Synthesis and application of molecular probe for detection of hydroxyl radicals produced by Na(125)I and gamma-rays in aqueous solution.
OBJECTIVE
To synthesize N-(3-(3-aminopropylamino)propyl)-2-oxo-2H-chromene-3-carboxamide (7), a novel DNA-binding, coumarin-based, fluorescent hydroxylradical ((*)OH) indicator and to assess its quantum efficiency compared with that of coumarin-3-carboxylic acid (1) and N1,N12-bis[2-oxo-2H-chromene-3-carbonyl]- 1,12-diamine-4,9-diazadodecane (9).


METHODS
Using computer-generated molecular modeling, 7 and 9 and their respective 7-hydroxylated derivatives 8 and 10 were docked onto DNA dodecamer d(CGCGAATTCGCG)2, the ligand-DNA complexes were energy minimized, and binding free energies and inhibition constants were calculated. Compound 7 was judged an appropriate target molecule and was synthesized. Compounds 1, 7, and 9 were incubated with Na(125)I or irradiated with (137)Cs gamma-rays, and the influence of pH, dose, type of radiation, and the concentration of indicator on fluorescence yield were determined.


RESULTS
Non-fluorescent 7 and 9 are converted to fluorescent, 7-hydroxylated derivatives 8 and 10 after interaction with (*)OH in aqueous solution. For 1, 7, and 9, hydroxylation yield increases linearly with both Na(125)I dose (0-700 x 10(6) decays) and (137)Cs dose (0-11.0 Gy). Fluorescence induction is significantly reduced at acidic pH and the fluorescent quantum yield of 8 is approximately 3 times that of 2 or 10 at pH 7.0. With Na(125)I incubation and gamma-ray irradiation, the fluorescence signal of 7 increases linearly with concentration and saturates at approximately 50 microM.


CONCLUSIONS
Compound 7 quantifies lower concentrations of (*)OH than do 1 and 9. This detector is therefore likely to be a good reporter of (*)OH produced within a few nanometers of DNA.
Singh
Amarjit
A
Department of Radiology, Harvard Medical School, Boston, Massachusetts, USA.
Yang
Yongliang
Y
Adelstein
S James
SJ
Kassis
Amin I
AI
D003374:Coumarins / Q000138:chemical synthesis* / Q000737:chemistry; D005456:Fluorescent Dyes / Q000138:chemical synthesis* / Q000737:chemistry; D005720:Gamma Rays; D006863:Hydrogen-Ion Concentration; D017665:Hydroxyl Radical / Q000032:analysis*; D007457:Iodine Radioisotopes; D015335:Molecular Probes / Q000138:chemical synthesis* / Q000737:chemistry; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000737:chemistry* / Q000528:radiation effects; D012974:Sodium Iodide / Q000737:chemistry*; D012996:Solutions; D013095:Spermidine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry; D013096:Spermine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry; D014867:Water
D003374:Coumarins; D005456:Fluorescent Dyes; D007457:Iodine Radioisotopes; D015335:Molecular Probes; C539576:N-(3-(3-aminopropylamino)propyl)-2-oxo-2H-chromene-3-carboxamide; C539577:N1,N12-bis(2-oxo-2H-chromene-3-carbonyl)-1,12-diamine-4,9-diazadodecane; D009841:Oligonucleotides; D012996:Solutions; D014867:Water; D013096:Spermine; D017665:Hydroxyl Radical; D012974:Sodium Iodide; D013095:Spermidine; C104227:coumarin-3-carboxylic acid