Development And Quality Research Of Arprinocid Solution

Arprinocid was the only nucleoside species of anticoccidial drugs, with no cross-resistance for other coccidiostats, and having good control effect to a variety of chicken coccidiosis. This study established an HPLC method for the determination of the arprinocid and developed new formulations of arprinocid by using orthogonal optimization test,and established an aother HPLC method to determine the solution,in addition, conducted its stability by stress testing, accelerated test and long-term of test.And microbial limits test was conducted. Details were as follows:1. Determination of arprinocid by HPLCTo establish an HPLC method for the determination of the arprinocid, the Unitary C18 column(4.6 mmx250 mm,5.0 μm) was used, and the mobile phase was composed of ultrapure water-acetonitrice(40:60), with a flow rate of 1.0 mL/min.The detection was conducted at 261 nm, and the column temperature was maintained at 30℃. The linear range of the calibration curve for arprinocid was 50-100 μg/mL(R=0.9998, n=5). The precision RSD is 0.66%, and the 90 d stability RSD is 1.85%. Content of three batches of the arprinocid drug is 99.10-99.89%.The method was convenient, sensitive, accurate, and reliable for the determination of arprinocid.2.Preparation and determination of arprinocid solutionDevelop new formulations of arprinocid, filterred out the best recipes of arprinocid solution,according to the single-factor test and orthogonal design method. The solution of the best combination of the factors was 40% solvent and 50% co-solvent and 0.2% antioxidant, and pH value was approximately 3.0. And To establish a mothod for determination of arprinocid solution by using high performance liquid chromatography(HPLC),with the proportion of acetonitrice, (0.1% phosphoric and 0.1% triethanolamine)and water was 21:79 by the mobile phase.The results showed that the linear range of the calibration curve for arprinocid was 50-100μg/mL (R=0.9993,n=5) and the RSD of precision and stability were 0.4% and 0.61%,and the average recovery rate was 98.56%(RSD=0.53%), and its signal content was 100.2%. The prescription was reasonable and the preparation technology was reliable. The HPLC method has high accuracy and sensitivity and can be used for arprinocid solution test.3. Stability of arprinocid solutionInorder to investigate the stability of prepared arprinocid solution, the factors affecting test (heat, light, temperature, freezing), accelerated test and long-term test were conducted. Stress testing results showed that the content, clarity and pH of the samples were not changed significantly at high temperatures, strong light, temperature, freeze-thaw conditions, but the samples darked. In bright light conditions, content changed volatilely in the acceptable range under freezing.The results indicated that its stability was better in light and low temperature, but a little sensitive to high temperature and freezing. So high temperature and frozen stimulation should to be avoid during storage. Prepared in 40 ± 2℃ conditions for 3 months, the samples showed characteristics, clarity, pH, content and the relevant substances did not change significantly. Long-term test results showed that characteristics, clarity, pH, content and related substances did not change significantly, indicating better stability of arprinocid solution. In summary, the stability of prepared arprinocid solution was good.4. Microbial limit test of arprinocid solutionBased on Chinese Veterinary Pharmacopoeia(2010),microbial limit test of arprinocid solution was conducted. Pre-microbial limits test-antibacterial activity test results showed that bacteria recovery of diluent control group were not less than 70%, but in the test group,the recovery of Staphylococcus aureus and Bacillus subtilis test group were less than 70%(32.9% and 55.4%). Using membrane filtration method, the recovery of each experimental group was not less than 70%, and the counting results showed that there were 2 cfu bacterial and 3 cfu mold and yeast in the arprinocid solution.Escherichia coli inspection verify method showed that experimental groups and the control groups detected positive and negative bacteria were not detected, indicating that the method is specific good. Control bacteria test results showed that detected in E. coli unexist. So, counting of bacteria and,mold and yeast test and control bacteria test showed that the microbial control of the solution met the limits on Pharmacopoeia.