Eosin Experiment

Abstract:

Once a drug (eosin) enters the body by intravenous injection, elimination begins. The kidneys enable the elimination of the drug that is present in the blood stream to occur by renal glomerular filtration. The aim for this experiment was to recreate the conditions in the kidney to see how long it takes a drug to be eliminated out the body by glomerular filtration from recording concentrations and absorbencies. Most drugs that are present inside the blood stream follow first order kinetics. This means that the natural logarithm of concentration against time is directly proportional to one another. By using a spectrophotometer, the absorbencies were calculated with a 516nm wavelength. This enabled us to understand the relationship

Eosin is an orange-pink dye that is used to stain tissues in the body while looking through a microscope (Bracegirdle, 2002). We mimicked the way in which blood flowed through the body by using a reservoir as the blood supply, attached to that, a plastic tubing with a clamp, which regulated the flow rate to which blood flowed in the body. This water running through the tubing, was caught in a large beaker where the eosin was added and further more being released into a smaller beaker. The way in which this experiment has been set up is similar to that of the clearance of a drug by glomerular filtration in the kidney. Glomerular filtration is the renal process whereby fluid in the blood is filtered across the capillaries of the glomerulus and urinary space of the bowman’s capsule (Mosby, 2009). By using this experimental technique, we aim to recreate the elimination conditions in the kidney by working out the absorbance of samples at different time intervals and therefore use that to find the final concentration of the drug left in the blood system after a 60 minute time period. A calibration graph is draw up to help us see how the drug concentration in the blood stream over the time interval was affected. Absorbance is the measure of a quantity of light being absorbed by a sample and is measured by a spectrophotometer. A

Start the timer for 1 minute, and using a measuring cylinder, collect an outflow of 17-23 ml min-1. Record this initial flow rate. With this flow rate, make sure the large beaker is filled to the top with water and place the magnetic stirrer inside. Using a piece of rolled tissue, insert it into the sprout positioned above the smaller beaker. Turn on the magnetic turner and remove 2 milliliters of water and place it in a cuvette using a 5-milliliter autopipette. This is the blank. Now add 2 milliliters of 2,5mg.ml-1 eosin dye into the large beaker. Once added, start the timer and at times: 1, 2.5, 5, 10, 20, 30, 40 and 60 minutes, 2 milliliters of the sample is removed from the large beaker and put in a clean cuvette and then 2 milliliters of distilled water is added back. At times 24 and 60 minutes, using new beaker, collect the volume for 2 minutes. Use this to determine you mid-and-end-experimental flow rates. Using the samples in the cuvettes, calculate the absorbance by inserting them into the spectrophotometer. Record these results and use them to calculate the concentration of the drug left in the blood, by using the calibration curve. To construct a calibration curve, make up solutions of Eosin and water in seven separate tubes using table 1 and calculate the absorbance of each. Plot the calibration curve with this absorbance vs. the final