Presentation description

Extracorporeal membrane oxygenation (ECMO) is a life support technology that takes over for the heart and lungs of critically ill patients. Using ECMO has risks, ranging from excess bleeding or clotting to poor blood flow to the limbs, resulting in a high mortality rate. This high mortality rate is also because of a lack of dosing guidelines due to drug adsorption to ECMO circuit components. Drug adsorption occurs when drugs adhere to the tubing or oxygenator fibers, removing a portion of the dose from the patient's bloodstream and making it difficult to know how much medication is in circulation. To combat drug adsorption, studies must be run on ECMO circuits to determine a drug's adsorption profile. This project aimed to create a protocol for extracting and quantifying Amphotericin B (AmpB) from human plasma samples taken during ex vivo studies. AmpB was dissolved in DMSO at 10 mg/mL. Methanol (MeOH) and Acetonitrile (ACN) were used as solvents to dilute AmpB further. Firstly, the UV absorption spectrum for AmpB was evaluated, and λmax = 409 nm was determined. Further, calibration curves were obtained for AmpB in MeOH (R2=0.9892) and ACN (R2=0.9816). Human plasma samples were then dosed with AmpB at different concentrations to obtain calibration curves for AmpB in human plasma. The drug was extracted by using MeOH or ACN to crash the plasma proteins. The samples were then centrifuged at 12,000 rpm for 20 mins. The supernatant was collected and evaluated using UV spectrophotometry to quantify the amount of AmpB. Calibration curves were obtained: MeOH (R2=0.9729) and ACN (R2=0.8917). The lower detection limit (LOD) for AmpB using UV was 10 µg/mL. To further reduce the LOD, High-Performance Liquid Chromatography (HPLC) was used to measure concentrations of AmpB in MeOH. A calibration curve was obtained (R2=0.9986) with a LOD of 2 µg/mL. Herein, we successfully developed a protocol to extract AmpB from plasma samples with a high recovery rate (~90%). Future studies include quantifying AmpB from plasma samples using HPLC and performing ex vivo ECMO studies.