This document summarizes the process of creating molecularly imprinted polymers (MIPs) and non-imprinted polymers (NIPs) for the detection of celecoxib. It describes the materials, synthesis, and testing methods used. The polymers were created using acrylamide as the functional monomer and ethylene glycol dimethacrylate as the cross-linker, with celecoxib as the template molecule. The binding capacity of the resulting MIPs and NIPs was tested through serial dilution binding experiments measuring absorbance with a NanoDrop spectrophotometer. The goal was to develop an imprinted polymer sensor for celecoxib detection.

1. STAJ RAPORU KAPAK SAYFASI
SUMMER TRAINING REPORT COVER PAGE
ÖĞRENCİNİN/ STUDENT;
STAJ YAPTIĞIKURUMUN/ INSTITUTION OF SUMMER TRAINING;
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2. SUMMARYOF THE INTERNSHIP PLACEAND ABOUT DR.PINARCAKIR HATIR
Istanbul Arel University has adopted as a principle to provide its students with a modern higher
education at international academic standarts, since this is the basic way to ensure that our
country earns the reputation it deserves among world countries. Istanbul Arel University aims
to raise individuals who are devoted to principles and reforms of Atatürk; believe in existence
of Republic of Turkey as a secular, democratic and social law state; who research, think
globally, respect universal values; and who are adhered to national values and open to scientific,
technologic developments.(1)
Istanbul Arel University was founded on 18th May, 2007. University has 5 Faculties as Faculty
of Arts and Sciences, Faculty of Engineering and Architecture, Faculty of Communication,
Faculty of Economic and Administrative Sciences, Faculty of Fine Arts; 3 Schools as School
of Health Sciences, School of Applied Sciences, School of Foreign Languages; 1 Vocational
School; and 2 Institutes as Institute of Sciences and Institute of Social Sciences. Istanbul Arel
University has 2 campuses:
1. Tepekent Campus, includes Faculties and Schools that provide four-years education.
2. 2. Sefaköy Campus, within the Sefaköy Campus, there are Vocational School and Graduate
School programs.(2)
I worked with Assist. Prof. Dr. Pinar Cakir Hatir during my internship. Dr. Cakir, graduated
from Boğaziçi University Department of Chemistry in 2004. Since 2013, she works as
Assistant Professor in the department of Biomedical Engineering at İstanbul Arel University.
In addition, she has published articles in international peer-reviewed journals and there are
projects.
MOLECULARLYIMPRINTED POLYMER (MIP) & NON IMPRINTED POLYMER (NIP)
INTRODUCTION:
A Molecularly Imprinted Polymer (MIP) is a polymer that has been processed using the
molecular imprinting technique which leaves cavities in polymer matrix with affinity to a
chosen "template" molecule. The process usually involves initiating the polymerization of
monomers in the presence of a template molecule that is extracted afterwards, thus leaving
complementary cavities behind. These polymers have affinity for the original molecule and
have been used in applications such as chemical separations, catalysis, or molecular sensors.(3)

3. Figure 1: Polymerization and Molecular Imprinting
In this study, as illustrated above, functional monomers and template (CELECOXIB) using
polymerization make and was obtained from the polymer removed the template, ultimately was
obtained the imprinted polymer.
What is celecoxib?
Celecoxib is a nonsteroidal anti-inflammatory drug (NSAID). It works by reducing hormones
that cause inflammation and pain in the body.
Celecoxib is used to treat pain or inflammation caused by many conditions such as arthritis,
ankylosing spondylitis, and menstrual pain.(4)
MATERIALS AND METHOD:
POLYMERIZATION STEP
Firstly, obtain two clean vial and reagents put in the amounts given values:
AAm (Acrylamide) : 0.2 mmol
EGDMA (Ethylene glycol dimethacrylate) :1 mmol
DMPA (Dimethoxy-2-phenylacetophenone) : 0.0088 mmol
CELEC (Celecoxib): 0.025 mmol
ACN (Acetonitrile) : 1920 mg
For first vial, I put all above reagents, but for second vial put all above, without Celecoxib.
Secondly,again two clean vial and reagents put in the amounts given values:
AAm (Acrylamide) : 0.2 mmol
EGDMA (Ethylene glycol dimethacrylate) :1 mmol

4. DMPA (Dimethoxy-2-phenylacetophenone) : 0.0088 mmol
CELEC (Celecoxib): 0.025 mmol
Tol (Toluene) : 1920 mg
For first vial, I put all above reagents, but for second vial put all above, without Celecoxib.
Solution prepared in four seperate vials, to start the reaction vials were placed at UV cabine for
18 hour.
WASHING STEP
The polymers obtained were transferred into the falcons with a spatul. After, in another beaker
20 ml acetic acid and 80 ml of methanol solution was prepared. This solution was prepared, it
was placed on the polymer in the tube to be 10 milliliters. It was stored on the shaker for one
hour and after that, it was centrifuged for 10 minutes, 5000 rpm. The liquid portion remaining
on the top, the bottom was evacuated to collapsed polymers. This step was performed 4 times,
respectively. Fourth washing, after centrifugation received 1 milliliter of the supernatant, the
absorbance value was measured in the NanoDrop. (1 ml was taken from each tube.) By the way
methanol was used to blank the NanoDrop.
To remove acetic acid, methanol was introduced into the falcons containing the polymer (to be
10 ml). It was stored on the shaker for 45 minutes and after that, it was centrifuged for 10
minutes, 5000 rpm. The liquid portion remaining on the top, the bottom was evacuated to
collapsed polymers. This step was performed 3 times, respectively.
After all these operations, cover of the tubes were opened, they were covered with napkin. It
was placed in a vacuum oven overnight.
[To give you some information about the reagents used: AAm: Functional Monomer ;
EGDMA:Cross-Linker ; DMPA: Initiator ; CELECOXIB: Template ; ACN: Solvent ; Tol :
Solvent]
FOR CALIBRATION STAGES:

5. Figure 2: Serial Dilution
First, to prepare a stock solution and then the amounts shown in fig-1, 6 pieces of prepared in
Eppendorf tubes. Then absorbance values were measured with the NanoDrop.(for measured
blank ACN)
As illustrated above, instead of ACN, using Toluene (same amounts) was prepared calibration.
FOR BINDING STAGES:
Figure 3: Binding Stage
For the first binding:
As shown in the above figures (Figure-3);
MIP & NIP polymer:
For 1st eppendorf tube : 5 mg MIP(ACN) + 1 mL ACN
For 2nd eppendorf tube : 5 mg NIP(ACN) + 1 mL ACN
For 3th eppendorf tube : 5 mg MIP(ACN) + 1 mL of 100 uM CELECOXIB,ACN
For 4th eppendorf tube : 5 mg NIP(ACN)+ 1 mL of 100 uM CELECOXIB,ACN

6. For 5th eppendorf tube : 5 mg MIP(Toluene) + 1 mL ACN
For 6th eppendorf tube : 5 mg NIP(Toluene) + 1 mL ACN
For 7th eppendorf tube : 5 mg MIP(Toluene) + 1 mL of 100 uM CELECOXIB,ACN
For 8th eppendorf tube : 5 mg NIP(Toluene) + 1 mL of 100 uM CELECOXIB,ACN
For 9th eppendorf tube : 1 mL of 100 uM CELECOXIB,ACN
It was mixed in given amounts and shaker was left overnight. Next day, eppendorf tubes lined
centrifuge machine and the centrifuge machine was set to be 10 minutes and 14.000 rpm. After
centrifugation from the supernatant, the Eppendorf tube 1.5 ul taking the absorbance values
were measured on the NanoDrop.
All of these stages, it was performed for MIP & NIP polymer experiments. However in these
experiments, to use 9 seperate vial.
Also, for the preparation of the 100 uM Celecoxib solution;
Figure 4: To prepare 100 uL of Celecoxib Solution
For stock solution 4 mg celecoxib was stirred with 5.2 ml acetonitrile. Then,
Add the 1st Falcon tube: 800 uL of Stock Sol.+ 800 uL ACN 1 mM of Celec.
Add the 2nd Falcon tube: 800 uL of 1 mM Sol.+800 uL ACN 0.5 mM of Celec.
Add the 3th Falcon tube: 800 uL of 0.5 mM Sol.+800 uL ACN 0.25 mM of Celec.
Add the 4th Falcon tube:800 uL of 0.25 mM Sol.+800 uL ACN 0.125 mM of Celec.
Add the 5th Falcon tube:200 uL of 0.125 mM Sol.+800 uL ACN 0.025 mM of Celec.

7. Add the 6th Falcon tube:200 uL of 0.025 mM Sol.+300 uL ACN 0.01 mM of Celec.
It was mixed in given amounts and shaker was left overnight. Next day, eppendorf tubes lined
centrifuge machine and the centrifuge machine was set to be 10 minutes and 14.000 rpm. After
centrifugation from the supernatant, the Eppendorf tube 1.5 ul taking the absorbance values
were measured on the NanoDrop.
The obtained result of absorbans value in the NanoDrop, to calculate Binding Calculation.
For the second binding;
In the meanwhile, acetonitrile were used solutions prepared in and to obtain the desired result
Toluene was used instead of acetonitrile.
MIP&NIP polymer :
For 1st eppendorf tube : 5 mg MIP(ACN) + 1 mL Toluene
For 2nd eppendorf tube : 5 mg NIP(ACN) + 1 mL Toluene
For 3th eppendorf tube : 5 mg MIP(ACN) + 1 mL of 100 uM CELECOXIB,Toluene
For 4th eppendorf tube : 5 mg NIP(ACN)+ 1 mL of 100 uM CELECOXIB,Toluene
For 5th eppendorf tube : 5 mg MIP(Toluene) + 1 mL Toluene
For 6th eppendorf tube : 5 mg NIP(Toluene) + 1 mL Tolene
For 7th eppendorf tube : 5 mg MIP(Toluene) + 1 mL of 100 uM CELECOXIB,Tolene
For 8th eppendorf tube : 5 mg NIP(Toluene) + 1 mL of 100 uM CELECOXIB,Toluene
For 9th eppendorf tube : 1 mL of 100 uM CELECOXIB,Toluene
The solutions prepared in acetonitrile, toluene was used but not to obtain the desired result
acetonitrile instead of 5% acetic acid, 100 uL Celec. + Toluene was used.
For the third binding:
MIP&NIP polymer :
For 1st eppendorf tube : 5 mg MIP(ACN) + 1 mL Toluene(5% Asetic Acid)
For 2nd eppendorf tube : 5 mg NIP(ACN) + 1 mL Toluene(5% Asetic Acid)
For 3th eppendorf tube : 5 mg MIP(ACN) + 1 mL of 100 uM CELEC,Toluene(5% Asetic Acid)

8. For 4th eppendorf tube : 5 mg NIP(ACN)+ 1 mL of 100 uM CELEC,Toluene(5% Asetic Acid)
For 5th eppendorf tube : 5 mg MIP(Toluene) + 1 mL Toluene(5% Asetic Acid)
For 6th eppendorf tube : 5 mg NIP(Toluene) + 1 mL Tolene(5% Asetic Acid)
For 7th eppendorf tube : 5 mg MIP(Toluene) + 1 mL of 100 uM CELEC,Tolene(5% Asetic Acid)
For 8th eppendorf tube : 5 mg NIP(Toluene) + 1 mL of 100 uM CELEC,Toluene(5% Asetic
Acid)[not 100 uM, calculation error was made.So 500 uM was used.]
For 9th eppendorf tube : 1 mL of 100 uM CELECOXIB,Toluene(5% Asetic Acid) [not 100 uM,
calculation error was made.So 500 uM was used.]
The obtained result of absorbance value in the NanoDrop, to calculate Binding Calculation.
For the fourth binding;
For starters, 2 MM stock solution was prepared.
3.8 mg CELECOXIB + 5 ml ACN
Afterwards, was made dilutions:
0.5 MM 2 ml of 2MM stock solution + 6 ml ACN
0.250 MM 4 ml of 0.5 MM solution + 4 ml ACN
0.125 MM 4 ml 0f 0.250 MM solution + 4 ml ACN
0.025 MM 2 ml of 0.125 MM solution + 8 ml ACN
0.001 MM 2 ml of 0.025 MM solution + 3 ml ACN
Subsequently, for the binding process
NIP (ACN):
For the 1st epp.tube: 1 ml of 0.5 MM Solution + 5 mg polymer
For the 2nd epp.tube: 1 ml of 0.25 MM Solution + 5 mg polymer
For the 3 th epp.tube: 1 ml of 0.125 MM Solution+5 mg polymer
For the 4th epp.tube: 1 ml of 0.025 MM Solution + 5 mg polymer
For the 5th epp.tube: 1 ml of 0.01 MM Solution + 5 mg polymer

9. For the 6th epp.tube: 1 ml of ACN + 5 mg polymer
MIP (ACN):
For the 1st epp.tube: 1 ml of 0.5 MM Solution + 5 mg polymer
For the 2nd epp.tube: 1 ml of 0.25 MM Solution + 5 mg polymer
For the 3 th epp.tube: 1 ml of 0.125 MM Solution+5 mg polymer
For the 4th epp.tube: 1 ml of 0.025 MM Solution + 5 mg polymer
For the 5th epp.tube: 1 ml of 0.01 MM Solution + 5 mg polymer
For the 6th epp.tube: 1 ml of ACN + 5 mg polymer
Also, to draw the calibration curve[2nd Calibration]; 6 piece of each tube, the solution was
added.
For the first epp.tube: Add 1 ml of 0.5 MM
For the second epp.tube: Add 1 ml of 0.250 MM
For the third epp.tube: Add 0.125 MM
For the fourth epp.tube: Add 0.025 MM
For the fifth epp.tube: Add 0.001 MM
It was mixed in given amounts and shaker was left overnight. Next day, eppendorf tubes lined
centrifuge machine and the centrifuge machine was set to be 10 minutes and 14.000 rpm. After
centrifugation from the supernatant, the Eppendorf tube 1.5 ul taking the absorbance values
were measured on the NanoDrop.
The obtained result of absorbans value in the NanoDrop, to calculate Binding Calculation.
For the fifth binding;
Firstly, 2 MM stock solution was prepared.
3.8 mg CELECOXIB + 5 ml ACN
Subsequently, dilution was made.
0.5 MM 2 ml of 2MM stock solution + 6 ml ACN
0.250 MM 4 ml of 0.5 MM solution + 4 ml ACN

10. 0.125 MM 4 ml 0f 0.250 MM solution + 4 ml ACN
0.025 MM 2 ml of 0.125 MM solution + 8 ml ACN
0.001 MM 2 ml of 0.025 MM solution + 3 ml ACN
After, for the binding process
NIP (Toluene):
For the 1st epp.tube: 1 ml of 0.5 MM Solution + 10 mg polymer
For the 2nd epp.tube: 1 ml of 0.25 MM Solution + 10 mg polymer
For the 3 th epp.tube: 1 ml of 0.125 MM Solution+10 mg polymer
For the 4th epp.tube: 1 ml of 0.025 MM Solution + 10 mg polymer
For the 5th epp.tube: 1 ml of 0.01 MM Solution + 10 mg polymer
For the 6th epp.tube: 1 ml of ACN + 10 mg polymer
P 172-2 MIP (Toluene)
For the 1st epp.tube: 1 ml of 0.5 MM Solution + 10 mg polymer
For the 2nd epp.tube: 1 ml of 0.25 MM Solution + 10 mg polymer
For the 3 th epp.tube: 1 ml of 0.125 MM Solution+10 mg polymer
For the 4th epp.tube: 1 ml of 0.025 MM Solution + 10 mg polymer
For the 5th epp.tube: 1 ml of 0.01 MM Solution + 10 mg polymer
For the 6th epp.tube: 1 ml of ACN + 10 mg polymer
In addition, to draw the calibration curve[3th Calibration]; 6 piece of each tube, the solution was
added.
For the first epp.tube: Add 1 ml of 0.5 MM
For the second epp.tube: Add 1 ml of 0.250 MM
For the third epp.tube: Add 0.125 MM
For the fourth epp.tube: Add 0.025 MM
For the fifth epp.tube: Add 0.001 MM

11. It was mixed in given amounts and shaker was left overnight. Next day, eppendorf tubes lined
centrifuge machine and the centrifuge machine was set to be 10 minutes and 14.000 rpm. After
centrifugation from the supernatant, the Eppendorf tube 1.5 ul taking the absorbance values
were measured on the NanoDrop. The obtained result of absorbans value in the NanoDrop, to
calculate Binding Calculation.
RESULT& DISCUSSION:
Graph-1: Graph of Calibration Celecoxib+ ACN [first calib.]
Graph-2: Graph of Calibration Celecoxib+ ACN [second calib.]
The result of first binding:
MIP(ACN): -4% ; NIP(ACN): -8.4% ; MIP(Toluene): 19.802% ; NIP(Toluene): -5.9%
The result of second binding:
MIP(ACN): 42.28% ; NIP(ACN): 45.14% ; MIP(Toluene): 44.19% ; NIP(Toluene): 46.66%
The result of third binding:
MIP(ACN): %17.08 ; NIP(ACN): 21.79% ; MIP(Toluene): 28.27% ; NIP(Toluene): 22.53%
y = 2.339x
R² = 0.9993
0
0.2
0.4
0.6
0.8
1
1.2
1.4
0 0.1 0.2 0.3 0.4 0.5 0.6
Abs.Value@251nm
Concentration of Celecoxib+ACN
CALIBRATION
y = 0.9582x + 0.0002
R² = 0.9976
0
0.1
0.2
0.3
0.4
0.5
0.6
0 0.1 0.2 0.3 0.4 0.5 0.6
ABS.VALUE@281nm
Concentration of Celecoxib+Toluene
Calibration

12. The result of fourth binding:
MIP(ACN)(500 uM Celec): -2.89% ; MIP(ACN)(250 uM Celec): -6.66% ; MIP(ACN)(125 uM
Celec):3.46% ; MIP(ACN)(25 uM Celec): 1.98% ; MIP(ACN)(10 uM Celec):-11.1 %
NIP(ACN)(500uM Celec): -6.08% ; NIP(ACN)(250uM Celec): -1.04% ; NIP(ACN)(125uM Celec):-
3.86% ; NIP(ACN)(25 uM Celec): -12.87% ; NIP(ACN)(10 uM Celec):2.22 %
The result of fifth binding:
MIP(Toluene)(500 uM Celec):-9.11% ; MIP(Toluene)(250 uM Celec):-2.19% ; MIP(Toluene)(125 uM
Celec):4.65% ; MIP(Toluene)(25 uM Celec):24% ; MIP(Toluene)(10 uM Celec):-22.5%
NIP(Toluene)(500 uM Celec):-4.64% ; NIP(Toluene)(250 uM Celec):-1.51% ; NIP(Toluene)(125 uM
Celec):-10.46% ; NIP(Toluene)(25 uM Celec):-20.37% ; NIP(Toluene)(10 uM Celec):-85%
Some of the results seem to be no binding(minus ones). The results of some seems to be very
little binding. According to the results obtained we could not obtain the molecular imprinting
polymer. The reagents used may be contaminated, it may have been faulty scales weighing the
reagents. Error in measurement can be made with the NanoDrop.
REFERENCES:
1) http://www.arel.edu.tr/about-the-university/mission-and-vision
2) http://www.arel.edu.tr/about-the-university/foundation-and-campuses
3) https://en.wikipedia.org/wiki/Molecularly_imprinted_polymer
4) https://www.drugs.com/celecoxib.html
APPENDIX:
Binding Calculation:
Example: 0.045-0.044=0.001
100% 0.045 x= 2.22 % NIP(ACN)(10 uM Celec)
X 0.001