Development of amperometric biosensor for choline determination
Aim. Development of an amperometric biosensor for measuring choline concentration in water samples. Methods. A bioselective element of the biosensor was created using choline oxidase which was covalently immobilized by glutaraldehyde crosslinking with bovine serum albumin on the surface of an ampero...
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Інститут молекулярної біології і генетики НАН України
2016
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| Cite this: | Development of amperometric biosensor for choline determination / D.Yu. Kucherenko, D.V. Siediuko, D.V. Knyzhnykova, O.O. Soldatkin, A.P. Soldatkin // Вiopolymers and Cell. — 2016. — Т. 32, № 3. — С. 229-234. — Бібліогр.: 9 назв. — англ. |
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Digital Library of Periodicals of National Academy of Sciences of Ukraine| _version_ | 1859750540778405888 |
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| author | Kucherenko, D.Yu. Siediuko, D.V. Knyzhnykova, D.V. Soldatkin, O.O. Soldatkin, A.P. |
| author_facet | Kucherenko, D.Yu. Siediuko, D.V. Knyzhnykova, D.V. Soldatkin, O.O. Soldatkin, A.P. |
| citation_txt | Development of amperometric biosensor for choline determination / D.Yu. Kucherenko, D.V. Siediuko, D.V. Knyzhnykova, O.O. Soldatkin, A.P. Soldatkin // Вiopolymers and Cell. — 2016. — Т. 32, № 3. — С. 229-234. — Бібліогр.: 9 назв. — англ. |
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| description | Aim. Development of an amperometric biosensor for measuring choline concentration in water samples. Methods. A bioselective element of the biosensor was created using choline oxidase which was covalently immobilized by glutaraldehyde crosslinking with bovine serum albumin on the surface of an amperometric platinum disk electrode. Results. The conditions of the bioselective element formation (the enzyme and glutaraldehyde concentrations, time of procedure) were optimized. The biosensor developed was characterized by good response reproducibility over hours of continuous operation. The linear range of substrate determination ranged from 10 µM to 1000 µM, a limit of choline detection – 1–3 µM, the biosensor sensitivity was 25–30 nA/mM. An effect of interfering substances was significantly reduced by the application of an additional semipermeable poly-m-phenylenediamine (PPD) membrane. Conclusions. The developed biosensor is well-suited for choline determination in water samples.
Мета. Розробка амперометричного біосенсора для визначення концентрацій холіну у водних зразках. Методи. Для створення біоселективного елементу біосенсора використовували холін оксидазу, що була іммобілізована ковалентною зшивкою глутаровим альдегідом з бичачим сироватковим альбуміном на поверхню амперометричного дискового платинового електроду. Результати. Було проведено оптимізацію умов формування біоселективного елементу на поверхню перетворювача (концентрація ферменту і глутарового альдегіду та час іммобілізації). Біосенсор характеризується доброю відтворюваністю відгуків впродовж декількох годин безперервної роботи. Лінійний діапазон визначення субстрату знаходився в межах від 10 мкМ до 1000 мкМ, мінімальна межа визначення холіну – 1–3 мкМ, чутливість біосенсора 25–30 нА/мМ. Завдяки використанню додаткової напівпроникної мембрани з полі-m-фенілендіаміну (ПФД) було значно зменшено вплив інтерферентів на роботу біосенсора. Висновки. Показано, що розроблений біосенсор добре підходить для визначення холіну у водних зразках.
Цель. Разработка амперометрического биосенсора для определения концентраций холина в водных образцах. Методы. Для создания биоселективного элемента использовали холин оксидазу, иммобилизованную ковалентной сшивкой глутаровым альдегидом с бычьим сывороточным альбумином на поверхность амперометрического дискового платинового электрода. Результаты. Была проведена оптимизация условий формирования биоселективного елемента на поверхность преобразователя (концентрация фермента и глутарового альдегида та время иммобилизации). Биосенсор характеризируется хорошей воспроизводимостью откликов на протяжении нескольких часов непрерывной работы. Линейный диапазон определения субстрата находился в пределах от 10 мкМ до 1000 мкМ, минимальная граница определения холина – 1–3 мкМ, чувствительность биосенсора 25–30 нА/мМ. Благодаря использованию дополнительной полупроницаемой мембраны с поли-м-фенилендиамина (ПФД) было значительно уменьшено влияние интерферентов на работу биосенсора. Выводы. Показано, что разработанный биосенсор хорошо применим для определения холина в водных образцах.
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229
D. Yu. Kucherenko, D. V. Siediuko, D. V. Knyzhnykova
© 2016 D. Yu. Kucherenko et al.; Published by the Institute of Molecular Biology and Genetics, NAS of Ukraine on behalf of Biopolymers and Cell.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/),
which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited
UDC 543.06 + 577.15 + 543.553
Development of amperometric biosensor for choline determination
D. Yu. Kucherenko1,2, D. V. Siediuko3, D. V. Knyzhnykova2,
O. O. Soldatkin1,2, A. P. Soldatkin1,2
1 Institute of Molecular Biology and Genetics, NAS of Ukraine
150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03680
2 Institute of High Technologies, Taras Shevchenko National University of Kyiv
2, korp.5, Pr. Akademika Hlushkova, Kyiv, Ukraine, 03022
3 National Aviation University
1, Komarova, Kyiv, Ukraine, 03058
didukh.d@gmail.com
Aim. Development of an amperometric biosensor for measuring choline concentration in water samples.
Methods. A bioselective element of the biosensor was created using choline oxidase which was covalently
immobilized by glutaraldehyde crosslinking with bovine serum albumin on the surface of an amperometric
platinum disk electrode. Results. The conditions of the bioselective element formation (the enzyme and gluta-
raldehyde concentrations, time of procedure) were optimized. The biosensor developed was characterized by
good response reproducibility over hours of continuous operation. The linear range of substrate determination
ranged from 10 µM to 1000 µM, a limit of choline detection – 1–3 µM, the biosensor sensitivity was 25–
30 nA/mM. An effect of interfering substances was significantly reduced by the application of an additional
semipermeable poly-m-phenylenediamine (PPD) membrane. Conclusions. The developed biosensor is well-
suited for choline determination in water samples.
K e y w o r d s: biosensor, amperometric transducer, immobilized enzyme, choline oxidase, choline.
Introduction
Choline (2-oxyethyltrimethyl ammonium hydro-
xide) belongs to the vitamin-like substances [1]. It is
an important ingredient for the nervous system due
to the synthesis of neurotransmitter acetylcholine.
The m onitoring of choline in blood serum is helpful
to identify neurodegenerative diseases: Alzheimer’s
disease, Parkinson’s disease, myasthenia gravis and
disruption of cholinergic neurotransmission [2].
The choline deficiency leads to delays in the de-
velopment and growth of organism, an increase in
blood cholesterol level, fatty infiltration of the liver,
varicose veins, high blood pressure and excess body
weight [1, 3, 4].
In clinical analysis, the choline quantification in
biological samples is of great importance. Spectro-
photometry, gas chromatography [5], mass spec-
trometry and chromatography-mass spectrometry
[6] are known as mostly used physical and chemical
methods for the choline identification. However,
they are also known to have significant drawbacks:
complicated and expensive equipment, requirement
for qualified personnel, and laboriousness.
The way to overcome these challenges is the us-
age of new bioanalytical devices – biosensors. Many
enzyme-based sensors have been developed for the
choline detection, such as the amperometric enzyme
sensor, electrochemiluminescent biosensor, ampero-
metric aqueous sol-gel biosensor and chemilumines-
Molecular and Cell
Biotechnologies
ISSN 1993-6842 (on-line); ISSN 0233-7657 (print)
Biopolymers and Cell. 2016. Vol. 32. N 3. P 229–234
doi: http://dx.doi.org/10.7124/bc.000924
mailto:didukh.d@gmail.com
230
D. Yu. Kucherenko, D. V. Siediuko, D. V. Knyzhnykova et al.
cent flow sensor [7]. The amperometric biosensors
are considered the most promising and successful
among the electrochemical biosensors.
Thus, this work was aimed at the development of
an amperometric biosensor for the quantitative anal-
ysis of choline in aqueous solutions. Moreover, this
biosensor could be a part of the biosensor array for
the simultaneous detection of several substances in
future investigation.
2. Materials and methods
2.1. Materials
In the work we used the enzyme choline oxidase
(ChOx), activity 15 unites/mg, from Sigma-Aldrich
(Japan). Bovine serum albumin (BSA, fraction V),
glycerol, HEPES, 50 % aqueous solution of glutaral-
dehyde (GA), choline chloride and m-phenylenedi-
amine were obtained from Sigma-Aldrich Chemie
(Germany). KH2PO4 and other compounds were of
Ukrainian production and of analytical purity grade.
2.2. Preparation of bioselective elements
In the work, the polyphenylenediamine (PPD) mem-
brane was prepared by the method described in [8].
The procedure was as follows. The bare working
electrodes were immersed in 5 mM m-phenylenedi-
amine solution, afterwards 10–12 cyclic voltammo-
grams were obtained. Next, the enzyme membrane
was deposited onto the PPD membrane.
The bioselective elements of biosensors were ob-
tained by immobilization of the enzyme and addi-
tives on the surface of amperometric transducer with
the PPD membrane .
The gel stock solution for preparation of the bi-
oselective membrane contained 8 % (hereinafter –
w/w) of ChOx, 4 % of BSA, 10 % of glycerol in
100 mM phosphate buffer, pH 6.5. Glycerol was
added to stabilize the enzymes during immobiliza-
tion to prevent early drop drying and to improve the
membrane adhesion to the transducer surface. Before
deposition onto the transducer surface, the enzyme
gel was mixed with 1.6 % aqueous solution of glu-
taraldehyde (crosslinking agent) in a 1:1 ratio.
Immediately thereafter, the mixture was deposited
onto the working surfaces of transducers and kept
for 10 min in air at room temperature. After immobi-
lization, the biosensors were washed in the working
buffer from unbound components of a bioselective
membrane and excess glutaraldehyde.
2.3. Measuring procedure
Three-electrode scheme of amperometric analysis
was used. Platinum disc electrodes of our own fab-
rication [9] served as amperometric transducers.
The working electrodes, auxiliary platinum elec-
trode and Ag/AgCl reference electrode were con-
nected to the potentiostat PalmSens (Palm
Instruments BV, Netherlands). The 8-channel de-
vice (CH-8 multiplexer from the same manufac-
turer) connected to the potentiostat allowed simul-
taneous registration of the signals from eight work-
ing electrodes, but actually only two or three work-
ing electrodes were attached.
The measurements were carried out at room tem-
perature in an open measuring cell, 3.5 ml in vol-
ume, with constant stirring at a constant potential of
+0.6 V vs Ag/AgCl reference electrode. The work-
ing buffer was 25 mM HEPES, pH 7.4. All experi-
ments were performed in triplicate. The substrate
concentrations in the working cell were set by add-
ing aliquots of the substrate stock solution.
3. Results and discussion
3.1. Principle of biosensor functioning
The functioning of amperometric biosensor for the
choline determination is based on the enzymatic re-
action (1), which takes place in a bioselective mem-
brane resulting in choline oxidation and formation of
electrochemically active hydrogen peroxide. While
applying a positive potential, the hydrogen peroxide
decomposion occurs on the electrode (2), which
causes the current changes directly recorded by the
amperometric transducer.
Choline oxidase (1)
Choline + O2 Betaine aldehyde + H2O2
231
Development of amperometric biosensor for choline determination
+0,6B (2)
H2O2 2H+ + O2 + 2e-
First, the biosensor was submerged in the 3.5 ml
measuring cell filled with 25 mM HEPES buffer
(pH 7.4) and kept there for several minutes to obtain
a stable baseline. Then aliquots of the stock choline
solution were added and the signals were obtained.
The data were automatically processed and graphi-
cally displayed on the computer monitor.
3.2. Choice of optimal conditions of choline
oxidase immobilization
The procedure of bioselective element immobilization
is an important factor, which affects analytical charac-
teristics and operation of the biosensor. The functioning
of enzyme biosensors depends on the composition of
bioselective element (the enzyme and GA concentra-
tions and their ratio) and time of the immobilization.
As the first step, an optimal GA concentration was
determined. The bioselective elements were pre-
pared with the GA mass fraction in the membrane of
0.1 %, 0.2 %, 0.4 %, 0.8 %, 1 % and 2.5 %. At low
GA concentrations, the biosensor responses were
low (Fig. 1), and they quickly decreased during re-
peated use. This was due to the weak binding of the
enzyme and its rapid leaching out the bioselective
membrane. At high GA concentrations, the respons-
es to choline were also low, this time because the
cross-linking was too strong and the enzyme could
not normally interact with the substrate.
In the further work, the membrane contains 0.8 %
of GA since at this concentration the biosensor re-
sponses to choline were the highest and most stable
during the measurement.
Another important parameter, which influences
the biosensor characteristics, is the enzyme concen-
tration in biomembranes. To determine the optimal
enzyme concentration, the bioselective elements
with ChOx mass fractions of 1 %, 2 %, 4 % and 6 %
were used. As seen (Fig. 2), the highest responses to
choline were observed at concentrations of 4 % and
6 %. Since these response values were about identi-
cal, in the further experiments the ChOx concentra-
tion in the bioselective element was 4 %.
The biosensors were also tested depending on the
duration of bioselective element formation ranging
from 5 to 40 min. The highest responses to choline
were observed when immobilization lasted 10 min,
at longer procedure the responses were significantly
lower (Fig. 3).
Fig. 1. Dependence of biosensor responses on GA concentration
in bioselective membrane. Choline concentrations – 100 µM
and 1000 µM. Measurements were carried out in 25 mM HEPES
buffer, pH 7.4, at constant potential of +0.6 V vs Ag/AgCl refer-
ence electrode.
Fig. 2. Dependence of biosensor responses on ChOx concentra-
tion in bioselective membrane. Choline concentrations: 100 µM
and 1000 µM. GA concentration in membrane – 0.8 %. Mea-
surements were carried out in 25 mM HEPES buffer, pH 7.4, at
constant potential of +0.6 V vs Ag/AgCl reference electrode.
232
D. Yu. Kucherenko, D. V. Siediuko, D. V. Knyzhnykova et al.
3.3. Reproducibility of biosensor responses
Reproducibility of the biosensor response is one of
the main characteristics of the efficiency of its opera-
tion. Therefore, the next step was to test the repro-
ducibility of biosensor response during continuous
operation. For the purpose, the responses to choline
concentrations of 10 µM, 100 µM and 1000 µM
were measured over one working day (Fig. 4). One
measurement lasted about 5 min, interval between
measurements was about 10 min (during this time
the biosensor was washed from the substrates chang-
ing the working buffer several times).
No notable decrease in the response value was ob-
served during 4 h continuous operation (15 measure-
ments); the relative standard deviation was 5 - 7 %.
3.4. Selectivity of biosensor
The mediatorless biosensor with relatively high operat-
ing potential (+0.6 V vs Ag/AgCl reference electrode)
was used, which made possible oxidation of a number
of electroactive compounds (for example, ascorbic
acid, uric acid and other) on the electrode surface.
Therefore, the selectivity of amperometric electrode
was improved by deposition of an additional polymer
membrane, which limits the diffusion of interfering
substances to the electrode surface. In the work, the
poly-m-phenylenediamine (PPD) membrane was used.
To confirm the improvement of selectivity of the
modified transducer, the electrodes sensitivity to-
wards interfering substances was tested. The bare
transducer reacted to certain substances quite strong-
ly, which could be a problem while analyzing the real
samples. The deposition of PPD membrane resulted
in essential decrease or even complete absence of the
biosensor responses to interferents whereas the sensi-
tivity to hydrogen peroxide remained almost the same
(Table 1). It was also found that the reproducibility of
the biosensor signals to choline is significantly im-
proved with the PPD membrane deposition due to
better adhesion of biomembrane to the electrode.
3.5 Analytical characteristics of biosensor for
choline determination
The linear working range of the biosensor and mini-
mum limit of choline detection were determined.
The minimum limit of choline detection was de-
fined as the choline concentration, the biosensor re-
sponse to which was three times higher than the
Fig. 3. Dependence of biosensor responses on time of immobili-
zation. Choline concentrations: 100 µM and 1000 µM. GA and
ChOx concentrations in membrane – 0.8 % and 4 %, respec-
tively. Measurements were carried out in 25 mM HEPES buffer,
pH 7.4, at constant potential of +0.6 V vs Ag/AgCl reference
electrode.
Fig. 4. Reproducibility of biosensor response. Choline concen-
trations - 10 µM, 100 µM and 1000 µM. Measurements were
carried out in 25 mM HEPES buffer, pH 7.4, at constant poten-
tial of +0.6 V vs Ag/AgCl reference electrode.
233
Development of amperometric biosensor for choline determination
baseline noise; it was found to be 1–3 µM. This val-
ue slightly varied for different biosensors but in-
creased as the biosensor was further used. The linear
working range was from 10 µM to 1000 µM, the bio-
sensor sensitivity 25–30 nA/mM. The typical cali-
bration curve of the biosensor for choline determina-
tion is shown in Fig. 5.
Conclusions
The biosensor based on choline oxidase has been de-
veloped for the choline determination in aqueous so-
lutions. The optimum conditions of the enzyme im-
mobilization by glutaraldehyde crosslinking with
bovine serum albumin on the transducer surface
were found to be as follows: GA concentration –
0.8 %, ChOx concentration – 4 %, time of immobili-
zation – 10 min. Reproducibility of the biosensor
response to choline was investigated over one work-
ing day, the relative standard deviation was 5–7 %. It
has been shown that the application of PPD mem-
branes on the transducer surface before the bioselec-
tive element deposition resulted in a significant de-
crease in an impact of the main possible in biological
fluids interferents on the biosensor operation.
The basic analytical characteristics of the devel-
oped biosensor were determined. They were as fol-
lows: linear range of choline determination – 10–
1000 µM, minimum limit of detection – 1–3 µM,
sensitivity – 25–30 nA/mM. This biosensor is
planned to be further used for the measurement of
choline concentration in real biological samples. It
could be also a part of the biosensor array for simul-
taneous measurement of several substances.
Funding
The authors gratefully acknowledge the financial sup-
port of this study by the National Academy of Sciences
of Ukraine in the frame of Scientific and Technical
Government Program “Sensor systems for medico-
ecological and industrial-technological requirement:
metrological support and experimental operation”.
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3. Zeisel SH. “Vitamin-like” molecules. In: Eds. Shils ME,
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Table 1. Selectivity of amperometric transducer before and
after deposition of PPD membrane
Possible interferent
Respose, nA
without PPD
membrane
with PPD
membrane
Hydrogen peroxide, 50 µM 34.7 ± 2.6 27.6 ± 0.8
Ascorbic acid, 500 µM 33.2 ± 1.7 0.9 ± 0.5
Dopamine, 20 µM 14.8 ± 1.3 1.2 ± 0.3
Uric acid, 100 µM 10.6 ± 1.8 0
Paracetamol, 100 µM 7.3 ± 1.2 0
Cysteine, 100 µM 2.8 ± 0.4 0
Citric acid, 500 µM 0 0
Sodium chloride, 1 mM 0 0
Potassium chloride, 1mM 0 0
Calcium chloride, 1mM 0 0
Fig. 5. Calibration curve of biosensor for choline determination.
Measurements were carried out in 25 mM HEPES buffer,
pH 7.4, at constant potential of +0.6 V vs Ag/AgCl reference
electrode.
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Розробка амперометричного біосенсора для визначення
холіну
Д. Ю. Кучеренко, Д. В. Сєдюко, Д. В. Книжникова,
О. О. Солдаткін, О. П. Солдаткін
Мета. Розробка амперометричного біосенсора для визначен-
ня концентрацій холіну у водних зразках. Методи. Для ство-
рення біоселективного елементу біосенсора використовували
холін оксидазу, що була іммобілізована ковалентною зшив-
кою глутаровим альдегідом з бичачим сироватковим альбумі-
ном на поверхню амперометричного дискового платинового
електроду. Результати. Було проведено оптимізацію умов
формування біоселективного елементу на поверхню перетво-
рювача (концентрація ферменту і глутарового альдегіду та
час іммобілізації). Біосенсор характеризується доброю від-
творюваністю відгуків впродовж декількох годин безперерв-
ної роботи. Лінійний діапазон визначення субстрату знахо-
дився в межах від 10 мкМ до 1000 мкМ, мінімальна межа
визначення холіну – 1–3 мкМ, чутливість біосенсора 25–
30 нА/мМ. Завдяки використанню додаткової напівпроникної
мембрани з полі-m-фенілендіаміну (ПФД) було значно змен-
шено вплив інтерферентів на роботу біосенсора. Висновки.
Показано, що розроблений біосенсор добре підходить для ви-
значення холіну у водних зразках.
К л юч ов і с л ов а: біосенсор, амперометричний перетворю-
вач, іммобілізований фермент, холін оксидаза, холін.
Разработка амперометрического биосенсора для
определения холина
Д. Ю. Кучеренко, Д. В. Седюко, Д. В. Книжникова,
А. А. Солдаткин, А. П. Солдаткин
Цель. Разработка амперометрического биосенсора для опре-
деления концентраций холина в водных образцах. Методы.
Для создания биоселективного элемента использовали холин
оксидазу, иммобилизованную ковалентной сшивкой глутаро-
вым альдегидом с бычьим сывороточным альбумином на по-
верхность амперометрического дискового платинового элект-
рода. Результаты. Была проведена оптимизация условий фор-
мирования биоселективного елемента на поверхность преоб-
разователя (концентрация фермента и глутарового альдегида
та время иммобилизации). Биосенсор характеризируется хоро-
шей воспроизводимостью откликов на протяжении несколь-
ких часов непрерывной работы. Линейный диапазон определе-
ния субстрата находился в пределах от 10 мкМ до 1000 мкМ,
минимальная граница определения холина – 1–3 мкМ, чув-
ствительность биосенсора 25–30 нА/мМ. Благодаря использо-
ванию дополнительной полупроницаемой мембраны с по-
ли-м-фенилендиамина (ПФД) было значительно уменьшено
влияние интерферентов на работу биосенсора. Выводы.
Показано, что разработанный биосенсор хорошо применим
для определения холина в водных образцах.
К л юч е в ы е с л ов а: биосенсор, амперометрический пре-
образователь, иммобилизированный фермент, холин оксидаза,
холин.
Received 25.04.2016
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| id | nasplib_isofts_kiev_ua-123456789-152821 |
| institution | Digital Library of Periodicals of National Academy of Sciences of Ukraine |
| issn | 0233-7657 |
| language | English |
| last_indexed | 2025-12-01T23:13:23Z |
| publishDate | 2016 |
| publisher | Інститут молекулярної біології і генетики НАН України |
| record_format | dspace |
| spelling | Kucherenko, D.Yu. Siediuko, D.V. Knyzhnykova, D.V. Soldatkin, O.O. Soldatkin, A.P. 2019-06-13T07:40:44Z 2019-06-13T07:40:44Z 2016 Development of amperometric biosensor for choline determination / D.Yu. Kucherenko, D.V. Siediuko, D.V. Knyzhnykova, O.O. Soldatkin, A.P. Soldatkin // Вiopolymers and Cell. — 2016. — Т. 32, № 3. — С. 229-234. — Бібліогр.: 9 назв. — англ. 0233-7657 DOI: http://dx.doi.org/10.7124/bc.000925 https://nasplib.isofts.kiev.ua/handle/123456789/152821 543.06 + 577.15 + 543.553 Aim. Development of an amperometric biosensor for measuring choline concentration in water samples. Methods. A bioselective element of the biosensor was created using choline oxidase which was covalently immobilized by glutaraldehyde crosslinking with bovine serum albumin on the surface of an amperometric platinum disk electrode. Results. The conditions of the bioselective element formation (the enzyme and glutaraldehyde concentrations, time of procedure) were optimized. The biosensor developed was characterized by good response reproducibility over hours of continuous operation. The linear range of substrate determination ranged from 10 µM to 1000 µM, a limit of choline detection – 1–3 µM, the biosensor sensitivity was 25–30 nA/mM. An effect of interfering substances was significantly reduced by the application of an additional semipermeable poly-m-phenylenediamine (PPD) membrane. Conclusions. The developed biosensor is well-suited for choline determination in water samples. Мета. Розробка амперометричного біосенсора для визначення концентрацій холіну у водних зразках. Методи. Для створення біоселективного елементу біосенсора використовували холін оксидазу, що була іммобілізована ковалентною зшивкою глутаровим альдегідом з бичачим сироватковим альбуміном на поверхню амперометричного дискового платинового електроду. Результати. Було проведено оптимізацію умов формування біоселективного елементу на поверхню перетворювача (концентрація ферменту і глутарового альдегіду та час іммобілізації). Біосенсор характеризується доброю відтворюваністю відгуків впродовж декількох годин безперервної роботи. Лінійний діапазон визначення субстрату знаходився в межах від 10 мкМ до 1000 мкМ, мінімальна межа визначення холіну – 1–3 мкМ, чутливість біосенсора 25–30 нА/мМ. Завдяки використанню додаткової напівпроникної мембрани з полі-m-фенілендіаміну (ПФД) було значно зменшено вплив інтерферентів на роботу біосенсора. Висновки. Показано, що розроблений біосенсор добре підходить для визначення холіну у водних зразках. Цель. Разработка амперометрического биосенсора для определения концентраций холина в водных образцах. Методы. Для создания биоселективного элемента использовали холин оксидазу, иммобилизованную ковалентной сшивкой глутаровым альдегидом с бычьим сывороточным альбумином на поверхность амперометрического дискового платинового электрода. Результаты. Была проведена оптимизация условий формирования биоселективного елемента на поверхность преобразователя (концентрация фермента и глутарового альдегида та время иммобилизации). Биосенсор характеризируется хорошей воспроизводимостью откликов на протяжении нескольких часов непрерывной работы. Линейный диапазон определения субстрата находился в пределах от 10 мкМ до 1000 мкМ, минимальная граница определения холина – 1–3 мкМ, чувствительность биосенсора 25–30 нА/мМ. Благодаря использованию дополнительной полупроницаемой мембраны с поли-м-фенилендиамина (ПФД) было значительно уменьшено влияние интерферентов на работу биосенсора. Выводы. Показано, что разработанный биосенсор хорошо применим для определения холина в водных образцах. The authors gratefully acknowledge the financial support of this study by the National Academy of Sciences of Ukraine in the frame of Scientific and Technical Government Program “Sensor systems for medicoecological and industrial-technological requirement: metrological support and experimental operation”. en Інститут молекулярної біології і генетики НАН України Вiopolymers and Cell Molecular and Cell Biotechnologies Development of amperometric biosensor for choline determination Розробка амперометричного біосенсора для визначення холіну Разработка амперометрического биосенсора для определения холина Article published earlier |
| spellingShingle | Development of amperometric biosensor for choline determination Kucherenko, D.Yu. Siediuko, D.V. Knyzhnykova, D.V. Soldatkin, O.O. Soldatkin, A.P. Molecular and Cell Biotechnologies |
| title | Development of amperometric biosensor for choline determination |
| title_alt | Розробка амперометричного біосенсора для визначення холіну Разработка амперометрического биосенсора для определения холина |
| title_full | Development of amperometric biosensor for choline determination |
| title_fullStr | Development of amperometric biosensor for choline determination |
| title_full_unstemmed | Development of amperometric biosensor for choline determination |
| title_short | Development of amperometric biosensor for choline determination |
| title_sort | development of amperometric biosensor for choline determination |
| topic | Molecular and Cell Biotechnologies |
| topic_facet | Molecular and Cell Biotechnologies |
| url | https://nasplib.isofts.kiev.ua/handle/123456789/152821 |
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