Transcript Summer Fellow Poster template

Ketorolac tromethamine and Zosteric acid for the Prevention
of Postoperative Pelvic Adhesions
Maureen Cheung, The University of Akron, Chemical and Biomolecular Engineering
Results or Accomplishments
Abstract
Purpose: Our long term goal is to create a polymeric microsphere encapsulated drug that can be easily applied in both open and
laparoscopic surgeries to decrease the occurrence, severity, and development of adhesions. The polymeric system offers the
opportunity to deliver drugs to promote healing, prevent infection, and manage pain.
The preliminary research will focus on the synthesis, characterization, and encapsulation of a non-toxic anti-fouling agent,
zosteric acid. The analgesic and anti-inflammatory properties of ketorolac tromethamine, a cox-2 inhibitor, will also be
explored. Among the reasons for utilizing ketorolac tromethamine are its hydrophilic nature which is similar to the
hydrophilicity of zosteric acid. Other potential candidates for encapsulation and successful adhesion prevention include statins,
ibuprofen, and other NSAIDs.
Objectives:
2
Calibration for KT in PBS at 323 nm
2.5
Sample 2 UV/VIS
1.8
1.6
2
Absorbance
Introduction: Following pelvic and abdominal surgeries, postoperative adhesions are unavoidable, occurring in 90% of major
abdominal surgery patients and in 55%-100% of women following pelvic surgery [1,2]. A reliable method to decrease pelvic
adhesions would be a useful addition to current surgical practice; preventing the need for subsequent surgeries to break the
adhesions and preventing some postoperative complications created by adhesions such as, small bowel obstructions, pain, and
infertility [1,2]. There are two clinically approved industry standard adhesion barriers. However, there efficacy and utility in
laparoscopic surgery is limited because they are both difficult to work with.
Absorbance
P# 15
Division of Surgical Research, Summa Health System
Bradford Fenton, M.D. Ph.D.; Bi-min Newby, Ph.D.; Michelle Evancho-Chapman, B.S
1.5
1
1.2
1
0.8
0.6
y = 17760x
R2 = 0.9897
0.5
1.4
0.4
0.2
0
0.E+00
0
2.E-05
4.E-05
6.E-05
8.E-05
1.E-04
Molarity KT (mol/L)
1.E-04
200
1.E-04
250
300
350
Wavelength (nm)
400
450
Figure 1. Calibration curve for ketorolac tromethamine (KT) in
phosphate buffered saline (PBS) at a wavelength of 323.
Figure 2. Representative graphical representation of UV/VIS
spectroscopy of a PLGA microsphere encapsulating KT.
KT Microsphere Yield
Loading Efficiency of KT Microspheres
1) Zosteric acid will be organically synthesized [3]. This method will be refined to ensure consistency and purity.
2) Successfully encapsulate ketorolac tromethamine into PLGA microspheres. We will use a previously established and
successful method of encapsulation [4].
80
3) Successfully encapsulate zosteric acid into PLGA microspheres.
Discussion: The successful encapsulation of ketorolac tromethamine and the successful synthesis of zosteric acid are important
steps in the development of a potential pharmaceutical agent for adhesion prevention. As is obvious from the results there is
still much ‘bench-work’ that must be done prior to in-vitro and in-vivo testing and this will be the focus of the remainder of the
summer.
Efficiency\
Result: Zosteric acid was successfully synthesized with consistency and the qualitative purity verified by mass spectrometry. A
quantitative purity will be determined by nuclear magnetic resonance spectroscopy. Ketorolac tromethamine was successfully
encapsulated into PLGA microspheres and the yield, encapsulation efficency, and total loading were determined. Zosteric acid
microspheres were synthesized but characterization is pending.
14%
70
Yield (%)
4) The microspheres will be characterized using scanning electron microscope, laser light scattering to determine size and size
distribution, and drug release rates will be determined at 37°C. Additionally, the yield, encapsulation efficiency and total
loading will be determined.
16%
60
50
40
12%
10%
8%
6%
30
4%
20
2%
10
0%
0
1% PVA 2% PVA 5% PVA 1% PVP 2% PVP 5% PVP
Figure 3. Yield of KT microspheres.
5% PVA
2% PVA 1% PVA 5% PVP 2% PVP
External aqueous phase
1% PVP
Figure 4. Loading Efficiency of KT microspheres.
Future Research: Following characterization, tissue culturing will be used to determine whether it is worth pursuing in vivo
studies. We will utilize fibroblasts as our culture cell because studies have shown that adhesion cells are very similar to
fibroblasts in both growth and connection [1].
If tissue culturing does not provide evidence of cell growth inhibition, than we will return to encapsulation step utilizing another
drug with the appropriate properties.
If tissue culturing provides evidence of cell growth inhibition and the absence of cytotoxicity, than we will pursue animal
testing. With the approval of IACUC, we will use the model developed by Summa Health System’s to determine the efficacy of
the microspheres in preventing adhesions.
Materials and Methods
Zosteric Acid Synthesis:
Figure 5. Mass spectrometry results for a representative batch of ZA. Left: Negative ion polarity. Right: Positive ion polarity.
Discussion
From Figure 5, it is concluded that ZA was successfully synthesized. Since only a qualitative purity and consistency has been established
through mass spectrometry, a quantitative assessment is needed utilizing nuclear magnetic resonance spectroscopy. At this time, ZA
microspheres have been made but assessment is pending.
Microsphere Encapsulation:
1) Dissolve drug in dH2O. Dissolve PLGA in DCM
2) Add aqeuous drug to PLGA solution. Emulsify.
3) Add emulsified solution to external aqueous phase
(PVP or PVA).
4) Allow solvent to evaporate at 1 atm with stirring.
5) Centrifuge and wash.
6) Lyphilize.
Ketorolac tromethamine was successfully encapsulated into PLGA microspheres and the yield, encapsulation efficency, and total loading
were determined. The yields and encapsulation efficiency are consistent with those expected for a hydrophilic agent such as KT.
The successful encapsulation of ketorolac tromethamine and the successful synthesis of zosteric acid are important steps in the development
of a potential pharmaceutical agent for adhesion prevention. As is obvious from the results there is still much ‘bench-work’ that must be
done prior to in-vitro and in-vivo testing and this will be the focus of the remainder of the summer.
References
[1] Liakakos T, Thomakos N, Fine PM, Dervenis C, Young RL. Peritoneal adhesions: etiology, pathophysiology, and clinical significance. Recent advances in prevention and
management. Dig Surg. 2001;18(4):260-73.
[2] El-Mowafi D, Diamon, M. Are Pelvic Adhesions Preventable? Surgical Technology International 2003;11: 222-235.
[3] Alexandratos, Spiro D. Synthesis and purification of zosteric acid. The University of Tennessee Research Corporation, assignee. Patent 5990336. 23 Nov. 1999. Print.
[4] Jain, RA. The manufacturing techniques of various drug loaded biodegradable poly(lactide-co-glycolide) (PLGA) devices. Biomaterials, 2000. 21(23):2475-90.
July 2009