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高效液相色譜法測定生物樣本中頭孢噻肟和舒巴坦的濃度
作者:顧宜,王曉娟,王榮,白娟娟,高蘇莉【關(guān)鍵詞】 頭孢噻肟;舒巴克坦;色譜法,高壓液相;血漿;尿
【Abstract】 AIM: To establish a determination method of concentrations in human biologic sample following intravenous administration of cefotaxime and sulbactam. METHODS: A RPHPLC method was developed and validated to determine cefotaxime and sulbactam in plasma and urine. The mobile phase of cefotaxime and sulbactam consisted of 1 g/L triethylamine (regulated with phosphate acid to pH=6.2) and methanol in a 68∶32 and 84∶16 (v/v) respectively and pumped at a flow rate of 1 mL/min. The plasma samples were deproteinated by addition of methanol, the urine samples were diluted with super pure water, then 20 μL of supernatant was injected onto a C18 column and monitored with UV detector at 254 nm and 220 nm respectively. RESULTS: The assay of cefotaxime in human plasma and urine were linear over the range of 5.3-530.0 mg/L and 5.45-545.00 mg/L respectively, with the detection limit of 1 mg/L and 0.25 mg/L respectively. The assay of sulbactam in human plasma and urine were linear over the range of 1.04-208.00 mg/L, with the detection limit of 0.50 mg/L and 0.25 mg/L respectively. The average recovery of the method was (89-110)%, and the RSD within and between days were all less than 5%. The cefotaxime and sulbactam sample of plasma and urine stored in a -20℃ refrigerator became stable within 3 d. CONCLUSION: The assay method is rapid, accurate and usable for the determination of cefotaxime and sulbactam concentrations in plasma and urine.
【Keywords】 cefotaxime; sulbactam; chromatography, high pressure liquid; plasma; urine
【摘要】 目的: 為靜滴注射用頭孢噻肟鈉舒巴坦鈉后在血漿和尿液中的藥物濃度分析提供檢測方法. 方法: 采用HPLC測定頭孢噻肟鈉血漿和尿液中濃度. 在血樣中加入甲醇沉淀蛋白后,尿樣用超純水稀釋后取上清液進(jìn)樣,色譜分析條件: 以C18反相柱作分析柱;柱溫:室溫;流速: 1 mL/min;頭孢噻肟和舒巴坦的流動相分別為:1g/L的三乙胺水溶液(用磷酸調(diào)pH值6.2)∶甲醇=68∶32和84∶16;頭孢噻肟和舒巴坦的紫外檢測波長分別為254 nm和220 nm. 結(jié)果: 頭孢噻肟血樣和尿樣的線性范圍分別為5.3~530.0 mg/L和5.45~545.00 mg/L,最低檢測濃度分別為1.00 mg/L和0.25 mg/L;舒巴坦血樣和尿樣的線性范圍為1.04~208.00 mg/L,最低檢出濃度分別為0.5 mg/L和0.25 mg/L;方法平均提取回收率在 (89~110)%之間;日內(nèi)、日間RSD均小于5%. 頭孢噻肟和舒巴坦血漿和尿液樣品置于-20℃冰箱3 d內(nèi)穩(wěn)定. 結(jié)論: 本方法具有快速、簡便、靈敏、準(zhǔn)確等特點,適合于頭孢噻肟和舒巴坦血漿和尿液濃度測定.
【關(guān)鍵詞】 頭孢噻肟;舒巴克坦;色譜法,高壓液相;血漿;尿
0引言
由湘北威爾曼制藥有限公司開發(fā)研制的一類復(fù)方新藥注射用頭孢噻肟鈉舒巴坦鈉已經(jīng)國家食品藥品監(jiān)督管理局批準(zhǔn)進(jìn)行Ⅰ期臨床試驗(臨床研究批件號:2005L00727). 頭孢噻肟鈉屬第三代半合成頭孢菌素類,其抗菌譜廣,對革蘭陰性菌、革蘭陽性菌、需氧菌和某些厭氧菌均有較好的抗菌活性,特別對革蘭陰性菌的殺滅作用更強(qiáng). 隨著頭孢噻肟在臨床的廣泛使用,許多臨床病原菌通過產(chǎn)生β內(nèi)酰胺酶對頭孢噻肟產(chǎn)生耐藥性而導(dǎo)致臨床治療方案的失。1-2]. 舒巴坦為半合成β內(nèi)酰胺酶抑制劑,與頭孢噻肟合用時出現(xiàn)協(xié)同作用,使對頭孢噻肟耐藥的細(xì)菌的最低抑菌濃度下降到敏感范圍之內(nèi). 為了評價靜滴注射用頭孢噻肟鈉舒巴坦鈉后該藥品在健康志愿受試者的耐受性以及藥代動力學(xué)特點,我們采用高效液相色譜法測定了生物樣本中頭孢噻肟和舒巴坦的濃度.
1材料和方法
1.1材料Beckman公司System gold高效液相色譜儀(125型二元梯度泵,7725i手動進(jìn)樣器,20 μL定量環(huán),168型紫外檢測器,32 Karat TM Software). 頭孢噻肟、舒巴坦對照品(中國藥品生物制品檢定所),甲醇為色譜純(迪馬公司),水為自制超純水,其余試劑均為分析純.
1.2方法
1.2.1色譜條件色譜柱: 迪馬公司 Diamonsil C18(150 mm×4.6 mm, 5 μm);柱溫: 室溫;流速: 1 mL/min;進(jìn)樣量: 20 μL. 頭孢噻肟流動相: 1 g/L的三乙胺水溶液(用磷酸調(diào)pH值6.2)∶甲醇=68∶32;頭孢噻肟紫外檢測波長: 254 nm. 舒巴坦流動相: 1 g/L的三乙胺水溶液(用磷酸調(diào)pH至6.2)∶甲醇=84∶16;舒巴坦紫外檢測波長: 220 nm.
1.2.2樣品處理血樣處理: 精密量取血漿0.2 m
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