Preparation Of Compound Timicosion And Trimethoprim Lactate Nanoemulsion And Its Pharmacodynamics

In this research,we made nanoemulsion as carrier of compatibility of Timicosion(TMS) and Trimethoprim Lactate(TMPL).The formulations of Compound Timicosion and Trimethoprim Lactate Nanoemulsion (TMS-TMPL-NE) were optimized by protract the pseudoternary phase diagrams to prepare TMS-TMPL-NE. Then the stability, safety, vitro antibacterial effect,pharmacy of TMS-TMPL-NE were studied providing a theoretical basis for it in the veterinary clinical application.1. Analytical method establishment of TMS-TMPL-NEThe content determination of TMS and TMPL analytical methods were respectively established established by ultraviolet-visible spectrophotometer. The results showed that two good linearity were obtained by TMS in the range of 5～50μg·mL-1 and by TMPL in the range of 1～30μg·mL-1, and the average recovery, relative standard deviation(RSD), RSD of the with-in-day precision, RSD of the day-to-day precision of TMS and TMPL were (99.18±1.89)% and (99.68±2.23)%, 1.91% and 2.24%, 0.62% and 1.32%,0.65% and 1.42%, respectively. The analytical methods of ultraviolet spectrophotometry (UV) which established had high recovery rate, good repetitiveness and precision in this research. It could offer a good specificity method of content determination of TMS–TMPL-NE.2. Preparation and quality evaluation of TMS-TMPL-NEThe effect of formation of the nanoemulsion factors was investigated by pseudo-ternary phase diagrams. The composition of nanoemulson was determined initially. The optimal match of TMS and TMPL was determined by Kirby-Baueer test. The prepared prescription of TMS-TMPL-NE was ascertained finally. The structure type of TMS-TMPL-NE was judged by centrifugation and staining method.Its appearance and particle diameter distribution was investigated by transmission electron microscope and laser particle size analysator respectively. Through photostability testing by strong light,accelerated testing and long-term testing verify its stability.The final prepared prescription of TMS-TMPL-NE was w(tween-80) =24.93%,w(ethanol and propylene glycol)=16.62%,w(phosphori cacid)=1.5%, w(IPM) =4.57%,w(TMS)=5.98%,w(TMPL)=0.66%,w(H2O)=45.74%.The determined match between TMS and TMPL was 1:1. The prepared TMS-TMPL-NE was the type of oil-in-water and the liquid was amber, transparent and homogeneous. The nanoemulsion drop presented spherical shape, and the drop size averaged 12.4 nm with a polydispersity index of 0.051. TMS-TMPL-NE had well stability and the expiration date was 20 months. The results showed that the quality of TMS-TMPL-NE was stable,achieving requirement of clinical medication.3. Safety evaluation of TMS-TMPL-NEThe safety of TMS-TMPL-NE was evaluated by acute toxicity test and skin irritation test. The median lethal dose(LD50) of TMS-TMPL-NE was 2346 mg.kg-1, so it belong to Low toxicity drug.Serious irritation didn't appear when TMS-TMPL-NE was used on complete and damaged skin of rabbits respectively.The results showed that TMS-TMPL-NE was safe by oral and intramammary administration.4. Combination antibacterial effect of TMS-TMPL-NE in vitroCombination antibacterial effect of TMS-TMPL-NE against three clinical strains in vitro was investigated by single drug minimum inhibitory concentration(MIC) determination, combination susceptibility test and Kirby-Baueer disc diffusion test. The results showed that synergistic effect was generated when TMS and TMPL were combined. MIC of the mixed TMS and TMPL against Staphylococcus aureus was 1/4 and 1/8 times compared with single drug respectively, MIC against Streptococcus agalactiae was both 1/4 times, and MIC against Escherichia coli was both 1/4 and 1/16 times. Each of fractional inhibitory concentration(FIC) index was not more than 0.5. The disc diffusion zone diameters of TMS-TMPL-NE against bacteria were significant(P<0.05)compared with positive control group,and highly significant difference(P<0.01)other each group. The results suggested that combination antibacterial activity of TMS-TMPL-NE against three pathogenic bacteria was strong.5. Therapeutic efficacy of TMS-TMPL-NE against chronic respiratory disease of chickenThe high, middle and low dose of TMS-TMPL-NE and TAI KE XING was respectively dosed by drinking freely to treat chronic respiratory disease of chicken. The results indicated that cure rate of TMS-TMPL-NE in high, middle and low dose group was 98%,88% and 72% respectively, and effective rate was 100%,98% and 89% respectively. The cure rate and effective rate of Tai Ke Xing was 74% and 91% respectively. The results of the statistical analysis showed the therapeutic efficacy of high and middle dose group were both obviously better than low dose group and TAI KE XING group(P<0.05). Clinical recommended dose was middle on these grounds when TMS-TMPL-NE was used to treat chronic respiratory disease of chicken. Administration dose could increase fairly for serious one.The prepared TMS-TMPL-NE had a high safety,good stability and high–perf ormance, That could provide a new type of nano-antibacterial drugs for veterinary clinical medication.