Faculty of Bioscience and Technology for Food, Agriculture and Environment, University of Teramo, via Renato Balzarini 1, 64100, Teramo, Italy; Laboratory of Process Engineering and Environment, Faculty of Sciences and Techniques, Hassan II University of Casablanca, Mohammedia, Morocco.
Faculty of Bioscience and Technology for Food, Agriculture and Environment, University of Teramo, via Renato Balzarini 1, 64100, Teramo, Italy.
Talanta. 2022 Apr 1;240:123195. doi: 10.1016/j.talanta.2021.123195. Epub 2021 Dec 30.
Several methods involving molecularly imprinted polymers (MIPs) devoted to extracting and analyzing sulfonamides from different matrices are reported in literature; however, the unresolved analytical issue is obtaining intra-class selectivity between sulfonamides. Here is presented for the first time a method coupling MIPs and enzymatic inhibition assay for the sensitive and selective determination of acetazolamide (ACZ) in biological samples. The MIPs were synthesized by thermal initiated polymerization in acetone, using acrylamide as functional monomer, ethylene glycol dimethacrylate as cross-linker and ACZ as template molecule. The developed MIPs/enzymatic inhibition based rapid colorimetric method was applied for the determination of ACZ in biological samples. The MIPs were used as sorbent phase in dispersive solid-phase extraction (MIPs-dSPE), and the optimal working parameters were selected. Liquid chromatography-tandam mass spectrometry (LC-MS/MS) analysis confirmed the MIPs ability to extract ACZ. Finally, to obtain a selective and sensitive method, the MIPs-dSPE was combined with an enzymatic inhibition colorimetric assay based on the carbonic anhydrase, an enzyme inhibited by specific sulfonamides. The developed combined method allowed the determination of ACZ in serum, blood and Diamox (a drug containing ACZ), with good recovery (85-96%). Furthermore, a significant correlation with LC-MS/MS analysis was achieved, with relative error ≤15%. In the proposed strategy, the double selectivity giving by MIPs and enzymatic inhibition allowed to obtain a method able to determine selectively ACZ in biological and pharmaceutical samples quantitatively.
有几种涉及分子印迹聚合物（MIPs）的方法被用于从不同基质中提取和分析磺胺类药物，这些方法在文献中有报道；然而，未解决的分析问题是在磺胺类药物之间获得类内选择性。本文首次提出了一种将 MIPs 与酶抑制测定法相结合的方法，用于在生物样品中灵敏和选择性地测定乙酰唑胺（ACZ）。MIPs 通过在丙酮中引发聚合反应合成，使用丙烯酰胺作为功能单体、乙二醇二甲基丙烯酸酯作为交联剂，ACZ 作为模板分子。所开发的基于 MIPs/酶抑制的快速比色法用于测定生物样品中的 ACZ。MIPs 被用作分散固相萃取（MIPs-dSPE）中的吸附相，并选择了最佳的工作参数。液相色谱-串联质谱（LC-MS/MS）分析证实了 MIPs 提取 ACZ 的能力。最后，为了获得选择性和灵敏的方法，将 MIPs-dSPE 与基于碳酸酐酶的酶抑制比色测定法结合，碳酸酐酶是一种被特定磺胺类药物抑制的酶。所开发的组合方法允许在血清、血液和 Diamox（一种含有 ACZ 的药物）中测定 ACZ，回收率良好（85-96%）。此外，与 LC-MS/MS 分析实现了显著的相关性，相对误差≤15%。在所提出的策略中，MIPs 和酶抑制提供的双重选择性允许获得一种能够在生物和药物样品中定量选择性地测定 ACZ 的方法。
