正电子发射计算机断层扫描（PET）技术是一种高灵敏度的分子成像技术，在诊断和指导治疗脑重大疾病及肿瘤等方面具有重要意义。随着PET技术的飞速发展，临床上对PET药物的需求也日益旺盛。然而，传统的评价和筛选PET药物的放射性方法常常受到放射性标记困难，筛选程序繁琐等限制。因此，免标记的快速高通量筛选候选化合物的新方法对加快新型放射性药物的开发具有重要意义。 高效液相色谱串联质谱（LC-MS/MS）技术具有高灵敏度、高分离能力和高选择性的特征。将LC-MS/MS分析方法引入放射性药物的研究，不仅能够减少放射性的使用，而且可以加快放射性药物的筛选速度，提高放射性药物的研究水平。另外，放射性方法无法区分放射性来源，一次实验只能评价一种候选PET显像剂，而LC-MS/MS技术能够实现多组分高通量分析，因此可以实现高通量药物筛选。盒式给药又称作N in 1给药，是将多种化合物混合后对同一只动物给药，然后进行样品处理和分析。将盒式给药方法与LC-MS/MS技术结合起来，同时测定多个化合物在生物体内的性质，可以提高药物筛选通量，大大缩短样品处理及分析时间，减少实验动物数量，是加快药物筛选及研发过程的有效方法。 本论文首次将基于盒式给药的LC-MS/MS技术方法运用到PET药物的筛选评价方面，成功建立了高通量的LC-MS/MS分析方法，验证了方法的可行性和可靠性，为PET药物的高通量筛选奠定了基础。 主要研究内容和结果如下： 1. 建立LC-MS/MS分析方法，根据美国FDA对生物样品检测的相关规定，对所建立的LC-MS/MS分析方法进行方法学验证，评价研究方法的准确性和科学性。 2. 进行体内受体占据实验，确定给药剂量，既满足检测灵敏度，又保证盒式给药的情况下，药物之间在体内不发生竞争抑制。 3. 大鼠随机分成三组，分别进行单一给药（FPBM），2 in 1盒式给药（FPBM、FPBM2）和3 in 1盒式给药（FPBM、FPBM2、AV-133），运用所建立的LC-MS/MS方法分析生物样品，研究三种给药方式下，各化合物在大鼠脑中的生物分布情况，验证盒式给药方法的可行性及可靠性。 本研究首次将基于盒式给药的LC-MS/MS分析方法运用于PET显像剂筛选。通过LC-MS/MS分析得到的生物分布数据与标准的放射性方法得到的结果匹配得很好。此外，盒式给药与单一给药之间没有显著性差异。实验结果表明，非标记的LC-MS/MS方法研究PET药物是准确可靠的，盒式给药可以用于放射性药物的早期高通量筛选。该方法将从根本上提高放射性药物早期筛选的通量，加速放射性药物的研究。基于盒式给药的LC-MS/MS技术是一种很有前景PET药物的高通量筛选方法。

Positron emission tomography (PET) is a very sensitive molecular imaging technique. However, the utility of this method is limited by the availability of suitable radiopharmaceuticals to probe specific targets and biology related to disease process. A complex radiolabeling procedure that is time consuming, expensive and technically difficult may deter the development of PET radiopharmaceuticals. Methods to screen candidate compounds prior to radiolabeling would speed up the discovery of novel radiopharmaceuticals. The unique application of the high-performance liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) method has been critical for reducing bottlenecks in radiopharmaceutical discovery by enabling the evaluation of non-labeled small molecules in vivo as potential radiopharmaceutical candidates. By circumventing the need for radiolabeled molecules, this method may be efficacious and safe for the research and development of new PET imaging agents. The label-based techniques are limited in their inability to distinguish which chemical compound the radioactivity is from, so only one imaging agent can be evaluated in one experiment. Besides its high sensitivity, LC-MS/MS provides the possibility to analyze multiple compounds simultaneously, which make it possible to enhance the throughput of PET imaging agent screening. Cassette dosing is also known as N-in-One dosing. Several compounds were simultaneously administrated to a single animal and then the samples were rapidly detected by LC-MS/MS. The use of cassette dosing is a successful strategy to enhance the efficiency of drug discovery and reduce animal usage. The purpose of this study is to develop an approach combining cassette microdosing with LC-MS/MS to enhance the throughput for screening radiopharmaceutical biodistribution in the rat brain. The experiments and results are as follows: 1. An LC-MS/MS analysis method was established and validated by a set of parameters that were in compliance with the reference guidelines defined in FDA documents. 2. The receptor occupancy experiment was performed to determine the dosage, ensuring that there was no competition inhibition among the drugs. 3. The rats were divided into three groups: (1) the group 1 were only administered FPBM; (2) the group 2 were given a mixture of FPBM and FPBM2; (3) the group 3 were given a cocktail of FPBM, FPBM2 and AV-133.The regional distribution was measured by LC-MS/MS. The results approached by LC-MS/MS matched very well with the values obtained by standard radioactivity measurements. Moreover, no significant differences between discrete microdosing and cassette microdosing were observed. In this study, a cassette microdosing with LC-MS/MS method was first applied to the screening of PET imaging agents. The cassette microdosing with LC-MS/MS strategy is feasible and reliable. The combination of cassette microdosing and LC-MS/MS would be a reasonably high throughput screening tool at an early stage in the discovery/development process of PET imaging agents.