BioChemistry > Lab Report > Lab Bio – 103 Lab Report 1: Spectrophotometry Determination of Concentration by Light Absorbance (All)

Lab Bio – 103 Lab Report 1: Spectrophotometry Determination of Concentration by Light Absorbance

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Lab Report 1: Spectrophotometry Determination of Concentration by Light Absorbance This experiment introduced the direct measurement approach to absorption spectrophotometry. Purpose: Recognize the... proper functioning of the spectrophotometer, acquire knowledge of the absorption of light in some compounds, understand quantitative relationships between transmittance, absorption and concentration, and to use spectrophotometric data to quantify copper (II) ion concentration is an unknown sample. Introduction: Visible light spectrum extends between 400 to 700 nm wavelength, contains all the colors of the rainbow and is the part of the electromagnetic radiation that can be perceived by the human eye. What we perceived as color is the ability of a substance to absorb and reflect light in a certain part of the visible spectrum. The amount of light absorbed by a substance is determined with the use of a spectrophotometer, which is a laboratory equipment that measures the amount of light intensity absorbed, after passing a specific wavelength through a sample solution. Spectrophotometer measurements yield light absorbance numerical values, which are used to calculate the concentration of a solution in accordance to the Beer-Lambert Law, which expresses the relationship between light absorbance and concentration in a solution A= εcl. That is, the amount of light absorbed by a solution is directly proportional to its concentration, therefore; the concentration of a solution can be determined by measuring its absorption of light at a specific wavelength and rearranging the equation: C = A εI . In addition, the concentration of test samples can also be determined by means of a standard curve, which is a graph that relates known concentrations of substances with the absorbance of each of them at the maximum wavelength. Methods: Prepare six test tubes, place only distilled water in test tube one which will blank the spectrophotometer, place standard copper sulfate (II) and water in test tubes two to five and unknown copper sulfate (II) in test tube six, following the table below: Test Tube # 1 2 3 4 5 6 Water (mL) 10.0 8.0 6.0 4.0 2.0 0 Standard copper (II) sulfate (mL) 0 2.0 4.0 6.0 8.0 - Unknown copper (II) sulfate (mL) - - - - - 10.0 Once all six test tubes are ready, the next step is to calibrate the spectrophotometer by adjusting the absorbance to zero, which means that the colorimeter is left blank. First, connect and turn on the unit, a colorimeter display should appear on the screen with transmittance values (% value) and absorbance values of the colors, red (660 nm), green (565 nm), blue (468 nm) and orange (610 nm). Then, pour the wate [Show More]

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