ABSTRACT
OBJECTIVE@#To reveal the effect and mechanism of Jiaotai Pill (, JTP) on insomniac rats.@*METHODS@#The insomniac model was established by intraperitoneal injection of p-chlorophenylalanine (PCPA). In behavioral experiments, rats were divided into control, insomniac model, JTP [3.3 g/(kg•d)], and diazepam [4 mg/(kg•d)] groups. The treatment effect of JTP was evaluated by weight measurement (increasement of body weight), open field test (number of crossings) and forced swimming test (immobility time). A high performance liquid chromatography-electrochemical detection (HPLC-ECD) method was built to determine the concentration of monoamine transmitters in hypothalamus and peripheral organs from normal, model, JTP, citalopram [30 mg/(kg•d)], maprotiline [40 mg/(kg•d)] and bupropion [40 mg/(kg•d)] groups. Expressions of serotonin transporter (SERT), dopamine transporter (DAT), and norepinephrine transporter (NET) were analyzed by quantitative polymerase chain reaction (qPCR) and Western blot in normal, model and JTP groups. A high performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-ESI-MS/MS) method was established to determine the pharmacokinetics, urine cumulative excretion of metformin in vivo, and tissue slice uptake in vitro, which were applied to assess the activity of organic cation transporters (OCTs) in hypothalamus and peripheral organs.@*RESULTS@#Compared with the insomniac model group, the body weight and spontaneous locomotor were increased, and the immobility time was decreased after treatment with JTP (P<0.01). Both serotonin and dopamine contents in hypothalamus and peripheral organs were increased (P<0.01). The norepinephrine content was increased in peripheral organs and decreased in hypothalamus (P<0.05 or P<0.01). At the same time, SERT, DAT, OCT1, OCT2, and OCT3 were down-regulated in hypothalamus and peripheral organs (P<0.05). NET was down-regulated in peripheral organs and up-regulated in hypothalamus (P<0.05 or P<0.01). Moreover, the activity of OCTs in hypothalamus and peripheral organs was inhibited (P<0.05).@*CONCLUSION@#JTP alleviates insomnia through regulation of monoaminergic system and OCTs in hypothalamus and peripheral organs.
ABSTRACT
To predict the mechanism of liver injury induced by Genkwa Flos, we investigated the effect of chloroform extract on UGTs and UGT1A1 activities of the liver microsomes in rat and human. In the present study, 4-nitrophenol(4-NP) and β-estradiol were elected as substrates to determine activities of UGTs and UGT1A1 by UV and HPLC. The results showed that there were 1.00% of apigenin, 6.40% of hydroxygenkwanin and 18.38% of genkwanin in chloroform extract; and total diterpene mass fraction was 31.40%. Compared with the control group, chloroform extract could significantly inhibit the activity of UGTs in rat liver microsomes(RLM) system, while the inhibitory effect was not obvious in human liver microsomes(HLM) system. UGT1A1 activity was inhibited by chloroform extract in rat liver microsomes and human liver microsomes (based on genkwanin, IC₅₀=8.76, 10.36 μmol•L⁻¹). The inhibition types were non-competitive inhibition(RLM) and uncompetitive inhibition(HLM). In conclusion, the results indicated that chloroform extract showed different inhibitory effects on UGTs and UGT1A1 activity, which may be one of the mechanisms of liver injury induced by Genkwa Flos.
ABSTRACT
In the present study, the effects of six Coptidis alkaloids (berberine, epiberberine, coptisine, jatrorrhizine, palmatine and magnoflorine) on liver microsomes UGTs and UGT1A1 activities in rats and mice were investigated in vitro and in vivo to study the mechanism of metabolic drug-drug interactions of Coptidis Rhizoma with other drugs. In vitro rat and mice liver microsomal incubation systems combined with UDPGA were applied, as well as mice liver microsomes after administration of six Coptidis alkaloids. 4-Nitrophenol and β-estradiol were selected as substrates to determine activities of UGTs and UGT1A1 by UV and HPLC, respectively. According to the in vitro rat study, berberine, epiberberine, coptisine and jatrorrhizine significantly inhibited rat liver microsome UGTs activity, particularly epiberberine showed the strongest inhibition. UGT1A1 activity was lowly inhibited by jatrorrhizine, with IC₅₀ at about 227 μmol•L⁻¹, whereas coptisine and magnoflorine significantly activated UGT1A1. According to the in vitro mice study, berberine, coptisine, jatrorrhizine and palmatine significantly inhibited mice liver microsome UGTs activity, and the six alkaloids all significantly activated UGT1A1. According to the in vivo mice study, UGTs activity was significantly activated only in berberine group, while UGT1A1 activity was significantly activated only in jatrorrhizine group. In conclusion, the effects of Coptidis alkaloids on UGT activity showed significant differences in species and between in vitro and in vivo. Meanwhile, the changes in structures of Coptidis alkaloids also have a big impact on UGT activity, which may be one of the causes for the drug-drug interactions between Coptidis Rhizoma and other drugs.
ABSTRACT
To predit the mechanism of metabolic drug-drug interactions of hydroxygenkwanin with other drugs, we investigated the inhibition inhibitory effect of hydroxygenkwanin on UGTs and UGT1A1 activities of different liver microsomes. In the present study, 4-nitrophenol (4-NP) and β-estradiol were elected as substrates to determine activities of UGTs and UGT1A1 by UV and HPLC, respectively. The results showed that, hydroxygenkwanin significantly inhibited UGTs activity in rat, mouse and human liver microsomes. UGT1A1 activity was inhibited by hydroxygenkwanin to varying degrees, with IC₅₀ about 190, 10.93, 20.07, 76.31 μmol•L⁻¹ in mouse liver microsome(MLM), rat liver microsome (RLM) and recombinant UGT1A1, and human liver microsome (HLM), respectively. The inhibition types were competitive inhibition (RLM, HLM) and linear mixed-typed linear inhibition (recombinant UGT1A1). The order for the inhibitory intensity was RLM>rUGT1A1>HLM>MLM. In conclusion, hydroxygenkwanin has an inhibitory effect on UGTs and UGT1A1 activities of different liver microsomes, with differences in species, indicating its potential drug interactions based on UGT1A1 enzyme. This study aims to provide a reliable experimental basis for its further research and development of hydroxygenkwanin, and provide theoretical reference for the clinic drug combination research.