Department of Engineering for Innovation, University of Salento, via per Arnesano Km 1, 73100 Lecce, Italy.
Biosens Bioelectron. 2013 Feb 15;40(1):336-41. doi: 10.1016/j.bios.2012.07.074. Epub 2012 Aug 11.
A highly selective water compatible molecularly imprinted polymer (MIP) for 3-nitro-L-tyrosine (3NT), an oxidative stress marker associated with neurodegenerative disorders, was prepared and its use as solid-phase extraction (SPE) sorbent material was demonstrated. The MIP was prepared by bulk polymerization using methacrylic acid as functional monomer and acetonitrile as porogen with traces of acetic acid and trifluoroacetic acid. In order to evaluate its binding properties, the MIP was analyzed by batch rebinding experiments and subsequently used as SPE sorbent for the selective clean-up and pre-concentration of 3NT from standard solutions and spiked human urine samples. The results obtained from batch rebinding experiments showed the presence of two association constants corresponding to high-affinity (Ka 4.20×10(3) M(-1)) and low-affinity (Ka 0.79×10(3) M(-1)) binding sites. Standard mixture solution loaded on MIP-SPE cartridge gave a recovery of 95% for 3NT, while the other compounds were totally eluted during washing step. Percentage of recovery higher than 90%, with relative standard deviation of 2%, was also obtained when a maximum of 55 μg of 3NT is used in spiked urine sample and loaded into the cartridge. Validation of the analytical method for 3NT quantification in human urine gave 0.7 μg mL(-1) of limit of detection, a linear range of 2.5-55 μg mL(-1) with a relative standard deviation of 2%.
一种对 3-硝基-L-酪氨酸（3NT）具有高选择性的亲水性分子印迹聚合物（MIP）被制备出来，3NT 是一种与神经退行性疾病相关的氧化应激标志物。该 MIP 是通过本体聚合制备的，使用甲基丙烯酸作为功能单体，乙腈作为致孔剂，并加入痕量的乙酸和三氟乙酸。为了评估其结合性能，通过批量再结合实验对 MIP 进行了分析，随后将其用作固相萃取（SPE）吸附剂，用于从标准溶液和加标人尿样中选择性地净化和预浓缩 3NT。批量再结合实验的结果表明，存在两种结合常数，分别对应于高亲和力（Ka 4.20×10(3) M(-1))和低亲和力（Ka 0.79×10(3) M(-1))结合位点。将标准混合物溶液加载到 MIP-SPE 柱上，3NT 的回收率为 95%，而其他化合物在洗涤步骤中完全洗脱。当在加标尿样中使用多达 55 μg 的 3NT 并将其加载到柱上时，回收率也高于 90%，相对标准偏差为 2%。在人尿中对 3NT 进行定量分析的方法验证得到了 0.7 μg mL(-1)的检测限，线性范围为 2.5-55 μg mL(-1)，相对标准偏差为 2%。
