| Milbemycin Oxime-Praziquantel Tablets are a compound insecticide,which is composed of Milbemycin Oxime and Praziquantel and has been approved in foreign countries for the removal of parasitic worms such as worms,worms,whips,and tapeworm in the canine gastrointestinal tract.Milbemycin Oxime and Praziquantel have been combined to expand the insect repellent spectrum.Based on the HPLC method for the determination of the content of Milbemycin Oxime and Praziquantel in canine plasma,this study according to the biological equivalence test of veterinary chemical,the bioequivalence test of domestic imitation was carried out by using the original research drug of Milbemycin Oxime-Praziquantel Tablets as reference product.To determine whether the domestic Milbemycin Oxime-Praziquantel Tablets and the reference listed drug has a clinical replaceable efficacy,and provide basis for clinical rational drug use of domestic Milbemycin Oxime-Praziquantel Tablets.1.A method for the determination of Milbexime in Dog Plasma by HPLCThe plasma of Milbemycin Oxime was abstracted and purified by acetonitrile,determined by high performance liquid chromatography with ultraviolet detection,and quantified by external standard method.The mobile phase is acetonitrile-water solution(85:15).The UV detection wavelength is 244 nm.The blank plasma supplemented with milbeloft has good linearity in the range of 0.05-10 μg/mL(R2>0.999).The limit of detection and the limit of quantification are 0.04 pg/mL and 0.05 μg/mL,respectively.The intra-and inter-assay coefficients of variation were less than 8.971%and 8.675%,respectively,when the blank plasma levels of Milbemycin Oxime were between 0.05 and 10.0 μg/mL.This method stipulates the extraction and purification of Milbemycin Oxime in plasma and is applicable to the determination of Milbemycin Oxime in dog plasma.2.A method for the determination of Praziquantel in Dog Plasma by HPLCThe plasma of praziquantel was abstracted and purified by acetic ether,determined by HPLC with UV detection,and quantified by external standard method.The mobile phase is acetonitrile-water solution(49:51).The UV detection wavelength is 223 nm.The blank plasma supplemented with praziquantel had good linearity in the range of 0.1-10 μg/mL(R2>0.999).The detection and quantitation limits were 0.05 μg/mL and 0.1 μg/mL,respectively.When the praziquantel plasma level was 0.1-10.0 μg/mL,the intra-and inter-assay coefficients of variation were less than 7.707%and 10.536%,respectively.This method stipulates the extraction and purification of praziquantel in plasma and is suitable for the determination of praziquantel in dog plasma.3.Bioequivalence test of Milbemycin Oxime-Praziquantel tablets in dogs18 male Beagle dogs were randomly divided into 2 groups with 9 in each group.Cross-test design is adopted,In the first stage,animals in the first group were given oral test products,and animals in the second group were given oral reference products.In the second stage,animals in the first group were given oral reference products,and animals in the second group were given oral test products.The two-stage drug delivery interval was 2 weeks.Fasting for 12 hours before administration and collecting a blank blood sample.after the drug,according to the scheduled time from the dog’s forelimbs under the subcutaneous veins to collect blood samples.The contents of Milbemycin Oxime and Praziquantel in canine plasma were respectively determined by HPLC using a verified canine plasma of Milbemycin Oxime and Praziquantel.The plasma concentration-time data of the measured Milbemycin Oxime and Praziquantel were analyzed by using the data analysis software of WINNONLIN version 6.4,and the pharmacokinetic parameters and Bioequivalence parameters were calculated by the method of non room model.The results of pharmacokinetic parameter fitting show that the average elimination half-life(T1/2)of Milbemycin Oxime was 29.39 H and 31.35 H respectively,and the average peak time(Tmax)was 2.61 H and 2.78 H respectively,and the average retention time was 11.83,respectively,after single oral administration of Praziquantel tablets and reference products in dogs.H and 11.36 H,peak concentrations(Cmax)are 0.294μg/ml and 0.421 μg/ml,respectively,from 0 to the last measurable sampling point under the Curve area(AUClast)respectively 4.51 μg*h/l and 4.25μg*h/l The average elimination half-life(T1/2)of Praziquantel was 1.68 H and 1.71 H,and the mean peak time(Tmax)was 1.11 H and 1.56 H respectively,and the mean residence time was 2.08 H and 2.34 h respectively,peak concentration(Cmax)were 1.035μg/ml and 1.135 respectively.MG/ML,the area under the curve(AUClast)was 2.72μg*h/l and 2.75μg*h/l respectively.The relative bioavailability of Milbemycin Oxime and Praziquantel was 106.12%and 98.91%,respectively,in the sample of the original research product of the Milbemycin Oxime-Praziquantel tablets.Pharmacokinetic studies showed that Milbemycin Oxime was slow to eliminate in dogs and that Praziquantel was eliminated quickly.Statistical analysis showed that there was no significant difference(p>0.05)between the main pharmacokinetic parameters of the two active components of Milbemycin Oxime tablets and reference products.The Winnonlin 6.4 version of the calculation software was used to test the bioequivalence of Cmax(logarithmic conversion),AUC0-t(logarithmic conversion)and AUC0-∞(logarithmic conversion)of the Praziquantel tablets and the reference products.The 90%confidence intervals of the geometric mean ratio of AUC(0-60h)in the test and reference group were 94.03%-115.79%,and the 90%confidence interval of Cmax geometric mean ratio was between 71.64%-105.85%and The 90%confidence intervals of the AUC(0-60h)geometric mean ratio of Praziquantel in subjects and reference groups were respectively 83.43%-98.75%of the 90%confidence interval for the 87.59%-105.71%,cmax geometric mean ratio.The 90%confidence intervals of the AUClast and Cmax geometric mean ratios of Milbemycin Oxime and Praziquantel are in the set range.The results showed that the test samples and reference products were bioequivalent and can be clinically substituted for each other. |
