TY - JOUR
T1 - Use of high-performance liquid chromatography to quantitative thymine-containing pyrimidine dimers in DNA
AU - Love, Jack D.
AU - Friedberg, Errol C.
N1 - Funding Information:
We gratefully acknowledge LaVonne McConkie and Waters Associates for supplying us with the @tyragel column. We also thank Linda Clogg, of Syntex, Inc., for her helpful advice during these investigations; Eric Radany and Dr. Thomas Bonura for their critical review of the manuscript; and IMS. Gina Johnson for her assistance in its preparation. These studies were supported by research grant CA 1242.8 from the U.S.P.H.S. and by contract DE AS03 76SF00326 with the U.S. Department of Energy. J.D.L. is supported by U.S.P.H.S. postdoctoral fellowship CA 06441.
PY - 1982/5/21
Y1 - 1982/5/21
N2 - We have developed two high-performance liquid chromatographic systems for the measurement of pyrimidine dimers in hydrolysates of DNA. Normal-phase chromatography on an NH2 column in methanol-ethyl acetate (3:97) at an elution rate of 2.0 ml/min allowed quantitaion of thymine-containing (thymine-thymine plus thymine-uracil) pyrimidine dimers at levels as low as 0.1% of the total radioactivity as thymine in DNA. This system was unaffected by the presence of up to 1 mg of contaminating protein (bovine serum albumin) or 40 μg of DNA in hydrolysates prepared for chromatography. Reversed-phase chromatography on a μBondapak C18 column allowed measurement of thymine-thymine dimers at concentrations as low as 0.02% of the total radioactivity. With 0.1% tetrahydrofuran in wateras the solvent at a flow-rate of up to 0.6 ml/min, thymine-thymine, thymine-uracil, and uracil-uracil dimers were completely resolved. We were not able to quantitate the latter two dimeric forms, however, owing to the presence of other radioactive components of undefined origin that eluted concomitantly with the uracil-containing dimers.
AB - We have developed two high-performance liquid chromatographic systems for the measurement of pyrimidine dimers in hydrolysates of DNA. Normal-phase chromatography on an NH2 column in methanol-ethyl acetate (3:97) at an elution rate of 2.0 ml/min allowed quantitaion of thymine-containing (thymine-thymine plus thymine-uracil) pyrimidine dimers at levels as low as 0.1% of the total radioactivity as thymine in DNA. This system was unaffected by the presence of up to 1 mg of contaminating protein (bovine serum albumin) or 40 μg of DNA in hydrolysates prepared for chromatography. Reversed-phase chromatography on a μBondapak C18 column allowed measurement of thymine-thymine dimers at concentrations as low as 0.02% of the total radioactivity. With 0.1% tetrahydrofuran in wateras the solvent at a flow-rate of up to 0.6 ml/min, thymine-thymine, thymine-uracil, and uracil-uracil dimers were completely resolved. We were not able to quantitate the latter two dimeric forms, however, owing to the presence of other radioactive components of undefined origin that eluted concomitantly with the uracil-containing dimers.
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U2 - 10.1016/S0021-9673(00)99626-2
DO - 10.1016/S0021-9673(00)99626-2
M3 - Article
C2 - 7047548
AN - SCOPUS:0020317751
SN - 0021-9673
VL - 240
SP - 475
EP - 487
JO - Journal of Chromatography A
JF - Journal of Chromatography A
IS - 2
ER -