title: Effects of chemical carcinogens and physicochemical factors on the UV spectrophotometric determination of DNA.
abstract: The ultraviolet (UV) absorbance ratio of 260/280 nm has been used as an indicator of DNA purity. However, the A260/A280 ratio may be beyond the normal range (1.8-1.9) due to physicochemical alterations produced by pH and temperature, and carcinogenic chemical modification. When the pH of the DNA solution buffer increased from 3 to 11, the A260/A280 ratio changed significantly from 1.5 to 2.2 in mixtures of DNA bases [A:T:C:G = 28.5:28.5: 21.5:21.5, i.e., (A + T)/(all four bases) = 57%, expressed as mole percent], of deoxyribonucleosides (adenosine:thymidine:cytidine:guanosine= 28.5:28.5:21.5:21.5, as mole percent), or of deoxyribonucleotides (dAMP:dTMP:dGMP:dCMP = 28.5:28.5:21.5:21.5, as mole percent) examined. The A260/A280 ratio increased with RNA contamination and exceeded 1.9 when RNA concentration was >30%, as mole percent. In contrast, the A260/A280 ratio was linearly reduced by increasing the protein concentration. Phenol (>0.02%) contamination also reduced the A260/A280 ratio to below 1.8. Benzo[a]pyrene diol epoxide (BPDE), a reactive carcinogen metabolite of benzo[a]pyrene (BaP), decreased the A260/A280 ratio correlated with the degree to which it modified the DNA. These results suggest that the UV A260/A280 ratio is significantly affected by pH and the presence of contaminating species of macromolecules and chemicals.
authors.affiliation: Laboratory of Molecular Toxicology, College of Pharmacy, Pusan National University, Jangjun-dong, Gumjung-ku, Pusan, South Korea.
authors.forename: Hyung Sik
authors.initials: HS
authors.lastname: Kim
authors.forename: Soo Hyun
authors.initials: SH
authors.lastname: Byun
authors.forename: Byung Mu
authors.initials: BM
authors.lastname: Lee
mesh_terms: D016604:Aflatoxin B1; D001482:Base Composition; D001564:Benzo(a)pyrene; D001580:Benzopyrenes; D002273:Carcinogens; D002725:Chloroform; D004247:DNA / Q000032:analysis*; D000431:Ethanol; D006863:Hydrogen-Ion Concentration; D009705:Nucleosides; D009711:Nucleotides; D000439:Pentanols; D019800:Phenol; D011506:Proteins; D012313:RNA; D013056:Spectrophotometry, Ultraviolet; D013696:Temperature
chemical_list: D001580:Benzopyrenes; D002273:Carcinogens; D009705:Nucleosides; D009711:Nucleotides; D000439:Pentanols; D011506:Proteins; D019800:Phenol; D001564:Benzo(a)pyrene; D000431:Ethanol; C072019:benzo(a)pyrene diolepoxide I; D012313:RNA; D002725:Chloroform; D004247:DNA; D016604:Aflatoxin B1; C029683:isopentyl alcohol