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Quantitative Fluorescence

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    1. The concentration of acetylsalicylic acid, C9H8O4, in aspirin tablets can be determined by hydrolyzing to the salicylate ion, C7H5O2–, and determining the concentration of the salicylate ion spectrofluorometri­cally. A stock standard solution is prepared by weighing 0.0774 g of salicylic acid, C7H6O2, into a 1-L volumetric flask and diluting to volume with distilled water. A set of calibration standards is prepared by pipeting 0, 2.00, 4.00, 6.00, 8.00, and 10.00 mL of the stock solu­tion into separate 100-mL volumetric flasks containing 2.00 mL of 4 M NaOH and diluting to volume with distilled water. The fluorescence of the calibration standards was measured at an emission wavelength of 400 nm using an excitation wavelength of 310 nm; results are listed in the following table.

      mL of stock solution

      emission intensity













      Several aspirin tablets are ground to a fine powder in a mortar and pestle. A 0.1013-g portion of the powder is placed in a 1-L volumetric flask and diluted to volume with distilled water. A portion of this so­lution is filtered to remove insoluble binders and a 10.00-mL aliquot transferred to a 100-mL volumetric flask containing 2.00 mL of 4 M NaOH. After diluting to volume the fluorescence of the resulting solu­tion is found to be 8.69. What is the %w/w acetylsalicylic acid in the aspirin tablets?


      1. You are determining the presence and concentration of cytokeratin, a tumor markers circulating in the blood of lung cancer patients, using a fluorescence assay you developed. After running your standard cytokeratin samples (concentrations in μM) to construct an external calibration curve you obtain a straight line with an R2 of 0.9884 and an equation of y=14591x + 204. You check the first patients blood sample and observe a fluorescence intensity of 56,643 RLU. What is the cytokeratin concentration in your patients blood?







      2. To make sure your results are accurate you decide to spike the sample with cytokeratin to confirm your results. You add cytokeratin to add a final concentration of an additional 2 μM to the sample from part a. You check the fluorescence again and the result is 73,188 RLU. Can you trust your original measurement of cytokeratin? Why or why not? How could you determine the actual concentration in your original unknown solution?







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