A novel automatic determination system for correction and simultaneous
quantification of peroxidase and ascorbate in biosamples
Abstract
Mastering synchronous changes of peroxidase (POD) activity concentration
(EPOD) and ascorbic acid contents (cAsA) in biosamples can be used for
estimation of ant-oxidative capability and healthy status of animals and
plants, however, in determination of EPOD, the coexisted AsA produced
serious interference. Therefore, to solve this problem and obtain true
EPOD, this study deeply investigated the AsA interference using a
guaiacol/POD/H2O2 reaction. Results manifest that instead of
inactivation of the enzyme, the AsA causes the extinction of the
chromogenic product formed in the reaction, resulting in the EPOD to
deviate true value; this phenomenon occurs in all methods for
quantification of EPOD. Because the interference of AsA is linear with
the EPOD, the utilization of the linear correlation realized the
correction of EPOD and the quantification of cAsA. Subsequently, the use
of the extinction and coloration phenomena in the reaction, a new
automatic analysis method and system were developed, and were used for
the simultaneous determination of EPOD and cAsA in biosamples. After
optimized, the variables of the system were obtained as follows: the
determination ranges are 75 to 1300 U•L-1 for EPOD and 5 to 100 mg•L-1
for cAsA; the detection limits are 22.5 U•L-1 for EPOD and 1.67 mg•L-1
for cAsA; the relative standard deviation is less than 1.2% (n = 11);
the sample throughput can reach 40 samples per hour; and the standard
addition recovery is in the range of 95 to 105%. This automatic method
and system has been used successfully for accurate quantifications of
EPOD and cAsA in various biosamples, and based on this, a new
specialized analyzer can be also fabricated for the study of biosamples
in biotechnology and bioengineering.