Development of Forensic Diagnosis of Acture Sarin Poisoning

合成了一种沙林类有机磷毒剂双（甲基）膦酸异丙酯（BIMP）。该制剂具有与沙林相同的膦酸酯基团，也具有与沙林相同的抗乙酰胆碱酯酶活性效力。制备实验性BMP暴露的人红细胞后，从红细胞膜上溶解BMP结合的乙酰胆碱酯酶（AChE），通过免疫亲和层析纯化，用胰蛋白酶消化，并通过碱性磷酸酶消化释放结合AChE的沙林水解产物。采用三甲基硅烷（TMS）衍生化，气相色谱-质谱（GC-MS）联用检测沙林水解产物。异丙基甲基膦酸和甲基膦酸，这是沙林的水解产物，在实验BIMP暴露的人红细胞中检测到。利用这种新的方法，我们可以检测到沙林水解产物的红细胞和福尔马林固定的小脑的东京地铁恐怖袭击的受害者。

A sarin-like organophosphorus agent, bis (isopropyl methyl) phosphonate (BIMP), was synthesized. This agent has the same phosphonate group as sarin and also has the same anti-acethylcholinesterase activity potency as sarin. After preparation of experimental BIMP-exposed human red blood cells, BIMP-bound acetylcholinesterase (AChE) was solubilized from erythrocyte membranes, purified by immunoaffinity chromatography, digested with trypsin, and the sarin hydrolysis products bound to AChE were released by alkaline phosphatasedigestion. The digested sarin hydrolysis products were subjected to trimethylsilyl (TMS) derivatization and detected by gas chromatography-mass spectrometry (GC-MS). Isopropyl methylphosphonic and methylphosphonic acids, which are the sarin hydrolysis products, were detected in experimental BIMP-exposed human red blood cells. Using this new method, we could detected the sarin hydrolysis products from erythrocytes and formalin-fixed cerebellums of victims of the Tokyo subway terrorist attack.