The Metabolism Of Amoxicillin-Sulbactam Heterozygous Molecule In Vivo And Its Pharmacokinetics In Mice

In this paper,an in vitro degradation study of a novel amoxicillin-sulbactam heterozygous molecules（AS）,and its metabolism and pharmacokinetics in mice were carried out.These studies provided basic data for further research and development of AS.The Ultra high performance liquid chromatography（UPLC）method was established to monitor the degradation of AS in artificial intestinal juice and artificial gastric juice.AS were incubated in two kinds of biological substrates at 37℃.The recovery rate of amoxicillin,sulbactam and AS in the biological matrix were between 85.20%and 95.33%.The limit of detection（LOD）were 1,5 and 1μg/m L,and the limit of quantification（LOQ）were 2,10 and 2μg/m L,respectively,which could meet the detection requirements of the in vitro degradation.The results showed that AS was degraded about 20%in artificial gastric juice for 4 hours,but AS was degraded in artificial intestinal juice within 4 hours completely,about 89%of which degraded to amoxicillin and sulbactam.At the same time,a small amount of other metabolites were produced.The results indicated that AS was mainly degraded in artificial intestinal juice,and the degradation products of AS were amoxicillin and sulbactam.The Ultra-high-pressure liquid chromatography/quadrupole-time of flight tandem mass spectrometry（UPLC/Q-TOF MS）method was used to isolate and identify the metabolites of AS in artificial intestinal fluid,mouse plasma,urine,feces,liver and kidney.The results showed that a large number of amoxicillin and sulbactam were detected in artificial intestinal fluid and mice.The metabolites（M1 and M2）of AS were also detected in artificial intestinal fluid.In addition,M1 was detected in mouse liver and urine.Amoxicillin-related metabolites were detected in artificial intestinal fluid and mice which was amoxicillin acid and amoxicillin diketopiperazine.The experimental indicated that the main metabolic mode of AS was the hydrolysis of diester bond to amoxicillin,sulbactam and a small amount of metabolite M1,which provides theoretical basis and support for later experiments.A total of 216 Kunming mice were randomly divided into 3 groups which was Amoxicillin,amoxicillin/sulbactam compound and AS.Blood samples were collected from orbital vein at 0.083,0.166,0.25,0.25,0.5,1,1.5,2,3,4,6 and 8 h after oral administration,respectively.The ultra-high performance liquid chromatography-tandem mass spectrometry（UPLC-MS/MS）method was established for the determination of amoxicillin and sulbactam in mice plasmas,and the pharmacokinetic parameters were calculated by Win Nonlin software.The results showed that the peak time(T_max)of amoxicillin in plasma of mice in each group was 0.25 h,the peak concentration(C_max)were3.06±0.48μg/m L,2.65±0.25μg/m L and 3.01±0.50μg/m L,The elimination half-life(t_1/2ke)was 0.65±0.15 h,0.67±0.11 h and 0.83±0.10 h,respectively,the average retention time(MRT_last)was 0.87±0.11 h,0.82±0.12 h and 1.00±0.09 h and the area under the concentration-time curve(AUC_all)was 2.31±0.60μg·h/m L,2.29±0.53μg·h/m L and2.20±0.45μg·h/m L,respectively.The t_1/2keof AS group was longer than that of amoxicillin alone and compound group,indicating that AS may have a sustained release effect.The T_maxof sulbactam in plasma of compound group and AS were 0.18±0.03 h and 0.20±0.04 h,C_maxwere 2.20±0.42μg/m L and 2.07±0.28μg/m L,t_1/2kewere 0.85±0.31 h and 0.81±0.38 h,MRT_lastwere 0.84±0.23 h and 1.02±0.17 h,and AUC_allwere 1.21±0.30μg·h/m L and1.53±0.16μg·h/m L,respectively.The AUC_allin AS group was higher than that in compound group,indicating that sulbactam was absorbed more after administration of AS than that of compound prescription.