Disposable cuvette test with integrated sensor layer for enzymatic determination of heavy metals

We report on a quick and simple test for detection of heavy metals based on the inhibition of free urease contained in a disposable cuvette. The wall of the cuvette has been covered with sensors for either ammonia or ammonium ion, and the kinetics of the enzymatic reaction has been monitored by monitoring the color of the sensing layer rather than that of the whole solution. In order to perform the test, buffer, enzyme and sample are placed in the cell. On addition of urea, enzymatic hydrolysis results in the formation of ammonia (and ammonium ion), and this is detected by one of the sensors. Heavy metals are detected by their inhibitory effect. This sensing scheme is new in that it enables even strongly colored and highly turbid samples to be analyzed, because it is the optical properties of the 4 mu m sensing layer on the wall that are monitored rather than the sample itself. Since sample cells can be made at low cost, they may be disposed of after use. Heavy metal ions are found to inhibit urease in the following decreasing order: Ag(I)>Hg(II)>Cu(II)much greater than Ni(II)>Co(II)>Cd(II)>Fe(III)>Zn(II)>Pb(II). The lowest limits of detection are found for Ag(I) (20 ppb), Hg(II) (70 ppb) and Cu(II) (250 ppb). A comparison of the ammonia gas-sensitive optode and the ammonium ion-sensitive optode showed the former to be more precise and appropriate for determination of heavy metals. The toxicity of the heavy metals correlates well with the stability constants of the corresponding metal sulfides. We also discuss the inhibitory effect of metal combinations on the activity of the enzyme and the effects of ionic strength and incubation time. (C) 1996 Elsevier Science Limited.