Spectrophotometric determination of l-asparagine by flow-injection analysis using l-asparaginase immobilized on an epoxy resin

✍ Scribed by M.J. Almendral; M.J. Porras; A. Alonso; M.D. Baez; C. Alonso

• Publisher: Elsevier Science
• Year: 1995
• Tongue: English
• Weight: 660 KB
• Volume: 308
• Category: Article
• ISSN: 0003-2670
• DOI: 10.1016/0003-2670(94)00623-t

✦ Synopsis

A continuous flow procedure for the determination of L-asparagine based on its L-asparaginase-catalyzed conversion to L-aspartate and ammonia is proposed. The ammonia formed is measured via the indophenol blue generated in Berthelot's reaction. The enzyme L-asparaginase is covalently bound to an epoxy resin (VA-Epoxy Biosynth, Riedel-De Hlen) packed in a glass column (50 X 3 mm i.d.). This enzyme reactor can be used for more than 5 months and maintains about 88% of its initial enzyme activity after 700 determinations. Under the proposed experimental conditions a linear calibration range is obtained for L-asparagine concentrations between 60 /_L.M and 15 mM (Abs = -4.29 X 10e3 + 8.05 X 10-4[Asp]; I = 0.999). The proposed procedure has a detection limit (3~) of 1.2 ppm (9 /IM) and a relative standard deviation of 1.9% (12 determinations at 30 ppm). Among the 22 amino acids investigated, only L-histidine (2 960 ppm) and L-methionine (2 640 ppm) were found to interfere.