TABLE 4

Comparison of kinetic parameters determined using human liver microsomes and recombinant enzymes for several substrates

The human liver microsomes used in these experiments were prepared in phosphate buffer. Kinetic parameters were determined using assay method A and activation by incubation for 15 min on ice with alamethecin (50 μg/mg protein). Each number represents an individual experiment or the mean ± standard deviation of three experiments (each kinetic determination was performed using nine substrate concentrations in duplicate).


Compound

Enzymes

Human Liver Microsomes

Recombinant Enzymes
Vmax
S50, Km
n
Vmax
S50, Km
n
pmol/min/mg μM pmol/min/mg μM
Estradiol-3-glucuronide UGT1A1 1,670, 810 50, 30 1.3, 1.3 310, 300 20, 22 1.4, 1.2
Buprenorphine UGT1A1 4,900 ± 1,400 57 ± 14 1.2 ± 0.01 2,300, 2,700 45, 68 1.2, 1.2
Ethinylestradiol UGT1A1 1,500, 1,200 21, 13 1.3, 1.2 700, 600 11, 9.7 1.2, 1.3
Anthraflavic Acid UGT1A1 170, 340 4, 5 1.3, 1.3 830, 810 12, 9.8 1.2, 1.2
Naphthol UGT1A6 17,000, 18,000 38, 45 1.0, 1.0 33,000, 27,000 78, 67 1.0, 1.0
Propofol UGT1A9 4,100, 3,500 190, 280 1.0, 1.0 3,200, 2,400 28, 45 1.0, 1.0
Morphine-3-glucuronide UGT2B7 2,500, 2,700 1,900, 3,100 1.0, 1.0 440, 410 290, 200 1.1, 1.2
Morphine-6-glucuronide UGT2B7 160, 140 690, 960 1.0, 1.0 81, 74 240, 240 1.2, 1.0
Androstanediol
UGT2B15
6,700, 5,800
25, 21
1.2, 1.2
980, 980
16, 13
1.1, 1.0