人工骨水泥中Aztreonam的釋出---體外及體內的研究

Release of Aztreonam from bone cement---in vitro and in vivo study

周心怡(Hsing-Yi Chou)

指導教授 ： 何意

臺北醫學大學/藥學院/藥學研究所/碩士(2005年)

摘要

摘 要 當病菌引發嚴重骨關節感染，而導致骨組織破壞時，須進行重建手術治療受感染的骨頭，必要時則需置換人工關節，在台灣每年已有超過6,000人，接受關節重建手術，其中以髖關節置換手術最常見，手術之成功與否以及感染的治療，對病患日後之生活品質影響甚鉅。以抗生素混合骨水泥應用於關節重建手術治療感染的給藥方法已行之有年，研究證實此投藥方式具有延長藥效的優點，二階段式的關節重建手術更能有效治療深度的骨感染，感染的治癒率高達82~96%，且含藥骨水泥兼具支持作用；然而，以含藥骨水泥粒的給藥方式迄今並未完整地被評估其抗菌效能及探討藥物釋出機制。 氮烯內醯胺(Aztreonam; AZ)，屬於monobactam抗菌劑，可有效治療由革蘭氏陰性菌(Gram-negative)需氧菌(Aerobes)所引起的各種感染症，投藥後衍生之副作用少而且過敏性低，可作為治療革蘭氏陰性菌引起感染症的首選藥物。本實驗之目的即在於探討氮烯內醯胺自人工骨水泥釋放-體外與體內的釋離情形研究。體外實驗部分係將AZ混入骨水泥製成含藥骨水泥粒，分別製作不同厚度、表面積以及處方之骨水泥粒，比較AZ釋離情形之差異，建立一完整之溶離模式，並評估AZ於骨水泥中的釋出機制。體內實驗部分欲開發一HPLC分析方法，可應用於檢測臨床髖關節置換手術後患者滑囊液中AZ濃度，並討論AZ自髖關節人工骨水泥粒釋出的藥物動力學特性，連結AZ體外的溶離機制與體內釋出的動態，並對於人工骨水泥的給藥模式提供預測以及建議。 以臨床所使用之藥物比例製作含藥實心骨水泥粒，溶離試驗結果顯示，AZ釋出量小於 5 mg，僅佔原骨水泥粒中藥物總量之3.6%，藥物釋出總量少；而不同包覆厚度之含藥骨水泥粒，AZ釋出總量相近，C.V.小於1%，利用統計學上之ANOVA檢定，亦無統計學上差異 (p>0.05)，因此推論AZ僅由表面釋出。不同表面積之含藥骨水泥粒溶離試驗結果顯示，隨著表面積加大，AZ釋出量即增加(R2> 0.9)，更可確定AZ之釋放機制係自骨水泥粒表面釋出。骨水泥之含藥比例增加，AZ的釋出總量呈兩段式線性關係，當含藥比例超過 4.30%，可釋出藥量明顯加大，釋放機制產生改變，對照掃瞄式電子顯微鏡之觀察結果，推論藥物可經骨水泥間孔隙釋出，因而增加釋離量。體外溶離數據之配適結果以Korsmeyer-Peppas model 經過修正後之配適公式配適性最佳，故選用其作為AZ自人工骨水泥中釋出的溶離配適模式，以配適所得之動力學常數以及擴散指數作為判斷藥物釋出情形的指標。將不同厚度的骨水泥粒之溶離結果作曲線配適，得到的擴散指數以及動力學常數相近，顯示厚度不影響藥物釋放；在酸鹼值為5.34及7.4之的溶離媒液中，動力學常數相近，惟AZ於其中之降解速率不同，造成溶離之差異。將骨粒之表面積與動力學常數作圖，可得一線性關係(R2=0.9924)；將含藥比例與溶離速率常數作圖，可得兩段式線性關係。 體內研究方面，檢品中AZ的測定係採用HPLC分析方式，選用逆相層析管柱，配合紫外光偵測，波長設定在310 nm。檢量線濃度範圍介於 0.025 ~ 25 μg/ml 間，線性良好，回內與回間分析之變異係數皆小於7 %，且平均回收率為80.76 %，顯示此分析方法是良好的AZ生物檢品檢測方法。此外，本研究亦探討AZ混合骨水泥所製成之人工關節，植入體內之後AZ的濃度變化，上述HPLC分析方法亦成功應用於檢測關節重建手術後病人之關節滑囊液檢品中AZ之濃度，由分析得知在病患住院期間AZ自髖關節部位的釋出速率常數0.2897±0.0028 (day-1)；而病患出院後至接受第二次手術之間的藥物平均輸移速率常數為0.0370±0.0027(day-1)，與靜脈注射給藥之後輸移速率常數1.60±0.57 (hr-1)相比較，顯示藥物自關節腔到中央室之輸移速率緩慢，不會有快速排除的現象，因此能夠於關節腔中維持高濃度狀態，有利於抗感染治療。又根據病人體內滑囊液中的濃度以及輸移速率常數k21，計算得知在置入手術後於病人體內維持最小抑菌濃度至少44天，占兩次手術間隔時間之42.62%，應可有效治癒感染，顯示此一給藥方式的確具有良好之治療效果。 總結以上體外試驗結果，得知AZ可自骨粒表面釋放，增加含藥骨粒的表面積或者是增加含藥比例皆有助於藥物釋出；於病人滑囊液之分析結果，則證實了含AZ骨水泥裝置應可長期且有效抗感染。但是對於此劑型應該略作改良，以含藥的薄層骨水泥，具有同樣的治療效果，使用加大表面積的植入裝置更可增加藥物之釋出量，既可達到抗菌之效果，並減少藥粉的浪費，最符合經濟效益。

並列摘要

Abstract The arthroplasty or revision surgery was used to treat osteomyelitis and infected bone destruction. Every year in Taiwan, more than 6000 patients receipted the implantation surgeries and the hip arthroplasty is mostly performed of all. Using antibiotics-loaded bone cement was against the infection in the arthroplasty. As for the higher hip score and the shorter hospital-stay treatments, the two-stage revision provided good efficacy. Aztreonam (AZ), a monocyclic β-lactam antibiotic, can be used against infections caused by gram-negative and aerobic bacterial. After administration of AZ, there were little adverse effects and low percentage of allergy. Nevertheless, the mechanisms of AZ releasing from bone cement and its clinical anti-bacterial efficacy were not well established. The aim of this study was to realize the mechanisms of AZ releasing from bone cement in vitro and pharmacokinetics of AZ in vivo. Compared the release profiles of AZ from AZ-loaded cement spheres between the coating thickness, surface area and formulation of the spheres. A HPLC method was developed to determine the AZ concentrations in human synovial fluid after the hip arthroplasty. Investigation AZ released from bone cement in vivo and estimated pharmacokinetics of AZ at hip. With the same formulation in clinical surgeries, the amounts of drug released from AZ-loaded solid cement spheres were only 3.6% of the total amounts of antibiotic incorporated into the cement spheres. After coated with different thickness of the cement spheres, the release-profiles of AZ in vitro were similar. With the extensive surface area of AZ-loaded cement spheres, the total amounts of AZ released from cement spheres increased linearly (r2=0.9692). It demonstrated AZ released from the surface of cement spheres. Adding the coating thickness of AZ-loaded cement spheres did not increase drug released. The spacer should be modified to the thin coating and the extensive area of AZ-loaded cement spacer with the efficiency and economy. Increasing the ratio of the drug in AZ-loaded cement spheres from 3.2 to 4.3 percent, the total release percentage increased linearly (Y = 0.286 X + 2.38, r2 = 0.9432). When the ratio was over 4.3 percent, the total percentage released significantly increased with the ratio of the drug (Y = 1.342 X – 2.30, r2=0.9793). Compared with the SEM results, the more loaded drugs and cracks on the surface of cement spheres, the more amounts of drug were released. The concentrations of AZ in biological fluid were determined by an accuracy and simple HPLC method consisted of a reversed column with UV detection at 310 nm. The standard curve of plasma samples showed good linearity in the concentration range of 0.025 ~ 25 μg/ml. The coefficients of variance and the relative errors were less than 7%. The average recovery of AZ was 80.76%. The HPLC method had been successfully applied to determine the concentration of AZ in human synovial fluid after the first stage revision hip arthroplasty with an AZ-loaded cement spacer. The average elimination rate constant from hip after surgery was 0.2897±0.0028 (day-1) at hostipial stay. The average distribution rate constant k21 was 0.0107 ± 0.0027 (day-1) until the second revision surgery. Because of low elimination rate at hip, it remained higher concentration of AZ than the MIC90. The durations of AZ concentration above the MIC90 were for months and covered more than 40% of the dosage interval in two-stage therapy (t>MIC90 > 40% dosage interval). AZ-loaded cement prosthesis could maintain AZ concentration above the MIC longer and provide anti-infection effect. In summary, with extensive surface area and modified formulation, AZ-loaded cement spheres could increase the drug release. The concentration of AZ was more than the MIC of pathogens in vivo. The spacer would provide sufficient local concentration of AZ to maintain the bactericidal action for months.

並列關鍵字

Aztreonam, HPLC, synovial fluid

參考文獻

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