An electrochemical sensor for Dimetridazole detection

1. Dimetridazole
 Dimetridazole (DMZ) is a nitroimidazole compound used in
veterinary medicine.
 The major metabolite of DMZ is 2-(hydroxymethyl)-l-methyl-5-
nitroimidazole (DMZOH), formed by hydroxylation of the 2-methyl
group.
Figure- Structure of DMZ

2. Use of DMZ
Dimetridazole was approved by the U.S. Food and Drug
Administration (FDA) in 1971 for use in pigs, poultry, turkeys, game
birds, pigeons and other caged birds.
The permitted uses for products containing DMZ were –
 treatment and prevention of blackhead in poultry and birds.
 treatment and prevention of swine dysentery in pigs.
 Treatment of canker in pigeons and caged birds.

3. Harmful Side Effect of DMZ on Human Beings
Carcinogenicity Mutagenicity

4. Importance of DMZ Detection
Due to the harmful side effect of DMZ, the usage of DMZ fully
restricted from different country
But DMZ are using in food additives, poultry feed and fish feed
illegally
DMZ ratio doesn’t damage after cooking
Therefore, the determination of DMZ residue is very necessary to protect
public health.

5. Technique of DMZ Detection
Several analytical methods including-
• high-performance liquid chromatography–mass spectrometry
• liquid chromatography
• gas chromatography
• capillary electrophoresis
• immunoassay
• adsorptive stripping voltammetry
• polarography

6. Drawbacks of the Following technique
Time consuming
Complexity
Costly
Less sensitivity and reproducibility
Less satisfactory limit of detection (LOD)

7. Electrochemical
sensor

8. Why Electrochemical Sensor???

9. Sensor

10. Electrochemical sensor
Transducer
Processing Display

11. MIP( Molecular Imprinting polymer) Sensor
/ MIP as Electrochemical sensor
• Molecularly Imprinted Polymers (MIPs) is a polymer that binds
together to form a specific binding site that is selective for certain
analyte.
• The template and functional monomers are mixed, a
prepolymerization complex is formed which is stable in a
predetermined solvent.
• Finally, the template is removed from the polymer, leaving a cavity
that is able to bind analyte selectively.

12. Materials & Methods

13. Electrode Preparation
Polishing GCE using aqueous slurry of aluminum of
0.05µm on micro cloth 1µm.
Washing GCE in Ultrasonic bath with dilute nitric acid
for 5 minute
Washing GCE in Ultrasonic bath with ethanol for 5
minute
Washing GCE in Ultrasonic bath with ultra pure water
for 5 minute
Acid treatment with 1M sulfuric acid at voltrametric
range of -1V to 1V at scan rate 100 for 20 cycle

14. Electrochemical Set up

15. Fabrication of MIP & NIP

16. Real sample Preparation
Weighting 1gm real sample (egg, milk, honey)
Taking 1gm of each sample in 15ml centrifuge tube
and adding 3ml of 10% trichloroacetic acid
Vortexing the mixture for 1 minute
Centrifuging the mixture at 5000 rpm for 5 minute
Filtering the mixture and separate the supernatant
Diluting 100 times using 0.1 ×10-3 mol L‒1 DMZ in
PBS

17. Result & Discussion

18. Electrochemical characterization of Proposed
Electrode
• Cyclic voltammetric characterization of GCE/P-arg@MIP
Bare GCE
GCE/P-Arg@NIP
GCE/P-Arg@MIP
Figure 1: Cyclic voltammograms of the different electrodes measured in (A) 0.1 mol L‒1 KCl including 5.0×10-3 mol
L‒1 Fe(CN)6
3‒/4‒ and (B) 0.1M PBS including 1mM DMZ.
A B

19. Electrochemical characterization of Proposed
Electrode cont.…..
• Effective surface area of different electrodes is calculated using the
Randles–Sevcik formula: Ip = (2.69×105)n3/2AD1/2Φ1/2C0
Where,
Ip= peak current (µA)
n= number of electrons
D= diffusion coefficient (cm2/s)
C0= the concentration of the redox probe (mol/cm3)
Φ= scan rate (mV/s)

20. Electrochemical characterization of Proposed
Electrode cont.…..
• For K3Fe(CN)6, n = 1, D = 7.6×10−6 cm2/s (0.1 M KCl). Based on the
slope of the linear relationship between Ip and Φ1/2.
For the bare GCE, effective surface area A = 0.112 cm2
For GCE/P-Arg@NIP, A= 0.198 cm2
For GCE/P-Arg@MIP, A=0.174 cm2

21. Electropolymerization of GCE/P-Arg@MIP
Sensor
Figure 2: Cyclic voltammograms for the electrochemical polymerization of 2.5 ×10-3 mol L-1 P-Arg on GCE electrode (A)
and CV of P-Arg electropolymerization with 1×10-3 mol L-1 DMZ during the modification of GCE/P-Arg@MIP sensor (B).
Deposition condition: 0.1 mol L-1 PBS solution (pH 7.4) at the scan rate 100 mV/s for 12 cycles).

22. Surface topographical characterization of MIP
sensor
Figure 4: SEM images of GCE/P-Arg electrode (A), GCE/P-Arg@MIP electrode before (B), and after (C) removal of
DMZ template.

23. Optimization condition of modified electrode
• Effect of pH
2
5
8
11
4.5 5.5 6.5 7.5 8.5 9.5
I/µA
pH
Figure 5. The effect of pH on the oxidation peak current of 1×10-3mol L-1 DMZ at the GCE/MIP@P-Arg
in 0.1M PBS at different pH

24. Optimization condition of modified electrode cont.….
Figure 6: CV response obtained for GCE/P-Arg/MIP sensor different scan rates (from inner to outer): 10, 20, 30, 40, 50, 60,
70, 80, 90, 100, 150 and 200 mV/s in 0.1 mol L‒1 KCl including 5.0 ×10-3 mol L‒1 Fe(CN)6
3‒/4‒ solution

25. Electrochemical Detection of DMZ
Figure 7 : DPVs of 1×10-3 mol L-1 DMZ at bare GCE (a), GCE/P-Arg@NIP (b), and GCE/P-Arg@MIP electrode (c)
in pH 7.4 phosphate buffer at a scan rate of 100 mV s−1

26. Electrochemical Detection of DMZ cont.…
2.2
2.4
2.6
2.8
3
3.2
3.4
3.6
3.8
4
-1.15 -1.05 -0.95 -0.85 -0.75 -0.65 -0.55 -0.45 -0.35
I/µA
E/V vs Ag/AgCl
y = 0.0104x + 0.3378
R² = 0.9859
0
0.5
1
1.5
0 50 100 150
Figure 7: DPVs obtained for detection of DMZ using GCE/P-Arg@MIP sensor in pH 7.4 phosphate buffer at scan
rate of 100 mV s−1 with a wide range of concentrations from 0.1 ×10-9 mol L‒1 to 100 ×10-6 mol L‒1. Inset shows the
calibration curves for all concentrations.

27. Reproducibility of the Sensor
The reproducibility of the sensor was investigated for a fixed
concentration of 10µmol/L by recording repeatedly.
No deviation in current response was found for several sample for the
same electrode.

28. Sensitivity of the Sensor
 The selectivity of the modified electrode was studied with different
compound and metal ions such as Na+, K+, Ca2+, Mg2+, Fe2+, Cu2+, Zn2+, Al3+,
Ascorbic acid and Uric acid.
 Some nitroimidazole analogues including Ornidazole, Metronidazole and
Ronidazole were also performed for evaluating interface.
 Limited interface (less than 5%) were exhibited by the modified electrode.
 The result was found in 10 µmol/L of DMZ and 200 fold of the following
compound.

29. Stability of the Sensor
• The DPV curve of 10 µmol/L of DMZ was recorded weekly to
examine the stability of the proposed sensor after its preservation at
4˚C in refrigerator.
• High stability of the sensor was obtained even after two week.

30. Comparison of the proposed sensor with others
Electrodes Methods Linear range
(µmolL−1)
Detection limit
(nmolL−1)
Reference
GCE/ AuNPs@MIP DPV 0.002-250 0.5 [1]
GCE/ Chitosan Amperometric 0.02-150 2.7 [2]
Cu2O/ErGO DPV 0.03-0.15 3.6 [3]
MIS-CPE CV 1-10 3.6 [4]
GCE/P-Arg@MIP DPV 0.0001-100 0.1 This work

31. Real sample Analysis
The feasibility and applicability of this sensor was evaluated by
detecting DMZ in milk powder, egg, and honey samples.
. The proposed MIP sensor exhibited recovery ranged from 94.2% to
101.8% suggests excellent feasibility for practical application in real
sample analysis
Sample Added (µM) Found (µM) Recovery (%)
Egg 10 9.42 94.2
Milk 10 10.18 101.8
Honey 10 9.55 95.5

32. Conclusion
Highly effective electrochemical MIP sensor developed for
ultrasensitive detection of dimetridazole.
 GCE/P-Arg@MIP electrode was fabricated via electrochemical
deposition technique.
 The proposed sensor exhibited a wide linear detection range with an
LOD of 0.1 nM.
 The GCE/P-Arg@MIP sensor was applied to detect analyte in egg,
milk and honey.

33. Future plan

34. Achievement
• This research article successfully
published in Analytica Chimica Acta
(IF-5.977) that’s available in online
from 5 May 2020.