Dissertation > Medicine, health > Pharmacy > Pharmacology

Studies on Pharmacy and Pharmacokinetics of Compound Butafosfan

Author DuXiaoYan
Tutor JiangJinShu;ZhuZuoLei
School China Agricultural University
Course Preventive Veterinary Medicine
Keywords compound butafosfan HPLC-RID LC-MS pharmacokinetics horse serum
Type PhD thesis
Year 2004
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Compound butafosfan injection (trade name: Catosal) is an aqueous solution. Both of butafosfan and vitamin B12 are the main ingredients of the compound. Butafosfan is an organic phosphorus compound and is active in stimulating metabolism and modulating the immune system. The preparation has been used as metabolic stimulant and tonic in a wide variety of species. The indications are metabolic disorder through malnutrition or faulty husbandry, developmental and nutritional disorder in young animals, metaphylaxis of sterility and puerperal diseases, tetany and pareses as adjunct to calcium and magnesium therapy and as effctive tonic in cases of over-exertion and exhaustion. In order to develop this drug in our country, our research group had synthesized butafosfan and developed two preparations. In our studies, the injection, one of the two preparations and its pharmacokinetics has been studied.For the determination of butafosfan, electronic diluting method and HPLC assay after derivatization with dansyl chloride (Dns-Cl) were developed. The butafosfan was diluted by tetrabutylammoniumoidide (0.1 mol/L). Electronic diluting method was simple and convenient and the C.V.% of intra-day and inter-day was 0.8-2.3% and 2.2%, respectively. The Dns-Cl derivatized butafosfan was identified with HPLC, TLC and MS and the reaction conditions were optimized. The results show that butafosfan was derivatied by Dns-Cl successfully and at pH9.8, the reaction between butafosfan and Dns-Cl acetonitrile solution can be finished after 60 min.The products can be seperated on an inertsil ODS column, acetonitrile-water (1.6:l,v/v) as mobile phase with ultraviolet detection at 254 nm. The peak area of derivative and concentration of butafosfan has good linear (r=0.9988) between 0.2mg/mL~2.0mg/mL. A TLC assay was used in this study to identify impurity in butafosfan.A kind of compound butafosfan injection was prepared based on the formula of the Bayer company after certain changes. For the determination of both injectable butafosfan and vitamin B12, HPLC-RID and HPLC-UV assay were developed, respectively. Thermal stability and the light stability test were undergone. At the end of the thermal stability test, the remains of butafosfan and vitamin B12 were 96.58%and 91.16%, respectively.The experimental results for stability study of the compound showed that our injection was do stable to heat and humidity during the testing period of 6 months, but unstable to strong light. The injection should be stored avoiding the direct sunshine.A specific and rapid LC-MS method to quantitate butafosfan concentration in horse serum was developed. Methanol was applied to extract butafosfan from lmL aliquots of serum and deposit proteins. The extraction was cleaned-up by selected C18 SPE column after evaporation and added 1.5mL buffer (pH3.5). The elute was dried and dissolved with water and injected onto LC column, which was eluted using a methanol/acetic acid gradient (pH3.0) over 10 min. Butafosfan was detected by a mass spectrometer operated in the positive single ion monitoring mode using ESI as an ionization process at m/z 114. The assay is linear between 0.02 and 10ug/mL and the determination limit of the assay was 0.01ug/mL calculated to be five times the baseline noise. The recoveries of the analyte in serum at the fortified level of 0.1, 0.5, 2.0 and 10 ug/mL ranged from 94.51%~ 114.23%. The coefficients of variation are less than 8.23%.The pharmacokinetics of the compound butafosfan injection administrated intravenously at dose rate of 5 mg/kg bw and 2.5 mg/kg bw to 6 horses was studied (3 horses per dose). Serum samples were taken at 1,6,12,30 min and 1,2,4,8,16,24,32h and determined by the LC-MS assay. The butafosfan concentrations in serum were calculated by the software 3P97. The results suggested that the kinetics of the injection was fitted to two compartments. Serum levels of butafosfan declined rapidly from the mean peak levels of 87.811ug/mL for high dose group and 10.224 ug/mL for the low dose group at the first sampling time (1

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