(a) Explain why fluorescence spectroscopy gives a more accurate determination of vitamin B1 than absorption spectroscopy. (b) Provide an annotated sketch of the excitation/emission spectrum as well as an annotated energy profile diagram of the vitamin B1 derivative, thiochrome.

Curren'S Math For Meds: Dosages & Sol
11th Edition
ISBN:9781305143531
Author:CURREN
Publisher:CURREN
Chapter13: Dimensional Analysis/units Conversion
Section: Chapter Questions
Problem 28SST
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Part 1
Since the UV-Vis method outlined in Question 1 above is not a standard method of analysis of the vitamins you
will have to compare your UV-Vis method to a standard HPLC or GC method. In your reading you found that
although vitamin B1 absorbs UV light at 240 nm, it is best converted to thiochrome by the scheme shown in Figure
2 before analysis by HPLC with fluorescence detection, using an excitation and emission wavelengths of 360 nm
and 450 nm, respectively.
он
он
NH:
K, Fe(CN,)
NaOH
Figure 2: Oxidation of vitamin B1, thiamine to thiochrome.?
(a) Explain why fluorescence spectroscopy gives a more accurate determination of vitamin B1 than
absorption spectroscopy.
(b) Provide an annotated sketch of the excitation/emission spectrum as well as an annotated energy profile
diagram of the vitamin B1 derivative, thiochrome.
Transcribed Image Text:Part 1 Since the UV-Vis method outlined in Question 1 above is not a standard method of analysis of the vitamins you will have to compare your UV-Vis method to a standard HPLC or GC method. In your reading you found that although vitamin B1 absorbs UV light at 240 nm, it is best converted to thiochrome by the scheme shown in Figure 2 before analysis by HPLC with fluorescence detection, using an excitation and emission wavelengths of 360 nm and 450 nm, respectively. он он NH: K, Fe(CN,) NaOH Figure 2: Oxidation of vitamin B1, thiamine to thiochrome.? (a) Explain why fluorescence spectroscopy gives a more accurate determination of vitamin B1 than absorption spectroscopy. (b) Provide an annotated sketch of the excitation/emission spectrum as well as an annotated energy profile diagram of the vitamin B1 derivative, thiochrome.
Part 2
Since the HPLC is down, your company would have to outsource the fluorescence method outlined in Part 1
above to one of its competitors which may be detrimental to your place of employment and your job in the future.
Upon further investigation, you found a GC method for analysis of vitamin B1; and fortunately, the GC is fully
functional. Both of the 2Bs vitamins are highly water soluble. In order to quantify vitamin B1 in the supplement,
you must first convert thiamine to the less polar derivative, 4-methyl-5-(2-hydroxyethyl)thiazole, shown in Figure
3 below, with subsequent extraction and cleanup, before GC analysis. The procedure yields the derivative in high
purity (~90%), provided a suitable extraction and cleanup procedure is followed.
H3C
HO
Figure 3: Structure of 4-methyl-5-(2-hydroxyethyl)thiazole
(c) Describe a suitable extraction and cleanup procedure for the GC method described in Part 2 above.
In your response include justifications for the extraction solvent chosen.
(d) The derivative mixture was analyzed by GC on a 10m capillary column. The solvent eluted after 75
seconds. The derivative and a second unknown peak eluted at 407 seconds and 420 seconds,
respectively. The width of the peaks at half height for the derivative and unknown are 8 seconds and
10 seconds, respectively.
(i)
Calculate:
the column resolution.
the number of theoretical plates required for a resolution of 1.5
(ii)
Without changing the GC column, explain how you can adjust your GC method to improve
separation of the two compounds in (d) above.
Transcribed Image Text:Part 2 Since the HPLC is down, your company would have to outsource the fluorescence method outlined in Part 1 above to one of its competitors which may be detrimental to your place of employment and your job in the future. Upon further investigation, you found a GC method for analysis of vitamin B1; and fortunately, the GC is fully functional. Both of the 2Bs vitamins are highly water soluble. In order to quantify vitamin B1 in the supplement, you must first convert thiamine to the less polar derivative, 4-methyl-5-(2-hydroxyethyl)thiazole, shown in Figure 3 below, with subsequent extraction and cleanup, before GC analysis. The procedure yields the derivative in high purity (~90%), provided a suitable extraction and cleanup procedure is followed. H3C HO Figure 3: Structure of 4-methyl-5-(2-hydroxyethyl)thiazole (c) Describe a suitable extraction and cleanup procedure for the GC method described in Part 2 above. In your response include justifications for the extraction solvent chosen. (d) The derivative mixture was analyzed by GC on a 10m capillary column. The solvent eluted after 75 seconds. The derivative and a second unknown peak eluted at 407 seconds and 420 seconds, respectively. The width of the peaks at half height for the derivative and unknown are 8 seconds and 10 seconds, respectively. (i) Calculate: the column resolution. the number of theoretical plates required for a resolution of 1.5 (ii) Without changing the GC column, explain how you can adjust your GC method to improve separation of the two compounds in (d) above.
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