Fast quantification of the urinary marker of oxidative stress 8-hydroxy-2 '-deoxyguanosine using solid-phase extraction and high-performance liquid chromatography with triple-stage quadrupole mass detection
T. Renner et al., Fast quantification of the urinary marker of oxidative stress 8-hydroxy-2 '-deoxyguanosine using solid-phase extraction and high-performance liquid chromatography with triple-stage quadrupole mass detection, J CHROMAT B, 738(2), 2000, pp. 311-317
Exogenous and endogenous oxidants constantly cause oxidative damage to DNA.
Since the reactive oxidants itself are not suitable for analysis, oxidized
bases like 8-hydroxy-2'-deoxyguanosine (80HdG) are used as biomarkers for
oxidative stress, either in cellular DNA or as elimination product in urine
. A simple, fast and robust analytical procedure is described for urinary 8
OHdG as an indicator of oxidative damage in humans. The adduct was purified
from human urine by applying a single solid-phase extraction step on LiChr
olut EN(R). After evaporation of the eluate, the residue was resolved and a
n aliquote was injected into a HPLC system with a triple quadrupole mass sp
ectrometer. The limit of detection was 0.2 ng ml(-1) (7 fmol absolute) when
using one product ion as quantifier and two further product ions as qualif
ier. The coefficient of variation was 10.1% (n=5 at 2.8 ng ml(-1) urine). T
he sample throughput was about 50 samples a day. Thus, this method is more
sensitive and much faster than the common method using HPLC with electroche
mical detection. The results of a study with nine volunteers investigated a
t six time-points each over 5 days are presented. The mean excretion of 80H
dG was 2.1 ng mg(-1) creatinine (range 0.17-5.9 ng mg(-1) creatinine; 4 of
53 samples were below the LOD). A relatively large intra(relative SD 66%) a
nd inter-individual (relative SD 71%) variation in urinary 8OHdG excretion
rates was found. (C) 2000 Elsevier Science B.V. All rights reserved.