Developments in the synthesis of a water compatible molecularly imprinted polymer as artificial receptor for detection of 3-nitro-l-tyrosine in neurological diseases

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 4.20×103 M−1) and low-affinity (Ka 0.79×103 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%.