M. Almendral et al., SPECTROPHOTOMETRIC DETERMINATION OF L-ASPARAGINE BY FLOW-INJECTION ANALYSIS USING L-ASPARAGINASE IMMOBILIZED ON AN EPOXY-RESIN, Analytica chimica acta, 308(1-3), 1995, pp. 170-177
A continuous now 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 g
enerated in Berthelot's reaction. The enzyme L-asparaginase is covalen
tly bound to an epoxy resin (VA-Epoxy Biosynth, Riedel-De Haen) packed
in a glass column(50 X 3 mm i.d.). This enzyme reactor can be used fo
r more than 5 months and maintains about 88% of its initial enzyme act
ivity after 700 determinations. Under the proposed experimental condit
ions a linear calibration range is obtained for L-asparagine concentra
tions between 60 mu M and 15 mM (Abs = -4.29 X 10(-3) + 8.05 X 10(-4)[
Asp]; r = 0.999). The proposed procedure has a detection limit (3 sigm
a) of 1.2 ppm (9 mu M) and a relative standard deviation of 1.9% (12 d
eterminations at 30 ppm). Among the 22 amino acids investigated, only
L-histidine (greater than or equal to 960 ppm) and L-methionine (great
er than or equal to 640 ppm) were found to interfere.