UF CHM 3120L - Lab 6: Fluorescence Determination of Quinine

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Lab 6: Fluorescence Determination of Quinine CHM3120LNovember 4, 2021IntroductionQuinine is a widely used compound in flavoring substances, such as tonic water. It is found in the bark of the cinchona tree and was originally developed as a medicine to fight against malaria.The structure of quinine includes phosphors, which gives it the ability to re-emit absorbed energyas fluorescent light. For this reason, the concentrations of quinine in tonic water and in Cinchona bark can be determined utilizing fluorescence spectroscopy. 1 In order to measure quinine concentrations, one can utilize its unique physical property of molecular fluorescence, which is the emission of light by molecules upon excitation with a light of shorter wavelength, or high energy. The molecule that absorbs energy is originally in the ground-state, then gets excitation by the absorption of radiation, and then goes to the ground state again. The radiation emitted from this change in states appears at a long wavelength in the observed fluorescence. These measurements are highly selective, meaning that it is very specific to the molecule, which results in minimal interference from other molecules present in the solution. Along with this, fluorescence is a very sensitive method, suggesting that it can measure very low concentrations of a substance. Compared to absorption measurements, fluorescence detects smaller changes because it measures light intensity against a dark background rather than a high background on absorption. 2The method of standard addition is utilized by adding small known amounts to the sample being analyzed. The emission spectrum is recorded to find the intensity after each addition. By adding small known concentrations of quinine in several samples of known concentrations of a stock solution of quinine sulfate, a plot of fluorescent intensity against amount of quinine in the standard addition can be created in order to determine the unknown concentration at a specific fluorescence emission. This method is used in this experiment in order to determine the concentration of quinine in various tonic water samples and in a sample of quinine extracted from cinchona bark.ExperimentalPart1:100 μL of 1.0 M sulfuric acid, 1.00 mL of tonic water 1 (Canada Dry), and 1.00 mL of DI water were added and mixed in a clean cuvette using a micropipette. The cuvette was placed into the fluorescence spectrometer and 5 readings of the background intensity were recorded at a wavelength of 771.281 nm. The mean and standard deviation of these 5 values was calculated. Then, the cuvette was spiked with eight 50 μL spikes of a 0.4 mM quinine sulfate stock solution. The intensity was recorded 5 times at 440 nm for the solution along with each spike and the average and standard deviation were calculated for each spike of quinine sulfate. These values were inserted into a standard addition plot with a linear regression line in order to calculate the intersection at the abscissa.The same procedure was repeated for Tonic Water samples 2,3, and 4 (Schweppes, Publix and Seagram, respectively). The only different step for these samples was recording the net intensity,as it was done at different wavelengths. These values were recorded at 718.036 nm for Tonic 2, 800.297 nm for Tonic 3, and 694.517 nm for Tonic 4. Part 2: 0.2006 g of powdered Cinchona bark was precisely weighed and added to 100 mL of water, which was set to gently boil for 30 minutes. Then, this solution was left to sit for a bout 15 minutes to cool down and allow the solids to settle onto the bottom. A 1 mL aliquot of this solution, along with 100 μL of 1.0 M sulfuric acid, and 1.00 mL of DI water were added and mixed in a clean cuvette using a micropipette. The cuvette was placed in a fluorescence spectrometer and the background was recorded at 726.67 nm. Then, the intensity of the sample was recorded 5 times, along with the intensity after eight 50 μL spikes of quinine sulfate solutionwas added at 440 nm. The spectrometer was set at 10 scans to average, 5 msec data upate rate, and an integration time of 100 ms. The mean and standard deviation were calculated, and the intensity and concentrations of quinine were plotted in a graph. The concentration of quinine wasfound using the linear regression equation and then converted to wight percent.ResultsPart 1:M quinineadded0 1.86E-05 3.64E-05 5.33E-05 6.96E-05 8.51E-05 0.0001 0.00011 0.000131985.43 2323.54 2552.58 2770.89 2858.73 2960.13 3049.21 3150.26 3226.381984.02 2357.12 2552.08 2754.8 2894.71 2979.53 3066.52 3129.16 3238.391985.95 2344.45 2562.75 2762.83 2908.1 2985.7 3053.44 3133.48 3230.961987.59 2353.1 2564.05 2752 2897.74 2974.75 3052.01 3131.55 3222.511985.89 2355.6 2563.79 2752.9 2900.22 2975.24 3070.74 3153.47 3227.16mean 1985.78 2346.762 2559.05 2758.684 2891.9 2975.07 3058.38 3139.58 3229.08stdev 1.27839 13.876016 6.15628541 8.05282124 19.1957873 9.43694071 9.59295 11.3717 6.00732Table 1: Measurement of mean and standard deviation for the net intensity readings for Tonic1 (Canada Dry) sample.Figure 1: Standard Addition plot of quinine concentration in molarity vs intensity in Tonic 1 (Canada Dry)Table 2: Measurement of mean and standard deviation for the net intensity readings for Tonic2 (Schweppes) sample.μL of quinine added0 50 100 150 200 250 300 350 400M quinine added0 1.86E-05 3.64E-05 5.33E-05 6.96E-05 8.51E-05 0.0001 0.00011 0.000132144.56 2536.66 2470.8 2841.02 3021.08 3084.22 3157.67 3272.2 3286.752147.06 2530.57 2463.85 2843.67 3005.22 3056.62 3158.99 3280.89 3286.172152.25 2535.69 2461.77 2847.79 3008.22 3055.12 3163.09 3282.58 3272.042139.91 2527.45 2461.7 2850.6 3018 3065.85 3151.22 3272.47 3286.892147.4 2527.47 2465.73 2833.31 3016.67 3055.98 3158.01 3262.53 3275.34Mean 2146.24 2531.568 2464.77 2843.278 3013.838 3063.558 3157.8 3274.13 3281.44stdev 4.50081 4.40644074 3.76011303 6.68174902 6.77533911 12.3382868 4.2649 8.03338 7.17359Figure 2: Standard Addition plot of quinine concentration in molarity vs intensity in Tonic 2 (Schweppes)Table 3: Measurement of mean and standard deviation for the net intensity readings of Tonic 3 (Publix)µL of quinine added0.00 50.00 100.00 150.00 200.00 250.00 300.00 350.00 400.00M quinine added0 1.86047E-05 3.64E-05 5.33E-05 6.96E-05 8.51E-05 0.0001 0.000114 0.0001281654.34 1834.97 2153.89 2380.41 2327.23 2540.17 2705.94 2664.51 2750.71691.82 1829.95 2161.64 2381.67 2351 2508.1 2703.08 2653.64 2727.221657.94 1837.22 2133.36 2382.54 2359.18


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UF CHM 3120L - Lab 6: Fluorescence Determination of Quinine

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