Magdalena Janicka

Polish Academy of Sciences, Warszawa, Masovian Voivodeship, Poland

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Publications (24)80.15 Total impact

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    ABSTRACT: 2-Thiouracil-containing nucleosides are essential modified units of natural and synthetic nucleic acids. In particular, the 5-substituted-2-thiouridines (S2Us) present in tRNA play an important role in tuning the translation process through codon-anticodon interactions. The enhanced thermodynamic stability of S2U-containing RNA duplexes and the preferred S2U-A versus S2U-G base pairing are appreciated characteristics of S2U-modified molecular probes. Recently, we have demonstrated that 2-thiouridine (alone or within an RNA chain) is predominantly transformed under oxidative stress conditions to 4-pyrimidinone riboside (H2U) and not to uridine. Due to the important biological functions and various biotechnological applications for sulfur-containing nucleic acids, we compared the thermodynamic stabilities of duplexes containing desulfured products with those of 2-thiouracil-modified RNA and DNA duplexes. Differential scanning calorimetry experiments and theoretical calculations demonstrate that upon 2-thiouracil desulfuration to 4-pyrimidinone, the preferred base pairing of S2U with adenosine is lost, with preferred base pairing with guanosine observed instead. Therefore, biological processes and in vitro assays in which oxidative desulfuration of 2-thiouracil-containing components occurs may be altered. Moreover, we propose that the H2U-G base pair is a suitable model for investigation of the preferred recognition of 3'-G-ending versus A-ending codons by tRNA wobble nucleosides, which may adopt a 4-pyrimidinone-type structural motif. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.
    Nucleic Acids Research 02/2015; 43(5). DOI:10.1093/nar/gkv109 · 9.11 Impact Factor
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    ABSTRACT: Homopurine phosphorothioate analogs of DNA, possessing all phosphorus atoms of RP configuration ([All-RP-PS]-DNA), when interact with appropriate complementary RNA or (2’-OMe)-RNA templates, form parallel triplexes or parallel duplexes of very high thermodynamic stability. The present results show that T-LNA or 5-Me-C-LNA units introduced into the parallel Hoogsteen-paired (2’-OMe)-RNA strands (up to four units in the oligomers of 9 or 12 nt in length) stabilize these parallel complexes. At neutral pH, dodecameric parallel duplexes have Tm’s of 62-68°C, which are by 4-10°C higher than Tm for the reference duplex (with no LNA units present), while for corresponding triplexes, Tm’s exceeded 85°C. For nonameric parallel duplexes, Tm’s of 38-62°C were found and (2’-OMe)-RNA oligomers containing 5-Me-C-LNA units stabilized the complexes more efficiently than the T-LNA containing congeners. In both series the stability of the parallel complexes increased with an increasing number of the LNA units present. The same trend was observed in experiments of reverse transcription RNA->DNA (using AMV RT reverse transcriptase) where formation of parallel triplexes (consisting of an RNA template, [All-RP-PS]-DNA nonamer and Hoogsteen-paired (2’-OMe)-RNA strands containing the LNA units) led to efficient inhibition of the process. Under the best conditions checked (four 5-Me-C-LNA units, two-fold excess over the RNA template) the inhibition was 94% effective, compared to 71% inhibition observed in the reference system with the Hoogsteen-paired (2’-OMe)-RNA strand carrying no LNA units. This kind of complexation may “arrest” harmful RNA oligomers (e.g., viral RNA or mRNA of unwanted proteins) and, beneficially, exclude them from enzymatic processes, otherwise leading to viral or genetic diseases.
    Organic & Biomolecular Chemistry 12/2014; 13(8). DOI:10.1039/C4OB02364A · 3.49 Impact Factor
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    ABSTRACT: Hydrazone hesperetin Schiff base (HHSB) − N-[(±)-[5,7-dihydroxy-2-(3-hydroxy-4-methoxy-phenyl)chroman-4-ylidene]amino]benzamide has been synthesized and its crystal structure was determined. This compound was used for the formation of Cu(II) complexes in solid state and in solution which were characterized using different spectroscopic methods. The analyses of potentiometric titration curves revealed that monomeric and dimeric complexes of Cu(II) are formed above pH 7. The ESI MS (electrospray ionization-mass spectrometry) spectra confirmed their formation. The EPR and UV-visible spectra evidenced the involvement of oxygen and nitrogen atoms in Cu(II) coordination. Hydrazone hesperetin Schiff base can show keto-enol tautomerism and coordinate Cu(II) in the keto (O−, N, Oket) and in the enolate form (O−, N, O−enol). The semi-empirical molecular orbital method PM6 and DFT (density functional theory) calculations have revealed that the more stable form of the dimeric complex is that one in which the ligand is present in the enol form. The CuHHSB complex has shown high efficiency in the cleavage of plasmid DNA in aqueous solution, indicating its potential as chemical nuclease. Studies on DNA interactions, antimicrobial and cytotoxic activities have been undertaken to gain more information on the biological significance of HHSB and copper(II)-HHSB chelate species.
    Journal of Inorganic Biochemistry 11/2014; 143. DOI:10.1016/j.jinorgbio.2014.11.005 · 3.27 Impact Factor
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    ABSTRACT: Small interfering RNAs (siRNAs) with phosphorodithioate modifications (PS2-RNA) possess favourable properties for use as RNAi therapeutics. Beneficial here is the combining of PS2 and 2′-O-methyl modifications (MePS2). SiRNAs with MePS2 moieties in the sense strand show promising efficacies in vitro and in vivo. Crystal structures of PS2- and MePS2- modified RNAs reveal subtle changes in geometry and hydration compared with natural RNA. A model of an MePS2-RNA:PAZ domain complex points to a hydrophobic effect as the source of the higher affinity of MePS2-RNA for Ago2.
    RSC Advances 11/2014; 4(110). DOI:10.1039/C4RA10986D · 3.71 Impact Factor
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    ABSTRACT: Chemically synthesized small interfering RNAs (siRNAs) have been widely used to identify gene function and hold great potential in providing a new class of therapeutics. Chemical modifications are desired for therapeutic applications to improve siRNA efficacy. Appropriately protected ribonucleoside-3'-yl S-[β-(benzoylmercapto)ethyl]pyrrolidino-thiophosphoramidite monomers were prepared for the synthesis of siRNA containing phosphorodithioate (PS2) substitutions in which the two non-bridging oxygen atoms are replaced by sulfur atoms. A series of siRNAs containing PS2 substitutions have been strategically designed, synthesized, and evaluated for their gene silencing activities. These PS2-siRNA duplexes exhibit an A-form helical structure similar to unmodified siRNA. The effect of PS2 substitutions on gene silencing activity is position-dependent, with certain PS2-siRNAs showing activity significantly higher than that of unmodified siRNA. The relative gene silencing activities of siRNAs containing either PS2 or phosphoromonothioate (PS) linkages at identical positions are variable and depend on the sites of modification. 5'-Phosphorylation of PS2-siRNAs has little or no effect on gene silencing activity. Incorporation of PS2 substitutions into siRNA duplexes increases their serum stability. These results offer preliminary evidence of the potential value of PS2-modified siRNAs.
    ACS Chemical Biology 04/2012; 7(7):1214-20. DOI:10.1021/cb300078e · 5.36 Impact Factor
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    ABSTRACT: The condensation of the 5'-O-DMT-3'-deoxy-3'-aminothymidine with 3'-O-TBDMS-thymidine-5'-aldehyde, followed by reduction of the resultant imine derivative and removal of tert-butyldimethylsilyl (TBDMS) protecting group, provided a dimer (denoted as TNHT), which is a congener of dithymidine phosphate with the phosphate linkage 3'-O-P(O)(OH)-O-5' replaced with an amino group (-NH-). After phosphitylation of the 3'-OH group, the dimer TNHT was introduced (by the standard phosphoramidite approach) into a central part of the nonadecathymidylate. This oligomer exhibited lower affinity to the complementary single and double stranded DNA complements as compared to unmodified T-19 oligonucleotide. The cleavage of modified oligomer with the snake venom and calf spleen phosphodiesterases was completely suppressed at the site of modification. RNA oligomers containing the TNHT dimer were used for preparation of siRNA molecules directed towards mRNA of BACE1 (beta-site amyloid precursor protein cleaving enzyme). The presence of the TNHT units at the 3'-ends of the RNA strands of the siRNA molecule (the siRNA itself is an effective gene expression inhibitor for BACE1) preserved the gene silencing activity and improved the stability of the modified siRNA in 10% fetal bovine serum.
    Collection of Czechoslovak Chemical Communications 12/2011; 76(12):1471-1486. DOI:10.1135/cccc2011090 · 1.14 Impact Factor
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    ABSTRACT: The 2-thiouridine (S2U) unit in the RNA strand is predominantly desulfured with H(2)O(2) to 4-pyrimidinone nucleoside (H2U). The resulting H2U-RNA exhibits significantly lower binding affinity to its complementary strand and in certain conditions undergoes strand scission. These results may explain the tRNA loss of biological function in oxidative stress conditions.
    Chemical Communications 03/2011; 47(17):4914-6. DOI:10.1039/c1cc10973a · 6.72 Impact Factor
  • Journal of Biotechnology 11/2010; 150:32-32. DOI:10.1016/j.jbiotec.2010.08.092 · 2.88 Impact Factor
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    ABSTRACT: Nucleoside 5'-O-phosphorothioates are formed in vivo as primary products of hydrolysis of oligo(nucleoside phosphorothioate)s (PS-oligos) that are applied as antisense therapeutic molecules. The biodistribution of PS-oligos and their pharmacokinetics have been widely reported, but little is known about their subsequent decay inside the organism. We suggest that the enzyme responsible for nucleoside 5'-O-monophosphorothioate ((d)NMPS) metabolism could be histidine triad nucleotide-binding protein 1 (Hint-1), a phosphoramidase belonging to the histidine triad (HIT) superfamily that is present in all forms of life. An additional, but usually ignored, activity of Hint-1 is its ability to catalyze the conversion of adenosine 5'-O-monophosphorothioate (AMPS) to 5'-O-monophosphate (AMP). By mutagenetic and biochemical studies, we defined the active site of Hint-1 and the kinetic parameters of the desulfuration reaction (P-S bond cleavage). Additionally, crystallographic analysis (resolution from 1.08 to 1.37 Å) of three engineered cysteine mutants showed the high similarity of their structures, which were not very different from the structure of WT Hint-1. Moreover, we found that not only AMPS but also other ribonucleoside and 2'-deoxyribonucleoside phosphorothioates are desulfurated by Hint-1 at the following relative rates: GMPS > AMPS > dGMPS ≥ CMPS > UMPS > dAMPS ≫ dCMPS > TMPS, and during the reaction, hydrogen sulfide, which is thought to be the third gaseous mediator, was released.
    Journal of Biological Chemistry 10/2010; 285(52):40809-18. DOI:10.1074/jbc.M110.162065 · 4.60 Impact Factor
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    ABSTRACT: The synthesis of a water-soluble phosphorus-containing dendrimer possessing a fluorophore (maleimide-type) linked to the core is described. This dendrimer is found brightly fluorescent in CH(2)Cl(2), but poorly fluorescent in water. The cytotoxicity of this compound is relatively low towards HeLa and A549 cells, and less toxic after 48 hours than after 24 hours. Association of this dendrimer with plasmid DNA (BACE-GFP) analyzed with circular dichroism (CD) indicates a possible disturbing of the helical B-type structure of DNA. The strength of this association (a "dendriplex") with BACE-GFP (also with HygEGFP) was measured by electrophoresis.
    Nucleosides Nucleotides &amp Nucleic Acids 03/2010; 29(3):155-67. DOI:10.1080/15257771003708462 · 0.89 Impact Factor
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    ABSTRACT: The use of synthetic short interfering RNAs (siRNAs) is currently a method of choice to manipulate gene expression in mammalian cells. Efforts aimed at improving siRNA biological activity, including increased silencing properties, higher substrate specificity and cellular stability, lower cytotoxicity, and improved target delivery, have been made through the introduction of various chemical modifications into the siRNA strands. In these studies, we present the synthesis of oligoribonucleotides with the single replacement of a cytidine unit for 2′,2′-difluoro-2′-deoxycytidine (gemcitabine, dFdC) and the use of them in a series of siRNAs for gene silencing experiments. The dFdC modifications are located in six different positions of the antisense strand, which are crucial for siRNA silencing activity. The results indicate a position-dependent tolerance for the dFdC modification. Gemcitabine units present in the “seed region”, at positions 1 or 8, resulted in only a 15% silencing activity in the corresponding duplexes. The dFdC unit at position 10 virtually switched off the silencing activity (below 10%), while the dFdC unit at the positions 2, 4 or 5 produced duplexes of silencing potential comparable to that of the non-modified duplex (70% silencing). The dFdC modification had little impact on the structure of the siRNA duplexes, as determined by circular dichroism analysis, while melting experiments showed their lower thermal stability.
    New Journal of Chemistry 01/2010; 34(5). DOI:10.1039/b9nj00746f · 3.16 Impact Factor
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    ABSTRACT: Homopurine deoxyribonucleoside phosphorothioates possessing all internucleotide linkages of R(P) configuration form a duplex with an RNA or 2'-OMe-RNA strand with Hoogsteen complementarity. The duplexes formed with RNA templates are thermally stable at pH 5.3, while those formed with a 2'-OMe-RNA are stable at neutrality. Melting temperature and fluorescence quenching experiments indicate that the strands are parallel. Remarkably, these duplexes are thermally more stable than parallel Hoogsteen duplexes and antiparallel Watson-Crick duplexes formed by unmodified homopurine DNA molecules of the same sequence with corresponding RNA templates.
    Biophysical Journal 12/2007; 93(10):3567-74. DOI:10.1529/biophysj.107.108183 · 3.83 Impact Factor
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    ABSTRACT: A series of nucleobase-modified siRNA duplexes containing "rare" nucleosides, 2-thiouridine (s(2)U), pseudouridine (Psi), and dihydrouridine (D), were evaluated for their thermodynamic stability and gene silencing activity. The duplexes with modified units at terminal positions exhibited similar stability as the nonmodified reference. Introduction of the s(2)U or Psi units into the central part of the antisense strand resulted in duplexes with higher melting temperatures (Tm). In contrary, D unit similarly like wobble base pair led to the less stable duplexes (DeltaTm 3.9 and 6.6 degrees C, respectively). Gene-silencing activity of siRNA duplexes directed toward enhanced green fluorescent protein or beta-site APP cleaving enzyme was tested in a dual fluorescence assay. The duplexes with s(2)U and Psi units at their 3'-ends and with a D unit at their 5'-ends (with respect to the guide strands) were the most potent gene expression inhibitors. Duplexes with s(2)U and Psi units at their 5'-ends were by 50% less active than the nonmodified counterpart. Those containing a D unit or wobble base pair in the central domain had the lowest Tm, disturbed the A-type helical structure, and had more than three times lower activity than their nonmodified congener. Activity of siRNA containing the wobble base pair could be rescued by placing the thio-nucleoside at the position 3'-adjacent to the mutation site. Thermally stable siRNA molecules containing several s(2)U units in the antisense strand were biologically as potent as their native counterparts. The present results provide a new chemical tool for modulation of siRNA gene-silencing activity.
    RNA 09/2007; 13(8):1301-16. DOI:10.1261/rna.538907 · 4.62 Impact Factor
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    ABSTRACT: The detailed analysis of the interaction between PAMAM G4 dendrimer and serum albumins was performed using circular dichroism, isothermal titration calorimetry, capillary electrophoresis, zeta-potential and fluorescence polarization. It was shown that serum albumins and PAMAM G4 dendrimer form the complex with stoichiometry of 4-6:1 for G4:HSA and 4-5:1 for G4:BSA molar ratio. The possible sites of PAMAM G4 dendrimers binding to protein surface were discussed. Also, it has been proposed that dendrimer does not significantly affect the protein secondary structure studied by circular dichroism.
    Biochimica et Biophysica Acta 08/2007; 1774(7):946-51. DOI:10.1016/j.bbapap.2007.04.016 · 4.66 Impact Factor
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    ABSTRACT: Synthetic procedures applied for the preparation of a new diastereomerically pure sulfinate and amidosulfite and a few enantiomerically pure sulfinyl derivatives, including sulfoxides functionalized with a perfluorocumyl substituent, are presented. Attempts to polymerize optically active 2-(3-thienyl)ethyl p-tolyl sulfoxide are also mentioned. Mechanistic and stereochemical aspects of the presented protocols are discussed. Structural studies, which include the X-ray, circular dichroism, and vibrational circular dichroism--based determinations of the absolute configurations at the stereogenic sulfur atoms and calculations of conformational equilibria for a few model sulfur-containing compounds are also described. The use of optically active t-butylphenylthiophosphinic acid as a chiral solvating agent is illustrated. © 2007 Wiley Periodicals, Inc. Heteroatom Chem 18:527–536, 2007; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.20335
    Heteroatom Chemistry 07/2007; 18(5):527 - 536. DOI:10.1002/hc.20335 · 1.26 Impact Factor
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    ABSTRACT: Homopurine deoxyribonucleoside phosphorothioates, as short as hexanucleotides and possessing all internucleotide linkages of RP configuration, form a triple helix with two RNA or 2'-OMe-RNA strands, with Watson-Crick and Hoogsteen complementarity. Melting temperature and fluorescence quenching experiments strongly suggest that the Hoogsteen RNA strand is parallel to the homopurine [RP-PS]-oligomer. Remarkably, these triplexes are thermally more stable than complexes formed by unmodified homopurine DNA molecules of the same sequence. The triplexes formed by phosphorothioate DNA dodecamers containing 4-6 dG residues are thermally stable at pH 7.4, although their stability increases significantly at pH 5.3. FTIR measurements suggest participation of the C2-carbonyl group of the pyrimidines in the stabilization of the triplex structure. Formation of triple-helix complexes with exogenously delivered PS-oligos may become useful for the reduction of RNA accessibility in vivo and, hence, selective suppression/inhibition of the translation process.
    Biophysical Journal 05/2007; 92(7):2507-15. DOI:10.1529/biophysj.106.099283 · 3.83 Impact Factor
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    ABSTRACT: The antisense or antigene properties are exerted not only by natural oligo- nucleotides, but also by their different analogs. Among them, phosphorothioate oligo- nucleotides (PS-DNA), in which the sugar-phosphate backbone is modified due to substitu- tion of the sulfur atom for one of the nonbridging oxygens, are much more resistant toward nucleases and, simultaneously, maintain good hybridization properties. However, the substi- tution induces the P-chirality of dinucleoside phosphorothioate moiety and even short PS-DNA synthesized using standard chemical methods exist as a mixture of hundreds or thousands of diastereoisomers. Diastereomerically pure oligomers of (PS)-d(CG)4 and (PS)-d(GC)4 series, obtained using the oxathiaphospholane method, were investigated with respect to their ability to adopt the left-handed conformation at high sodium chloride con- centration. Obtained data allow us to postulate the formation of a strong intramolecular hy- drogen bond with anionic sulfur atom as an acceptor. Homopurine (All-RP-PS)-oligos, but not (All-SP-PS)-oligos, form with the RNA templates extremely stable triplex structures, so far not described in the literature. Most likely, the triplexes are stabilized by hydrogen bonds or water bridges with the participation of sulfur atoms of internucleotide linkage. Notably, target RNA molecules are "arrested" by properly designed (All-RP-PS)-DNA probes at sub- micromolar concentration, and as the result, they are not recognized by reverse transcriptase.
    Pure and Applied Chemistry 05/2006; 78(5):993-1002. DOI:10.1351/pac200678050993 · 3.11 Impact Factor
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    ABSTRACT: The SP-isomer of 5′-OH-N4-benzoyl-2′-deoxycytidine-3′-O-(2-thio-4,4-pentamethylene-1,3,2-oxathiaphospholane) undergoes DBU-promoted intramolecular cyclization providing as a sole product SP-deoxycytidine cyclic 3′,5′-O,O-phosphorothioate. Unexpectedly, the RP-counterpart yields a mixture of products consisting of RP-deoxycytidine cyclic 3′,5′-O,O-phosphorothioate and macrocyclic oligo(deoxycytidine phosphorothioate)s. The results of molecular modeling indicate that the dychotomy observed for the RP substrate may result from remarkably higher energy of the corresponding transition states, caused by the presence of bulky ‘spiro’ pentamethylene substituent at the position C4 in the oxathiaphospholane ring.
    Tetrahedron 03/2006; 62(11):2698-2704. DOI:10.1016/j.tet.2005.12.022 · 2.82 Impact Factor
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    ABSTRACT: S(C) and R(C) diastereomers of 5'-C-(O,O-diethyl)-phosphonylthymidine ((R)T and (S)T) were used for the synthesis of the dimers T(R)T and T(S)T, respectively. These dimers were incorporated at selected sites in oligonucleotide constructs. Melting temperature (Tm) experiments demonstrated that relative to the unmodified oligodeoxyribonucleotide, the presence of the (R)T moiety reduced the thermal stability of the duplexes by approximately 5.0 degrees C per modification, whereas their (S)T counterparts only slightly destabilized the duplex structure (deltaTm < or = 1 degree C/modification). The stability of the triple-helical complexes containing one, two, or three modified thymidines is slightly higher than that of the parent complex. Nuclease resistance studies performed with snake venom phosphodiesterase, calf spleen phosphodiesterase, and 3'-exonuclease from human plasma showed that cleavage of the oligonucleotides at the site of the modification was completely suppressed regardless of the stereochemistry of the 5'-C-chiral center. The influence of the (R)T and (S)T modification in the recognition sequence of HindIII, EcoRI, and HpaI restriction endonucleases was also investigated. Although the catalytic activity of HindIII was not affected by the presence of the 5'-C-ethoxyphosphonyl modification, the activities of the two remaining restriction enzymes were partially suppressed depending on the site of modification or the stereochemistry of the modification or both ((R)T vs. (S)T).
    Oligonucleotides 02/2006; 16(1):68-82. DOI:10.1089/oli.2006.16.68 · 3.08 Impact Factor
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    ABSTRACT: Stereoregular chimeric oligonucleotides modified with diastereomerically pure methylphosphonothioate linkages interact with complementary DNA or RNA templates in a stereodependent manner. There is an unprecedented large difference in the stability of duplexes of oligonucleotides modified with alternating methylphosphonothioate linkages (ΔTm = 34 °C) that is dependent only on the stereochemistry of the methylphosphonothioate phosphorus centres. The nature of these interactions was studied by circular dichroism and NMR spectroscopy, and a thermodynamic analysis was performed. The collected data indicate that the absolute configuration at the P-chiral methylphosphonothioate linkages in chimeric 2′-O-methylribonucleotides could, in some cases, determine the global conformation of single–strand oligonucleotides and constrain the conformation of hybrid duplexes formed with RNA. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)
    European Journal of Organic Chemistry 12/2005; 2005(24):5189 - 5197. DOI:10.1002/ejoc.200500395 · 3.15 Impact Factor