Please help to answer 3c, and 3d. Fluorescence spectroscopy has better detection

Question

Please help to answer 3c, and 3d.

Fluorescence spectroscopy has better detection limits than UV-Vis spectroscopy. Why? (An answer without an explained reason would carry zero points) (b) Two fluorescent compounds X and Y are present in a mixture How would you quantify the components X and Y in a mixture, give steps clearly; (i) If excitation spectrum peaks of X and Y with peaks at lambda_ex, X = 300nm and lambda_ex, Y = 350nm at lambda_cm, X = 400nm and lambda_cm, Y = -500nm (ii) If excitation spectrum peaks of X and Y with peaks at lambda_ex, x = 300nm and lambda_ex, Y = 305 nm at lambda_, X = 400nm and lambda_ Y = 500nm. The Excitation and emission spectra of quinine and anthracene arc given on the next Use only those labels shown on the plots. appropriate to your answer to refer to. emission maxima or any other spectral parameters that you would use to the at hand

Explanation / Answer

3C . Let think about anthracene first, prepare a known concentration solution (min 5 set) excite at 280 nm and collect the emission spectra. Note the emission intensity at point 8 (where quinine doesn't have emission) and plot it against concentration. Similarly, take the emission of the unknown composition of quinine and anthracene and collect at 8 and the concentration can be found from the linear plot made earlier.

Now you have the concentration of anthracene with you. (Here we can excite at point 1)

Subtract the contribution of anthracene emission at point 3, in the unknown solution.

and make a plot of quinines emission intensity Vs known concentration at 3 (As did in the case of anthracene)

From this linear graph, we will get the concentration of the quinine in the unknown mixture

3D. We know that phosphorescence is a delayed process. If we are using continuous excitation, fluorescence will come and mask the phosphorescence. Thus, for phosphorescence measurements, a pulsed excitation source is needed in order to remove the fluorescence contribution. Also, we will keep a delay time after excitation about 50-100 ns for the complete removal of the fluorescence.

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