【作者】 李英；

【导师】 乔海灵； 郭玉忠；

【作者基本信息】 郑州大学 ， 药理学， 2008， 硕士

【摘要】 β—内酰胺类抗生素（包括青霉素、头孢菌素类和非典型β—内酰胺类等）目前在世界抗生素市场占主导地位。头孢拉定为第一代头孢菌素类抗生素,可口服、肌注、静注及静滴,抗菌效果好,临床应用广泛,主要用于呼吸道、泌尿道、皮肤、骨等部位的感染。该药毒代动力学研究国内外尚未见报道,研究其毒代动力学,有利于探讨该药毒性作用的发生机制,指导临床合理用药。目的研究注射用头孢拉定在Beagle犬体内的毒代动力学,阐述在毒性试验条件下药物所达到的全身暴露与剂量、时间的内在联系,并结合头孢拉定组织分布的浓度,解释毒理试验数据的价值,对药物的临床前安全性进行全面和综合的评价。方法1.给药方案及标本采集健康、成年Beagle犬18只,雌雄各半,随机分为低(200 mg·kg^-1)、中(1000 mg·kg^-1)、高(1800 mg·kg^-1)三个给药剂量组,每组6只。静脉滴注头孢拉定,每日一次,共35天。给药首日,三个剂量组分别于给药前0h,给药后0.167,0.333,0.667,1,1.167,1.333,1.667,2,2.5,3,4,5,6,7h抽取后肢静脉血0.8 ml;给药第34天,三个剂量组分别于给药前0h,给药后0.333,0.667,1,1.333,2,3,5,7h抽取后肢静脉血0.8 ml。分离血清于-20℃保存待测。最后一天用药结束后立即处死动物,解剖采集脑、心、肺、肝、脾、肾、胰腺、子宫和卵巢组织。制备匀浆,取上清液,-20℃保存待测。2.药物浓度测定样品中药物浓度按抗生素微生物琼脂平板扩散法测定,检测菌为藤黄八叠球菌28001。配制0.78、1.56、3.125、6.25、12.5、25mg·L^-1的头孢拉定标准曲线溶液,以标准液抑菌圈直径与药物浓度的对数作线性回归,求出回归方程。以未知样品的抑菌圈直径带入回归方程,求得样品的药物浓度。3.方法学考察用犬空白混合血清将头孢拉定对照品配成浓度分别为1.56、6.25和25 mg·L^-1的待测液,按“标准曲线”方法测定日内、日间变异和回收率。并进行稳定性考察。结果1.方法学研究标准曲线方程为:1gC=0.0729 D-0.96,相关系数r=0.9968,线性范围为0.78～25 mg·L^-1。经精密度及回收率考察显示,RSD均小于10%。经稳定性考察显示,头孢拉定在室温放置10h,-20℃冰冻2周和冻融（-20℃）3次的情况下均稳定。2.单次用药毒代动力学Beagle犬静脉滴注（1h）头孢拉定高、中、低三个剂量组后,主要毒代动力学参数为:T_1/2分别为1.01±0.13 h,0.93±0.07 h和0.90±0.06h;Ke分别为0.69±0.08（1/h）,0.75±0.06（1/h）和0.77±0.06（1/h）;AUC₀¹分别为805.78±111.37 mg·L^-1·h,3900.38±1165.07 mg·L^-1·h和9072.59±2355.43mg·L^-1·h;AUC₀^∞分别为823.34±110.85 mg·L^-1·h,3960.79±1207.28 mg·L^-1·h和9203.31±2395.67 mg·L^-1·h;Vd分别为0.39±0.07 L,0.39±0.12 L和0.29±0.06L;CL分别为0.27±0.03 L·h^-1,0.29±0.09 L·h^-1和0.22±0.06 L·h^-1;C_max分别为415.97±99.01 mg·L^-1,2250.86±510.38 mg·L^-1和5162.32±1119.54 mg·L^-1。结果显示单次静脉滴注给药后,头孢拉定在Beagle犬体内呈线性动力学过程,体内暴露量（AUC）和峰浓度(C_max)与给药剂量均呈很好的正相关性（r=1.000）。3.多次用药毒代动力学Beagle犬静脉滴注（1h）头孢拉定高、中、低三个剂量组后,主要毒代动力学参数为:T_1/2分别为0.98±0.11 h,1.03±0.24h和0.94±0.05 h;Ke分别为0.72±0.08（1/h）,0.71±0.19（1/h）和0.74±0.04（1/h）;AUC₀¹分别为1224.92±112.13 mg·L^-1·h,6313.88±2098.97mg·L^-1·h和9322.36±2512.75mg·L^-1·h;AUC₀^∞分别为1248.05±115.36 mg·L^-1·h,6403.88±2080.21 mg·L^-1·h和9400.51±2531.69mg·L^-1·h;Vd分别为0.31±0.09 L,0.35±0.20 L和0.37±0.18L;CL分别为0.21±0.04 L·h^-1,0.22±0.09 L·h^-1和0.27±0.11 L·h^-1;C_max分别为852.60±56.21 mg·L^-1,3880.51±1160.66 mg·L^-1和6469.96±704.57 mg·L^-1。结果显示多次静脉滴注给药后,头孢拉定在Beagle犬体内呈线性动力学过程,体内暴露量（AUC）和峰浓度(C_max)与给药剂量呈很好的正相关（r=1.000）。4.多次用药组织分布浓度Beagle犬静脉滴注（1h）头孢拉定低、中、高三个剂量组后,其组织浓度分别为肾:0.482±0.164,1.563±0.278,2.538±1.159mg·g^-1;肝:0.204±0.084,0.865±0.712,1.448±1.173 mg·g^-1;肺:0.073±0.006,0.45±0.076,0.800±0.176mg·g^-1;脑:0.012±0.011,0.013±0.007,0.046±0.009 mg·g^-1;心:0.030±0.008,0.183±0.031,0.252±0.033 mg·g^-1;脾:0.032±0.005,0.130±0.008,0.249±0.038 mg·g^-1;胰腺:0.031±0.007,0.159±0.044,0.299±0.040 mg·g^-1;卵巢:0.087±0.007,0.700±0.478,0.999±0.695 mg·g^-1;子宫:0.096±0.035,0.721±0.454,1.050±0.837 mg·g^-1。结果显示,在三个剂量组中,肾的浓度最高,其次是肝和子宫、卵巢,在脑中的浓度最低。且各组织中（除脑组织外）分布浓度与剂量呈较好的正相关（r=1.000）。结论1.Beagle犬静脉滴注200 mg·kg^-1,1000 mg·kg^-1和1800 mg·kg^-1的头孢拉定后体内毒代动力学行为符合静脉滴注一房室模型。2.头孢拉定单次和多次静脉滴注给药后,在Beagle犬体内呈线性动力学过程,体内暴露量（AUC）和Cmax与给药剂量呈正相关。而t_1/2、Vd、CL与给药剂量及给药时间均无关。3.Beagle犬静脉滴注200 mg·kg^-1,1000 mg·kg^-1和1800 mg·kg^-1的头孢拉定后在肾中的浓度最高,其次是肝脏和子宫、卵巢,在脑中的浓度最低。且各组织中（除脑组织外）分布浓度与剂量呈较好的正相关（r=1.000）。更多还原

【Abstract】 Beta-lactam antibiotics（including penicilin,cephalosporins,atypical beta-lactam antibiotics,etc.） play a significant role in world-wide application of antibiotics.As the first generation cephalosporins,cefradine can administered by oral,intramuscular injection,or intravenous injection and intervenous drop infusion with satisfactory effectiveness.Cefradine was extensively used in infection of respiratory system, urinary system,skin and bone,etc.No toxicokinetics study was reported at domestic or abroad about this drug.It is helpful to discuss the toxic action of cefradine to instruct clinical rational administration.Objective Expound the relationship between exposure and dose under venenous test conditions by research toxicokinetics of cefradine injection in beagles. Expound the value of venenous test data from cefradine’s concentration in tissue, quibus evaluating preclinical safety.Methods1.Project of administration and specimen collection 18 healthy and ripe beagles,9 male and 9 female,were divided into 3 groups randomly,using dose of 200 mg·kg^-1（high）,1000 mg·kg^-1（middle） and 1800 mg·kg^-1（low）.Each group has 6 beagles,intervenous drop infusion of cefradine once daily totally 35 days.First day, each group was taken suction for 0.8ml of hind legs venous blood at the point of 0h before administration,0.167h,0.333h,0.667h,1h,1.167h,1.333h,1.667h,2h,2.5h, 3h,4h,5h,6h,7h after administration.34^th day each group was taken suction for 0.8ml of hind legs venous blood at the point of 0h before administration,0.333h, 0.667h,1h,1.333h,2h,3h,5h,7h after administration.All blood serum were conserved in -20℃before test.Last day sacrifice all the beagles and collect the brains, the hearts,the lungs,the livers,the spleens,the kidneys,the pancreases,the uteruses and the ovarys.Make homogenate and get supernatant,conserving in -20℃for being tested.2.Determine the drug concentration The exemplar drug concentration was determined by antibiotics microbial agar diffusion method in which sarcina lutea was used.Microbiological method was used to determine drug concentration in blood serum.To prepare 0.78,1.56,3.125,6.25,12.5,25 mg·L^-1 standard curve of cefradine solution,we extract the regression equation by linear regression of inhibition zone’s diameter of standard curve and drug concentration’s logarithm.The drug concentration was calculated accord to inhibition zone’s diameter of unknown exemplar by regression equation.3.Technology study To prepare the test solution of 1.56,6.25 and 25 mg·L^-1 with cefradine reference substance by beagles blank pooled serum.To determine the intra-day and Inter-day deviation and recovery by standard curve and perform stability study.Results1.Technology research Standard curve equation:1gC=0.0729D-0.96, r=0.9968,linear range:0.78～25mg/L.Investigated by degree of precision and recovery rate,we get RSD＜10%.Put cefradine under room temperature for 10h, -20℃freezing for 2 weeks and freeze thawing（-20℃） for three times,results suggest the cefradine are stabilized substance.2.Toxicokinetics of administration of single Toxicokinetics indexes of 3 groups（high,middle,low） after given cefradine intervenous drop infusion（1h） were calculated.The T_1/2 was 1.01±0.13h,0.93±0.07h,0.90±0.06h.The Ke was 0.69±0.08（1/h）,0.75±0.06（1/h）,0.77±0.06（1/h）.The AUC₀¹ was 805.78±111.37 mg·L^-1·h,3900.38±1165.07 mg·L^-1·h,9072.59±2355.43mg·L^-1·h.The AUC₀^∞was 823.34±110.85 mg·L^-1·h,3960.79±1207.28mg·L^-1·h,9203.31±2395.67 mg·L^-1·h.The Vd was 0.39±0.07L,0.39±0.12L,0.29±0.06L.The CL was 0.27±0.03 L·h^-1, 0.29±0.09 L·h^-1,0.22±0.06 L·h^-1.The C_max was 415.97±99.01 mg·L^-1, 2250.86±510.38 mg·L^-1,5162.32±1119.54 mg·L^-1.From the result of administration once,the cefradine act as linear dynamics process in beagles.The AUC and C_max act as positive correlation with dose.3.Toxicokinetics of administration of multiple Toxicokinetics indexes of 3 groups（high,middle,low） after given cefradine intervenous drop infusion（1h） were calculated.The T_1/2 was 0.98±0.11h,1.03±0.24h,0.94±0.05h.The Ke was 0.72±0.08 （1/h）,0.71±0.19（1/h）,0.74±0.04（1/h）.The AUC₀¹ was 1224.92±112.13 mg·L^-1·h, 6313.88±2098.97 mg·L^-1·h,9322.36±2512.75 mg·L^-1·h.The AUC₀^∞was 1248.05±115.36 mg·L^-1·h,6403.88±2080.21mg·L^-1·h,9400.51±2531.69mg·L^-1·h.The Vd was 0.31±0.09L,0.35±0.20L,0.37±0.18L.The CL was 0.21±0.04 L·h^-1, 0.22±0.09 L·h^-1,0.27±0.11 L·h^-1.The C_max was 852.60±56.21 mg·L^-1, 3880.51±1160.66 mg·L^-1,6469.69±704.57 mg·L^-1.From the result of administration for multiple,the cefradine act as linear dynamics process in beagles.The AUC and C_max act as positive correlation with dose.4.Drug distribution in tissue Drug concentration was measured in tissue from 3 groups of beagles（high,middle,low） which were given intervenous drop infusion of cefradine（1h）.The concentration in kidney was 0.482±0.164 mg·g^-1, 1.563±0.278 mg·g^-1,2.5384±1.159 mg·g^-1.The concentration in liver was 0.204±0.084 mg·g^-1,0.865±0.712 mg·g^-1,1.448±1.173 mg·g^-1.The concentrationin lungs was 0.073±0.006 mg·g^-1,0.45±0.076 mg·g^-1,0.800±0.176 mg·g^-1.The concentration in brain was 0.012±0.011 mg·g^-1,0.012±0.007 mg·g^-1,0.046±0.009 mg·g^-1.The concentration in heart was 0.030±0.008 mg·g^-1,0.183±0.031 mg·g^-1,0.252±0.033 mg·g^-1.The concentration in spleen was 0.032±0.005 mg·g^-1,0.130±0.008 mg·g^-1, 0.249±0.038 mg·g^-1.The concentration in pancreas was 0.031±0.007 mg·g^-1, 0.159±0.044 mg·g^-1,0.2994±0.040 mg·g^-1.The concentration in ovary was 0.087±0.007 mg·g^-1,0.700±0.478 mg·g^-1,0.999±0.695 mg·g^-1.The concentration in uterus was 0.096±0.035 mg·g^-1,0.721±0.454 mg·g^-1,1.050±0.837 mg·g^-1.The result shows that the concentration in tissue ranks from high to low is kidney, liver,uterus,ovary,brain.Conclusion1.According to result of cefradine intervenous drop infusion to beagles with dose of 200 mg·kg^-1,1000 mg·kg^-1 and 1800 mg·kg^-1,the behavior of toxicokinetics acts as one compartment model. 2.No matter intervenous drop infusion of cefradin for once or for multiple,the cefradine act as linear dynamics process in beagles.The AUC and C_max act as positive correlation with dose.But t_1/2,Vd,CL have no relevance with dose and administration time.3.After intervenous drop infusion of cefradine to beagles with dose of 200 mg·kg^-1, 1000 mg·kg^-1 and 1800 mg·kg^-1,the maximal concentration appears in kidney, secondly in liver,uterus and ovary.The minimum concentration is in the brain.更多还原