1. Rajvi Mehta Chem 4101, Fall 2011 December 9, 2011 DEHP Leaching from PVC into Contents of Medical Devices

2. Analyte: DEHP Di-2-ethylhexyl Phthalate (DEHP) is a phthalate ester plasticizer that is used to make PVC medical devices soft and flexible. It was shown that with a longer storage time, there was a larger accumulation of DEHP in the contents of the medical devices. Large amounts of DEHP can cause a variety of symptoms and diseases, including birth defects, decreased fetal weight, miscarriages, and liver cancer, and it can also act as an endocrine disruptor. 1,2

3. Analytical Problem and Hypothesis Analytical Problem o The use of plastics for the development and fabrication of medical devices has increased recently, the most used plastic being poly(vinyl chloride) (PVC). DEHP has been leaching from the medical devices and into their contents, such as plasma and blood. 1 Hypothesis o With continued exposure to DEHP, many more adverse health effects could manifest and be detrimental to the human body.

4. Studies Needed Identify medical devices that are fabricated with PVC. Measure DEHP accumulation levels in blood stored in said medical devices over time. Measure DEHP accumulation levels in blood stored in medical devices that are (a) known to be made using DEHP, as a positive control, and (b) not made using DEHP, as a negative control.

5. Method of Choice The method of choice for this analytical problem is HPLC with a fluorescence detector. The concentration of DEHP in whole blood is, on average, 0.0238mg/mL. 3 The limits of detection for the fluorescence detector is sufficient for the detection of DEHP. Fluorescence was chosen as a detection method, even though it was not explicitly used in the literature, due to high selectivity and sensitivity. HPLC was chosen as a separation method due to shorter analysis times and better resolution, as compared with electrophoretic methods. 4,5

6. 1260 Infinity Quaternary LC System Pressure range up to 600 bar Flow rate up to 5 mL/min ZORBAX Extend –C18 Column (20mm x 2.1mm i.d.) 1.8 micron particle size 6

7. 1260 Infinity Fluorescence Detector LOD = 1.3fg S/N = 3000:1 Multi-wavelength detection to improve sensitivity and selectivity 90 o geometry helps prevent the light from the source reaching the detector 7

8. Sample Preparation This sample preparation procedure is for use in LC: Centrifugation: The whole blood was centrifuged at 4,200g for 10 minutes, making PPP (platelet-poor plasma). Extraction: 1mL of PPP was removed from PVC bags and extracted with 3mL of acetonitrile, 1mL of NaOH (1 N), and 100 μ L of an internal standard solution. Centrifugation II: Shake for 5 minutes, and then centrifuge at 4,000g for 10 minutes. The supernatant is then ready for injection onto the column. 8

9. Hypothetical Results These are HPLC chromatograms of the PPP sample: (A) blank plasma sample; (B) blank plasma sample spiked with 112.50 μ g/mL DEHP; (C) supernatant of PPP stored in bag for 14 days containing 512.55 μ g/mL DEHP. 8

10. Other Methods

11. Conclusions By analyzing samples of blood, the accumulation of DEHP in the contents of PVC medical devices over time can be ascertained, and perhaps in the future an alternative can be found. In the future, efforts should be made to study MEHP, the monoester metabolite of DEHP, and its accumulation and effect on the human body.

12. References 1. Jaeger, R.J.; Rubin, R.J. Migration of a Phthalate Ester Plasticizer from Polyvinyl Chloride Blood Bags into Stored Human Blood and its Localization in Human Tissues. New Engl. J. Med. 1972, 287, 1114-1118. 2. Mitani, K. ; Narimatsu, S. ; Izushi, F. ; Kataoka, H. Simple and Rapid Analysis of Endocrine Disruptors in Liquid Medicines and Intravenous Injection Solutions by Automated In-Tube Solid-Phase Microextraction/High Performance Liquid Chromatography. J. Pharm. Biomed. Anal. 2003, 32, 469-478. 3. Valeri, C.R. ; Contreras, T.J. ; Feingold, H. ; Sheibley, R.H. ; Jaeger, R.J. Accumulation of Di-2-ethylhexyl Phthalate (DEHP) in Whole Blood, Platelet Concentrates, and Platelet-Poor Plasma: 1. Effect of DEHP on Platelet Survival and Function. Environ. Health Perspect. 1973, 103-118. 4. Guo, B. ; Wen, B. ; Shan, X. ; Zhang, S. ; Lin, J. Separation and Determination of Phthalates by Micellar Electrokinetic Chromatography. J. Chromatogr. A. 2005, 1095, 189-192. 5. Takeda, S. ; Wakida, S. ; Yamane, M. ; Kawahara, A. ; Higashi, K. Migration Behavior of Phthalate Esters in Micellar Electrokinetic Chromatography With or Without Added Methanol. Anal Chem. 1993, 65, 2489- 2492. 6. http://www.columbia.edu/cu/chemistry/ugrad/hssp/EXP_8_files/image-23.png 7. http://s4jkuch.edu.glogster.com/fluorescence-spectroscopy/ 8. Dine, T. ; Luyckx, M. ; Cazin, M. ; Brunet, C. ; Cazin, J.C. ; Goudaliez, F. Rapid Determination by High Performance Liquid Chromatography of Di-2-ethylhexyl Phthalate in Plasma Stored in Plastic Bags. Biomed. Chromatogr. 1991, 5, 94-97.