Development And Bioavailability Evaluation In Hens Of Florfenicol Nanocrystals

As a broad-spectrum antibiotic for animals,florfenicol（FFC）has many advantages,such as fast absorption,wide distribution,fast metabolism,good antibacterial effect,wide safety range,low side effects,difficulty to develop drug resistance,etc.FFC is widely used in the prevention and treatment of bacterial diseases in cattle,sheep,pigs,aquatic animals,and poultry.However,FFC is a low-solubility/high-permeability drug,which is slightly soluble in water.Making FFC into nanocrystals will improve its solubility in water and other physical properties,as well as its bioavailability.The purpose of this study is to prepare florfenicol nanocrystal（FFC-NC）by emulsion solvent evaporation method.The preparation process was optimized by single factor test,and the physicochemical characterization and bioavailability of the FFC-NC were evaluated.FFC exerts its efficacy in the form of nanocrystals,which will improve its solubility and physical properties,and increase its bioavailability.1.Development and process optimization of FFC-NCBy single factor test analysis,the hydrodynamic size of FFC-NC was used as the main parameter for screening the optimal preparation conditions.The results showed that the best condition for the preparation of FFC-NC was as follows:volume ratio of organic phase and aqueous phase of 1:2,concentration of P188 of 5‰,speed of homogenization at 7000 r/min,time of homogenization of 5 min,pressure of homogenization of 300 bar,and number of homogenization cycles as 3.The average particle size of FFC-NC prepared under the best condition was 226.1±11.3 nm and the average polymer dispersity index（PDI）was 0.29±0.03.The particle size of most particles ranges 141-243 nm,and the particle size distribution was narrow.2.Physicochemical characterization of FFC-NCPhysicochemical characterization of FFC-NC prepared under the optimal preparation condition included the following tests:Scanning electron microscope inspection（SEM）,fourier-transform infrared spectroscopy（FTIR）,X-ray diffraction detection（XRD）,and differential scanning calorimetry（DSC）.The solubility and dissolution rate of FFC-NC was evaluated.SEM results showed that the morphology of FFC-NC was an irregular sphere,and the particle size distribution was uniform.FTIR results showed that the chemical structure of FFC-NC had not changed.XRD and DSC results showed that the crystal structure of FFC-NC was changed.The solubility of FFC-NC was（3.156±0.097）mg/m L,2.5 times higher compared to FFC.Results of dissolution rate test showed that the accumulated dissolution of FFC-NC reached 100.17%at 10 min,but the accumulated dissolution of FFC was 78.57%.Results of potential measurement showed that the suspension of florfenicol nanocrystals had good stability.3.Bioavailability evaluation of FFC-NCCross-over design method was used to evaluate the bioavailability of FFC and FFC-NC in hens.After administration,blood samples were collected from wing veins of hen at different time.The concentration of FFC in plasma samples were determined by high performance liquid chromatography（HPLC）.Obtained data were analyzed by DAS 2.0;the plasma concentration–time curve was plotted by Graph Pad Prism 8.0;the relevant pharmacokinetic parameters were analyzed by IBM SPSS Statistics 25.0.Results showed that the established HPLC chromatogram showed a stable baseline,and the plasma peak was completely separated from the florfenicol peak.Results of recovery rate and precision met the requirements,and the repeatability was good.This method was suitable for the determination of FFC concentration in hen plasma.Results of pharmacokinetic showed that compared with the FFC group,value of time to reach peak concentration(T_max)of FFC-NC group was（0.875±0.137）h,significantly reduced.Values of area under the concentration-time curve(AUC_（0-∞）)and peak concentration(C_max)of FFC-NC group were（23.957±2.338）μg/m L·h and（8.249±0.713）μg/m L,respectively,which were significantly improved.The relative bioavailability of FFC-NC group was 3.6 times higher compared to FFC group.The pharmacokinetic characteristics of FFC-NC are significantly improved compared with FFC.In conclusion,the particle size of the optimized FFC-NC prepared using emulsified solvent evaporation method was small,and the distribution range of FFC-NC was narrow.Results of physicochemical characterization showed that the chemical structure of FFC-NC did not change,and the crystal structure was changed.The solubility and bioavailability of the prepared nanocrystals were improved.This study breaks through the traditional preparation technology of FFC-NC,provides a new idea for the solubilization of florfenicol,and is of great significance to the clinical use of florfenicol.