Characterization of nuciferine metabolism by P450 enzymes and uridine diphosphate glucuronosyltransferases in liver microsomes from humans and animals
Acta Pharmacologica Sinica
Characterization of nuciferine metabolism by P450 enzymes and uridine diphosphate glucuronosyltransferases in liver microsomes from humans and animals
Yan-liu LU 1
Yu-qi HE 0
Miao WANG 1
Li ZHANG 1
Li YANG 0
Zheng-tao WANG 0
Guang JI 1
0 Institute of Chinese Materia Medica, Shanghai University of Traditional Chinese Medicine , 1200 Cai Lun Road, Shanghai 201203 , China
1 Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine , Shanghai 200032 , China
Aim: To characterize the metabolism of nuciferine by P450 enzymes and uridine diphosphate glucuronosyltransferase (UGT) in liver microsomes from humans and several other animals including rats, mice, dogs, rabbits and monkeys. Methods: Nuciferine was incubated with both human and animal liver microsomal fractions containing P450 or UGT reaction components. Ultra performance liquid chromatography coupled with mass spectrometry was used to separate and identify nuciferine metabolites. Chemical inhibition was used to identify the involved isozymes. Species difference of nuciferine metabolism in human and various animals were investigated in the liver microsomal incubation system. Results: Among the nuciferine metabolites detected and identified, seven were catalyzed by P450 and one by UGT. Ketoconazole inhibited the formation of M292, M294 and M312. Furafylline, 8-methoxypsoralen and quercetin inhibited the formation of M282. Hecogenin showed a significant inhibitory effect on nuciferine glucuronidation. While the P450-catalyzed metabolites showed no species differences, the glucuronidation product was only detected in microsomes from humans and rabbits. Conclusion: The isozymes UGT 1A4, CYP 3A4, 1A2, 2A6 and 2C8 participated in the hepatic metabolism of nuciferine. Based on the observed species-specific hepatic metabolism of nuciferine, rats, mice, dogs and even monkeys are not suitable models for the pharmacokinetics of nuciferine in humans.
nuciferine; metabolism; cytochrome P450; UGT; species differences; rats; mice; rabbits; monkeys; human
Lotus is a common plant that grows worldwide. This plant
has long been used as an herb in traditional Chinese
medicine. In recent years, significant pharmacological activities
have been observed for lotus, which displays beneficial effects
on hyperlipidemia, obesity, arrhythmia and
atherosclerosis . Most of the beneficial effects have been attributed to the
main constituent of lotus, the aporphine alkaloid nuciferine.
Given its salutary effects, nuciferine is a promising drug
candidate. However, the metabolism and in vivo kinetics of
nuciferine have not been investigated until now.
Cytochrome P450 (CYP) enzymes participate in 70%–80%
of the known phase I metabolism of drugs and, as such, are
among the most important drug-metabolizing enzymes. In
recent years, there has been increased interest in in vitro
metabolism studies involving P450 enzymes during preclinical drug
development[6–8] in order to identify drugs with undesirable
metabolites and to prevent failure in clinical trials. In
addition to CYP enzymes, UDP-glucuronosyltransferases (UGTs)
participate in nearly half of the known phase II metabolism of
the top 200 prescribed drugs in the United States. Thus, like
CYPs, UGTs are important drug-metabolizing enzymes.
In the present study, both P450- and UGT-mediated
metabolism of nuciferine were investigated in order to learn the
general structure of the metabolites, to identify the isozymes
involved in nuciferine metabolism and to examine the
species differences in nuciferine metabolism in humans and other
commonly used experimental animals.
Materials and methods lows: 296 to 235 for nuciferine; 294 to 248 and 294 to 279 for
Chemicals and materials M294-1 and M294-2, respectively; 292 to 246 for M292; 282 to
Nuciferine, glucose-6-phosphate (G-6-P), G-6-P dehydro- 191 and 282 to 250 for M282-1 and M282-2, respectively; 312 to
genase, alamethicin, uridine diphosphate glucuronic acid, 250 and 312 to 248 for M312-1 and M312-2, respectively; 472 to
β-nicotinamide-adenine dinucleotide phosphate (NADP), 265 for M472.
sulfaphenazole, quinidine, clomethiazole, furafylline, 8-meth- Another alkaloid, senecionine, was used as the internal
oxypsoralen, hecogenin, fluconazole, androsterone and phe- standard, which was also detected in MRM mode with ion
nylbutazone were purchased from Sigma-Aldrich (St Louis, transition of 336 to 138. The accuracy for determination of
MO, USA). Ketoconazole and S-mephenytoin were obtained nuciferine was more than 90% and less than 105%. The
precifrom ICN Biomedicals Inc (Aurora, OH, USA) and Toronto sion for determination of nuciferine and metabolites was
meaResearch Chemicals Inc (North York, Canada), respectively. sured, and the residual standard deviation (RSD) values were
Pooled human liver microsomes (HLM) were prepared using less than 15%.
tissue from 13 Chinese donors and stored in phosphate buffer
(100 mmol/L, pH 7.4). Except the pooled HLM, 6 HLM
samples (male) and male cynomolgus monkey liver microsomes
were purchased from Rild Research Institute for Liver
Diseases (Shanghai, China). Experiments involving human
subjects were approved by the local governmental ethics
authorities and were pursuant to the Helsinki Declaration. All other
reagents were of HPLC grade or the highest grade
Microsomal incubation system for P450-mediated metabolism
A standard incubation system for P450-mediated
metabolism (System P450) included HLM (0.5 g/L, 10 µL), G-6-P
(1 mmol/L, 20 µL), G-6-P dehydrogenase (1 unit/mL, 20
µL), phosphate buffer (100 mmol/L, pH 7.4, 108 µL), MgCl2
(4 mmol/L, 20 µL), and nuciferine (200 μmol/L, 2 µL).
Nuciferine was dissolved in methanol; all other reagents were
dissolved in phosphate buffer. The total volume of the
incubation system was 200 μL, and the total organic volume was less
than 1% of the system. The reaction was initiated by adding
NADP (1 mmol/L, 20 µL). After incubation for 60 min, 200
μL of acetonitrile was added to stop the reaction. The stopped
incubation system was centrifuged for 10 min at 20 000 g
(4 °C) and a 2-µL aliquot of the supernatant was analyzed as
Preparation of liver microsomes
Livers from rats (Sprague-Dawley, male, n=6), mice (Swiss,
male, n=6), rabbits (New Zealand white, male, n=6) and dogs
(Beagle, male, n=6) were obtained from healthy animals at the
Experimental Animal Center of Shanghai University of
Traditional Chinese Medicine (SUTCM, Shanghai, China). The use
of livers in the present study was approved by the ethics
committee of SUTCM. Liver samples were pooled by species and
stored in liquid nitrogen immediately after being harvested.
Microsomes were prepared from pooled frozen liver tissue
by differential ultracentrifugation as described previously.
Protein concentration was determined using bovine serum
albumin as reference. Liver microsomes were diluted to 10
mg/mL and stored at -80 °C.
Microsomal incubation system for UGT-mediated metabolism
A standard incubation system for UGT-mediated
metabolism (system UGT) with a total volume of 200 µL contained
HLM (0.5 g/L, 10 µL), alamethicin (25 µg/mg protein, 10 µL),
UDPGA (5 mmol/L, 20 µL), MgCl2 (4 mmol/L, 20 µL),
TrisHCl buffer (50 mmol/L, pH 7.4) and nuciferine (200 μmol/L,
2 µL). Reactions were started by adding UDPGA at 37 °C and
stopped after 60 min by adding ice-cold 10% trichloroacetic
Parameters for chromatography and mass spectrometry acid (200 µL). After stopping the reaction, the incubation
mixAn ultra performance liquid chromatography (UPLC) system ture was centrifuged for 10 min at 20 000 g (4 oC). Aliquots of
coupled with triple quadrupole mass spectrometry (Acquity- the supernatant were subsequently analyzed.
Premier, Waters, Milford, MA) and an electrospray ionization
(ESI) source was used in the present study. A Waters bridged Identification of metabolites
ethyl hybrid (BEH) C18 (50×2.1 mm, 1.7 μm) column was used Three experimental groups were used in the present study
for separation. Acetonitrile and formic acid solution (0.1% to identify P450-catalyzed nuciferine metabolites: a reaction
in water) were used as mobile phase components A and B, group, which included nuciferine, HLM, NADP and the other
respectively. The mobile phase elution gradient was as fol- components described above; a negative control group, which
lows: 0 to 0.5 min, 5% A; 0.5 to 1.5 min, 5% to 50% A; 1.5 to 3 included nuciferine and the other reaction components except
min, 50% to 80% A; 3 to 4 min, 80% to 95% of A. The mobile- NADP (which was replaced by phosphate buffer); and a
phase flow rate was 0.3 mL/min. blank group, which included all reaction components without
The positive ion monitor mode was adopted, and mass nuciferine (but with the appropriate volume of methanol).
parameters for UPLC-MS were set as follows: capillary volt- Similarly, three experimental groups were used in the
presage, 3.2 kV; cone voltage, 40 V; extractor voltage, 1.59 V; ent study to identify glucuronidated nuciferine metabolites:
source and desolvation temperatures, 100 and 350 °C, respec- a reaction group, which included nuciferine, HLM, UDPGA
tively; cone and desolvation gas flow, 50 and 550 L/h, respec- and the other components described above; a negative control
tively. Multiple reaction monitor mode (MRM) was used to group, which included nuciferine and the other reaction
comdetermine the metabolites and ion transitions were set as fol- ponents except UDPGA (which was replaced by phosphate
buffer); and a blank group, which included all reaction
components without nuciferine (but with the appropriate volume
Peaks appearing only in the reaction group, but not in the
negative control and blank groups, were considered to
correspond to metabolites of nuciferine. The molecular weight and
the MS/MS spectra of the metabolites were compared with
those of nuciferine to determine their structures.
Chemical inhibition of P450-mediated metabolism of nuciferine in HLM
Selective inhibitors, including furafylline (10 μmol/L),
8-methoxypsoralen (2.5 μmol/L), quercetin (10 μmol/L),
sulfaphenazole (10 μmol/L), quinidine (10 μmol/L),
clomethiazole (50 μmol/L), and ketoconazole (1 μmol/L), were used
to inhibit CYP 1A2, 2A6, 2C8, 2C9, 2D6, 2E1, and CYP3A4,
respectively, in HLM[12–14]. All of the inhibitors added to the
incubation system were dissolved in 1 μL of methanol, which
was less than 0.5% of the total incubation volume; other
components in the incubation system were the same as those
described above. An incubation without any inhibitor, but
with the dissolving medium was used as a solvent control.
The reaction system was pre-incubated for 3 min at 37 °C, and
the reaction was initiated by the addition of nuciferine. The
quantities of metabolites in the inhibited incubations were
compared with those in the solvent control incubations, which
were normalized to 100%. The compared value was defined
as the remaining enzyme activity (% of control) and used as a
parameter to evaluate the catalytic capacity of an isozyme.
Species differences in nuciferine metabolism in liver microsomes from humans and other animals
To compare the species differences in nuciferine metabolism
and to identify a suitable animal species for pharmacokinetic
studies, nuciferine was incubated in the P450 and UGT
systems described above but with liver microsomes from human,
rat, mouse, dog, rabbit, or monkey. The concentrations of
nuciferine used in the incubation systems were 50, 100, and
200 µmol/L. The protein concentration of the microsomes
used in the incubation systems was 0.5 g/L. The incubation
time was 60 min. The quantities of metabolites formed by
human and the various non-human microsomes were
compared. Microsomes used to determine species differences in
nuciferine metabolism were different from other experiments
of the present study. The microsomes were from six animals,
and their preparation was the same as for the pooled HLMs
purchased from the Rild Institute for Liver Disease in
Shanghai. Average values of the six individuals were used.
All data were acquired from three replicates. Student’s t-tests
(group) were performed to evaluate the statistical significance
of differences between two experimental groups using a
probability value (P) threshold of 0.05 or 0.01. Inhibitory potency
(IC50) values were obtained by non-linear regression of
inhibition-versus-log (inhibitor) plots using the “sigmoidal fit”
function built into Origin (OriginLab Corporation, Northampton,
Chemical inhibition of glucuronidation of nuciferine in HLM Identification of metabolites
Fluconazole, phenylbutazone[16, 17], androsterone[16, 17], and In the HLM P450 system, seven NADPH-dependent
metabohecogenin were used to inhibit nuciferine glucuronidation lites were observed (Figure 1). According to the molecular
in HLM. Fluconazole and hecogenin showed a potent and weight values, these metabolites were identified as M-312-1,
selective inhibitory effect on UGT 2B7 and UGT 1A4, respec- M-312-2, M-282-1, M-282-2, M-294-1, M-294-2, and M-292. In
tively[15, 17]. Androsterone showed a non-selective inhibitory the HLM UGT system, a unique UDPGA-dependent
metaboeffect on several UGT isozymes including UGT 2B7, 2B15, 1A1, lite was detected and identified as M-472.
1A3, 1A4, and 1A9, with UGT 1A9 being inhibited strongly. The molecular weight value indicated that M-312-1 and
Phenylbutazone showed non-selective effect on several M-312-2 were probably hydroxylated nuciferine or nuciferine
isozymes including UGT 1A1, 1A3, 1A6, 1A7, 1A8, 1A9, 1A10, N-oxide. In the nuciferine molecule (Figure 2, A), C1 to C9
and 2B15; however, the compound showed no inhibitory effect and the N-atom are the most likely sites for oxidation. The
on UGT 1A4. A range of concentrations including 1, 10, 50, MS/MS spectrum showed that the fragment ions at m/z 179
and 200 µmol/L were used for all of the inhibitors. The con- and 191 were observed for nuciferine, M-312-1 and
M-312centration of nuciferine in the reactions was 100 µmol/L. The 2, indicating that the hydroxylation sites were not C2 to C9.
reaction time and protein concentrations were set at 60 min Hydroxylation increases water-solubility and thus shortens
and 0.5 g/L, respectively. All other reaction parameters were the retention time of a compound on a C18 column; however,
the same as those in the standard incubation system described M-312-1 was retained slightly longer than nuciferine,
indiabove. The concentration of organic solvent dissolving inhibi- cating that M-312-1 was not a hydroxylated product but an
tors was less than 0.5% of the total incubation volume. An N-oxide (Figure 3). Thus, M-312-2 was a C1-hydroxylated
experimental group without inhibitors but dissolving sol- product of nuciferine (Figure 3).
vents was used as the solvent control, in which the activities The molecular weight value indicated that M-282-1 and
of nuciferine glucuronidation were normalized to 100%. The M-282-2 were demethylated products (Figure 3). However,
activities of nuciferine glucuronidation in the inhibited sam- the demethylated sites could not be distinguished between
ples were compared with the solvent control to calculate the M-288-1 and M-288-2 based on the present data. The
metaboremaining enzyme activity (% of control). lites M-294-1 and M-294-2 had a molecular weight 2 Da less
than that of nuciferine, indicating they were dehydrogenated
products (Figure 3). In the nuciferine molecule, the likely
dehydrogenation sites are C1-C2 and C8-C9. However, the
dehydrogenation sites could not be distinguished between
M-294-1 and M-294-2. The product M-292 had a molecular
weight 4 Da smaller than that of nuciferine, indicating it was a
di-dehydrogenation product. The dehydrogenation sites were
both C1-C2 and C8-C9 as they were the only possible sites for
dehydrogenation (Figure 3).
The metabolite M-472 was a unique UDPGA-dependent
product of nuciferine. Both its molecular weight value and
its MS/MS fragment ions indicated it was a glucuronidation
product of nuciferine (Figure 2D). Because the N-atom in
nuciferine was the only possible glucuronic acid conjugation
site, M-472 was identified as nuciferine N-glucuronide (Figure
Chemical inhibition of P450-mediated metabolism of nuciferine in HLM
Furafylline, 8-methoxypsoralen, and quercetin significantly
inhibited the formation of M-282-1 and M-282-2 (Figure 4A),
indicating that CYP 1A2, 2A6, and 2C8 participated in the
demethylation of nuciferine. As for the formation of M-282-1,
quercetin inhibited it slightly, indicating that CYP 2C8
contributed less to its production than did CYP 1A2 and 2A6.
Ketoconazole was the most effective and selective inhibitor of
CYP 3A4. In the present study, the formation of M-312-1,
M-312-2, M-294-1, M-294-2, and M-292 was significantly
inhibited by ketoconazole (Figure 4B), indicating that CYP 3A4
participated in the N-oxidation, hydroxylation and
dehydrogenation of nuciferine.
Chemical inhibition of nuciferine N-glucuronidation in HLM
Four potent chemical inhibitors of major UGT isozymes
including fluconazole, androsterone, phenylbutazone and
hecogenin were tested for inhibition of nuciferine
N-glucuronidation. Each drug was assayed at four concentrations: 1, 10,
50 and 200 µmol/L. As shown in Figure 5, hecogenin was
the most potent and concentration-dependent inhibitor of
nuciferine N-glucuronidation, reducing it more than 75% at
1 µmol/L. The IC50 for hecogenin inhibition of nuciferine
N-glucuronidation was calculated to be 0.63±0.15 µmol/L.
Fluconazole was an effective and selective inhibitor of UGT
2B7. In the present study, this inhibitor showed no inhibitory
effect on nuciferine N-glucuronidation at concentrations up
to 200 µmol/L, indicating that UGT 2B7 did not participate
in this metabolism. Androsterone and phenylbutazone, two
additional UGT inhibitors with a broad inhibitory effect on
several UGT isozymes, did not significantly inhibit nuciferine
N-glucuronidation. Together, the results showed that UGT
1A4 was the main enzyme involved in the N-glucuronidation
Species differences of nuciferine metabolism in human and animal liver microsomes
To investigate the species differences in nuciferine
metabolism, nuciferine (at 50, 100, and 200 µmol/L) was incubated
with human, rat, mouse, dog and rabbit liver microsomes
in both the P450 and the UGT systems. In the P450 system,
microsomes from all species generated the metabolites
identified in the HLM incubation system described above (Figure
6). For most of the P450-catalyzed metabolites, microsomes
from all species showed comparable levels. However, for the
metabolite M-282-1, rabbit liver microsomes generated
significantly higher production than did microsomes from humans,
rats, mice and dogs. As concerns the N-glucuronidation
product M-472, only human and rabbit liver microsomes generated
that nuciferine metabolite.
Identification of the structure of M-312-1 was based on its
chromatographic characteristics. Usually, in the acidic mobile
phase, the alkaloid compound would show high water
solubility and would be eluted from the C18 column early.
However, when the N-atom was blocked, the basic portion of the
alkaloid could no longer accept a proton, leading to a longer
retention time than that of the free alkaloid. This
enon has already been reported for other alkaloids. Thus,
based on its longer retention time (Figure 1) compared with
nuciferine, M-312-1 was identified as nuciferine N-oxide. In
addition, hydroxylation would increase the water-solubility of
a compound, leading to a shorter retention time than the
parent compound. Thus, M-312-2 (Figure 1) was reasoned to be
a hydroxylated product of nuciferine. Theoretically, both the
C1 (Figure 2A) and the N-methyl group of nuciferine could be
hydroxylated; however, N-methyl hydroxylation would
generate an extremely unstable product. Because M-312-2 was
stable for more than 6 days in our incubation system (data not
shown), we reasoned that the metabolite was hydroxylated at
In addition to the two selective inhibitors fluconazole (for
UGT 2B7) and hecogenin (for UGT 1A4), the inhibitors
androsterone and phenylbutazone were also used to inhibit
nuciferine N-glucuronidation in the present study.
Androsterone was a potent inhibitor of UGT 1A9, 2B7, and 2B15.
However, even at a high concentration, androsterone only
slightly inhibited nuciferine glucuronidation. Thus, UGT 1A9,
2B7, and 2B15 are not likely to participate in glucuronidation
of nuciferine. Phenylbutazone inhibited UGT 1A1, 1A3, 1A6,
1A7, 1A8, 1A9, 1A10, and 2B15 significantly, but did not
inhibit UGT 1A4, suggesting that nuciferine
N-glucuronidation was not likely catalyzed by the phenylbutazone-sensitive
isozymes. Combining these results, UGT 1A4 is the sole
isozyme responsible for nuciferine N-glucuronidation, which
is consistent with reports that UGT 1A4 is the most effective
isozyme for N-glucuronidation of compounds with tertiary
amine groups. In addition, because UGT1A4 was shown to
be deficient in human intestines[20, 21], the liver is likely the
predominant organ responsible for glucuronidation of nuciferine
Metabolism can alter the activity or even toxicity of drugs
in vivo. To avoid failure in clinical studies, a suitable animal
model with metabolic properties similar to those of humans
should be useful. In vitro metabolism studies could also be
used to help rapidly and conveniently screen animal
species. Rats, mice and dogs are commonly used experimental
animals, and their P450 enzyme systems are very similar to
that of humans. Indeed, P450 enzymes are highly conserved
in these mammals. However, N-glucuronidation of tertiary
amines, an important phase II metabolic process, is deficient in
the aforementioned animal models. Of the UGT isozymes,
UGT 1A4 is known to have the greatest effect on
N-glucuronidation of compounds with tertiary amine groups[23–25], and
the corresponding UGT 1A4 genes in rats, mice and dogs are
pseudogenes. Thus, microsomes from these animals did not
catalyze the N-glucuronidation of nuciferine, as demonstrated
in Figure 6. Among all of the animal species investigated in
the present study, rabbits are the only species possessing the
same nuciferine metabolic activity as humans. Thus, rabbits
may be more suitable than rats, mice and dogs for further
pharmacokinetic studies of nuciferine in vivo. However,
potential differences in nuciferine absorption between humans
and rabbits should be considered.
In the present study, two demethylated metabolites, two
dehydrogenated products, a hydroxylated species, an N-oxide
and a di-dehydrogenated derivative of nuciferine were
observed in the HLM P450 system, and a unique
N-glucuronidation product of nuciferine was observed in the HLM UGT
system. The isozymes CYP 3A4, 1A2, 2A6, and 2C8 were
responsible for the P450-mediated metabolism of nuciferine,
and UGT 1A4 was the most effective isozyme involved in
nuciferine N-glucuronidation. The species difference
studies on the metabolism of nuciferine showed that rabbits, with
the nuciferine metabolism most similar to that in humans,
might be useful for advanced pharmacokinetic studies in vivo.
Furthermore, the present study demonstrated that rats, mice 9 Williams JA, Hyland R, Jones BC, Smith DA, Hurst S, Peterkin TCGV,
and dogs are not suitable for such studies because they lack et al. Drug-drug interactions for UDP-glucuronosyltransferase
subthe important phase II metabolism of nuciferine that occurs in strates: a pharmacokinetic explanation for typically observed low
humans. exposure (AUCi/AUC) ratios. Drug Metab Dispos 2004; 32: 1201–8.
10 Walsky RL, Obach RS. Validated assays for human cytochrome p450
activities. Drug Metab Dispos 2004; 32: 647–60.
Acknowledgements 11 Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement
The authors are grateful for financial support from the with the Folin phenol reagent. J Biol Chem 1951; 193: 265–75.
National Natural Science Foundation of China (No 30873260), 12 Bjornsson TD, Callaghan JT, Einolf HJ, Fischer V, Gan L, Grimm S, et
the Talents Scheme from the Science and Technology Commis- al. The conduct of in vitro and in vivo drug-drug interaction studies: A
sion of Shanghai Municipality (No 09XD1403800), the China PhRMA perspective. Drug Metab Dispos 2003; 31: 815–32.
Postdoctoral Science Foundation (No 20090460645), the Sup- 13 Huang SM, Temple R, Throckmorton DC, Lesko LJ. Drug interaction
porting Project for Elitists in the New Century from the Minis- studies: study design, data analysis, and implications for dosing and
try of Education (No NCET-07563) and the Shanghai Leading labeling. Clin Pharmacol Ther 2007; 81: 298–304.
Academic Discipline Project (No J50305 and E3008). 14 Harris JW, Rahman A, Kim BR, Guengerich FP, Collins JM. Metabolism
of taxol by human hepatic microsomes and liver slices: participation
of cytochrome P450 3A4 and an unknown P450 enzyme. Cancer Res
Author contribution 1994; 54: 4026–35.
Yan-liu LU, Yu-qi HE and Guang JI designed the research. 15 Liu HX, He YQ, Hu Y, Liu Y, Zhang JW, Li W, et al. Determination of
Yan-liu LU and Yu-qi HE performed the main experiments UDP-glucuronosyltransferase UGT2B7 activity in human liver
microand wrote the paper. Miao WANG, Li ZHANG, and Chang- somes by ultra-performance liquid chromatography with MS detection.
hong WANG prepared the animal liver microsomes, Li YANG J Chromatogr B 2008; 870: 84–90.
participated in the UPLC-MS analysis of the metabolites, 16 Liu HX, Liu Y, Zhang JW, Li W, Liu HT, Yang L.
UDP-glucuronosylZheng-tao WANG participated in the identification of the transferase 1A6 Is the major isozyme responsible for protocatechuic
aldehyde glucuronidation in human liver microsomes. Drug Metab
metabolite structures. Dispos 2008; 36: 1562–9.
17 Uchaipichat V, Mackenzie PI, Elliot DJ, Miners JO. Selectivity of
subReferences strate (trifluoperazine) and inhibitor (amitriptyline, androsterone,
1 Commission CP (China). Pharmacopoeia of the People’s Republic of canrenoic acid, hecogenin, phenylbutazone, quinidine, quinine, and
China. Beijing: People’s Medical Publishing House; 2005. sulfinpyrazone) «probes» for human udp-glucuronosyltransferases.
2 Ho HH, Hsu LS, Chan KC, Chen HM, Wu CH, Wang CJ. Extract from Drug Metab Dispos 2006; 34: 449–56.
the leaf of nucifera reduced the development of atherosclerosis via 18 Xiong A, Yang L, He Y, Zhang F, Wang J, Han H, et al. Identification
inhibition of vascular smooth muscle cell proliferation and migration. of metabolites of adonifoline, a hepatotoxic pyrrolizidine alkaloid,
Food Chem Toxicol 2010; 48: 159–68. by liquid chromatography/tandem and high-resolution mass
spec3 Ono Y, Hattori E, Fukaya Y, Imai S, Ohizumi Y. Anti-obesity effect of trometry. Rapid Commun Mass Spectrom 2009; 23: 3907–16.
Nelumbo nucifera leaves extract in mice and rats. J Ethnopharmacol 19 Fu PP, Xia Q, Lin G, Chou MW. Genotoxic pyrrolizidine alkaloids —
2006; 106: 238–44. mechanisms leading to DNA adduct formation and tumorigenicity. Int
4 Xu Q, Guo RX, Wang CY, Hu XY. Application of activated glassy carbon J Mol Sci 2002; 3: 948–64.
electrode for the detection of nuciferine in lotus leaves. Talanta 2007; 20 King CD, Rios GR, Green MD, Tephly TR.
UDP-glucuronosyltrans73: 262–8. ferases. Curr Drug Metab 2000; 1: 143–61.
5 Nebert DW, Dalton TP. The role of cytochrome P450 enzymes in 21 Zhang W, Liu W, Innocenti F, Ratain MJ. Searching for tissue-specific
endogenous signalling pathways and environmental carcinogenesis. expression pattern-linked nucleotides of UGT1A isoforms. PLoS One
Nature 2006; 6: 947–60. 2007; 2: e396.
6 Bachmann KA, Ghosh R. The use of in vitro methods to predict in vivo 22 Hasler JA, Estabrook R, Murray M, Beaune P. Human cytochromes
pharmacokinetics and drug interactions. Curr Drug Metab 2001; 2: P450. Mol Aspects Med 1999; 20: 1–137.
299–314. 23 Chiu SH, Huskey SW. Species differences in N-glucuronidation. Drug
7 Howgate EM, Rowland Yeo K, Proctor NJ, Tucker GT, Rostami-Hodjegan Metab Dispos 1998; 26: 838–47.
A. Prediction of in vivo drug clearance from in vitro data. I: impact of 24 Green MD, Tephly TR. Glucuronidation of amines and hydroxylated
inter-individual variability. Xenobiotica 2006; 36: 473–97. xenobiotics and endobiotics catalyzed by expressed human UGT1.4
8 Obach RS, Walsky RL, Venkatakrishnan K, Gaman EA, Houston JB, protein. Drug Metab Dispos 1996; 24: 356–63.
Tremaine LM. The utility of in vitro cytochrome P450 inhibition data in 25 Kassahun K, Mattiuz E, Franklin R, Gillespie T. Olanzapine
10-Nthe prediction of drug-drug interactions. J Pharmacol Exp Ther 2006; glucuronide. A tertiary N-glucuronide unique to humans. Drug Metab
316: 336–48. Dispos 1998; 26: 848–55.