i. The selectivity of fluorimetry is greater than that of absorption spectrophotometry as it depends on excitation, absorption and emission spectra. As for fluorimetry, when molecule absorbs radiation and become excited, fluorimetry detects the intensity of light emitted from the molecule returns to ground state from excited state. Fluorescence is good at rigid molecules as it absorbs UV radiation. When two compounds are excited at the same wavelength radiation, they would emit different wavelength radiation as different compound has its unique spectrum. ii. Second, the sensitivity of fluorescence is greater than absorption spectrophotometry.
As for absorption spectrophotometry, the concentration is directly proportional to the absorbance. However, in fluorimetry, concentration is directly related to the luminescent radiant power which is measured against a very small background. Moreover, flourimetry is easier to measure the small difference. Disadvantages: i. Quenching is resulted in a variety of processes such as excited state reactions, energy transfer, complex formation and collisional quenching occur. This will decreases the fluorescence intensity of a given substance and affects the experimental result.
ii. Standard solutions or the fluorescent cells may be contaminated, leading to absorption some energy of the incident radiation and transmitted radiation. It will interfere the experimental results. iii. The reflection or scattering of the incident or transmitted radiation in the fluorescent cell may influence the accuracy of the intensity of the fluorescent. iv. Fluorescent substance will undergo decomposition in fluorimetry which is related to the solvent which control the pH, temperature and pressure interfering the excitation-relaxation fluorescence process.
The line will be broadened and fused together to give a less structured spectrum in the presence of solvent. 2. Define the term “quantum efficiency” for fluorescence processes. How does temperature affect quantum efficiency? “Quantum efficiency” for fluorescence processes is the ratio of the number of molecules that luminescence to the total number of excited molecules. For some species which do not have propriety of fluorescent, the quantum efficiency will approach to zero. The quantum efficiency of a highly fluorescent molecule may approach unity under certain conditions.
The quantum efficiency of fluorescence in a lot of molecules will decrease with increasing temperature. It is because the kinetic energy of the molecules and hence the frequency of collisions will be increased with increasing temperature. Some molecules will convert the excitation energy to heat instead of emitting light. So the number of molecules emits fluorescence and hence the fluorescence intensity will be decreased. Result: A calibration curve with fluorescent intensity against concentration of quinine hydrochloride in ppm was plotted by using least square method.
The least square equation of the calibration curve is y = 3629. 7x – 12. 157. From this equation, the amount of quinine hydrochloride in the sample was calculated to be 78. 645 ppm. Conclusion: The least square equation of the calibration curve was y = y = 3629. 7x – 12. 157 and by calculation, the amount of quinine hydrochloride in Schweppes tonic water was 78. 645 ppm. According to the US Food and Drug Administration (FDA), the limit of quinine content in tonic water is 83 ppm. The quinine concentration in Schweppes has not exceeded the limit.