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Highly Sensitive Determination Method for Orally Active Antitumor Platinum Complexes of Next Generation and Their Active Metabolites

新一代口服活性抗肿瘤铂配合物及其活性代谢物的高灵敏测定方法

基本信息

批准号：
07672312
负责人：
KIZU Ryoichi
金额：
$ 1.41万
依托单位：
Kanazawa University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

This study was conducted to develop highly sensitive method for determining orally active antitumor platinum complexes of the next generation and their active metabolites. The complexes used were trans, cis, cis-bis (n-butyrato) (1R,2R-cyclohexanediamine) (oxalato) platinum (IV) (C4-OHP) and trans, cis, cis-bis (n-valerato) (1R,2R-cyclohexanediamine) (oxalato) platinum (IV) (C5-OHP), which are expected as promising oral agents because of their high antitumor activities in screening tests. Oral antitumor activities of C4-OHP and C5-OHP were first evaluated in mice bearing mouse leukemia L 1210. Significant increase in life span was observed with C5-OHP-treated group but not with C4-OHP-treated group. Therefore, subsequent study was made mainly on C5-OHP.Then, HPLC method for determining C5-OHP in biological samples was developed. C5-OHP was extracted with ethyl acetate and then subjected to reversed-phase HPLC.C5-OHP was chromatographed on an ODS column with water/methanol eluent and spectrophotometrically detected by at 210 nm. This method was found to be highly sensitive, giving the detection limit of 50 nM.C5-OHP in plasma, urine and cultured cell samples could be determined. Next, examination of active metabolites of C5-OHP was made by means of HPLC,revealing that (1R,2R-cyclohexanediamine) (oxalato) platinum (II) (OHP) was yielded from C5-OHP and the production was almost quantitative. Therefore, HPLC method for determining OHP in biological samples was developed. OHP was chromatographed in reversed-phase mode followed by post-column derivatization by sodium bisulfite and spectrophotometric detection at 290 nm. This method was also sensitive, giving the detection limit of 50 nM.The method required no pretreatment of samples but deproteinization by ultrafiltration and applicable to plasma, urine and cultured cell samples.

本研究旨在建立检测下一代口服抗肿瘤铂配合物及其活性代谢物的高灵敏度方法。所使用的配合物为反式、顺式、顺式-双（正丁酸）（1R， 2r -环己二胺）（草酸）铂（IV）（C4-OHP）和反式、顺式、顺式-双（n-戊酸）（1R， 2r -环己二胺）（草酸）铂（IV）（C5-OHP），由于在筛选试验中具有较高的抗肿瘤活性，有望成为有前景的口服药物。C4-OHP和C5-OHP的口服抗肿瘤活性首次在小鼠白血病l1210小鼠中进行了评价。c5 - ohp治疗组寿命明显延长，而c4 - ohp治疗组寿命无明显延长。因此，后续的研究主要针对C5-OHP。建立了HPLC法测定生物样品中C5-OHP的方法。用乙酸乙酯提取C5-OHP，反相高效液相色谱法测定。C5-OHP在ODS色谱柱上以水/甲醇洗脱，在210 nm处进行分光光度检测。该方法灵敏度高，检测限为50 nM。可测定血浆、尿液及培养细胞标本中的C5-OHP。接下来，通过HPLC检测C5-OHP的活性代谢产物，发现C5-OHP得到（1R， 2r -环己二胺）（草甘膦）铂（II）（OHP），产量几乎是定量的。为此，建立了HPLC法测定生物样品中OHP的方法。用反相色谱法对OHP进行色谱分析，然后用亚硫酸氢钠进行柱后衍生化，在290 nm处进行分光光度检测。该方法灵敏度高，检出限为50 nM。该方法无需样品预处理，只需超滤脱蛋白即可，适用于血浆、尿液和培养细胞样品。