Class: BE-210
Group: W5
Members: George Wall Bell, Aqiyla Muhammad, Satyam (Tom) Sarma, Jeanette Yung
Date: April 30, 1997
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Abstract:
Hemoglobin is an important biological molecule used in the transport of respiratory gases. The absorption spectra of oxy and deoxyhemoglobin were determined using a spectrophotometer (Milton Roy Spectronic-20D) from a range of 400 nm to 700 nm. Hemoglobin was extracted from the red blood cell lysate of horse blood. Twenty-two trials of the absorption spectrum were run. The spectra were analyzed for peaks and molecular extinction coefficients. Both oxy and deoxyhemoglobin were found to have peak wavelengths at 420 nm and 540 nm. Actual literature values gave peaks for oxyhemoglobin at 420 nm, 540 nm, and 576 nm and deoxyhemoglobin peaks at 430 nm and 555 nm. The peaks for oxyhemoglobin compared well to literature values with the exception of the 576 nm peak which could not be resolved. Instead, a plateau was seen in the region from 560 nm to 580 nm. Deoxyhemoglobin peaks also corresponded well to literature values. The experimental values obtained for molecular extinction coefficients were significantly different from literature values The experimental values for the molecular extinction coefficient were: for deoxy and oxyhemoglobin (in L/(mol*cm)) at 420 nm and deoxy and oxyhemoglobin at 540 nm, 1.42 E5 + 1.37 E4, 1.68 E5 + 2.25 E3, 3.57 E4 + 9.1 E3, and 4.72 E4 + 4.39 E3, respectively. While there was a large percentage deviation from the literature values, the absorbance spectrum was consistent within itself, showing the same percentage shift in absorption values. In ten trials, oxyhemoglobin exhibited a significantly higher molecular extinction coefficient than the deoxy- samples. However, 12 trials indicated that no significant differences arose between oxy- and deoxyhemoglobin. Attempts were made to bring experimental values to literature values by purifying the hemoglobin solution. Chloroform was used to extract organic molecules from the red blood cell lysate. Absorbance spectrum were collected for the chloroform treated samples. Although it was expected that removal of organic molecules would bring the value for molecular extinction coefficient closer to literature values, the values deviated more than untreated samples.