TABLE 1

Validation of SPE method Compounds were incubated for 120 min with 0.5 mg/ml DLM. Deproteinized reaction mixtures were loaded on SPE plates that were eluted with 4 × 1 ml of water and 2 × 0.5 ml of MeOH. Results represent the percentage of radioactive counts eluting in the water and MeOH fractions relative to the total [14C]UDPGA present in incubations. MeOH fractions from the first incubation were dried, reconstituted in water, and loaded on a second extraction plate that was eluted with water followed by MeOH. Results represent the percentage of radioactivity recovered in the MeOH eluate of the second plate relative to the total input. Results are mean values ± S.D. (n = 3) except for the blank and time zero controls (S.E.M., n = 15).


Compound

Concentration

Radioactivity Eluted in First Extraction

Recovery of Glucuronide in Second Extraction
Water Fraction
MeOH Fraction
Total
mM % of total [14C]UDPGA in incubation %
4MU 0.5 19.5 ± 7.3 70.3 ± 5.8 89.8 69 ± 3
HFC 1 98.7 ± 2.4 8.5 ± 2.4 107.2 86 ± 25
Octyl gallate 0.5 74.5 ± 1.8 8.3 ± 0.5 82.8 73 ± 10
1NP 1 45.4 ± 22.9 64.6 ± 16.0 110.0 95 ± 6
5α-Androstane-3α,17α-diol 0.25 99.9 ± 15.8 4.9 ± 0.4 104.8 89 ± 6
5α-Androstane-3α,17β-diol 0.25 80.4 ± 2.1 6.6 ± 0.3 87.0 94 ± 4
Hyodeoxycholic acid 1 75.0 ± 10.0 9.2 ± 0.1 84.2 86 ± 16
β-Estradiol 1 101.7 ± 3.1 5.8 ± 0.4 107.6 87 ± 7
Ketoprofen 1 84.0 ± 10.9 21.7 ± 4.7 105.6 82 ± 13
Naproxen 1 82.0 ± 3.3 14.1 ± 0.4 96.0 89 ± 12
Naloxone 1 97.4 ± 2.5 12.8 ± 0.5 110.3 78 ± 14
Furosemide 1 82.5 ± 6.3 10.2 ± 1.4 92.8 76 ± 8
Gemfibrozil 1 45.6 ± 2.3 41.7 ± 1.7 87.0 84 ± 1
Valproic acid 1 62.0 ± 4.1 29.7 ± 1.1 91.7 69 ± 1
17α-Ethynylestradiol 0.25 N.D. 4.6 ± 0.2 N.D. 115 ± 25
None (blank) 98.8 ± 1.8 0.72 ± 0.07 99.4
None (time zero)

100.1 ± 1.8
0.11 ± 0.02
100.2

  • N.D., not determined.