfonic acid to form a diazonium ion, which gives an orange color at pH 1.5 with maximum absorption at 310 nm, or an orange-red color at pH 4 with maximum absorption at 485 nm. Diazotation is fast at pH 1.5 but is stable only for 30 min after which the color fades continuously. However, at pH 4, the color develops after 1 hr and remains stable for >40 hr. The time of color development at pH 4 can be reduced to 30 min when heated to 60–70 °C for 5 min, but is followed by reduction of its intensity to half. The described method is found suitable for the determination of 0.1–5.0 ppm of the titled ion.     

Dynamic Vapochromic Behaviors of Organic Crystals Based on the Open–Close Motions of S‐Shaped Donor–Acceptor Folding Units

The first vapochromic organic crystals are described with respect to their preparation, color change, adsorption/desorption properties, crystal structures, and color‐change mechanism. Non‐solvatochromic, 1,4,5,8‐naphthalene‐tetracarboxylic diimide (NDI) derivatives 1 a bearing two pyrrole imine (PI) tethers have been used as a motif for the crystal packing template. Red‐purple vapochromic solid 3 was prepared by evacuation of orange crystals 2 (equivalent to 1 a ?2 MeOH), obtained by recrystallization of 1 a from MeOH. Solid 3 showed high‐adsorption ability and unprecedented vapor‐dependent color changes upon exposure to a variety of organic vapors, whereas light brown amorphous solid 1 a , did not show vapo‐ or solvatochromic behavior toward any organic solvent. The strong adsorption capability of 3 was confirmed by TGA experiments and adsorption/desorption isotherms. Analysis of the solid‐state UV/Vis analysis revealed that the vapor‐dependent color changes of 3 were owed to the specific interference of solvent vapors with its broad CT absorbance at λ=450–650 nm. Packing structures of 1 a in orange crystals 2 , red‐purple solid 3 , and regenerated orange solid 2 were unequivocally established by single crystal and synchrotron powder X‐ray diffraction, respectively. Molecular structures and arrays of 1 a in these materials indicated that 1) unit 1 a had an S‐shaped folded conformation in 2 and 3 by intramolecular donor–acceptor interactions between NDI and two PI units; 2) inclusion of the guest vapor into the S‐shaped template decreased the intramolecular PI‐NDI interactions, accompanied by increasing intermolecular NDI‐NDI and PI‐PI interactions; and 3) such flexible, open–close motions of the S‐shaped template could be repeated during reversible adsorption/desorption processes without degradation of crystal packing. The adsorption properties and mechanism of molecular shape‐dependent vapochromic behavior of 3 are discussed with reference to experimental results, crystallographic data, and theoretical calculations.     

Hybridization‐Sensitive On–Off DNA Probe: Application of the Exciton Coupling Effect to Effective Fluorescence Quenching

The design of dyes that emit fluorescence only when they recognize the target molecule, that is, chemistry for the effective quenching of free dyes, must play a significant role in the development of the next generation of functional fluorescent dyes. On the basis of this concept, we designed a doubly fluorescence‐labeled nucleoside. Two thiazole orange dyes were covalently linked to a single nucleotide in a DNA probe. An absorption band at approximately 480 nm appeared strongly when the probe was in a single‐stranded state, whereas an absorption band at approximately 510 nm became predominant when the probe was hybridized with the complementary strand. The shift in the absorption bands shows the existence of an excitonic interaction caused by the formation of an H aggregate between dyes, and as a result, emission from the probe before hybridization was suppressed. Dissociation of aggregates by hybridization with the complementary strand resulted in the disruption of the excitonic interaction and strong emission from the hybrid. This clear change in fluorescence intensity that is dependent on hybridization is useful for visible gene analysis.     

Spectrophotometric determination of cationic and anionic surfactants with anionic dyes in the presence of nonionic surfactants,part I: A general aspect

The behavior of color development of anionic azo dyes, methyl orange and its analogues, was examined in aqueous media by changing the microenvironment of the dyes. The addition of alcohols, organic onium ions, anionic surfactants and nonionic surfactants brought about a decrease of the band at wavelengths near 480 nm and an increase of the band at wavelengths near 420 nm. Such a shift toward the shorter wavelengths in spectra was attributed to the change of the micro-environment around the dyes from a polar field to a less polar field; that is, the shift is caused by the change of the contribution of the following resonance forms, On the basis of the color change phenomena, the spectrophotometric methods for the determination of organic onium ions and anionic surfactants were proposed.     

Modulation of pyrene fluorescence in DNA probes depends upon the nature of the conformationally restricted nucleotide

The DNA probes (ODNs) containing a 2'-N-(pyren-1-yl)-group on the conformationally locked nucleosides [2'-N-(pyren-1-yl)carbonyl-azetidine thymidine, Aze-pyr (X), and 2'-N-(pyren-1-yl)carbonyl-aza-ENA thymidine, Aza-ENA-pyr (Y)], show that they can bind to complementary RNA more strongly than to the DNA. The Aze-pyr (X) containing ODNs with the complementary DNA and RNA duplexes showed an increase in the fluorescence intensity (measured at lambda em approximately 376 nm) depending upon the nearest neighbor at the 3'-end to X [dA ( approximately 12-20-fold) > dG ( approximately 9-20-fold) > dT ( approximately 2.5-20-fold) > dC ( approximately 6-13-fold)]. They give high fluorescence quantum yields (Phi F = 0.13-0.89) as compared to those of the single-stranded ODNs. The Aza-ENA-pyr (Y)-modified ODNs, on the other hand, showed an enhancement of the fluorescence intensity only with the complementary DNA (1.4-3.9-fold, Phi F = 0.16-0.47); a very small increase in fluorescence is also observed with the complementary RNA (1.1-1.7-fold, Phi F = 0.17-0.22), depending both upon the site of the Y modification introduced as well as on the chemical nature of the nucleobase adjacent to the modification site into the ODN. The fluorescence properties, thermal denaturation experiments, absorption, and circular dichroism (CD) studies with the X- and Y-modified ODNs in the form of matched homo- and heteroduplexes consistently suggested (i) that the orientation of the pyrene moiety is outside the helix of the nucleic acid duplexes containing a dT-d/rA base pair at the 3'-end of the modification site for both X and Y types of modifications, and (ii) that the microenvironment around the pyrene moiety in the ODN/DNA and ODN/RNA duplexes is dictated by the chemical nature of the conformational constraint in the sugar moiety, as well as by the nature of neighboring nucleobases. The pyrene fluorescence emission in both X and Y types of the conformationally restricted nucleotides is found to be sensitive to a mismatched base present in the target RNA: (i) The X-modified ODN showed a decrease ( approximately 37-fold) in the fluorescence intensity (measured at lambda em approximately 376 nm) upon duplex formation with RNA containing a G nucleobase mismatch (dT-rG pair instead of dT-rA) opposite to the modification site. (ii) In contrast, the Y-modified ODN in the heteroduplex resulted in a approximately 3-fold increase in the fluorescence intensity upon dT-rG mismatch, instead of matched dT-rA pair, in the RNA strand. Our data corroborate that the pyrene moiety is intercalated in the X-modified mismatched ODN/RNA (G mismatch) heteroduplex as compared to that of the Y-modified ODN/RNA (G mismatch) heteroduplex, in which it is located outside the helix.     

White Light Luminescence from a Homo-conjugated Molecule with Thermally Activated Delayed Fluorescence

Luminophores with tunable emission properties are appealing due to various applications. Among those properties, thermally activated delayed fluorescence (TADF) has been attracting enormous research interests. Herein, we synthesized a 9,9’-spirobifluorene based homo-conjugated molecule 1 , which connects a diphenylamino moiety as electron donor and a naphthalimide group as electron acceptor via 2,2’-positions of spirofluorene. Compound 1 displays dual emission behaviour with both blue and orange fluorescence. The one orange fluorescence around 555 nmshows sensitivity to oxygen and a prolonged lifetime of 284 ns in degassed toluene. Such characteristics imply TADF nature for this emission from a charge-transfer excited state. The other emission at 440 nm with blue colour displayed resistance to oxygen quenching and a normal fluorescence lifetime of 1.5 ns. Compared with control molecule, this emission band is assigned as conventional fluorescence from a localized excited state. In addition, dual emission property allows molecule 1 to be modulated to emit white photoluminescence in thin film with a CIE color coordinate of (0.25, 0.33).     

LED用Sr(1-1.5x)Mo0.8Si0.2O3.8:Eu3+x红色荧光粉的制备及其荧光性能

通过高温固相法合成了LED用红色荧光粉Sr(1-1.5x)Mo0.8Si0.2O3.8∶Eu3+x（x=0.1,0.2,0.3,0.4,0.5）。 通过XRD、激发光谱和发射光谱测试了材料的物相组成以及发光性能。 x=0.1样品的XRD谱与JCPDS 08-0482(SrMoO4)的标准卡片相同。 Eu3+代替晶格中Sr2+的位置成为发光中心。 随着Eu3+含量x的增加,593 nm处的5D0-7F1跃迁和614 nm处的5D0-7F2跃迁发射强度会相互转换：当x≤0.4时,以磁偶极5D0-7F1跃迁为主,发射橙色光;而当x=0.5时,以电偶极5D0-7F2跃迁发射为主,发射红光。 可能是过量掺杂的Eu3+离子,只能存在于晶格空位形成缺陷,无法占据SrMoO4中Sr2+的格位中,Eu3+在晶格中占据非对称中心的格位,导致电偶极跃迁变成允许跃迁,从而增加了5D0-7F2跃迁,减弱了5D0-7F1跃迁。 因此,可以通过调节激活剂的含量获得不同发光色的荧光粉。 Eu3+掺杂的硅钼酸锶体系,614 nm激发下,在368 nm处出现宽的基质吸收峰和467 nm处7F0-5D2的跃迁峰,且这2处的吸收峰在x=0.5时比x=0.4时强3倍左右。 材料能非常好的吸收368 nm波长的光,产生颜色可调的橙红色。 与近紫外光LED芯片匹配良好。     

Ratiometric, colorimetric and fluorescent chemosensor for "turn-on" detection of cyanide (CN-)

Herein, we unveiled the first zinc complex based chemosensor for cyanide (CN(-)). The changed visual color from light reddish orange to dark reddish orange, the enhanced fluorescence intensity at approximately 600 nm and the near-linear correlation A(495)/A(325)versus the cyanide concentration were observed after adding CN(-).     

Synthesis of novel axially chiral cyclic benzopolysulfides

Novel axially chiral benzopentathiepins were synthesized by sulfurization of dithiastannole. Naphthyl moiety was introduced near the pentathiepin ring by Suzuki-Miyaura cross-coupling reaction. Pentathiepins were found as diastereomeric mixture. Rotational energy barrier for C-C bond was estimated by theoretical calculation. Energy barrier for the inversion of pentathiepin ring was experimentally determined by variable temperature ¹H NMR.     

Synthesis, structure and thermal stability of fully hydrophobic porphyrin-DNA conjugates

The DNAs modified with tetraphenyl porphyrin at the center of 13mer oligonucleotide were synthesized using phosphoramidite chemistry and automated DNA synthesis. When the porphyrin modified oligonucleotide was annealed with its complementary strand, they formed a standard B-form duplex. The porphyrin moiety intercalated in the duplex, and moderately lowered the thermal stability.     

Spermine moiety attached to the C-5 position of deoxyuridine enhances the duplex stability of the phosphorothioate DNA/complementary DNA and shows the susceptibility of the substrate to RNase H

Novel phosphorothioate-modified oligodeoxynucleotides (S-ODNs) containing a deoxyuridine derivative bearing a spermine moiety at the C-5 position were synthesized. The study of the thermal stability and the thermodynamic stability showed that the modified S-ODNs have been able to form the stable duplexes with the complementary DNA. It was also found that the duplex composed of the modified S-ODN and its complementary RNA strand is the substrate for Escherichia coli RNase H, and the cleavage of the RNA strand by the enzyme was almost similar as in the case of the unmodified one.     

193 NM LIGHT INDUCES SINGLE STRAND BREAKAGE OF DNA PREDOMINANTLY AT GUANINE

Irradiation of DNA with 193 nm light results in monophotonic photoionization, with the formation of a base radical cation and a hydrated electron (φ_P1 = 0.048–0.065). Although >50% of the photoionization events initially occur at guanine in DNA, migration of the “hole” from the other bases to guanine occurs to yield predominantly its radical cation or its deprotonated form. From sequence analysis, the data reveal that 193 nm light induces single strand breaks (ssb) in double-stranded DNA preferential 3’ to a guanine residue. However, it has previously been reported that 193 nm light yields very low yields of ssb (<2% of the yield of eaq). The distribution of these ssb at guanine is nonrandom, showing a dependence on the neighboring base moiety. The efficiency of ssb formation at nonguanine sites is estimated to be at least one order of magnitude lower. The preferred cleavage at guanine is consistent with migration and localization of the electron loss center at guanine. It is argued that singlet oxygen and the photoionized phosphate group of the sugar moiety are not major precursors to ssb. At present, the mechanisms of strand breakage are not known although a guanine radical or one of its products remain potential precursors.     

Real-time Imaging of Nascent DNA in Live Cells by Monitoring the Fluorescence Lifetime of DNA-Incorporated Thiazole Orange-Modified Nucleotides

A modified 2′-deoxycytidine triphosphate derivative ( dC^TOTP ) bearing a thiazole orange moiety tethered via an oligoethylene glycol linker was designed and synthesized. The nucleotide was incorporated into DNA by DNA polymerases in vitro as well as in live cells. Upon incorporation of dC^TOTP into DNA, the thiazole orange moiety exhibited a fluorescence lifetime that differed significantly from the non-incorporated (i.e. free and non-covalently intercalated) forms of dC^TOTP . When dC^TOTP was delivered into live U-2 OS cells using a synthetic nucleoside triphosphate transporter, it allowed us to distinguish and monitor cells that were actively synthesizing DNA in real time, from the very first moments after the treatment. We anticipate that this probe could be used to study chromatin organization and dynamics.