BIO202H5S: Oxygen Dissociation Curves & Bohr Effect Lab Report

This lab report investigates the effect of varying partial pressure of oxygen (PO2) on the percentage of oxygen saturation of haemoglobin and the influence of different pH levels on the haemoglobin-oxygen affinity of deoxygenating sheep blood. The experiment involves recording percentage transmittance using a spectrophotometer at different PO2 levels and pH values of 6.8 and 7.4. Standard curves are constructed to relate transmittance to haemoglobin saturation, and oxygen dissociation curves are plotted to illustrate the relationship between PO2 and haemoglobin saturation. The results indicate that pH affects the oxygen-binding affinity of haemoglobin, demonstrating the Bohr effect, with higher pH values generally leading to increased oxygen saturation at a given PO2. Desklib provides access to this and other solved assignments for students' reference.

Running head: LAB REPORT ON ANIMAL PHYSIOLOGY
Studying the effect of changing PO2 on % oxygen saturation of Haemoglobin and
observe the varying pH’s effect on Haemoglobin- oxygen affinity of deoxygenating
sheep (Ovis aries) blood.
Name of the Student
Name of the University
Author’s Note:

1LAB REPORT ON ANIMAL PHYSIOLOGY
Introduction:
Every living animal needs oxygen to survive and fast enough circulation of oxygen to
sustain bodily actions. In order to achieve fast enough oxygen supply, animals have
developed dedicated organs (such as lungs) for oxygen diffusion as oxygen diffusion does not
remain effective if the living tissues become more than 1 mm thick (Gillooly et al., 2016).
Although, developing of organ is not enough, as it also need blood pigment to successfully
transport oxygen to the cells. Haemoglobin (Hb) present in the vertebrate’s red blood cells
are act as the transportation molecule (Liu et al., 2016). This Haemoglobin (Hb) transfer
oxygen to the tissues and cells after absorbing oxygen from lungs while returning carbon-di-
oxide from tissues and cells to lungs. The iron ion present in Haemoglobin (Hb) is
responsible for the releasing and binding of carbon-di-oxide and oxygen (Liu et al., 2016).
The relationship between oxygen and haemoglobin can be expressed through Oxygen
Disassociation Curve where percentage of oxygen bound to haemoglobin is illustrated at
various partial pressure (PO2) (Collins et al., 2015). P50 is used as a traditional measurement
standard for the healthy individuals and can be defined as partial oxygen saturation at 50 per
cent saturation. This oxygen affinity of the haemoglobin varies due to various reason and
decrease and increase of pH is one of them. Low oxygen partial pressure and increment of
carbon-di-oxide is responsible for decrement in oxygen affinity and vice versa. This
phenomenon is known as Bohr Effect (Jensen, Storz & Fago, 2016). Many studies have
investigated the Bohr Effect in animal and reported that an inversely related Bohr Effect is
present between pH and oxygen affinity (Okonjo, 2018).
Therefore, the purpose of this experiment is to study the effect of changing PO2 on %
oxygen saturation of Haemoglobin and observe the varying pH’s effect on Haemoglobin-
oxygen affinity of deoxygenating sheep (Ovis aries) blood. In order to achieve this %
transmittance will be recorded using spectrophotometer at different variation of PO2.

2LAB REPORT ON ANIMAL PHYSIOLOGY
Results:
A standard curve was constructed by plotting % transmittance against % of Hb
saturation at two different pH 6.8 and 7.4. Vacuum pressure was obtained using a Manometer
and percentage of transmittance was collected using spectrophotometer from deoxygenating
sheep (Ovis aries) blood. The constructed standard curve is presented below in the Figure 1.
0 20 40 60 80 100 120
0
10
20
30
40
50
60
f(x) = 0.54 x + 0.739999999999998
R² = 0.965689796593142
f(x) = 0.538 x + 2.4
R² = 0.970267530545037
pH 6.8
Linear (pH
6.8)
pH 7.4
Linear (pH
7.4)
Hb Saturation (%)
Transmittance (%)
Figure 1: This figure presents the constructed standard curve of haemoglobin at pH 6.8 and
7.4.
From the above graph two standard equation have been obtained for each of the pH 6.8 and
7.4. These equation are mentioned below:
For pH 6.8, Y = 0.538X + 2.4
For pH 7.4, Y = 0.54X + 0.74 where X= % of Hb saturation and Y= % transmittance.
The above equations were used to convert the % Transmittance to % of Hb saturation.

3LAB REPORT ON ANIMAL PHYSIOLOGY
Manometer readings were converted to Partial Pressure of Oxygen using the following
equation:
Partial Pressure O2 (mmHg) = 0.21(D – W – M); where W = water vapour pressure in mmHg,
D= barometer pressure in mmHg and M = vacuum pressure in mmHg.
0.00 20.00 40.00 60.00 80.00 100.00 120.00 140.00 160.00
0.00
20.00
40.00
60.00
80.00
100.00
120.00
Hb Saturation (%) pH 6.8
Linear (Hb Saturation (%) pH
6.8)
Hb Saturation (%) pH 7.4
Linear (Hb Saturation (%) pH
7.4)
Partial Pressure of Oxygen (mmHg)
Hb Saturation (%)
Figure 2: This figure presents the Oxygen Disassociation Curve of Hb at pH 6.8 and 7.4. It
depicts the relationship between Partial Pressure of Oxygen and percentage saturation of Hb.
From the above figure, it can be observed that the Oxygen Disassociation Curve for
both pH produces same type of sigmoid curve. However, the curve for pH 7.4 deviates a lot
whereas the curve for pH 6.8 follows the similar trend. From the figure, it can be established
that the percentage of oxygen saturation in haemoglobin is higher in case of pH 7.4 in
comparison with pH 6.8. This is also true for the value percentage of oxygen saturation in
haemoglobin at P50. In a nutshell, it can be said that the both the curve follows the similar
trends despite difference in value.

4LAB REPORT ON ANIMAL PHYSIOLOGY
References:
Collins, J. A., Rudenski, A., Gibson, J., Howard, L., & O’Driscoll, R. (2015). Relating
oxygen partial pressure, saturation and content: the haemoglobin–oxygen dissociation
curve. Breathe, 11(3), 194-201.
Gillooly, J. F., Gomez, J. P., Mavrodiev, E. V., Rong, Y., & McLamore, E. S. (2016). Body
mass scaling of passive oxygen diffusion in endotherms and ectotherms. Proceedings
of the National Academy of Sciences, 113(19), 5340-5345.
Jensen, B., Storz, J. F., & Fago, A. (2016). Bohr effect and temperature sensitivity of
hemoglobins from highland and lowland deer mice. Comparative Biochemistry and
Physiology Part A: Molecular & Integrative Physiology, 195, 10-14.
Liu, Y., Gong, J., Wu, W., Fang, Y., Wang, Q., & Gu, H. (2016). A novel bio-nanocomposite
based on hemoglobin and carboxyl graphene for enhancing the ability of carrying
oxygen. Sensors and Actuators B: Chemical, 222, 588-597.
Okonjo, K. O. (2018). Bohr effect and oxygen affinity of carp, eel and human hemoglobin:
Quantitative analyses provide rationale for the Root effect. Biophysical
chemistry, 242, 45-59.