馬兜鈴酸在家兔體內之藥物動力學與腎毒性研究
Pharmacokinetics and nephrotoxicity of aristolochic acid in rabbits
中文摘要 馬兜鈴酸(AA)是馬兜鈴科(Aristolochiaceae)植物的主成分，為馬兜鈴酸Ⅰ(AAⅠ) 及馬兜鈴Ⅱ(AAⅡ)所構成的混合物，研究指出 AA 具有腎毒性及腫瘤毒性，但對 於 AA 在藥物動力學方面的研究報告則是相當有限。本研究採用高效液相層析法 (HPLC)作為分析動物血漿檢品中 AAⅠ、AAⅡ含量測定的工具，且分析法之確 效試驗結果均符合生物檢品分析準則。 本研究分別以單次靜脈投予不同劑量的 AANa (0.25, 0.5, 1.0, 2.0 mg/kg)至四組家 兔體內進行藥物動力學研究(每組均使用 6 隻家兔)，由於偵測極限的關係所以投 予 0.25 mg/kg AANa 所得到的結果是以一室模式作配適，其他劑量則均使用二室 模式作配適；除了 0.25mg/kg 的劑量之外，在 0.5、1.0、2.0 mg/kg 的劑量範圍內 得到 AAⅠ與 AAⅡ的藥物動力學參數無統計上差異(P>0.05)，AAⅠ清除率依序 為 349.27 112.92、236.99 48.92、228.47 31.04、226.99 19.42 mL/h/kg， AAⅡ清除率依序為 224.20 34.01、199.58 41.93、156.76 30.30、185.42 29.81 mL/h/kg，其給藥劑量(Dose)與藥物血中濃度曲線下面積(AUC)呈線性關 係(AAⅠ: Y= 4754.23X-188.20, r2=0.980, P<0.001; AAⅡ: Y= 5728.56X-21.57, r2=0.911, P<0.001)。 另外使用六隻家兔，以多次靜脈注射不同劑量(0.5、1.0、2.0 mg/kg) 的 AANa 到 此六隻家兔體內，在這組研究中的每隻家兔每次給藥至少相隔七日，藥物動力學 參數在靜脈給予 0.5~2.0 mg/kg 之 AANa 的範圍內會改變，AAⅠ的清除率依序 為 236.99 48.92、 198.37 55.09、102.10 22.39 mL/h/kg，AAⅡ的清除率 依序為 199.58 41.93、97.63 23.22、42.31 7.64 mL/h/kg；其給藥劑量與曲 線下面積得非線性藥物動力學的特徵(AAⅠ:Y=13618.09X2-875.43X + 469.52, r2=0.908, P<0.001; AAⅡ: Y=26583.73X2 - 3779.03X + 489.38, r2=0.948, P<0.001 )。非線性藥物動力學結果可能與 AAⅠ、AAⅡ所造成的腎損傷導致清除 率改變有關。 在口服投予的研究則口服給予 1.0 mg/kg 之 AANa 至家兔體內，AA 在胃酸中不 安定，會受到胃酸的強酸性而降解導致血漿中 AAⅠ與 AAⅡ的含量低，吸收情 形沒有規則性，且個體間差異大，生體可用率不好。 家兔在投予 AANa 後的第一日及第八日進行犧牲取其腎臟進行組織型態學研 究，單次靜脈投予 AANa 得到的組織檢查如下所述：投予 0.25 mg/kg 後的第一 日腎的型態學並沒有顯著的改變，在 0.5、1.0、2.0 mg/kg 的劑量之下則是觀察到 輕微的細胞浸潤、腎小管上皮細胞變性萎縮的情形；在第八日觀察 0.25 mg/kg 的劑量下的損傷則是輕微的浸潤及腎小管上皮細胞萎縮情形，在 0.5、1.0、2.0 mg/kg 的劑量下則是發現有中度至嚴重的細胞間質浸潤、上皮細胞變性萎縮、間質纖維化及透明圓柱的沉積。口服投予 1.0 mg/kg 之 AANa 後，也於第一日及第 八日取其腎臟進行組織型態學研究，口服投藥後的第一日組織形態學並無顯著變 化，但到了第八日則觀察到輕微的上皮細胞變性萎縮及間質纖維化。根據組織形 態研究結果得知腎臟損傷的程度會隨單次投藥劑量的增加而增加，且 AA 所造成 的腎損傷有不斷在惡化的情形，但在腎絲球方面則未見損傷。 英文摘要
Aristolochic acid (AA) is the main component of Aristolochia plants. It is a mixture of aristolochic acid Ⅰ(AAⅠ) and aristolochic acid Ⅱ(AAⅡ). It has been proved by several researches that AA has nephrotoxicity and tumor toxicity. However, the pharmacokinetics of AA is still limited. A validated high performance liquid
chromatography (HPLC) method was used to determine the plasma concentration of AAⅠ and AAⅡ. The HPLC method used was met the guidance of bioanalytical method validation.
The pharmacokinetics of AAⅠ and AAⅡ was studied by respectively intravenous administration of four different doses(0.25, 0.5, 1.0, 2.0 mg/kg ) of aristolochic acid sodium salt (AANa) in four groups of rabbits (n=6 for each group). Because of detection limit, the results of AAⅠ and AAⅡ from 0.25 mg/kg AANa were fit with one compartment model. Others were fit with two compartment model. There was no significant difference (P>0.05) in pharmacokinetic (PK) parameters of AAⅠ and AA Ⅱ, except 0.25 mg/kg AANa-treated group. The system clearance of AAⅠ were 349.27 112.92, 236.99 48.92, 228.47 31.04, 226.99 19.42 mL/h/kg. Clearance for AAⅡ were 224.20 34.01, 199.58 41.93, 156.76 30.30, 185.42 29.81 mL/h/kg. The area under the curves(AUC) of AAⅠ and AAⅡ were calculated from zero to infinite. The AUCs of AAⅠ and AAⅡ were proportional to the dose administrated (AAⅠ: Y= 4754.23X-188.20, r2=0.980, P<0.001; AAⅡ: Y= 5728.56X-21.57, r2=0.911, P<0.001). It indicated that AAⅠ and AAⅡ might behave dose-independent pharmacokinetics between 0.25~2.0mg/kg of AANa IV injection.
In another study, six rabbits were intravenous administered with different doses (0.5, 1.0, 2.0 mg/kg) of AANa. All animal of this study were treated with AANa with 7 days interval. The PK parameters of AAⅠ and AAⅡ were changed between 0.5 ~ 2.0 mg/kg of AANa IV injection. The clearance of AAⅠ were 236.99 48.92, 198.37 55.09, 102.10 22.39 mL/h/kg. The clearance of AAⅡ were 199.58 41.93, 97.63 23.22, 42.31 7.64 mL/h/kg. The AUCs of AAⅠ and AAⅡ were not proportional to the increasing doses. The relationship of non-linear
pharmacokinetic properties between dose and AUC were obtained (AAⅠ:
3779.03X + 489.38, r2 = 0.948, P< 0.001 ). The non-linear pharmacokinetics might due to renal lesions caused by AAⅠ and AAⅡ.
After oral administration of AANa 1.0 mg/kg, the gastric acid would destroy AA Ⅰ and AAⅡ. The amount of AAⅠ and AAⅡ detected in plasma were very low. And the absorption of AAⅠand AAⅡ was not regularity and in high variation.
After treated with AANa, rabbits were sacrificed on day-1 and day-8 to obtain the kidney sections. The histological examination of intravenous administration of AANa was described as the following. On day-1, there was no significant alteration in renal morphology on 0.25 mg/kg AANa-treated group. Mild interstitial infiltration,
epithelial cell degeneration and atrophy were observed on 0.5, 1.0, and 2.0 mg/kg AANa-treated group. On day-8, mild degeneration of epithelium and atrophy were observed on 0.25 mg/kg AANa-treated group. Moderate to severe renal lesions including, interstitial infiltration, degeneration and atrophy of epithelium, interstitial fibrosis and hyaline cylinders were found on 0.5, 1.0 and 2.0 mg/kg group. Kidney sections were also obtained on day-1 and day-8 after oral treated with 1.0 mg/kg AANa to rabbits. In oral study, there was no significant change in renal morphology on day-1. But mild degeneration and atrophy of epithelium and interstitial fibrosis were observed on day-8. According to the histological examination, the extent of renal lesions increased with doses increasing was elucidated. The renal lesions caused by AA were continued to deteriorate. However, lesions in glomerulus were not observed.
