Then Effects Of Shuanghuanglian Injection On Cyp In Rats

Cytochrome P450(CYP),which is a kind of of mixed function oxidase,can catalyze many kinds of reactions in the body.This CYP enzyme takes part in drug metabolism of 75-80 percent and eliminates medicaments of 65-70 percent in the clinic.Omeprazole(OMP) is a proton pump inhibitor to be used in the treatment of peptic ulcer.In the liver,it is primarily metabolized by cytochrome P-450(CYP450) isoenzymes such as CYP2C19 and CYP3A4.5-Hyroxyomeprazole(5-OHOMP) and omeprazole sulfone(OMP-SFN) are the two major metabolites of OMP in human. While is primarily metabolized via hepatic CYP1A1/2,2D1/2,in male Sprague-Dawley rats.This study selected Omeprazole as reference to examine the consequence of Shuanghuanglian Injection on CYP.The main constitutes of Shuanghuanglian Injection are honeysuckle flower, Scutellaria baicalensis and fructus forsythia.Its bases is baicalin.It is a kind of complex prescription of Chinese drugs.It is mainly used for fever and coughing caused by affection of exogenous wind-heat and many types of the respiratory tract infection complications,having been the primary medicine of treating respiratory tract infection complications in Chinese drugs.This paper makes researches on the consequence of Shuanghuanglian Injection and baicalin on CYP.It does prediction of the mutual effects of metabolite medicines when Shuanghuanglian Injection and other medicines are used together,supplying reference to the rational administration of drug in the clinic. Materials and methods:1.Administration In accordance with the principle of randomization,the rats were set experimental group and control group.Every group had 9 male SD rats, respectively.The rats of experimental group were injected Shuanghuanglian Injection (10.42mL·kg^-1).While the rats of control group were dosed the same quantity of normal saline through vein every day.The rats were dosed continuously for 7 days.2.Preparation of liver microsome and determination of liver microsome protein The rats of the experimental group and the control group were killed by decapitation and the livers were cleaned.The liver microsome were prepared by the way of Calcium precipitation approach.Protein concentration of liver microsome was determined by Bradford approach.3.Incubation reaction system Incubation system contained NADPH,liver microsome,phosphate buffer(pH7.4) and probe drug(omeprazole,OPZ).The overall volume of incubation system is 200μL.The microsomal pretein was preincubated for 5min at 37℃.The reaction was started since NADPH was mixed.4.Species dealing and chromatographic conditions After 20min of reaction, adding the cold acetonitrile of 200μL is to end the reaction.Take 30μL to make analysis after 10min of 12000rpm's centrifugation.Chromatographic conditions:chromatographic column was 5μm DiamonsiL C18 column(particle size 5μm,4.6mm×200 mm).The mobile phase consisted of methanol-acetonitrile-0.1 mol·L^-1 ammonium acetate butter solution(10:35:55,v:v:v) at a flow rate of 1.0ml·min^-1.The detection of omeprazole was performed at wavelength of 302nm.The column tempreture was 25℃.The injection volume was 30μl.5.Inhibition of Shuanghuanglian Injection on CYP in vitro Added different volume of Shuanghuanglian Injection to blank incubation system of liver microsome. The CYP activity in vitro was measured by the determination of OPZ using HPLC.6.Inhibition of Baicalin on CYP in vitro Added different constituencies of Baicalin to incubation system of blank liver microsome.The CYP activity in vitro was measured by the determination of OPZ using HPLC.7.The influence of Shuanghuanglian Injection on CYP in vivo Determined the consistency of OPZ using HPLC in incubation system of the liver microsome of the experimental group and control group,respectively.Result1.The inhibitory experiment of Shuanghuanglian Injection on CYP in vitro When the concentrations of Shuanghuanglian Injection in incubation system were 0, 0.5,1,1.5,2,2.5,3.0μl·100μl(-1),TR values were 477.07±14.13,324.15±13.68,258.74±12.96,201.32±21.60,100.89±4.68,84.84±10.13,68.67±21.76 pmol·min^-1·mg^-1.CYP2C19 activity decreased with increasing concentration of Shuanghuanglian Injection.IC₅₀ was 0.96μl·100μl^-1.2.The inhibitory curve of Baicalin on CYP in vitro When the concentrations of Baicalin in incubation system were 0,6,10,15,20,25,30,35 mg·mL^-1,TR values were 419.47±27.52,298.89±9.20,279.55±16.09,211.21±10.91,163.31±63.54,115.71±19.56,102.29±22.24,21.81±3.50pmol·min^-1·mg^-1.CYP2C19 activity decreased with increasing concentration of Baicalin.IC50 was 12.89mg·mL^-1.3.The influence of Shuaughnanglian Injection on CYP in vivo The TR values of the experimental group and control group were 298.96±13.63pmol·min^-1·mg^-1 and 204.73±23.62pmol·min^-1·mg^-1.Shuanghuanglian Injection had marked inhibition on CYP activity,(P＜0.05).Conclusion1.Shuanghuanglian Injection and Baicalin can inhibit the activity of CYP in vitro.2.Shuanghuanglian Injection can also inhibit the activity of CYP in vivo.