Absorbance and Spectrophotometry

Experiment 2 Absorbance and Spectrophotometry ABSTRACT This was an investigation into the effect of different wavelengths of catch fire on methylene group group radical radical radical game and violent personnel casualty on the absorbance protect on a spectrophotometer. A spectrophotometer is practised to stripe swinging intensity by emitting a single bring tidy sum author done a cuvette of colou reddish resolving. The particles in the ancestor, which are tinted, absorb the light depending on how concentrated it is and this produces an electronic fileing from the photometer which is the absorbance value.The level best preoccupancy was found for both ascendants and was used to calculate the mill extinction coefficient of methylene grisly. An dark closeness of methylene savoury was calculated by utilize graphs produced in the dilution experiments prior. The results produced supported Beers Law because the absorbance was directly proportional to the immers ion, and so, we croupe be assured that the stringency of the unknown methylene depressed solution calculated is relatively accurate. INTRODUCTION A spectrophotometer is used to measure the absorbance of light by coloured solutions.The absorbance value is produced by a photometer that compares the light detected with a blank cuvette (a cuvette containing just urine/clear neutral solvent, which should be 0), with the amount of light detected with a test solution in this case, methylene blue or carmine red. Using Beers Law, we know that the absorbance is directly proportional to the concentration, therefore, knowing the absorbance of a solution rat be very useful as the concentration of the solution fuel be find by substituting known values into the equating Absorbance = k c t Where k = constant c = concentration of absorbing jettyecules = thickness of absorbing layer The aims of this experiment were to use solutions methylene blue and carmine red to confirm that Beers Law is authorized by finding the utter more or less preoccupancy value for each solution, and wherefore using this, find the ducking of methylene blue solution at various dilutions. By plotting these results on a calibration curve (concentration against absorbance), this allows the experimenter to read the concentration at a particular absorbance directly, such as the unknown concentration of methylene blue. METHOD A spectrophotometer was used throughout this experiment. RESULTS afterwards finding the ingress for 0. 005% methylene blue solution and 0. 0005% carmine red solution at different wavelengths of light, we plotted a graph to certify our findings to make it easier to render what region of wavelength the maximum absorption would occur at. divert refer to type 1. From this graph, we locoweed see that the maximum absorption for methylene blue is nigh 650nm-675nm as the peak on the hound for methylene is around these values for carmine red, we can see that the maximum abso rption for carmine red is 500-550nm. To obtain a more accurate wavelength value, I placed more cuvettes of methylene blue and carmine red around their regions of maximum absorption.After finding the absorption values around each region, I plotted the findings of each solution on separate graphs to show the maximum absorption value. protrude 2 shows that the maximum absorption of methylene blue is 665nm because this has the peak absorption of 0. 965. However, this is not as accurate a value as it could be because the spectrophotometer did not go to more accuracy than 5nm. From class 3, we can see that the maximum absorption of carmine red is 0. 207 at wavelength 520nm as this is the peak on the graph. Maximum Absorption at 0. 0005% methylene radical vipers bugloss Carmine Red 0. 965 0. 207 After we found the maximum absorption for methylene blue, 0. 965, at 665nm, we made up various dilutions of methylene blue and put each solution through the spectrophotometer at wavelength 6 65nm to find the molar extinction coefficient. I plotted these results on a graph (figure 6) and did the line of best fit through the points to find the gradient, which is the molar extinction coefficient. Figure 4. Graph showing dilutions of methylene blue and the absorptions each solution givesThe char line on figure 4 represents the regression line. We can use this to find the concentration of the unknown concentration of methylene blue solution by drawing a tangent to the regression line at absorbance 0. 262 (where the unknown absorbed) and reading down from that point on the graph to the concentration. The concentration of the unknown methylene blue is 4. 4 x 10-6 mol dm-3. We can find the molar extinction coefficient by substituting values of absorbance and the concentration of the unknown concentration of methylene blue into Beers laws equation.Absorbance = k c t k = absorbance / c t k = 0. 262 / 4. 4 x 10-6 x 1 k = 59545 mol dm-3 cm-3 Therefore, k, the molar extinction coef ficient is 59545 mol dm-3 cm-3. DISCUSSION The main objectives of this experiment was to find the unknown concentration of methylene blue by using a spectrophotometer. I found the maximum absorption for methylene blue and carmine red (please refer to figure 1) and using this I determined a more accurate maximum absorption value for each solution by taking shape up readings around the peak of each line to determine the maximum.However, the findings of maximum absorption for methylene blue and carmine red may not be as accurate as we think because there are outdoor(a) changeables that we can not necessarily control. One is that the outside of the cuvette may watch been dirty (however, this was controlled to an extent as I wiped each side down of the cuvette with a paper towel before placing it in the spectrophotometer) another variable is that the dial on the spectrophotometer only measured in wavelength intervals of 5nm, and so, we could not desexualize more accurate readings th an the ones we concluded with.From figure 1, we can also see that high (maximum) absorptions for carmine red occurs at around 475nm-550nm. This is because the light absorbs most light at this wavelength, and therefore, reflects light at approximately 675nm-725nm which are the wavelengths of the colour red, so we see red solution. The same can be employ to methylene blue solution because we can see from figure 1 that high absorptions for methylene blue occurs around 600nm-675nm the light absorbs most colours at this wavelength and reflects light at approximately 400nm-450nm which are the wavelengths of the colour blue, so we see blue solution.We could use the maximum absorption of methylene blue found to make dilutions of methylene blue with water to plot a graph proving that Beers Law is true that the absorbance is directly proportional to the concentration. This is confirmed by the graph produced as the line of best fit is accurate and goes through the origin. APPENDIX Finding the maximum absorbance Wavelength/nm Absorption methylene group Blue Carmine Red 350 0. 33 0. 156 375 0. 015 0. 018 400 0. 015 0. 046 425 0. 018 0. 048 450 0. 006 0. 127 475 0. 029 0. 093 500 0. 041 0. 65 525 0. 040 0. 186 550 0. 077 0. revenue 575 0. 186 0. 068 600 0. 476 0. 039 625 0. 622 0. 028 650 0. 800 0. 005 675 0. 95 0. 013 700 0. 102 0. 004 much accurate values of methylene blue to a greater extent accurate values of carmine red Methylene Blue Wavelength/nm Absorption 630 0. 623 640 0. 679 655 0. 885 660 0. 929 665 0. 965 670 0. 913 Carmine Red Wavelength/nm Absorption 510 0. 205 515 0. 204 520 0. 207 530 0. 191 540 0. 169 Table to a lower place shows the dilutions and the absorbance values of methylene blue at 665nm Tube Water Methylene Blue (ml) Absorption Concentration of methylene blue in water/mol dm-3 1 41 0. 171 3. 13 x 10-6 2 32 0. 376 6. 26 x 10-6 3 23 0. 595 9. 9 x 10-6 4 14 0. 762 12. 51 x 10-6 5 05 0. 963 15. 64 x 1 0-6 Blank 50 0. 000 0 Unknown solution absorbance 0. 262 Formula mass of methylene blue 319. 6 Working out concentration of methylene blue from % 1. 0. 0001% methylene blue so, 100/0. 001 = 1000000 so, 1/1000000 = 1 x 10-6 g cm-3 so, conc. = 1 x 10-6 g cm-3 / 319. 6 g mol-1 = 3. 13 x 10-6 mol dm-3 2. (3. 13 x 10-6) x 2 = 6. 26 x 10-6 mol dm-3 3. (3. 13 x 10-6) x 3 = 9. 39 x 10-6 mol dm-3 4. (3. 13 x 10-6) x 4 = 12. 51 x 10-6 mol dm-3 5. (3. 13 x 10-6) x 5 = 15. 64 x 10-6 mol dm-3 Figure 1. Methylene blue and carmine reds absorption at regular intervals of wavelengths Figure 2. to a greater extent accurate wavelengths to find the max. absorption for methylene blue Figure 3. More accurate wavelengths to find the max. absorption for carmine red