[1] Studies of Benzothiazole and Benzoselenazole Squaraines as Fluorescent Probes for Albumins Detection K. D. Volkova, V. B. Kovalska, M. Yu. Losytskyy, A. Bento, L. V. Reis, P. F. Santos, P. Almeida, S.M. Yarmoluk Journal of Fluorescence. – 2008. – Vol. 18, ¹5. – P. 877-882.
S.M. Yarmoluk, V.B. Kovalska, M.Yu. Losytskyy Biotechnic and Histochemistry. – 2008. – Vol. 83, Iss. 3-4. – P. 131-145.
T.R. Berkelman, S.M. Yarmoluk, V.B. Kovalska, M.Yu. Losytskyy, K.D. Volkova Patent Application WO 2008/027821 A1, 06.03.2008. Abstract Squaraine dyes are incorporated into a separation medium in which protein or polypeptide mixtures are separated, which medium also contains a detergent that forms a complex with the polypeptides or proteins. The dyes, upon excitation in the separation medium, exhibit a heretofore unrecognized selectivity in their fluorescent emissions by emitting a signal only when the dye molecules are associated with complexes of protein (or polypeptide) and detergent molecules, despite the additional presence of dyes in the bulk of the separation medium. The dyes are thus able to indicate the presence and locations of proteins or polypeptides in the separation medium without the need for removing unassociated dyes from the medium. [4] To the studies of (p-dimethylaminostyryl)pyridinium based homodimer to dsDNA binding mechanism M.Yu. Losytskyy, N. Akbay, V.B. Kovalska, A.O. Balanda, A. Boutorine and S.M. Yarmoluk Ukrainica Bioorganica Acta. – 2007. – Vol. 5, No. 2. – P. 52-55. Abstract With the aim to study interaction mode of (p-dimethylaminostyryl)pyridinium homodimer dye Dst-6 with dsDNA, the equilibrium constant of dye-dsDNA binding (K) and the number of dsDNA base pairs occupied with one bound dye molecule were estimated. Obtained data support the previously made suggestion about realization of groove-binding mechanism of dye-DNA interaction. [5] Specific fluorescent detection of fibrillar α-synuclein using mono- and trimethine cyanine dyes K.D. Volkova, V.B. Kovalska, A.O. Balanda, M.Yu Losytskyy, A.G. Golub, R.J. Vermeij, V. Subramaniam, O.I. Tolmachev and S.M. Yarmoluk Bioorganic and Medicinal Chemistry. – 2008. – V. 16, Iss. 3. – P.1452-1459. Abstract With the aim of searching of novel amyloid-specific fluorescent probes the ability of series of mono- and trimethine cyanines based on benzothiazole, pyridine and quinoline heterocycle end groups to recognize fibrillar formations of α-synuclein (ASN) was studied. For the first time it was revealed that monomethine cyanines can specifically increase their fluorescence in aggregated ASN presence. Dialkylamino-substituted monomethine cyanine T-284 and meso-ethyl-substituted trimethine cyanine SH-516 demonstrated the higher emission intensity and selectivity to aggregated ASN than classic amyloid stain Thioflavin T, and could be proposed as novel efficient fluorescent probes for fibrillar ASN detection. Studies of structure–function dependences have shown that incorporation of amino- or diethylamino- substituents into the 6-position of the benzothiazole heterocycle yields in a appearance of a selective fluorescent response to fibrillar α-synuclein presence. Performed calculations of molecular dimensions of studied cyanine dyes gave us the possibility to presume, that dyes bind with their long axes parallel to the fibril axis via insertion into the neat rows (so called ‘channels') running along fibril. [6] The mechanism of benzothiazole styrylcyanine dyes binding with dsDNA: studies by spectral-luminescent methods N. Akbay, M. Yu. Losytskyy, V. B. Kovalska, A. O. Balanda and S. M. Yarmoluk. Journal of Fluorescence. – 2008. – V.18, Iss.1. – P.139-147. Abstract In the presented work studies of the interaction mode of monomer and two homodimer benzothiazole styryl dyes containing spermine-like linkage/tail group with the double stranded (ds) DNA are reported. For these dyes, equilibrium constant of dye binding to DNA (K b), as well as the number of dsDNA base pairs occupied by one bound dye molecule (n) were determined. The data obtained show that the presence of spermine-like group containing quaternary nitrogen (Bos-5) results in increase of K b value as compared to this of unsubstituted analogue (Sbt). Besides, for the dimer dyes containing benzothiazole styryl chromophores, the K b value is either five times higher (DBos-13) or almost the same (DBsu-10) as compared to this of corresponding monomer Sbt, depending on the position in the benzothiazole ring where the linker is attached. Moreover, the n values for both dimers are significantly different as well, pointing to the bis-intercalative binding mechanism for DBos-13 and for the groove-binding one for DBsu-10. The conclusion about the dimer dyes-dsDNA binding mechanisms is also supported by the study of the fluorescent response of these dyes on the presence of AT- and GC-containing polynucleotides. [7] Modern techniques for protein detection on polyacrylamide gels: problems arising from the use of dyes of undisclosed structures for scientific purposes K.D. Volkova, V.B. Kovalska and S.M. Yarmoluk Biotechnic and Histochemistry. – 2007. – Vol. 82, Iss. 4&5. – P.201-208 [8] Cyanine dye–protein interactions: looking for fluorescent probes for amyloid structures K.D. Volkova, V.B. Kovalska, A.O. Balanda, R.J. Vermeij, V. Subramaniam, Yu.L. Slominskii and S.M. Yarmoluk Journal of Biochemical and Biophysical Methods. – 2007. – Vol.70, Iss. 5. – P.727-733. Abstract We ascertained the ability to detect fibrillar beta-lactoglobulin (BLG) of a series of mono-, tri-, penta-, and heptamethinecyanines based on benzothiazole and benzimidazole heterocycles, and of benzothiazole squaraine. Fluorescence properties of these cyanine dyes were measured in the unbound state and in the presence of monomeric and fibrillar BLG and compared with those for the commercially available benzothiazole dye Thioflavin T. The correlation between the chemical nature of the dye molecules and the ability of dyes to bind aggregated proteins was established. We found that meso-substituted cyanines with amino substituents in heterocycle in contrast to the corresponding unsubstituted dyes have a binding preference to fibrillar BLG and a noticeable fluorescence response in the presence of the aggregated protein. For the squaraines and benzimidazole penthamethinecyanines studied, fluorescence emission increased both in the presence of native and fibrillar protein. The trimethinecyanines T-49 and SH-516 exhibit specifically increased fluorescence in the presence of fibrillar BLG. These dyes demonstrated the same or higher emission intensity and selectivity to aggregated BLG as Thioflavin T, and are proposed for application in selective fluorescent detection of aggregated proteins. [9] Novel, monomeric cyanine dyes as reporters for DNA helicase activity C. Xu, M.Yu. Losytskyy, V.B. Kovalska, D.V. Kryvorotenko, S.M. Yarmoluk, S. McClelland and P.R. Bianco. Journal of Fluorescence. – 2007. – V. 17, Iss. 6. – P. 671-685. Abstract The dimeric cyanine dyes, YOYO-1 and TOTO-1, are widely used as DNA probes because of their excellent fluorescent properties. They have a higher fluorescence quantum yield than ethidium homodimer, DAPI and Hoechst dyes and bind to double-stranded DNA with high affinity. However, these dyes are limited by heterogeneous staining at high dye loading, photocleavage of DNA under extended illumination, nicking of DNA, and inhibition of the activity of DNA binding enzymes. To overcome these limitations, seven novel cyanine dyes (Cyan-2, DC-21, DM, DM-1, DMB-2OH, SH-0367, SH1015-OH) were synthesized and tested for fluorescence emission, resistance to displacement by Mg2+, and the ability to function as reporters for DNA unwinding. Results show that Cyan-2, DM-1, SH-0367 and SH1015-OH formed highly fluorescent complexes with dsDNA. Of these, only Cyan-2 and DM-1 exhibited a large fluorescence enhancement in buffers, and were resistant to displacement by Mg2+. The potential of these two dyes to function as reporter molecules was evaluated using continuous fluorescence, DNA helicase assays. The rate of DNA unwinding was not significantly affected by either of these two dyes. Therefore, Cyan-2 and DM-1 form the basis for the synthesis of novel cyanine dyes with the potential to overcome the limitations of YOYO-1 and TOTO-1. [8] Two-photon excited luminescent styryl dyes as probes for the DNA detection and imaging. Photostability and phototoxic influence on DNA V.M. Yashchuk, V.V. Gusak, I.M. Dmytruk, V.M. Prokopets, V.Yu. Kudrya, M.Yu. Losytskyy, V.P. Tokar, Ya.O. Gumenyuk, S.M. Yarmoluk, V.B. Kovalska, A.O. Balanda, D.V. Kryvorotenko Molecular Crystals and Liquid Crystals. – 2007. – V. 467, Iss.1. – P. 325-338. Abstract The photostability and phototoxic influence on DNA of several styryl dyes are investigated by studying the absorption, fluorescence and phosphorescence spectra of these dyes and DNA + dye systems. The dyes Bos-1, DBos-24, and DBos-30 exhibit a rather intensive fluorescence under the two-photon excitation. Changes of the optical density in the wavelength regions 250 ÷ 300 nm (DNA absorption) and 370 ÷ 650 nm (dye absorption) of DNA + dye solutions caused by the visible light irradiation are fixed. Bos-1 and DBos-30 dyes bound to DNA are more photostable than those in free state, being photochemically safe for DNA. DBos-24 and Dst-MdO dyes show a slight phototoxic effect on DNA. [9] The optical biomedical sensors for DNA detection and imaging based on two-photon excited luminescent styryl dyes: phototoxic influence on the DNA V.M. Yashchuk, V.Yu. Kudrya, M.Yu. Losytskyy, V.P. Tokar, S.M. Yarmoluk, I.M. Dmytruk, V.M. Prokopets, V.B. Kovalska, A.O. Balanda, D.V. Kryvorotenko, T.Yu. Ogul'chansky Proceedings of SPIE. – 2007. – Vol. 6796. – 67960M. – 14 pp. Abstract The optical absorption, fluorescence and phosphorescence of the novel styryl dyes developed for the fluorescent detection of DNA were investigated. The energy structures of dye molecules as well as spectral manifestations of the dyes aggregate formation and interaction with DNA were studied. The dramatic increase (up to 1000 times) of the fluorescence intensity of dyes in the presence of DNA was observed. The photostability and phototoxic influence on the DNA of several styryl dyes were studied by analyzing absorption, fluorescence and phosphorescence spectra of these dyes and dye-DNA systems. Changes of the optical density value of dye-DNA solutions caused by the visible light irradiation were fixed in the wavelength regions of the DNA absorption and of the dye absorption. Fluorescence emission of dye-DNA complexes upon two-photon excitation (TPE) at wavelength 1064 nm with the 20 ns pulsed YAG: Nd3+ laser and at 840 nm with the 90 fs pulsed Ti:sapphire laser was registered. The values of two-photon absorption cross-sections of dye-DNA complexes were evaluated. [10] Polymethine dyes – derivatives of the 7-N,N-dialkylaminocoumarines Ya.O. Prostota, Ya.O. Kachkovsky, A.V. Kropachev, M.Yu. Losytskyy, S.S. Tarnavskyy, S.M. Yarmoluk Ukrainica Bioorganica Acta. – 2007. – V. 5, ¹ 1. – P. 32-42. Abstract A series of new polymethine dyes — derivatives of the 7-N,N-diethylaminocoumarine substituted by polymethine chromophore at position 3 and its analogue with the fully fixed aminogroup (coumarine 343) were obtained. Absorption and emission spectra of these dyes were investigated. All these compounds were examined as possible fluorescent probes for nucleic acids and proteins detection. Dyes 5á and 6â are the most appropriate for the non-specific detection of proteins in the presence of SDS (sodium dodecylsulphate). Nevertheless, the main disadvantage of this type of dyes is their low chemical stability in buffered solutions, as well as in the presence of biological molecules. [11] Detection of polyamino acids using trimethincyanine dyes V.B. Kovalska, D.V. Kryvorotenko, M.Yu. Losytskyy, P. Nording, A. Rueck, B. Schoenenberger, S.M. Yarmoluk, F. Wah Patent US2006207881, 21.09.2006, Appl. 01.02.2005 Abstract The present invention is generally directed to a method for detecting polyamino acids. More specifically, the present invention is directed to a method for detecting polyamino acids using trimethincyanine dyes that interact non-covalently with polyamino acids to produce an optically detectable dye/polyamino acid complex. [12] Synthesis and spectral studies of styrylcyanines based on 4-oxo-4,6,7,8-tetrahydropyrrolo[1,2-a]thieno[2,3-d]pyrimidinium and 5-oxo-1,2,3,5-tetrahydropyrrolo[2,1-b]quinazolinium A.O. Balanda, K.D. Volkova, V.B. Kovalska, V.P. Tokar and S.M. Yarmoluk Ukrainica Bioorganica Acta. – 2006. – Vol. 4, ¹ 2. – P. 10-15 (in Ukrainian). Abstract A series of novel styrylcyanine dyes based on 4-oxo-4,6,7,8-tetrahydropyrrolo[1,2-a]thieno[2,3-d]pyrimidinium and 5-oxo-1,2,3,5-tetrahydropyrrolo[2,1-b]quinazolinium. Spectral-luminescent properties of obtained dyes in unbound state and in presence of nucleic acids and BSA (bovine serum albumin) were evaluated. p-dimethylaminostyryls based on 4-oxo-4,6,7,8-tetrahydropyrrolo[1,2-a]thieno[2,3-d]pyrimidinium appeared to selectively interact with RNA and thus could be used as RNA-specific fluorescent probes. Besides, for these dyes large values of two-photon absorption cross-sections were admitted in RNA presence. [13] Synthesis and spectral-luminescent studies of novel 4-oxo-4,6,7,8-tetrahydropyrrolo[1,2-a]thieno[2,3-d]pyrimidinium styryls as fluorescent dyes for biomolecules detection A.O. Balanda, K.D. Volkova, V.B. Kovalska, M.Yu. Losytskyy, V.P. Tokar, V.M. Prokopets and S.M. Yarmoluk Dyes and Pigments. – 2007. – V. 75, No. 1. – P. 25-31. Abstract A series of novel 4-oxo-4,6,7,8-tetrahydropyrrolo[1,2-a]thieno[2,3-d]pyrimidinium and 5-oxo-1,2,3,5-tetrahydropyrrolo[2,1-b]quinazolinium styryl dyes were synthesized. For preparing of studied dyes the standard method of styrylcyanines synthesis was modified. Spectral-luminescent properties of obtained dyes in free state and in the presence of nucleic acids and BSA were studied. It was shown that p-dimethylaminostyryls based on 4-oxo-4,6,7,8-tetrahydropyrrolo[1,2-a]thieno[2,3-d]pyrimidinium with aliphatic substituents in 2 and 3 positions demonstrated RNA-binding preference. These dyes in the presence of RNA significantly enhance emission intensity and could be used as RNA-specific fluorescent probes. Besides, the fluorescence emission after two-photon absorption of dye-RNA complexes in buffer solutions was measured. [14] Spectroscopic study of squaraines as protein-sensitive fluorescent dyes K.D. Volkova, V.B. Kovalska, A.L. Tatarets, L.D. Patsenker, D.V. Kryvorotenko and S.M. Yarmoluk Dyes and Pigments. – 2007. – V. 72, No. 3. – P. 285-292. Abstract A series of new symmetrical and asymmetrical squaraines were synthesised and efficiency of their use as fluorescent probes for the specific detection of proteins was studied. Spectral-luminescent properties of the squaraines were measured in presence of bovine serum albumin (BSA), human serum albumin (HSA), ovalbumin, avidin from hen egg white (AVI), lysozyme, and trypsin. All investigated squaraines show considerable (in 24-190 times) emission increase in the presence of BSA. At the same time the fluorescent response of the studied dyes in the presence of other albumins is significantly lower – emission enhances up to 24 times. The 3-oxo-substituted indolenine dye 9(74) demonstrates sufficient fluorescence increasing value and emission intensity level in the presence of BSA as well as of HSA and ovalbumin. Dyes containing N-carboxyalkyl group demonstrate sufficient emission enhancement (up to 16 times) and noticeable fluorescent signal in the presence of avidin from hen egg white. Squaraines slightly increase or even decrease their emission intensity in the presence of hydrolases lysozyme or trypsin. [15] Studies of monomeric and homodimeric oxazolo[4,5-b]pyridinium cyanine dyes as fluorescent probes for nucleic acids visualization V.B. Kovalska, V.P. Tokar, M.Yu. Losytskyy, T. Deligeorgiev, A. Vassilev, N. Gadjev, K.-H. Drexhage and S.M. Yarmoluk Journal of Biochemical and Biophysical Methods. – 2006. – V. 68, No. 3. – P. 155-165. Abstract The series of recently synthesized monomeric and homodimeric cyanine dyes based on monomethine cyanine chromophore with oxazolo[4,5-b]pyridinium and quinoline end groups [Vassilev A., Deligeorgiev T., Gadjev N., Drexhage K-H. Synthesis of novel monomeric and homodimeric cyanine dyes based on oxazolo[4,5-b]pyridinium and quinolinium end groups for nucleic acid detection, Dyes Pigm 2005;66:135–142] were studied as possible fluorescent probes for nucleic acids detection. Significant fluorescence enhancement and intensity level (quantum yield up to 0.75) was observed for all the dyes in the presence of DNA. The oxazolo[4,5-b]pyridinium cyanines demonstrated high sensitivity as fluorescent stains for post-electrophoretic visualization of nucleic acids in agarose gels upon both VIS and UV transillumination, and the visualized band contained 0.8 ng of dsDNA. [16] 6,6'-Disubstituted benzothiazole trimethine cyanines – new fluorescent dyes for DNA detection V.B. Kovalska, K.D. Volkova, M.Yu. Losytskyy, O.I. Tolmachev, A.O. Balanda and S.M. Yarmoluk Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy. – 2006. – V. 65, No. 2. – P. 271-277. Abstract The influence of methyl-, 2-hydroxyethyl-, dimethyl-, diethyl- and benzoyl-amino substituents in the 6,6'-positions of benzothiazole heterocycle of trimethine cyanines on their spectral-luminescent properties and behavior in presence of DNA, RNA and BSA was studied. It was shown that incorporation of 6,6'-substituents generally leads to the increase in dyes tendency to aggregation, resulting in the considerable decrease in the emission intensity of the disubstituted dyes as compared to the unsubstituted ones. Emission of the studied 6,6'-disubstited dyes in DNA presence is considerably more intensive than in presence of RNA, that points on the existing of DNA binding preference for the mentioned dyes. Insertion of benzoyl-amino groups into the 6,6'-positions permitted us to design the DNA-sensitive dyes on the basis of symmetric trimethine cyanines with unsubstituted polymethine chain, while typically such dyes slightly respond on the presence of biopolymers. 6,6'-Benzoyl-amino-disubstituted trimethine cyanines are proposed as efficient dyes for DNA detection. [17] Fluorescence of styryl dyes-DNA complexes induced by single- and two-photon excitation V.P. Tokar, M.Yu. Losytskyy, V.B. Kovalska, D.V. Kryvorotenko, A.O. Balanda, V.M. Prokopets, M.P. Galak, I.M. Dmytruk, V.M. Yashchuk and S.M. Yarmoluk Journal of Fluorescence. – 2006. – V. 16, No. 6. – P. 783-791. Abstract The series of novel monomer and homodimer styryl dyes based on (p-dimethylaminostyryl) benzothiazolium residues were synthesized and studied as possible fluorescent probes for nucleic acids detection. Spectral-luminescent and spectral-photometric properties of obtained dyes in the unbound state and in DNA presence were studied. Fluorescence emission induced by two-photon excitation of dye-DNA complexes in aqueous buffer solution was registered. Two-photon absorption cross section values of the studied dyes in DNA presence were evaluated. [18] New method for covalent fluorescent biomolecule labeling with hemicyanine dye O.M. Kostenko, V.B. Kovalska, K.D. Volkova, P. Shaytanov, I.O. Kocheshev, Y.L. Slominskiy, I.V. Pisareva and S.M. Yarmoluk Journal of Fluorescence. – 2006. – V. 16, No. 4. – P. 589-593. Abstract Fluorescent chromophore, alkylamino-(tetra-hydronaphthalenylidene)- benzothiazolium derivatives (HBTN dyes), are proposed as covalent labels for proteins via aliphatic amino groups. Spectral-luminescent properties of 3-methyl-2-{(E)-[7-(methylamino)-4,4a,5,6-tetra-hydronaphthalen-2(3H)-ylidene]methyl}-1,3-benzothiazol-3-ium chloride (HBTN, R=Me) and its predecessor, 2-[(E)-(7-methoxy-4,4a,5,6-tetrahydronaphthalen-2(3H)-ylidene)methyl]-3-methyl-1,3-benzothia-zol-3-ium chloride (ABTN), are studied for free dyes and in the presence of DNA and BSA. Considerable spectral-luminescent changes accompany the transformation of ABTN into HBTN that allows monitoring conjugation reaction. In presence of DNA and BSA the HBTN increases its emission in 15 and 4 times respectively and becomes strongly fluorescent. The conditions for labeling are developed and a model conjugate of HBTN dye with BSA is synthesized. It was shown that using of HBTN dye as a fluorescent label allows detection by eye of about 3 μg/band of BSA on polyacrylamide gel upon UV-irradiation. [19] Novel styrylcyanines and their dimers as fluorescent dyes for nucleic acids detection: synthesis and spectral-luminescent studies A.O. Balanda, K.D. Volkova, V.B. Kovalska, M.Yu. Losytskyy, S.S. Lukashov, S.M. Yarmoluk Ukrainica Bioorganica Acta. – 2006. – V. 4, No. 1. – P. 17-29. Abstract With the aim to design DNA-sensitive fluorescent probes, a series of novel (p-dimethylaminostyryl) benzothiazolium dyes with tetraalkylammonium residues were synthesized. Spectral-luminescent properties of obtained dyes in free state and in the presence of nucleic acids and BSA (bovine serum albumin) were studied. It was shown that dimeric styrylcyanines with linker containing two tetraalkylammonium residues demonstrated 3 orders of magnitude fluorescence increase in the presence of DNA. The interaction mechanism between synthesized styrylcyanines and dsDNA was studied and possible binding mechanism was suggested. The possibility to use styrylcyanines as probes for fluorescent nucleic acids in vivo visualization was revealed. [20] New highly sensitive fluorescent dye Barva NA for DNA visualization after the denaturing gradient gel electrophoresis O.O. Solovyov, L.A. Livshits, V.B. Kovalska, M.Yu. Losytskyy, Yu.L. Slominski, S.M. Yarmoluk Ukrainica Bioorganica Acta. – 2006. – V. 4, No. 1. – P. 30-33. Abstract The comparison of the efficiency of cyanine dye Barva NA (Otava, Ukraine), the analogue of SYBR Green, and ethidium bromide for visualization of PCR products of exon 20 CFTR gene after mutant alleles separation using denaturing gradient gel electrophoresis (DGGE) was performed. The higher sensitivity of Barva NA was shown. [21] Synthesis of novel monomeric cyanine dyes containing mercapto and thioacetyl substituents for nucleic acid detection T. Deligeorgiev, N. Gadjev, A. Vasilev, K.-H. Drexhage, S.M. Yarmoluk Dyes and Pigments. – 2006. – V. 70, No. 3. – P. 185-191. Abstract Fifteen novel mono-, di- and tricationic monomeric monomethine cyanine dyes, bearing mercapto and thioacetyl substituents, useful for nucleic acid detection are described. All derivatives absorb in the region 417-508 nm and have molar absorptivities of 61 000-101 300 l mol-1 cm-1. The products were characterized by 1H NMR, HPLC-MS and elemental analysis. [22] Cyanine dyes as intercalating agents: kinetic and thermodynamic studies on the DNA/Cyan40 and DNA/CCyan2 systems T. Biver, A. De Biasi, F. Secco, M. Venturini and S. Yarmoluk Biophysical Journal. – 2005. – V. 89. – P. 374-383. Abstract The interaction of cyanines with nucleic acids is accompanied by intense changes of their optical properties. Consequently these molecules find numerous applications in biology and medicine. Since no detailed information on the binding mechanism of DNA/cyanine systems is available, a T-jump investigation of the kinetics and equilibria of binding of the cyanines Cyan40 [3-methyl-2-(1,2,6-trimethyl-4(1H)pyridinylidenmethyl)-benzothiazolium ion] and CCyan2 [3-methyl-2-[2-methyl-3-(3-methyl-2(3H)-benzothiazolylidene)-1-propenyl]-benzothiazolium ion] with CT-DNA is performed at 25 °C, pH 7 and various ionic strengths. Bathochromic shifts of the dye absorption band upon DNA addition, polymer melting point displacement (ΔT = 8-10 °C), site size determination (n = 2), and stepwise kinetics concur in suggesting that the investigated cyanines bind to CT-DNA primary by intercalation. Measurements with poly(dA-dT)•poly(dA-dT) and poly(dG-dC)•poly(dG-dC) reveal fair selectivity of CCyan2 toward G-C basepairs. T-jump experiments show two kinetic effects for both systems. The binding process is discussed in terms of the sequence D + S ↔ D,S ↔ DS² ↔ DS²², which leads first to fast formation of an external complex D,S and then to a partially intercalated complex DS² which, in turn, converts to DS²², a more stable intercalate. Absorption spectra reveal that both dyes tend to self-aggregate; the kinetics of CCyan2 self-aggregation is studied by T-jump relaxation and the results are interpreted in terms of dimer formation. [23] Synthesis of novel monomeric and homodimeric cyanine dyes with thioacetyl substituents for nucleic acid detection T. Deligeorgiev, N. Gadjev, A. Vasilev, K.-H. Drexhage, S.M. Yarmoluk Dyes and Pigments. – 2007. – V. 72. – P.28–32. Abstract Several novel di-, tri- and tetracationic monomeric and homodimeric monomethine cyanine dyes for nucleic acid detection were synthesized by condensation of 3,4-dihydro-(2H)-1,3-thiazino[2,3-b]benzooxazolium bromide and appropriate 1-(ω-bromoalkyl)-4-methylpyridinium or 1-(ω-bromoalkyl)-4-methylquinolinium bromide followed by quaternization with N,N,N',N'-tetramethyl-1,3-propanediamine (TMPDA) or N,N,N,N',N'-pentamethyl-1,3-propanediammonium iodide, or bisquaternization with TMPDA or N,N,N',N'-tetramethyl-1,6-hexanediamine (TMHDA). [24] Synthesis of novel fluorescent styryl dyes based on the imidazo[1,2-a]pyridinium chromophore and their spectral-fluorescent properties in the presence of nucleic acids and proteins V.B. Kovalska, M.Yu. Losytskyy, D.V. Kryvorotenko, A.O. Balanda, V.P. Tokar, S.M. Yarmoluk Dyes and Pigments. – 2006. – Vol. 68, No. 1. – P. 39-45. Abstract The imidazo[1,2-a]pyridinium heterocyclic system was used to prepare styryl dyes. Improved synthetic methods were proposed for the parent imidazo[1,2-a]pyridines and their corresponding quaternary salts. The standard method for preparing styrylcyanines was modified for the synthesis of the target imidazo[1,2-a]pyridinium dyes. Spectral-luminescent properties of the obtained dyes in free state and in the presence of nucleic acids, BSA, and BSA/detergent system were studied. 7-[2-(4-Dihexylaminophenyl)-1-ethenyl]-2-(2,4-dimethoxyphenyl)-1-ethylimidazo[1,2-a]pyridin-1-ium iodide (SIP-8) containing C6 aliphatic tails and 2,4-methoxy substituents in the 2-phenyl ring exhibited specificity to BSA. [25] Fluorescent properties of pentamethine cyanine dyes with cyclopentene and cyclohexene group in presence of biological molecules M.Yu. Losytskyy, K.D. Volkova, V.B. Kovalska, I.E. Makovenko, Yu.L. Slominskii, O.I. Tolmachev, and S.M. Yarmoluk Journal of Fluorescence. – 2005. – Vol. 15, No. 6. – P. 849-857. Abstract A series of pentamethine cyanine dyes with cyclohexene or cyclopentene group in polymethyne chain, assumed as DNA groove-binders, were studied as fluorescent probes for nucleic acids as well as for native and denatured proteins. It was revealed that the presence of methyl or dimethyl substituent in 5 position of the cyclohexene group hinders the formation of dye-DNA fluorescent complex, while the methyl substituent in 2 position leads to the increasing of the dye-DNA complex fluorescence intensity. The dyes SL-251, SL-1041, and SL-1046 containing methyl group in the 2 position of the cyclic group, are reported as bright DNA-sensitive dyes. The study of the dyes DNA-binding specificity demonstrated significant AT-preference that points to the groove-binding interaction mode. At the same time, the dyes SL-251, SL-377, and SL-957 with the 2-methyl substituted cyclohexene group were shown to be sensitive fluorescent dyes both for nonspecific (in SDS presence) proteins detection and for native BSA. [26] Studies on the spectral-luminescent properties of the novel homodimer styryl dyes in complexes with DNA V. B. Kovalska, I. O. Kocheshev, D. V. Kryvorotenko, A. Balanda and S. M. Yarmoluk Journal of Fluorescence. – 2005. – Vol. 15, No. 3. – P. 215-219. Abstract Series of homodimer styryls containing on (p-dimethylaminostyryl) pyridinium residues that are connected with aliphatic linkage group was synthesized. Spectral luminescent properties of obtained dyes in free state and in nucleic acids presence were studied. It was shown that DNA binding affinity of the novel homodimers exceeds that of parent monomer (p-dimethylaminostyryl)pyridine iodide. For homodimers with the linkage 4–10 carbon atoms preference in binding to DNA than to RNA was observed. It could be concluded that parent monomer has different mechanisms of binding to nucleic acids than corresponding homodimer dye. [27] Studies of mutagenic activity of fluorescent DNA-sensitive monomethinecyanine and carbocyanine dyes in Ames test B.P. Matselyukh, D.Ya. Matselyukh, V.B. Kovalska, K.D. Volkova, D.V. Kryvorotenko, and S.M. Yarmoluk Ukrainica Bioorganica Acta. – 2005. – Vol. 3, No. 2. – P. 27-34. Abstract Cyanine dyes are widely used as fluorescent probes for visualization of nucleic acids in electrophoretic gels. Unfortunately, some of these dyes are known to have mutagenic and carcinogenic activity. Here, four cyanine dyes D-6, D-9, CPent V and CCyan 2-O, proposed before as fluorescent dyes for DNA visualization in gels, were evaluated for mutagenicity in the Salmonella typhimurium assay (Ames assay) and cytotoxicity. The most sensitive of the studied dyes, mesosubstituted carbocyanine CCyan 2-O, was selected for further examination in acute toxicity assay on laboratory animals. Results indicated that according to the ‘2-fold rule' of assessing mutagenicity in the Ames assay monomethinecyanines D-6 and D-9 are not mutagenic, and carbocyanines CCyan2-O and CPent-V appeared to be weak mutagens. Mutagenic activity of carbocyanines was up to two times higher as compared to monomethine dyes. Carbocyanines CCyan2-O and CPent-V demonstrated higher cytotoxicity, being tested against Salmonella cells strain TA98, than did monomethinecyanines D-6 and D-9. Acute toxicity test indicates that the oral LD50 of CCyan 2-O for albino mice and rats are equal to 53.0 and 56.5 mg/kg, respectively. The lowest dose causing lethality LDLO for both species of rodents amounted to 20 mg/kg. [28] Toxicity evaluation of the series of cyanine dyes used for nucleic acids and proteins detection N.N. Nizamov, Z.F. Ismailov, E.N. Kurtaliev, Sh.N. Nizamov, F.U. Khaydarova, G. Khodjayev, O.P. Kukharenko, A.O. Balanda, S.M. Yarmoluk Ukrainica Bioorganica Acta. – 2005. – Vol. 3, No. 2. – P. 35-38. Abstract Potential toxicity of some commercial and novel dyes specifically interacting with biopolymer molecules was comparatively studied on bioobjects such as protozoa Paramecium caudatum, fungus Rhizoctonia solani and E. coli. The toxicity and critical concentrations were determined for compounds Rhodamine C, Cyan-40, SIL, SBT, D-174. Paramecium caudatum revealed the highest sensitivity to studied dyes and can be used as sensitive indicator organism in the toxicity tests. [29] Fluorescent homodimer styrylcyanines:synthesis and spectral-luminiscent stidies in nucleic acids and protein complex Kovalska V.B., Kryvorotenko D.V., Balanda A.O., Losytskyy M.Yu., Tokar V.P., Yarmoluk S.M. Dyes and Pigments. – 2005. – Vol. 67, No. 1. – P. 47-54. Abstract A series of homodimer styrylcyanine dyes based on (p-dimethylaminostyryl)pyridinium, (p-dimethylaminostyryl)benzoxazolium, (p-dimethylaminostyryl)benzothiazolium, (p-dimethylaminostyryl)-1,3,3-trimethyl-3H-indolium residues were synthesized. Spectral-luminescent properties of the obtained homodimers in free state and in the presence of nucleic acids and BSA were studied. Homodimer styrylcyanines with the length of linkage group of 2 or more carbon atoms demonstrated a DNA-binding preference. Significant long-wave shifts of fluorescence and emission maxima of dyes with short linkage group could be explained by the interaction between chromophores due to the short distance between them, as it is the case for molecular aggregates. Homodimer dyes based on the (p-dimethylaminostyryl)pyridinium residue having linkage group of 5 or more carbon atoms interact with dsDNA with significant emission increase and could be used as DNA specific fluorescent probes. [30] Chameleon labels for staining and quantifying proteins Wetzl B.K., Yarmoluk S.M., Graig D.B., and Wolfbeis O.S. Angew. Chem. Int. Ed. – 2004. – Vol. 43. – P. 5400–5402. Abstract Herein we report an entirely new class of labels for gel electrophoresis and also for general protein quantitation. Pyrylium groups react with amines to form the respective pyridinium analogues. We have synthesized the new labels by attaching a pyrylium group to a small aromatic group (thereby forming one of the smallest blue and fluorescent chromophores known) and this resulted in the label Py-1. [31] Spectroscopic study of the fluorescent dyes interaction with DNA I.V. Valyukh, V.V. Vyshnyak, A.V. Slobodyanyuk, S.M. Yarmoluk Functional Materials. – 2003. – Vol. 10, No. 3. – P. 528–533. Abstract The interaction between DNA and fluorescent dyes 4-[2-(4-Dimethylaminophenyl)ethenyl]-1-methylpyridinium iodide and thiazole orange have been studied using absorption and fluorescence spectra. For both dyes, a drastic fluorescence intensity enhancement (up to 103 times) is observed in the presence of DNA. Fluorescence measurements under polarization have been used to evaluate the rigidity of dye-DNA complexes. The possible binding modes of these dyes to DNA are discussed. Fluorescence depolarization of the complexes studied was compared with polarization properties of dye molecules emitting in solutions with some amount of glucerin added to decelerate rotation. [32] Pat. US2003175988, DE 101 53 818 A 1. Method and compounds for the fluorescent labeling of biomolecules and polymer particles. 5.11.2001. Appl. 18.09.2003. 6 pp. Wolfbeis O.S., Kostenko O.M., Tolmachev O.I., and Yarmoluk S.M. Abstract A new efficient biomolecule labelling method was proposed as a simple and convenient alternative to known procedures. It was based on the use of pyrilium salts as amine-specific reactive group. Unlike usual amino reactive dyes pyrilocyanines, used in our method, are quite stabile in normal conditions and do not require any activation before reaction of conjugation. [33] Luminescence spectroscopic studies of trimethinecyanines substituted in polymethine chain with nucleic acids and proteins V.B. Kovalska, M.Yu. Losytskyy and S.M. Yarmoluk Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy. - 2004. - Vol. 60, No. 1-2. - P. 129-136. Abstract The series of symmetrical β-substituted and α,γ-substituted trimethinecyanine dyes were studied for their absorption and fluorescent characteristics in unbound state and in the presence of nucleic acids and proteins. It was shown that β-substituted and α,γ-bridged trimethinecyanines containing extended heterocyclic systems or N-phenyl as well as N-cyclohexyl substituents demonstrate increased affinity to proteins. At the same time the presence of both N-phenyl and N-cyclohexyl substituents leads to the decrease of the dye fluorescence intensity in complexes with nucleic acids. For trimethinecyanines similarly to unsymmetrical monomethines the presence of N-ω-hydroxy alkyl substituents results in the increase of fluorescence intensity of dye-DNA complex and the emission decrease of dye-RNA complex. [34] Interaction of cyanine dyes with nucleic acids: XXXI. Using of polymethine cyanine dyes for the visualization of DNA in agarose gels B.P. Matselyukh, S.M. Yarmoluk, A.B. Matselyukh, V.B. Kovalska, I.O. Kocheshev, D.V. Kryvorotenko, S.S. Lukashov Journal of Biochemical and Biophysical Methods, 2003, 57, 1, pp. 35-43. Abstract Fifteen polymethine cyanine dyes were studied as fluorescent stains for DNA in electrophoretic gels. Among studied cyanines, two dyes CPent V and CCyan 2-O most effectively visualized covalently closed and linear double-stranded DNA molecules in gels under standard conditions using UV-illumination, green filter and black-and-white photo film. Ethidium bromide was 1.2-1.6 times more effective as compared to cyanine dyes in staining of DNA in the concentration range of 8-18 ng, while studied cyanines were more sensitive to DNA quantity above 50 ng. [35] The mechanism of interaction of monomethine cyanine dye Cyan 40 with dsDNA: computer modelling O.Ya. Yakovenko, A.G. Golub, M.Yu. Losytskyy, V.G. Bdzhola, S.M. Yarmoluk Biopolymers and Cell, 2003, 19, 1, pp. 93-98 (in Ukrainian). Abstract The mechanism of interaction of cyanine dye Cyan 40 and DNA was studied by molecular modelling methods. The energy of interaction of the terminal parts of π-electrons clouds of aromatic heterocycles of DNA bases and Cyan 40 molecule was calculated by ab initio methods. We supposed that Cyan 40 interacted with dsDNA according to full-intercalation binding mode. Using combination of Amber, ÌÌ+, AM1, PM3 and ab initio calculation methods, the full-intercalation interaction of Cyan 40 with DNA was proved to be more favourable energetically than half-intercalation one. The series of the dye-DNA complex metastates were found, illustrating the hypothesis of three consequent stages of Cyan 40 interaction with DNA: (1) association of Cyan 40 with DNA minor groove, (2) interaction by half-intercalation mechanism, and (3) fixation of Cyan 40 between DNA bases pairs by the mechanism of full-intercalation. [36] Spectral properties of symmetrical trimethine cyanines with a-, g- substituted polymethine chain in the presence of nucleic acids I. Valyukh, A. Slobodyanyuk, V.B. Kovalska, A. Grangan, Y.L. Slominskii, S.M. Yarmoluk Journal of Physical Studies, 2002, 6, 2, pp.236-242. Abstract Spectral-luminescent properties of four symmetrical trimethine cyanine dyes with a-, g- substituted polymethine bridge in a free state and in the presence of nucleic asides (NA) have been investigated. It has been shown that replacement of ethylene bridge by vinylene one in a polymethine chain of the trimethine cyanines influenced meaningfully the spectral properties of these dyes. The bands of aggregates in absorption spectra of the aqueous solutions of free dyes were observed. CPentE, CPentV and CpentE-OMe form high fluorescence complexes in the NA presence. These dyes can be used as luminescent probes for NA detection in homogeneous assays. The introduction of methoxy groups into the residue of the tricarbocyane with ethylene bridge does not make worse its properties as probe for NA detection. The presence of these groups in an analog with the vinylene bridge has diminished the fluorescence intensity in the CPentE-OMe – NA complex. The obtained results for CpentE and CPentV indicate that these dyes are bound into the DNA groove. [37] Interaction of cyanine dyes with nucleic acids. Spectral-luminescent properties of pyrilium and pyridinium mono/trimethincyanines V.B. Kovalska, I.V. Valyukh, O.M. Kostenko, S.Yu. Dmitrieva, O.I. Tolmachov, S.M. Yarmoluk Biopolymers and Cell, 2002, 16, 6, pp.540-546 (in Ukrainian). Abstract The spectral-luminescent properties of series of pyrylium and pyridinium mono- and trimethincyanines as possible fluorescent dyes for the nucleic acids (NA) detection and labeling were studied. We propose 5,6-methilendioxy-benzothiazolomonomethinepyridocyanine MDO40 for using in homogeneous assay, as well as for the construction of oligonucleotides-based “light-up probes”. [38] Interaction of cyanine dyes with nucleic acids. New (pyrido)(thio)trimethincyanine dye CCyan 40 for fluorescent labeling of oligonucleotides S.M. Yarmoluk, M.Yu. Losytskyy, V.B. Kovalska, S.M. Tomachynski, T.Yu. Ogul'chansky, O.M. Kostenko, V.V. Kurdyukov, O.I. Tolmachev Biopolymers and Cell, 2002, 16, 4, pp. 340-346 (in Ukrainian). Abstract The spectral-luminescent characteristics of the CCyan 40 carbocyanine dye in free state and in the presence of DNA and synthetic polynucleotides were studied. The dye has high molar extinction coefficient (5.7x104 M-1cm-1) and average quantum yield value (0.042). In the presence of DNA, poly(dGC/dGC) and poly(dA/dT) the quantum yield of CCyan 40 increases in 5.5, 3.8 and 1.8 times respectively. We recommend the dye CCyan 40 for the fluorescent labeling of oligonucleotides using the reaction, proposed by us earlier. [39] Interaction of cyanine dyes with nucleic acids: XXVI. Intercalation of the trimethine cyanine dye Cyan 2 into double-stranded DNA: study by spectral luminescence methods S.M. Yarmoluk, S.S. Lukashov, M.Yu. Losytskyy, B. Akerman, O.S. KornyushynaSpectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2002, 58, 14, pp.3223-3232. Abstract The interaction between double-stranded (ds) DNA and the cyanine dye Cyan 2 has been studied with spectral luminescence methods. Binding constant values have been determined by fluorescence titration and dye distribution in the two-phase system ethyl acetate-water (3.6×104 and 1.5×104 M−1, respectively). Cyan 2 exhibits a small specificity for guanine-cytosine (GC) sequences in total DNA and synthetic polydeoxynucleotides poly(dA/dT) and poly(dGdC/dGdC). The DNA complexes with Cyan 2 are stable at high-ionic strength solution when NaCl is added. The dye molecule complexed with DNA is apparently shielded from the anionic quencher—iodide ion. The negative linear dichroism of the visible absorption band of aligned Cyan 2–DNA complexes indicates that the bound dye lies almost perpendicularly to the DNA helix axis. The linear dichroism of the absorption band at 260 nm suggests a considerable change in the DNA B-form. The results are consistent with an intercalative binding interaction between Cyan 2 and ds DNA. [40] Interaction of cyanine dyes with nucleic acids. Meso-substituted trimethincyanines for fluorescent detection of nucleic acids S.S. Lukashov, M.Yu. Losytskyy, Yu.P. Kovtun, S.M. Yarmoluk Biopolymers and Cell, 2002, 18, 3, pp. 243-252 (in Ukrainian). Abstract A series of trimethinecyanines with different meso-substituents in polymethine chain has been synthesized, and spectral luminescent properties of the dyes in the presence of double-stranded DNA, RNA and bovine serum albumin (BSA) protein have been examined. The dyes fluorescence enhances up to 270-fold in the presence of nucleic acids while in the presence of protein the increase is lower. The highest rise of fluorescence intensity has been observed for the dye-nucleic acid complexes of meso-methyl-substituted dyes. When large substituents like aryl- or tret-alkyl groups are linked to the meso-position of dyes polymethine chain by flexible chain from 1-3 CH2-groups, the fluorescent properties of these dyes are similar to those of the meso-methyl-substituted dyes. [41] An investigation of tricarbocyanines "Stains-All" and "iso-Stains-All" as fluorescent nucleic acids probes V.B. Kovalska, I.V. Valyukh, S.S. Lukashov, Yu.L. Slominskii and S.M. Yarmoluk Journal of Fluorescence, 2002, 12, 2, pp. 209-212. Abstract Luminescent properties of carbocyanine dyes Stains-All and its isomer iso-Stains-All were studied in the presence of nucleic acids. Both dyes show sufficient fluorescent intensity increase in the presence of DNA and RNA and may be used as fluorescent probes for nucleic acids (NA) detection in homogeneous assays. It was supposed that Stains-All and iso-Stains-All bind with nucleic acids through different interaction modes [42] New approach for fluorescent peptide labeling with pyrylium cyanine dyes O.M. Kostenko, S.Y. Dmitrieva, O.I. Tolmachev and S.M. Yarmoluk Journal of Fluorescence, 2002, 12, 2, pp.173-175. Abstract New approach for fluorescent peptide labeling with cyanine dyes utilizing the reaction of pyrylium salts with aliphatic aminogroup is proposed. The reaction of two pyrylium cyanines dye acids was investigated. Lysocyme was used as a model peptide for conjugation. The proposed method can be used as a simple and convenient alternative for the known procedures because it does not require preparation of the unstable amino-reactive intermediates from a carboxyl- or sulfo-derivative of cyanine dyes. [43] Influence of the aggregation of homo-N-mer cyanine dyes on the nucleic acids detection sensitivity S.M. Yarmoluk, V.B. Kovalska, I.O. Kocheshev Journal of Fluorescence, 2002, 12, 2, pp.155-157. Abstract Novel benzothiazolopyridinium homo-n-mer cyanine dyes are proposed for nucleic acid fluorescent detection. Dependence of the sensitivity of detection in solution from the dye molecules/DNA base pairs ratio was studied. It was shown that the presence of the dye excess could significantly decrease the detection limit. We believe this could be explained by the formation of the dye aggregates on DNA surface. [44] Davydov splitting in spectra of cyanine dye J-aggregates, formed on the polynucleotides M.Yu. Losytskyy, V.M. Yashchuk, S.S. Lukashov and S.M. Yarmoluk Journal of Fluorescence, 2002, 12, 1, pp.109-112. Abstract It was found in our previous work (T. Y. Ogul'chansky et al. (2001) Spectrochimica Acta Part A 57, 2705-2715) that the carbocyanine dye Cyan βiPr forms J-aggregates in the groove of poly(dA)-poly(dT). In the present paper we study in detail the spectral properties and energy levels structure of the Cyan βiPr J-aggregates by means of absorption and fluorescence spectroscopy and polarization measurements. The energy structure of an aggregate consists of at least two exciton zones, and dipole moments of the absorption transitions to these zones are oriented at an angle of about 90° one to another. It was supposed that the transition moment of the lower zone is parallel to the polynucleotide axis and the moment of the upper zone is perpendicular. The fluorescent transitions are possible only from the lower exciton zone, while the excitations of higher zone undergo nonradiative transitions to the lower one. [45] Spectroscopic studies of α,γ-disubstituted trimethine cyanine: new fluorescent dye for nucleic acids I.V. Valyukh, V.B. Kovalska, Y.L. Slominskii and S.M. Yarmoluk Journal of Fluorescence, 2002, 12, 1, pp.105-107. Abstract Spectral properties of 3-methyl-2-{3-[3-methyl-1,3-benzothiazolo-2(3H)-ylidene]-1,4-cylopentadien-1-yl}-1,3-benzothiazolo-3-ium tosylate (Cyan-Cpentd) in a free state and in the complexes with nucleic acids and synthetic polynucleotides have been investigated by absorption and fluorescence spectroscopy. Significant fluorescence intensity enhancement of dye-nucleic acids complexes is observed. For the first time Cyan-Cpentd is proposed as a new probe for nucleic acid detection. Binding mechanism of Cyan-Cpentd is discussed in view of the NA-ligand interaction models. [46] Nonradiative deactivation of the electronic excitation energy in cyanine dyes: influence of binding to DNA S.M. Yarmoluk, M.Yu. Losytskyy, V.M. Yashchuk Journal of Photochemistry & Photobiology, B: Biology, 2002, 67, 1, pp. 57-63. Abstract The processes of nonradiative deactivation of electronic excitation energy in cyanine dyes determine their quantum yield. Because of that, the study of the influence of cyanines binding to DNA on these processes can provide information on the causes leading to the cyanines fluorescence intensity enhancement in the presence of DNA. In the presented paper, the activation energies of nonradiative degradation of electronic excitation, quantum yields and rate constants of nonradiative transitions of several cyanines in free state and in the presence of DNA were established and compared. The mechanisms of nonradiative deactivation of dye excitation energy were discussed. [47] The interaction of cyanine dyes with nucleic acids. XXXIII. Meso-methylsubstituted trimethincyanines, as possible probes for fluorescent nucleic acid detection S.S. Lukashov, I.E. Makovenko, M.Yu. Losytskyy, Yu.L. Slominskii, S.M. Yarmoluk Biopolymers and Cell, 2001, 17, 5, pp.448-454 (in Ukrainian). Abstract
[48] Interaction of cyanine dyes with nucleic acids. Simulation of the aggregates structures and their electronic transitions for homo-n-mere benzothiazole cyanine dye using the quantum-mechanical methods S.M. Yarmoluk, V.B. Kovalska, I.O. Kocheshev, Yu.Yu. Kachkovskiy Current Studies of Biotechnology, 2002, V.II. - Environment, pp.117-123. Abstract Earlier we studied the aggregation of novel benzothiazolopyridinium homo-n-mere cyanines in water solution and in the DNA presence. We suggested the existence of associates that include two and more dye's chromophores and proposed their possible structures [Kochehev I. et al. Biopolymers and Cell, 2000, V.15, .557-567.]. In present paper we performed the computer simulation of electronic transitions of dye aggregates built according to the structure proposed. The results of simulations provide the evidence that proposed structures are similar to the homo-n-mere dyes' associates that are formed in solution in the presence of DNA. [49] Interacion of cyanine dyes with nucleic acids. XXIII. Computer simulation of "half-intercalative" interaction of monomethyne cyanine dyes with DNA G.O. Kachkovsky, S.S. Lukashov, S.M. Yarmoluk, G.Kh. Matsuka Biopolymers and Cell, 2001, 17, 4, pp. 331-336 (in Ukrainian). Abstract Computer simulation of interaction with DNA of six monomethyne cyanine dyes was carried out using MM+, AMBER, PM3 and CNDO/2 methods from Hyperchem 5.0 program packet. Geometry optimization of complexes was carried out, charge density for ground S0 and exited S1 states and potential energies of heterocycles' rotation around the bonds of methyne group for unbound dyes and dyes in complexes with DNA were calculated. Obtained energies of complexes are in agreement with observed fluorescent intensities of dyes in presence of DNA. Calculated electron density distribution changes in dyes' molecules upon DNA binding diverge with observed Stocks' shift values changes. Rotation potential energy values show that rigidity of fixation of dye's molecule planar conformation increases strongly upon binding to DNA. [50] The interaction of cyanine dyes with nucleic acids. 22. Spectral-luminescent properties of monomethine pyrylium and pyrydinium cyanines and their DNA-complexes S.S. Lukashov, G.O. Kachkovskyy, M.Yu. Losytskyy, S.M. Yarmoluk Biopolymers and Cell, 2001, 17, 3, pp. 242-248 (in Ukrainian). Abstract We obtained a number of pairs of monomethyne cyanines, dyes of each pair having similar structure and contrary direction of electric dipole moment, and investigated the spectral-luminescent properties of these dyes in free state and at presence of double-stranded DNA. The change of dyes stockes shifts values under interaction with DNA is the evidence for the location of dyes in complexes with DNA in dipole medium, that confirms the "half-intercalation" interaction model. Considering the change of dyes stockes shifts under interaction with the DNA to be the manifestation of changes in electronic assymethry of these dyes, we made an assumption about the location of each dye inside the DNA. As the result it was concluded, that the dye molecule orientation in DNA-complex is defined by the dye's geometry, and its dipole moment direction is of secondary importance. [51] Interaction of cyanine dyes with nucleic acids. XVIII. Formation of the carbocyanine dye J-aggregates in nucleic acid grooves T.Yu. Ogulchansky, M.Yu. Losytskyy, V.B. Kovalska, S.S. Lukashov, V.M. Yashchuk and S.M. Yarmoluk Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2001, 57, 13, pp. 2705-2715. Abstract Spectral properties of previously synthesized carbocyanine dye 3- methyl- 2- [3- methyl -2- (3- methyl- 2,3- dihydro- 1,3- benzothiazole- 2- iliden)- 1-butenyl]- 1,3- benzothiazole -3-il iodide (Cyan biPr) in solutions in the presence and in the absence of deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) have been studied within a wide concentration range. It has been established that Cyan biPr molecules bind to DNA mainly via groove binding mode. Under an increase of the dye concentration, Cyan biPr molecules form J-aggregates in the DNA-added solutions unlike RNA-added solutions. A study of sequence selectivity showed that J-aggregates of Cyan biPr molecules are situated in DNA minor grooves and have the structure, which causes the Davydov splitting of the J-aggregate absorption band. An increase in solution ionic strength leads to the release of dye molecules from DNA grooves and prevents J-aggregates. Based on the data obtained, the conclusion was made that formation of the dye molecule J-aggregates in DNA grooves can be applied to selective detection of DNA fragments. Strong dependence of the J-aggregate formation on the dA:dT sequence implies the possibility to use it in determination. [52] Proteins and cyanine dyes. Part III. Synthesis and spectroscopic studies of benzothiazolo-4-[1,2,6-trimethylpyridinium] monomethine cyanine dyes for fluorescent detection of bovine serum albumin in solutions S.M. Yarmoluk, D.V. Kryvorotenko, A.O. Balanda, M.Y. Losytskyy, V.B. Kovalska Dyes and Pigments, 2001, 51, 1, pp.41-49. Abstract The spectral-luminescent properties of a series of new cyanine dyes as possible probes for homogeneous assay of proteins have been studied. The fluorescent cyanine dyes development have been based on the principle of "affinity-modifying group" with the use of benzothiazolo-4-[1,2,6-trimethylpyridinium] monomethine as template. It was shown that the cyanine dye P-5 can be used as a sensitive and specific fluorescent probe for the detection of BSA. [53] Interaction of cyanine dyes with nucleic acids. VII. Carbocyanine dyes, substituted in polymethine chain, as possible probes for fluorescent nucleic acid detection S.S. Lukashov, M.Yu. Losytskyy, Yu.L. Slominskii, S.M. Yarmoluk Biopolymers and Cell, 2001, 17, 2, pp.169-177 (in Ukrainian). Abstract To extend the application of carbocyanine dyes in fluorescent nucleic acid detection, the use of dyes substituted in polymethine chain is proposed. A series of thiacarbocyanine derivatives with alkyl substituents in a polymethine chain have been synthesized, and the spectral luminescent properties of dyes in the presence of double-stranded DNA, RNA and bovine serum albumin (BSA) have been examined. The intrinsic fluorescence of all derivatives prepared is lower than that of unsubstituted thiacarbocyanine. The dyes show 1.75-76-fold fluorescence enhancement in the presence of nucleic acids. Lower enhancement (1.75-16.9 times) takes place in the presence of large excess of BSA. The highest fluorescence intensity for dye-nucleic acid complexes is observed for the b -methyl-substituted dye that is more than two times higher as compared to thiacarbocyanine. The substituents larger than methyl group disturb planarity of thiacarbocyanine fluorophore excessively, and the fluorescence intensities of nucleic acid complexes of corresponding dyes are lower. [54] Interaction of cyanine dyes with nucleic acids. XXVII. Synthesis and spectral properties of novel homodi- and homotrimeric monomethine cyanine dyes S.M. Yarmoluk, V.B. Kovalska, I.A. Kocheshev Dyes and Pigments, 2001, 50, 1, pp.21-28. Abstract Three new methods of preparation based on reactions of pyrylium salts with primary amines, acylation of polyamines with activated carboxyl derivatives of cyanine dyes, diisocyanate with aminoderivatives of cyanine dyes were used to obtain fluorescent homodi- and, for the first time, homotrimer cyanine dyes. Spectral-luminescent properties of synthesized dyes and their nucleic acid complexes were studied. [55] Interaction of cyanine dyes with nucleic acids. XXI. Arguments for half-intercalation model of interaction S.M. Yarmoluk, S.S. Lukashov, T.Yu. Ogul'chansky, M.Yu. Losytskyy, O.S. Kornyushyna Biopolymers, 2001, 62, 4, pp.219-227 Abstract The spectral luminescent properties of two groups of monomethine cyanine dyes were studied in the presence of DNA. The first group included five dyes with 5,6-methylenedioxy-[d]-benzo-1,3-thiazole heterocycle and their unsubstituted analogs. Five monomethine pyrylium cyanines and their N-methyl-pyridine analogs were included in the second group. In each pair the pyrylium and pyridine dyes had similar geometry but differed in charge density distribution. The results presented some evidence in favor of the half-intercalation interaction mode between the studied dyes and DNA. When the benzothiazole residue had the lowest electron donor ability between the two heterocycles in the dye molecule, its substitution with the bulky methylenedioxy group led to a significant decrease in fluorescence enhancement of the dye-DNA complex. On the contrary, when the substituents that create steric hindrance (e.g., methylenedioxy and methyl groups) were introduced into the heterocycle with the higher electron donor ability, the fluorescence enhancement value of the dye-DNA complex was virtually unchanged. The changes in the Stock's shift values upon the formation of the dye-DNA complexes were in agreement with the proposed half-intercalation model. Interestingly, in the dye-DNA complexes the pyrylium dyes probably resided in a place similar to the pyridine ones. It is possible that the benzothiazole (or benzooxazole) ring intercalated between the DNA bases and the pyrylium (or pyridine) residue was located in the DNA groove closer to the phosphate backbone. [56] Interactions of cyanine dyes with nucleic acids. XXIV. Aggregation of monomethine cyanine dyes in presence of DNA and its manifestation in absorption and fluorescence spectra T.Yu. Ogul'chansky, M.Yu. Losytskyy, V.B. Kovalska, V.M. Yashchuk & S.M. Yarmoluk Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2001, 57, 7, pp.1525-1532 Abstract Absorption, fluorescence emission and excitation spectra of benzothiazole cyanine dyes — thiazole orange (TO) and 7-methyl-6-(3-methyl-2,3-dihydro-1,3-benzothiazol-2-ylidenmethyl) [1,3] dioxolo [4',5':4,5] benzo [d] [1,3] thiazolium methylmethosulfate (Cyan 13) — were investigated over a wide concentration range. The dyes form aggregates with a 'sandwich'-like structure in water solution. At low dye to DNA concentration ratios, Cyan 13 and TO monomers appear to interact with the DNA. On increasing the dye to DNA concentrations ratio, free dye molecules aggregate with the DNA-bound ones. The spectra of the free dye aggregates and the aggregates formed on the DNA, are characterized by an anomalously large (more than 100 nm) Stokes shift. This suggests, that the -electron systems of the aggregates undergo substantial changes in excited state, compared to those of the monomers. The formation of aggregates consisting of the free and DNA-bound dye molecules can be explained using the half-intercalation model of the interaction of the cyanine dye monomers with the DNA. [57] Interaction of cyanine dyes with nucleic acids. XXV. Influence of affinity-modifying groups in the structure of benzothiazol-4-[2,6-dimethylpyridinium] dyes on the spectral properties of the dyes in the presence of nucleic acids S.M. Yarmoluk, V.B. Kovalska, D.V. Kryvorotenko, A.O. Balanda & T.Yu. Ogul'chansky Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2001, 57, 7, pp.1533-1540 Abstract Novel monomethine pyridinium cyanine dyes of similar structure and containing 'affinity-modifying' groups of different chemical nature were studied by spectral-luminescent methods as possible fluorescent probes for the nucleic acids detection. It was shown that the nature of the functional groups in the dye linker influences the fluorescent properties of the dye–nucleic acids complexes. Incorporation of a hydroxyl group into the linker structure leads to a significant increase in the fluorescence intensity of the dye — double-stranded DNA complexes relative to the parent dye Cyan 40. [58] Peptides and cyanine dyes. 2. Covalent labelling of amino-reactive pyrilium dye for the conjugation of amino acids and peptides by means of the reaction of pyrilium cyanine dyes with amines. O.M. Kostenko, S.U. Dmitrieva, S.M. Yarmoluk Biopolymers and Cell, 2001, 17, 1, pp. 80-84. Abstract Here we propose a new amino-reactive pyrilium dye for the conjugation with peptides through N-terminal and N-e -lysine's aminogroups. The reactivity of Cyan 39 towards eleven most interesting aminoacids has been investigated in triethylammonium acetate and sodium carbonate-bicarbonate buffers. The conjugation with 72-aminoacid model peptide has been carried out successfully. [59] Interaction of cyanine dyes with nucleic acids. XX. New methods for the preparation of fluorescent probes based on benzothiazol-4-[2,6-dimethylpyridinium] cyanine dyes S.M. Yarmoluk, D.V. Kryvorotenko, V.B. Kovalska Dyes and Pigments 2001, 48, 3, pp. 165-172. Abstract Two new methods of preparing fluorescent dyes containing affinity-modifying groups for nucleic acid detection are proposed. One method of preparation is based on the reaction of the pyrylium monomethine cyanine dye 2-(2,6-dimethyl-pyran-4-ilidenmethyl)-3-methyl-benzothiazolium perchlorate with primary amines. The second method of preparation involves the amidation of carboxy substituted dyes with appropriate amines. [60] Proteins and cyanine dyes. IV. Benzothiazol-4-[2,6-dimethylpyridinium] monomethine cyanine dyes for homogeneous detection of proteins D.V. Kryvorotenko, M.Yu. Losytskyy, A.O. Balanda, S.M. Yarmoluk Vistnyk of “Lvivs'ka Politechnika” University – Chemistry 2000, 414, pp. 127-135 (in Ukrainian). Abstract The spectral-luminescent properties of the series of novel monomethine cyanine dyes in presence of proteins and nucleic acids were studied. The P-5 dye, which contains the 3-indolylacetic acid template in its structure, increased its fluorescence intensity in the complexes with BSA in 3 orders of magnitude. The results of studies show us that binding of P-5 with BSA occurs through the interaction of dye with the specific protein site, formed at the level of tertiary structure. We consider the P-5 cyanine to be perspective fluorescent probe for the homogeneous protein detection. [61] Interaction of cyanine dyes with nucleic acids. XVII. Towards an aggregation of cyanine dyes in solutions as a factor facilitating nucleic acid detection T.Yu. Ogul'chansky, V.M. Yashchuk, M.Yu. Losytskyy, I.O. Kocheshev, S.M. Yarmoluk Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2000, 56, 4, pp. 805-814. Spectral properties of newly synthesized cyanine dyes, namely 1-[6-(4-{6-[2,6-dimethyl-4-(3-methyl-2,3-dihydro- 1,3-benzothiazol-2-ylidenmethyl)-1-pyridiniumyl]hexanoyl}piperazino)-6-oxohexyl]-2,6-dimethyl-4- (3-ethyl-2,3- dihydro-1,3-benzothiazol-2- ylidenmethyl)pyridinium (K-6) (bichromophoric dye) and 1-[5-di(3-{5-[2,6- dimethyl-4- (3-methyl-2,3-dihydro-1,3-benzothiazol-2-ylidenmethyl)-1-pyridiniumyl]pentylcarboxamido}propyl) carbamoylpentyl]-2,6-dimethyl-4-(3-methyl-2,3-dihydro-1,3-benzothiazol-2-ylidenmethyl) pyridinium (K-T) (trichromophoric dye) in solutions in the presence of and without deoxyribonucleic acid (DNA) were studied within a wide concentration range. It has been established that absorption, as well as fluorescence of investigated dye solutions, without DNA are mainly determined by H-aggregates of dye molecules. On the contrary, the fluorescence of dye solutions in the presence of DNA gives an intrinsic dye molecular fluorescence. H-aggregates are broken because of binding dye molecules with DNA. It has been suggested that both K-T and K-6 molecules bind mainly with DNA via the interaction of two chromophores. As the ratio of the number of dye molecules to that of DNA base pairs increases with an increase in dye concentration, a formation of dye molecule H-aggregates on DNA molecules are observed. Such aggregates have a different structure than those formed in the solutions without DNA. On the grounds of the data obtained, it is concluded that it is possible to use a dye aggregation capable of obtaining higher values for fluorescence enhancement of the DNA stains. [62] Electronic singlet excitations transfer in DNA-cyanine dye system M.Yu. Losytskyy, V.M. Yashchuk, S.M. Yarmoluk Functional Materials, 2000, 7, 4, pp. 638-642. Abstract The processes of singlet electronic excitation transfer in deoxyribonucleic acid (DNA)- cyanine dye system at room temperature were studied. The DNA nucleotide bases have been shown to absorb UV-radiation mainly as separate centers, so the excitations in DNA are localized. The singlet electronic excitation energy of DNA can be transferred to the dye fixed on DNA. The efficiency of such transfer depending on the DNA to dye concentration ratio has been estimated. Possible ways of the excitation transfer between DNA base pairs and the contribution of dipole-dipole and exchange interaction in the process are discussed. [63] Interaction of cyanine dyes with nucleic acids. Part 19: new method for the covalent labeling of oligonucleotides with pyrylium cyanine dyes. S.M. Yarmoluk, A.M. Kostenko, I.Y. Dubey Bioorganic and Medicinal Chemistry Letters, 2000, 10, 9, pp. 2201-2204. Abstract New chemistry for the fluorescent labeling of oligonucleotides with cyanine dyes is proposed. It is based on the use of pyrylium salts as amine-specific reagents. Monomethyne pyrylium cyanine dye was covalently linked to 5'-aminoalkyl modified oligonucleotide, with simultaneous conversion of the non-fluorescent dye into fluorescent pyridinium cyanine structure. [64] The interaction of cyanine dyes with nucleic acids. Õ²². Novel monomethyne cyanines based on the 5,6-methylenedioxy-benzothiazole and spectral-luminescent properties of their complexes with nucleic acids S.S. Lukashov, M.Yu. Losytskyy, S.M. Yarmoluk, Y.L. Slominskii Biopolymers and Cell, 2000, 16, 6, pp. 562-572 (in Ukrainian). Abstract Seven novel monomethyne cyanine dyes with the 5,6-methylenedioxy-benzothiazole ring fused with the heterocycles of different geometry and basisity were synthesized. Their optical properties in presence of dsDNA, RNA and protein were studied. Some of synthesized dyes are perspective objects for further investigations and applying in the NA analysis. The comparison of fluorescent properties of studied dyes and their analogues with unsubstituted benzothiazole ring in presence of nucleic acids allowed us to confirm the hypothesis of “half-intercalation” of monomethyne cyanines into dsDNA. [65] The interaction of cyanine dyes with nucleic acids. 14. Spectral peculiarities of several monomethyne benzothiazole cyanine dyes and their interaction with DNA T.Yu. Ogul'chansky, V.M. Yashchuk, S.M. Yarmoluk, M.Yu. Losytskyy Biopolymers and Cell, 2000, 16, 5, pp. 345-355 (in Ukrainian). Abstract Absorption, fluorescence emission and excitation spectra of monomethyne benzthiazole cyanine dyes – thiazole orange (TO), Cyan 40 and Cyan 13 – have been studied in a wide concentration range in the presence and without DNA. It has been established that both molecular and aggregate absorption as well as both molecular and aggregate fluorescence are observed for the solutions of the dyes investigated, the aggregates electron system being essentially transformed after photon absorption. It has been shown that the own fluorescence of dye molecules dominates in emission of investigated dye solutions with DNA. Its considerable quantum yield can be explained by the dye molecule fixation on the DNA molecule. The concentration quenching of this fluorescence has been found and it is accompanied with the appearance of a new absorption band. It has been supposed that that the aggregates consisting of dye molecules fixed on DNA and free dye molecules are formed. On the ground of the data obtained we have concluded that the dye molecules interact through half-intercalation mode with DNA molecules and that the model of full intercalation is inapplicable to the interaction of the dyes investigated with DNA molecules. [66] The interaction of cyanine dyes with nucleic acids. 16. New fluorescent benzothiazole-4-[2,6-dimethylpyridinium] monomethyne cyanine dyes D.V. Kryvorotenko, V.B. Kovalska, S.M. Yarmoluk Biopolymers and Cell, 2000, 16, 4, pp. 320-326 (in Ukrainian). Abstract New fluorescent probes for nucleic acids detection based on 2-[1-(5-carboxypentyl)- 2,6-dimethyl-1,4-dihydro-4-pyridinilidenmethyl]-3-methyl-1,3- benzothiazole-3-ium perchlorate have been synthesized. The spectral properties of the compounds obtained and their complexes with nucleic acids have been investigated. The influence of different functional substituents in the dye molecule on its interaction with nucleic acids has been studied. The results obtained show that the inclusion of hydroxy- groups and acridine fragment in the structure of cyanine increases the sensibility of the nucleic acids detection. [67] Proteins and cyanine dyes. 1. Asymmetrical carbocyanine dye 14 K - a new fluorescent probe for homogeneous detection of proteins in solution S.M. Yarmoluk Biopolymers and Cell, 2000, 16, 3, pp. 236-239 (in Ukrainian). Abstract An interaction of carbocyanine 14K (6,8-diethyl-7-[E-3-(8-ethyl-7,8-dihydro-[1,3]-thiazolo- [4'‚ 5', 3, 4], benzo-[d]-[1,2,3]-thiadiazolo-7-iliden)- 1-propenyl]-6H-imidazolo-[4'‚ 5'‚ 5, 6]tiodiazolo-8- ium iodide) with four proteins in both native and denatured states has been studied. The difference in fluorescent enhancement of this dye in the presence of various proteins has been shown. The fluorescence intensity of 14K in the presence of denatured proteins increased by 25 - 40 % in comparison with the fluorescence intensity of this dye in the complexes with corresponding native proteins. A possible mechanism of increasing the dye fluorescence under the interaction with proteins and nucleic acids has been proposed. The influence of different substances on fluorescent properties of 14K has been studied. [68] Interaction of cyanine dyes with nucleic acids. 10. Synthesis and spectral properties of novel benzothiazol-4-[2,6-dimethylpyridinium] monomethyne cyanine dyes D.V. Kryvorotenko, V.B. Kovalska, S.M. Yarmoluk Biopolymers and Cell, 2000, 16, 2, pp. 145-152 (in Ukrainian). Abstract A number of new derivatives of 2,6-dimethyl-4-(3-methyl-3H-benzothiazol-2-ilidenmethyl)- 1-alkyl-pyridinium perchlorate have been synthesized to develop novel fluorescent dyes for the nucleic acids detection. Spectral properties of the synthesized dyes, their complexes with nucleic acids and also in the presence of protein have been studied. The influence of different functional groups on the fluorescent properties of the novel cyanine dyes and their nucleic acids complexes have been investigated. [69] Interaction of cyanine dyes with nucleic acids. 9. The study of spectral properties of cyanine dyes-DNA complexes in the presence of organic solvents M.Yu. Losytskyy, V.B. Kovalska, S.M. Yarmoluk Biopolymers and Cell, 2000, 16, 1, pp. 75-81 (in Ukrainian). Abstract The influence of medium polarity on spectral-luminescent properties of monomethyne cyanine dyes and their DNA-complexes has been studied in the presented work. The presence of organic solvent in aqueous dye and DNA-dye solutions resulted in significant decrease in DNA-dye complex fluorescence and increase in fluorescent intensity of free dyes. The fluorescence intensity of DNA-Ethidium bromide complexes depends weekly on the nature of an organic solvent. This result can be explained using the intercalation interaction model. But for the cyanine dyes studied this dependence is much stronger. So, the considerable contact to the medium of the cyanine dyes bound to DNA has been shown. Nevertheless, the interaction of these dyes with DNA shows no reference to the AT-sequences. Thus, in this presentation we assume the interaction of the cyanine dyes studied with the DNA according to the half-intercalation mechanism. [70] Interaction of cyanine dyes with nucleic acids. 15. Spectral-fluorescent properties of novel benzothiazolmethylene-2,5-dimethylpyridine homodi- and homotrimere cyanine dyes and their complexes with nucleic acids I.A. Kocheshev, V.B. Kovalska, S.M. Yarmoluk, B. Akerman, G.Kh. Matsuka Biopolymers and Cell, 1999, 15, 6, pp. 557-567 (in Ukrainian). Abstract Spectral properties of novel homodi- and homotrimere cyanine dyes and their complexes with nucleic acids have been investigated. The dyes studied have shown the increasing of fluorescence intensity up to 1200 and 800 times under the binding with the DNA and with RNA respectively, which make them promising as fluorescent probes for nucleic acids detection. The nature and character of aggregation of free and complexed dyes have been studied. Possible aggregate structures have been proposed. [71] Interaction of cyanine dyes with nucleic acids. 11. Synthesis and spectral properties of novel 2,6-dimethyl-4-[(3-methyl-1,3-benzothiazol-3-ium-2-yl)methylene]-1,4-dihydro-1-pyridinium homodimer cyanine dyes I.A. Kocheshev, V.B. Kovalska, S.M. Yarmoluk Biopolymers and Cell. – 1999. – V. 15, 5. – P. 449-455 (in Ukrainian). Abstract A novel convenient method for cyanine homodimer dyes synthesis based on the reaction of primary amines with pirilium salts is proposed. The method allows obtaining homodimers with linkers of different length and chemical nature. Spectral-luminescent properties of the synthesized dyes and their complexes with nucleic acids, as well as a dependence of these properties on the length and nature of linker were studied. A novel homodimer K6 in complexes with DNA showed approximately 2000-fold increase in fluorescence intensity. We consider this dye to be perspective as a fluorescent probe for the nucleic acids detection. [72] Interaction of cyanine dyes with nucleic acids. XII. β-substituted carbocyanines as possible fluorescent probes for nucleic acids detection S.M. Yarmoluk, V.B. Kovalska, S.S. Lukashov, Y.L. Slominskii Bioorganic and Medicinal Chemistry Letters. – 1999. – V. 9, 12. – P. 1677-1678. Abstract Results of investigations of fluorescent properties of a β-substituted carbocyanine and its complexes with nucleic acids in comparison with those for the unsubstituted dye are present. Carbocyanine substituted in polymethine chain has shown promising properties for use as fluorescent probe in homogeneous systems of nucleic acids detection. [73] Interaction of cyanine dyes with nucleic acids. 8. Investigation of cyanine dyes as fluorescent probes for nucleic acids detection S.M. Yarmoluk Biopolymers and Cell, 1999, 15, 4, pp. 328-336 (in Ukrainian) Abstract The nineteen polymethine cyanine dyes with various heterocycles were investigated as possible fluorescent probes for the homogeneous nucleic acids detection in solution. Absorption and fluorescent properties of these dyes, their complexes with DNA and RNA and in presence of protein were studied. A possible model of “viscous environment” for intensity enhancement of cyanine dyes and their fluorescent complexes with nucleic acids was proposed. [74] Interaction of cyanine dyes with nucleic acids. 6. Synthesis and spectroscopic properties of thiazole orange- amino acids conjugates S.M. Yarmoluk, D.V. Kryvorotenko, Yu.S. Gerasymchuk Biopolymers and Cell. – 1999. – V. 15, 3. – P. 247-251. Abstract The synthesis of amino acids (L-Trp and L-Tyr) derivatives of thiazole orange, monomethine cyanine dye, which significantly increases fluorescence intensity when bound to nucleic acids is described. Hydroxysuccinimide ester of Cyan 6 (2-[4-N-Carboxyethyl-quinoline)-methyl]-3-methylbenzothiazole-1,4- p-toluenesulsonate) was used for the conjugation of dye with amino acids. Interaction of obtained conjugates with nucleic acids was investigated using spectroscopic methods. [75] Interaction of cyanine dyes with nucleic acids. 5. Towards model of “half intercalation” of monomethyne cyanine dyes into double-stranded nucleic acids S.M. Yarmoluk, V.B. Kovalska, Yu.P. Kovtun Biopolymers and Cell. – 1999. – V. 15, 1. – P. 75-82. Abstract Recently we have proposed a model of “half intercalation” of monomethine cyanine dye into the double-stranded helix. Benzothiazole terminal heterocycle intercalates, nestled between the adjacent base pairs just when second heterocycle is spatially fixed by nucleic acids groove. We consider that heterocycle with high basicity hits in the more nucleophilic groove whereas heterocycle with low basicity insert in more electophilic interbase space. The purpose of this study is to determine the possible fixation mode of second heterocycle in nucleic acids groove. We synthesized and investigated the series of structural-like cyanine dyes, which have different charge electronic distribution. We suppose that binding of second heterocycle of high basicity is caused both by spatial fixation and electrostatic interaction with phosphate group of nucleic acids. Monomethine pyridinium cyanine dye (Cyan 40; 4-[(1-methyl-benzothiazoleildene-2)methyl] –1,2,6 trimethylpyridinium p-toluenesulsonate interacts with native DNA and RNA with strong fluorescence enhancement. It seems to be perspective for development of new nucleic acids binding cyanine dyes. [76] Interaction of cyanine dyes with nucleic acids. 4. Efficient 5'-fluorescent labelling of oligonucleotides with monomehtyne pyrilium cyanine dye Cyan 39 S.M. Yarmoluk, O.M. Kostenko, O.S. Kornushyna, I.Ya. Dubey Biopolymers and Cell. – 1998. – V. 14, 1. – P. 82–86. Abstract A new system for fluorescent 5'-end labbeling of oligonucleotides with cyanine dyes is proposed. Reaction of primary amine with pyrilium salt was used for the efficient conjugation of monomethine pyrilium cyanine dye with 5'-aminoalkyl oligonucleotides with simultaneous conversation of the “brightless” pyrilium dye Cyan 39 into “bright” pyridinium one Cyan 40. [77] Interaction of cyanine dyes with nucleic acids. 3. The use of new cyanine dyes Cyan 13 and Cyan 40 for detection of nucleic acids in agarose gel S.M. Yarmoluk, I.Ya. Dubey Biopolymers and Cell. – 1997. – V. 13, 5. – P. 419–421. Abstract Detection of double-stranded DNA (dsDNA) and single stranded DNA (ssDNA) and RNA with two new cyanine dyes Cyan 13 and Cyan 40 is reported. Cyan 13 and Cyan 40 bind to nucleic acids to form stable fluorescent complexes and can be used for the detection of DNA and RNA samples separated by gel electrophoresis. Sensitivity of detection is comparable to that of ethidium bromide (EtBr), a common used nucleic acids staining dye. [78] Interaction of cyanine dyes with nucleic acids. 2. Spectroscopic properties of methyleneoxy analogues of Thiazole Orange S.M. Yarmoluk, V.B. Kovalska, T.V.Smirnova, M.P. Shandura, Yu.P. Kovtun, G.Kh. Matsuka Biopolymers and Cell. – 1996. – V. 12, 6. – P. 74-81. Abstract A series of asymmetric cyanine dyes based on methylenoxy benzothiazole terminal heterocycle was synthesized. Absorption and fluorescent properties of these dyes and their complexes with nucleic acids were investigated. The best results were obtained with 2-[(3-methyl-2(3H)-benzothiazolyliden)-methyl]-5,6-dioxymethylene –3-methylbenzthiazollium p-toluenesulfonate (Cyan 13). A possible model of binding of monomethine benzothiazole cyanine dyes with double stranded nucleic acids is proposed. [79] Interaction of cyanine dyes with nucleic acids. 1. Studies on monomethyne as possible fluorescent probes for the nucleic acids S.M. Yarmoluk, A.N. Zhyvoloup, V.B. Kovalska, I.V. Klimenko, A.P. Kukharenko, Yu.P. Kovtun, Yu.L. Slominsky Biopolymers and Cell. – 1996. – V. 12, 1. – P. 69-75. Abstract The series of monomethine cyanine dyes as possible fluorescent probes for DNA and RNA analysis were synthesized and their optical properties were studied. Two dyes interacted with native DNA and RNA with strong fluorescence enhancement (up to 350 and 1500 times for Cyan 3 and Cyan 6, respectively). The complex of Cyan 4 with nucleic acids had extremely large Stokes shift (154 nm). Methylenoxybenzothiazole residue seems to be perspective structural fragment for synthesis of new nucleic acids binding cyanine dyes. |
© ²ÄÄ²Ë ÊÎÌÁ²ÍÀÒÎÐÍί ղ̲¯