Garnuszek Piotr, Pawlak Dariusz, Maurin Michał, Jankovic Drina, Karczmarczyk Urszula, Mikołajczak Renata
National Centre for Nuclear Research — Radioisotope Centre POLATOM, Otwock, Poland.
Nucl Med Rev Cent East Eur. 2012 Aug 30;15(2):95-100.
The reliable method for determination of identity and radiochemical purity (RCP) is of great importance in radiopharmaceutical development. This is especially relevant when more than one form of radiometal/ligand complex can be formed during radiolabelling, such as complexes of 99mTc or 188Re with meso-2,3-dimercaptosuccinic acid (DMSA), where depending on the pH, metal can occur either at +3 or +5 oxidation state. The aim of our study was to evaluate possibilities for optimization of chromatographic systems leading to specific and reliable analytical method for determination of the identity and RCP of DMSA complexes with 99mTc or 188Re.
The commercial DMSA kits (POLATOM) were used for preparation of technetium-99m (III) and (V) complexes with DMSA. 99mTc(V)-DMSA complexes were prepared by addition of NaHCO3 to the kit vial prior to 99mTc-eluate to obtain pH ~8. 188Re(V)-DMSA was prepared either directly or using intermediate 188Re(III)-EDTA complex added to DMSA. RCP was evaluated by TLC using: ITLC-SG developed in methylethylketon, SG60 coated plates developed in: n-BuOH/H2O/CH3COOH and n-PrOH/H2O/CH3COOH systems, and in H2O. Comparative biodistribution studies were performed in normal Wistar rats.
Using silica gel plates and n-PrOH, H2O and acetic acid in the developing solution, we observed that 99mTc/188Re(III)-DMSA and 99mTc/188Re(V)-DMSA complexes could be well separated from each other and from the impurities in the form of free pertechnetate/perrhenate. In vivo studies showed quite different biodistribution of 99mTc(III)- and 99mTc(V)-DMSA. The trivalent complex accumulated mainly in kidneys (>40%ID), while 99mTc(V)-DMSA revealed high excretion with urine and relatively high concentration in osseous tissue (ca. 2 %ID/g). Accumulation of this complex in kidneys was very low (ca. 2.5 %ID). Biodistribution pattern of 188Re(V)-DMSA prepared directly was almost identical to that of 99mTc(V)-DMSA. Biodistribution results of the 188Re preparation obtained using 188Re(III)-EDTA intermediate indicated that the preparation contained the mixture of penta- and trivalent 188Re complexes. The quite high accumulation of radioactivity in kidneys (23 %ID) gave evidence of the presence of 188Re(III)-DMSA in this preparation, what was also confirmed by the results of TLC analysis performed using silica gel plate and n-propanol/water/acetic acid as developing system.
Based on our study, we have made recommendation on the suitable methods for investigations of RCP of DMSA complexes, i.e.: SG60 plates developed in the mixture of n-propanol/water/acetic acid, which enable determination of the tri- and pentavalent DMSA complexes, as well as, the pertechnetate/perrhenate impurity, and developed in water for determination of the colloidal residue.
可靠的放射性药物鉴定及放射化学纯度（RCP）测定方法在放射性药物研发中至关重要。当在放射性标记过程中可能形成不止一种形式的放射性金属/配体络合物时，这一点尤为重要，例如99mTc或188Re与内消旋-2,3-二巯基琥珀酸（DMSA）形成的络合物，其中根据pH值不同，金属可能以+3或+5氧化态存在。我们研究的目的是评估优化色谱系统的可能性，以建立一种特定且可靠的分析方法，用于鉴定99mTc或188Re与DMSA形成的络合物的身份及RCP。
使用市售DMSA试剂盒（POLATOM）制备99mTc（III）和（V）与DMSA的络合物。通过在99mTc洗脱液加入试剂盒小瓶之前加入NaHCO3使pH值达到约8来制备99mTc（V）-DMSA络合物。188Re（V）-DMSA可直接制备，也可通过将中间产物188Re（III）-EDTA络合物加入DMSA来制备。通过TLC评估RCP，使用在甲基乙基酮中展开的ITLC-SG、在n-丁醇/H2O/CH3COOH和正丙醇/H2O/CH3COOH体系以及在H2O中展开的SG60涂层板。在正常Wistar大鼠中进行了比较生物分布研究。
使用硅胶板以及展开液中的正丙醇、H2O和乙酸，我们观察到99mTc/188Re（III）-DMSA和99mTc/188Re（V）-DMSA络合物能够彼此良好分离，并且能与游离高锝酸盐/高铼酸盐形式的杂质分离。体内研究表明99mTc（III）-DMSA和99mTc（V）-DMSA的生物分布有很大差异。三价络合物主要蓄积在肾脏（>40%ID），而99mTc（V）-DMSA经尿液大量排泄，在骨组织中浓度相对较高（约2%ID/g）。该络合物在肾脏中的蓄积非常低（约2.5%ID）。直接制备的188Re（V）-DMSA的生物分布模式与99mTc（V）-DMSA几乎相同。使用188Re（III）-EDTA中间产物获得的188Re制剂的生物分布结果表明，该制剂含有五价和三价188Re络合物的混合物。肾脏中相当高的放射性蓄积（23%ID）证明该制剂中存在188Re（III）-DMSA，这也通过使用硅胶板和正丙醇/水/乙酸作为展开系统进行的TLC分析结果得到证实。
基于我们的研究，我们对DMSA络合物RCP的适宜研究方法提出了建议，即：在正丙醇/水/乙酸混合物中展开的SG60板，可用于测定三价和五价DMSA络合物以及高锝酸盐/高铼酸盐杂质；在水中展开用于测定胶体残留物。
