Conclusions:
Lithium ion adduction-based UPLC-MS/MS was applied to urinary steroid
analysis and thirteen 3-OH steroids were detected. Among them, 7-OH P5
and 7-OH DHEA were identified for the first time in human urine. A total
of 29 steroids and 19 free steroids were identified by combining
conventional methods to detect protonated ions. The urinary steroid
profile obtained in this study is useful for monitoring relatively
long-term physiological conditions and diagnosing chronic diseases.
Figure 1. Biosynthesis map of steroid hormones.
Urinary steroids identified in this study are shown, of which 3-OH
steroids are shown in green and newly identified steroids are shown in
red.
Figure 2. MRM chromatograms of newly identified 7-OH P5 (left)
and 7-OH DHEA (right) in human urine. Chromatograms were obtained for
standard (2 ng for each, topmost), female-total (second), male-total
(third), female-free (fourth), male-free (fifth) by the Li method (left)
and the H method (right). The values above the peaks indicate retention
times and the values in brackets indicate the peak-height ratio of the
two peaks in each chromatogram observed at the two MRM transitions set
for each compound (see Table S1). Typical fragmentation ions of
lithiated 3-OH steroids are depicted in their structures.
Figure 3. Pie chart of relative amounts of major steroids in
male (a) and female urine (b) and bar chart of free and total steroid
levels (ng/creatinine (mg)) obtained in male (top) and female (bottom)
urine (c). All the data were summarized in Table S5.