| Danofloxacin mesylate (DFM) is a fluoroquinolone antimicrobial drug, which is against a broad range of pathogens responsible for several disease syndromes of livestock.The spectrum of antimicrobial activity of DFM includes most Gram-negative bacteria, some Gram-positive bacteria, intracellular pathogens and mycoplasmas; but it has poor activity against anaerobes. We could only get soluble powder for oral administration or intramuscular administration of DFM, which with short elimination half-life in cattle, goats, sheep and pigs after intramuscular administration of DFM. The study aim was to develop a controlled-release and lung-targeting dosage form of DFM, which will be used for intramuscular or intravenous administration with a longer elimination half-life and stronger lung-targeting.In order to prepare DFM-GMS successfully, blank GMS was prepared at first. Gelatin was chosen as the carrier of microspheres because of its characteristics of biodegradationable, cheap and low toxicity, and the GMS was prepared using the method of chemical-crosslinking. Certain factors were considered when GMS was prepared: temperature of emulsifying procedure, stirring velocity, amount of emulsifying agent, gelatin concentration in aqueous solution, volume ratio of the W/O and the amount of crosslinking agent. The single factor test proved that four reaction conditions had crucial roles:stirring velocity; gelatin concentration in aqueous solution; amount of emulsifying agent and amount of crosslinking agent. An orthogonal test of L9(34) was conducted to optimize reaction conditions.In DFM-GMS preparing, microspheres with different drug content were obtained when various weight ratios of DFM/gelatin were tested, but drug content was low. Adjuvants were considered during DFM-GMS preparation to improve the affinity between drug and gelatin. PEG 6000 was selected because it could increase drug content of the microspheres and had little effect on microsphere characteristics. DFM-GMS prepared by optimized technology was regular microspheres, and the mean diameter was 10.25±0.69μm, drug loading was 33.30±8.12mg/g, encapsulation efficiency was 92.15±1.69%. DFM-GMS showed no change in appearance, drug content and encapsulation efficiency after acceleration test and strong light exposure test.In vitro release of DFM-GMS, the characteristics of DFM-GMS and DFM were studied. The release medium was pH7.4 phosphate buffer solution(PBS), the release profile of DFM-GMS fitted best to Higuchie quation, and DFM fitted best to zero-order release equation. It showed that there were significant differences between DFM-GMS and DFM.In the study of pharmacokinetics of DFM-GMS in pigs, the correlation of calibration curve was good, and the correlation coefficient was 0.9996. The average extraction recovery was more than 75% of DFM from plasma. The intra-day coefficient of variations were less than 5%, and the inter-day coefficient of variations were less than 10%. Pharmacokinetics of DFM-GMS was studied in pigs with conventional injection as the reference preparation. After intramuscular injection of DFM-GMS and DFM with a single dosage of 2.5 mg DFM/kg body weight in the DFM-GMS group and DFM group, respectively. The results revealed that the pharmacokinetic characteristics of DFM of DFM group in pig manifested rapid absorption and rapid elimination. However, the pharmacokinetic characteristics of DFM of DFM-GMS group in pig manifested slow absorption and slow elimination. The plasma concentration-time data of DFM-GMS and conventional injection all conformed to two-compartment open models.The absorption half-lives(T1/2ka) of DFM-GMS and DFM were 0.91±0.12 and 0.15±0.01h, respectively. The peak concentration(Cp) of DFM-GMS and DFM were 0.39±0.02μg/mL and 0.45±0.09μg/mL, respectively. The peak time(Tp) of DFM-GMS and DFM were 2.62±0.11 and 0.64±0.13h, respectively. The elimination half-lives(T1/2β) of DFM-GMS and DFM were 24.32±1.61 and 6.61±0.74h, respectively. The mean residence time(MRT) of DFM-GMS and DFM were computed as 31.54±0.69 and 10.01±0.85h, respectively. The areas under the concentration-time curve(AUC) of DFM-GMS and DFM were 9.97±0.81 and 3.34±0.43μg·h/mL, respectively. Statistical analysis showed that these follow parameters were significant differences between the DFM-GMS and DFM(P< 0.01):t1/2ka, Tp, t1/2β, MRT and AUC.In the study of DFM-GMS disposition of plasma and lung in rats, the correlation of calibration curve of DFM in plasma and lung were good, and the correlation coefficient were more than 0.9990. The average extraction recovery was more than 85%of DFM from plasma and more than 75%from lung. The intra-day coefficient of variations were less than 5%, and the inter-day coefficient of variations were less than 10%. With conventional injection as the reference preparation, plasma and lung concentration of DFM were gotten following single intravenous injection of DFM-GMS with a single dosage of 2.5 mg DFM/kg body weight in rats. The results revealed that DFM-GMS could increase the amout of DFM in lung, and showed the characteristic of lung-targeting.
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