Study Of The Effect Of Alcohol To Mangiferin Blood Concentration And Excretion Characteristics In Macaca Fascicularis

Objective: By comparing the blood concentration and excretion characteristics of the mangiferin tablets in alcohol and non-alcohol Macaca fascicularis,the effect on pharmacokinetics of alcohol will be revealed,the data will support to non-clinical research and guides clinical medication.Methods:(1)Establishing HPLC method of mangiferin and its metabolites in biological samples: The contents of mangiferin in blood and its metabolites M1,M2 and M3 in feces and urine wereinvestigated,including linear relationship,recovery rate,precision,repeatability,stability.(2)Pharmacokinetic study of mangiferin in Maraca fascicularis: Macaca fascicularis were given normal diet,drank water freely,every 3 Maraca fascicularis a dose;Macaca fascicularis were given 288 mg/kg,576 mg/kg and 864 mg/kg(mangiferin is accounted)by or aladministration.The blood sample was obtained at 0,1,2,4,6,9,11,13,16,22,26,28,30,32 h from Macaca fascicularis' s limb;Urine and feces sample were obtained during 0-120 h.According to “Establishing HPLC method of mangiferin and its metabolites in biological samples”,the content of mangiferin in the plasma and its metabolites in feces and urine were measured.The pharmacokinetic parameters were calculated by pharmacokinetic software DAS2.0;The excretion of mangiferin and its metabolites in feces and urine were calculated by each period of time(expressed as mole percent),and draw the excretion curve.(3)Pharmacokinetic study of mangiferin in alcohol drinking Maracafascicularis: Establishing the alcohol dependent model in Macaca fascicularis,alcohol was administered at a dose of 5% concentration in water of 30 days,the blood alcohol concentration was detected after 7?14?28 days,the level of plasma ACTH and cortisol was compared before/during drinking;To study the effect of alcohol to mangiferin pharmacokinetics and excretion in macaca fascicularis.(4)The effect of alcohol to mangiferin blood concentration and excretion characteristics in Macaca fascicularis: Comparing the pharmacokinetic parameters of mangiferin in alcohol and non-alcoholic maraca fascicularis,and the excretion and total excretion of mangiferin and its metabolites in feces and urine of alcohol and non-alcoholic maraca fascicularis.Result:(1)The chromatographic conditions for determination of mangiferin blood and mangiferin and its metabolites in blood,feces and urine as folow: Detection wavelength at 258nm;Column temperature in 30°C;Injection volume 10?l;The mobile phase consisted of methanol(A)and(B)0.1% formic acid solution.The analytical column was Shimadzu ODS C-18(250×4.6 mm,5?m).The linear equation of mangiferin and internal standard in Macaca fascicularis plasma was Y=27.681X-0.1461,r2=0.9984,the standard curve in the range of 0.504-50.4?g/ml had good linear relation.The linear equation of mangiferin and internal standard in Macaca fascicularis feces was Y=3.6249X-0.1309,r2=0.9942,the standard curve in the range of 1.68-336?g/ml had good linear relation;The linear equation of M1 and internal standard in Macaca fascicularis feces was Y=10.227X-0.1305,r2=0.9992,the standard curve in the range of 1.5-300?g/ml had good linear relation;The linear equation of M2 and internal standard in Macaca fascicularis feces was Y=8.4719X-0.0183,r2=0.9994,the standard curve inthe range of 1.11-333?g/ml had good linear relation;The linear equation of M3 and internal standard in Macaca fascicularis feces was Y=3.3714X-0.1644,r2=0.9956,the standard curve in the range of 1.66-664?g/ml had good linear relation;The linear equation of mangiferin and internal standard in Macaca fascicularis urine was Y=4.208X+0.348,r2=0.9957,the standard curve in the range of 1.68-336?g/ml had good linear relation;The linear equation of M1 and internal standard in Macaca fascicularis urine was Y=10.745X-0.1441,r2=0.9975,the standard curve in the range of 1.5-300?g/ml had good linear relation;The linear equation of M2 and internal standard in Macaca fascicularis urine was Y=9.2432X-0.0782,R2=0.9986,the standard curve in the range of 1.11-333?g/ml had good linear relation;The linear equation of M3 and internal standard in Macaca fascicularis urine was Y=3.5515X-0.1854,r2=0.9930,the standard curve in the range of M3 of 1.66-664?g/ml had good linear relation.The recoveries of mangiferin,M1,M2,M3 were between 85-115%,the RSD% of intra-day precision,repeatability,24-hour stability,frozen 14-day stability were less than 15%.(2)The main pharmacokinetic parameters of three doses(288mg/kg,576mg/kg,864m/kg)of mangiferin in non-alcoholic Macaca fascicularis respectively: Cmax was 2.0956±0.0329?g/ml,2.2763±0.1990?g/ml,3.2580±0.0447?g/ml;AUC(0-t)was 30.5690 ± 1.2869(?g/ml)·h,32.1697 ±1.077(?g/ml)·h,38.644 ± 0.5661(?g/ml)·h;AUC(0-?)was 53.6777 ±0.5956(?g/ml)·h,48.8297±2.5018(?g/ml)·h,57.8203±0.6486(?g/ml)·h;t1/2a was 2.7820±0.2780 h,2.3387±0.0950 h,2.2927±0.1204h;Ka was 0.382±0.0188h-1,0.39937±0.0156h-1,0.4000±0.0184h-1,K12 was 0.133±0.0225h-1,0.1523±0.0031h-1,0.1423±0.0097h-1;K10 was 0.0707±0.0021h-1,0.1047±0.00823h-1,0.1333±0.0111h-1.The total molarratio of mangiferin and its metabolites M1,M2,M3 in the non-alcoholic Macaca fascicularis feces and urine were 53.358±0.923%,52.602±1.368%,49.739±1.435% in 288,576 and 864mg/kg three-dose group;Of which the prototype mangiferin was 4.530±0.179%,6.411±0.336%,6.485±0.403%;In addition,the molar ratio of M1 excretion was: 7.447±0.721%,8.069±0.164%,6.741±0.626%;The molar ratio of M2 excretion was: 15.024±1.393%,14.336±0.467%,13.591±1.303%;The molar ratio of M3 excretion was: 16.834±1.589%,15.959±1.543%,13.918±0.968%.The excretion of mangiferinin feces was 2.557±0.141%,3.969±0.255%,3.710±0.455%,the excretion of M1 was 4.875±0.631%,3.814±0.128% 3.130±0.609%,the excretion of M2 was 6.319±0.952%,6.602±0.47%,6.770±0.651%,the excretion of M3 was 16.834±1.589%,15.959±1.543%,13.918±0.968%;The excretion of mangiferin in the urine was 1.973±0.05%,2.715±0.271%,2.775±0.189%,and the excretion of M1 was 2.572±0.291%,4.255±0.251%,3.611±0.2%,and the excretion of M2 was 8.705±0.521%,7.735±0.064%,6.854±1.000%,and the excretionof M3 was 9.524±1.231%,7.826±0.866%,9.003±0.352%.(3)There was no significant differences in the concentration of alcohol in the blood of the Macaca fascicularis(4.23±0.14mg/100 ml,4.30±0.17 mg/100 ml,4.19±0.11mg/100ml)(P>0.05).The level of plasma ACTH and cortisol have significant increase after ethanol withdrawal(P<0.05 or P<0.01).The main pharmacokinetic parameters of three dose(288mg/kg,576mg/kg,864m/kg)of mangiferin in alcohol Macaca fascicularis respectively: Cmax was 2.4396±0.0975?g/ml,2.9822±0.0266?g/ml,3.8512±0.1672?g/ml;AUC(0-t)was 33.8087±1.4440(?g/ml)·h,37.1883±0.3358(?g/ml)·h,42.0070±1.9510(?g/ml)·h,AUC(0-?)was58.5070±1.4791(?g/ml)·h,54.9657±1.6947(?g/ml)·h,61.3960±2.0153(?g/ml)·h;t1/2a was 2.1210±0.1637 h,1.9603±0.0680 h,1.9110±0.0705h;Ka is 0.4797±0.0057h-1,0.4657±0.0340h-1,0.4790±0.0242h-1,K12 is 0.1793±0.0156h-1,0.1717±0.0115h-1,0.1617±0.0064h-1,K10 is 0.0907±0.0084h-1,0.1417±0.0072h-1,0.1707±0.0091h-1.The total molar ratio of mangiferin and its metabolites M1,M2,M3 in the non-alcoholic Macaca fascicularis feces and urine were 52.860±1.313%,50.305±1.363%,49.705±1.324% in 288,576 and 864mg/kg three-dose group;Of which the prototype mangiferin was 4.686±0.177%,6.515±0.175%,6.307±0.397%;In addition,the molar ratio of M1 excretion was: 7.900±0.376%,7.369±0.640%,7.293±0.533%;The molar ratio of M2 excretion was: 15.611±0.377%,14.165±0.307%,12.989±0.180%;The molar ratio of M3 excretion was: 24.663±1.011%,22.256±0.675%,23.116±0.390%.The excretion of mangiferin in feces was 2.012±0.146%,3.022±0.060%,2.917±0.310%,the excretion of M1 was 6.084±0.275%,4.362±0.314%,4.545±0.471%,the excretion of M2 was 7.949±0.204%,7.507±0.108%,7.823±0.123%,the excretion of M3 was 12.374±0.302%,12.743±0.336%,11.813±0.627%;The excretion of mangiferin in the urine was 2.647±0.106%,3.493±0.157%,3.390±0.284%,and the excretion of M1 was 1.816±0.158%,3.007±0.388%,2.748±0.074%,and the excretion of M2 was 7.662±0.212%,6.658±0.212%,5.166±0.147%,and the excretion of M3 was 12.289±0.753%,9.513±0.355%,11.304±0.603%.(4)The results showed that there were significant differences in the main pharmacokinetic parameters between the alcoholic and non-alc oholic Macaca fascicularis: Cmax,AUC(0-t),AUC(0-?),Ka,K12,K10,V,C L,t1/2a,t1/2Ka,t1/2?(P<0.01 or P<0.0)had significant differences.The total excretion of M0,M1,M2,M3 in feces and urine,excretion ofM0,M1,M2,M3 in feces,excretion of M0,M1,M2,M3 in urine,the total excretion of M0 in feces and urine,the total excretion of M1 in feces and urine,the total excretion of M2 in feces and urine,and the total excretion of M3 in feces and urine(P>0.05),had no significant differences.There was significantdifference in the excretion of M0,M1,M2 and M3 in feces and urine(P<0.01 or P<0.05).Conclusion:(1)The RP-HPLC method for the analysis of mangiferin and its metabolites in the biological samples established in this experiment is sensitive,rapid and stable,which meets the basic requirements of biological samples.(2)The alcoholic model issuccessful.(3)Alcohol can increase the concentration of mangiferin and increase the bioavailability of mangiferin.Alcohol has no effect on the total excretionof mangiferin and its metabolites in Macaca fascicularis in feces and urine,but has effects on the excretion on feces and urine to single component.