Hemoglobin is composed of four identical heme groups and four protein (globin) chains; each heme group contains a single iron atom that can bind to one molecule of oxygen. The protines are not the same, and most adult hemoglobin molecules contain two alpha chains and two beta chains.
In the human circulatory system, blood with low levels, called ‘deoxygenated’ blood is sent to the lungs. Adequate lung ventilation insures a high level of oxygen in the alveoli and concentration gradient down which oxygen can diffuse from the air into the blood. While some oxygen dissolves in the plasma, most (about 98%) binds with hemoglobin, clearly, hemoglobin increases the oxygen carrying capability of blood. There are three essential properties of hemoglobin that permit this respiratory pigment to act as an oxygen carrier.
1. Hemoglobin reacts reversibly with oxygen
2. The amount of oxygen bound to hemoglobin depends upon the amount of oxygen available; when in graphical form, this relationship is called the oxygen dissociation curve.
3. The amount of oxygen bound to hemoglobin depends upon factors like temperature, pH, and the concentration of 2, 3-diphosphoglycerate, which is a by product of glycolysis and becomes elevated where oxygen levels are low.
In this lab, you will construct oxygen disassociation curves for hemoglobin samples by using a spectrophotometer to measure the color of the solution at a wavelength of 620mm. Color, or more specifically the amount of light transmitted through the solution, is assumed to be an indication of the amount of bound oxygen.
The atmosphere is composed of many gases. The partial pressure of a particular gas (Pgas) can be calculated as Pgas=%gas in the air x barometric pressure
At sea level the partial pressure of oxygen (PO2) IS
If the barometric pressure is changed, the value for PO2 will also change, therefore if you reduce the pressure (by applying a vacuum, for example) PO2 will be reduced, that is, less oxygen will be available (to bind to hemoglobin). In this lab, you will apply a vacuum to a hemoglobin solution and measure its color, which indicates the amount of oxygen bound to hemoglobin. Your data will allow you to construct oxygen dissociation curves and determine the effect of pH on the affinity of hemoglobin for oxygen.