Ratiometric fluorescent l‐arginine and l‐asparagine biosensors based on the oxazine 170 perchlorate‐ethyl cellulose membrane

Abstract

Ratiometric fluorescent l-arginine (Arg) and l-asparagine (Asn) biosensors were developed using an oxazine 170 perchlorate (O17) ethyl cellulose (EC) membrane and the enzymes entrapped into the matrix of EC and hydrogel polyurethane. The sensing principles were based on the hydrolysis reactions of urea and l-Arg under the catalysis of the urease and arginase to produce ammonia in the case of an l-Argsensing membrane and also on the hydrolysis reaction of l-Asn under the catalysis of asparaginase in the case of an l-Asn-sensing membrane. The O17-EC membrane reacted with the ammonia produced from the hydrolysis reactions and changed the fluorescence intensities at λ em = 565 and 625 nm. The ratio of the fluorescence intensities at λ em = 565 and 625 nm was proportional to the concentrations of l-Arg or l-Asn in the range of 0.1-10 mM. The LOD of the l-Arg-and l-Asn-sensing membranes was 0.082 ± 0.0014 and 0.074 ± 0.0023 mM, respectively. The sensing membranes also showed good quality in terms of response time, reversibility, and stability. The interference study demonstrated that some components such as amino acids had little negative effects on the performance of the sensing membranes for the detection of l-Arg and l-Asn. These simple and sensitive ratiometric fluorescent sensing membranes provide a basic or comprehensive method for detecting l-Arg and l-Asn in blood and urine samples as well as in the fermentation processes.Keywords: l-Arginine / l-Asparagine / O17-EC membrane / Ratiometric fluorescent biosensor Additional supporting information may be found in the online version of this article at the publisher's web-site

Recent Advances in Cellulose-Based Biosensors for Medical Diagnosis

“…An et al [95] reported recently the development of ratiometric fluorescent l-arginine and l-asparagine biosensors employing ethyl cellulose membrane, oxazine 170 perchlorate fluorophore, and enzymes immobilized in a matrix of ethyl cellulose membrane and polyurethane hydrogel. The sensory mechanism depended on the hydrolysis reaction of urea and l-arginine in presence of urease and arginase as catalysts.…”

“…The biosensing membranes displayed a high quality in terms of the reversibility, stability, and response time. The interference study showed that some constituents, such as amino acids, had slight negative impacts on the efficiency of the biosensing membranes in detecting l-arginine and l-asparagine in both urine and blood samples as well as in the fermentation processes [95,96]. Schyrr et al [97] investigated cellulose nanocrystals as an appropriate material for the simple integration of high-density biological sensors.…”

Recent advances in cellulose-based membranes for their sensing applications

“…The biosensor supplied a potential method for urea-sensing in food safety and clinical diagnostics. Similarly, a ratiometric fluorescent biosensor for l -arginine (Arg) and l -asparagine (Asn) detection was designed by Jong Il Rhee, which was combined oxazine 170 perchlorate (O17)-ethyl cellulose (EC) membrane with enzymes and hydrogel polyurethane (An et al 2017 ). Ammonium ions were produced by the hydrolysis reactions of urea and L-Arg under the catalysis of the urease and arginase, as well as the hydrolysis reaction of L-Asn under the catalysis of asparaginase.…”

Directed Evolution of Ornithine Cyclodeaminase Using an EvolvR-Based Growth-Coupling Strategy for Efficient Biosynthesis of l-Proline

“…Li et al screened asparaginase of ultrahigh thermostability by ammonia detection with Nessler’s reagent . An O17-EC sensing membrane, reacting with the ammonia generated from the reactions through enzyme hydrolysis and changing the proportional fluorescence intensity ratio of λ em 565/λ em 625, was used to detect of l -Arg and l -asparagine . However, ammonium detection is susceptible to environmental interference and affected greatly by cell growth fluctuation.…”