Novel QCM biosensing based on analyte-specific precipitation reactions

We describe a novel method for the determination of paracetamol, in solution, using a method of analyte-specific precipitation. Surface adsorption is detected using the quartz crystal microbalance (QCM) sensor. The method is based on the enzymatic hydrolysis of paracetamol using arylacylamidase (AAA) to give 4-aminophenol (4-AP). The 4-AP specifically reacts with excess of a given phenol (for example o-cresol) to form insoluble indophenol dye species. The o-cresol is para-unsubstituted and acts as a sacrificial phenol. The free para position is an important requirement for the phenol coupling reaction. The change in frequency and impedance characteristics due to surface adsorption of the analyte-specific precipitate can be directly related to 4-AP concentration and subsequently to paracetamol levels. Ammonium persulphate (APS), a free-radical initiator was used in order to improve the speed of reaction and analysis time. Moreover, it was found that the speed of QCM sensor response was further improved when A PS was first heated at a fixed temperature (between 50-70°C) for 55-65 minutes. The enzyme (AAA) was studied either in solution or chemically cross-linked between polycarbonate membranes. The paracetamol analysis time was in the order of 5-7 minutes. The detection range was 0.1-1.5 mM paracetamol.