Encapsulation of Urease in Molybdenum Trioxide Sol-Gel
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A6.2.1
ENCAPSULATION OF UREASE IN MOLYBDENUM TRIOXIDE SOL-GEL Prashant K. Jha and P. I. Gouma Department of Material Science State University of New York Stony Brook, NY 11794 ABSTRACT Molybdenum trioxide sol-gel matrices were tested for their ability to host biomaterials in their structures without adversely affecting the activity of the encapsulated biomolecules. The sol-gel composites were prepared at room temperature; the urease was added during the hydrolysis step to ensure proper encapsulation in the pore structure of the sol. Phosphate buffer solution was added during this stage to maintain the pH value of the sol in the bio-compatible range. The activity of incorporated urease was tested by exposing composite sol-gel material to standard urea test solutions and measuring the amount of gaseous ammonia (released by the reaction of urea with water in the presence of urease) using an ammonia electrode. The concentration of urea solutions ranged from 1mM to 10mM. An electronic olfactory system (EOS 835) was also employed to detect the presence of gaseous ammonia. Gas sensing tests were done to verify the sensitivity of the matrix to ammonia. Microstructural analysis was carried out by means of scanning and transmission electron microscopy. It was found that urease retained its activity inside molybdenum trioxide sol-gels that are sensitive to ammonia. These hybrid nanoporous composites are useful in the field of biosensors and fuel cells. INTRODUCTION Most of the urea biosensors are based on the enzymatic action of urease (equation 1) which specifically catalyzes hydrolysis of urea and the product/s of the reaction are detected by various techniques [1-6]. NH2CONH2 + H2O
Urease
CO2 + NH3
(1)
The potentiometric sensor [7], which is most widely used, suffers from many drawbacks like slow response to change in NH+ ion concentration and interference from other ions present in the solution like Na+, K+ [8]. In some sensors, urease is immobilized in passive media (such as natural and synthetic polymers and passive sol-gel matrices) and the change in electrical conductivity or potential is measured using a probe electrodes. The silica sol-gel method has emerged as a popular method of encapsulating bio-molecules as it is done at room temperature which is conducive for the bio-molecules [9]. The optical transparency of silica has been utilized for optical detection of pathogens [10-11]. Our approach is to utilize the gas sensitivity of the matrix to fabricate a resistive type bio-sensor, and this has not been tried before to the best of our knowledge. Molybdenum trioxide was chosen as the host for bio-molecules as it has been shown in our earlier works to be sensitive and selective towards ammonia [12]. Here we report successful incorporation of urease in MoO3 sol-gel matrix and sensitivity of bio-composite to ammonia.
A6.2.2
EXPERIMENTAL Urease solution was made by mixing 0.662g (10592 units) of urease [(EC 3.5.1.5) from SIGMA] in 25ml water + 25 ml glycerol. This procedure was based on reference [13]. Then for e
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