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Chapter 25: Q43P (page 711)

(a) List ways in which the resolution between two closely spaced peaks might be changed.

(b) After optimization of an isocratic elution with several solvents, the resolution of two peaks is 1.2How might you improve the resolution without changing solvents or the kind of stationary phase?

Short Answer

Expert verified

a) The resolution between two closely spaced peaks might be changed

- by changing the fraction of each solvent

- by changing the solvent

- by changing the PH

- by changing the stationary phase

- by changing the temperature

Step by step solution

01

Resolution of elution definition

The resolution of a elution is a quantitative measure of how well two elution peaks can be differentiated in a chromatographic separation.

02

Improving the resolution  

a) The resolution between two closely spaced peaks might be changed

- by changing the fraction of each solvent

- by changing the solvent

- by changing the PH

- by changing the stationary phase

- by changing the temperature

Question: (b) After optimization of an isocratic elution with several solvents, the resolution of two peaks is How might you improve the resolution without changing solvents or the kind of stationary phase?


Answer:

To improve the resolution without changing solvents or the kind of stationary phase we can use a different temperature, longer column, slower flow rate and smaller particle size.

03

Resolution of elution definition

The resolution of a elution is a quantitative measure of how well two elution peaks can be differentiated in a chromatographic separation.

04

Improving the Resolution  

To improve the resolution without changing solvents or the kind of stationary phase we can use a different temperature, longer column, slower flow rate and smaller particle size

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Most popular questions from this chapter

a. Why is high pressure needed in HPLC?

b. For a given column length , why do smaller particles give a higher plate number?

c. What is bonded phase in liquid chromatography?

In monolithic columns60 the stationary phase is a single porous piece of silica or polymer filling the entire column and synthesized within the column from liquid precursors. Monolithic columns offer similar plate height to HPLC particles, but with less resistance to flow. Therefore, faster flow or longer columns can be used. The figure shows separation of isotopic molecules on a long monolithic column. Packed columns have too much resistance to flow to be made so long.

Separation of isotopic molecules on a 440-cm-long monolithic C18-silica column eluted withCH3CN/H2O(30: 70 vol/vol) at 308C. [Data from K. Miyamoto, T. Hara, H. Kobayashi, H. Morisaka, D. Tokuda, K. Horie, K. Koduki, S. Makino, O. Nuñez, C. Yang, T. Kawabe, T. Ikegami, H. Takubo, Y. Ishihama, and N. Tanaka, “High-Efficiency Liquid Chromatographic Separation Utilizing Long Monolithic Silica Capillary Columns,” Anal. Chem. 2008, 80, 8741.]

(a) Unretained thiourea is eluted in 41.7 min. Find the linear velocity ux (mm/s).

(b) Find the retention factor k forC6D6

(c) Find the plate number N and plate height forC6D6

(d) Assuming that the peak widths forC6H5Dand C6D6are the same as that of C6D6, find the resolution of C6H5Dand C6D6.

(f) If we just increased the column length to increase N, what value of N and what column length would be required for a resolution of 1.000?

(g) Without increasing the length of the column, and without changing the stationary phase, how might you improve the resolution?

(h) When the solvent was changed fromCH3CN/H2O(30:70 vol/vol) toCH3CN/CH3OH/H2O(10:5:85), the relative retention for C6H5D andC6D6increased to 1.0088 and the retention factor for C6H6 changed to 17.0. If the plate number were unchanged, what would be the resolution?

Suppose that an HPLC column produces Gaussian peaks. The detector measures absorbance at 254 nm. A sample containing equal moles of compounds A and B was injected into the column. Compound A E254=2.26×104M-1cm-1has a height h=128mm and a half-width w1/2=10.1mm. Compound B E254=168×104M-1cm-1has w1/2=7.6mm. What is the height of peak B in millimeters?

A bonded stationary phase for the separation of optical isomers has the structure

To resolve the enantiomers of amines, alcohols, or thiols, the compounds are first derivatized with a nitroaromatic group that increases their interaction with the bonded phase and makes them observable with a spectrophotometric detector.

When the mixture is eluted with 20 vol% 2-propanol in hexane, the (R)-enantiomer is eluted before the (S)-enantiomer, with the following chromatographic parameters:

Resolution=Δtrwav=7.7

Relative retention(α)=4.53

k for (R)-isomer =1.35

tm=1.00min

wherewavis the average width of the two Gaussian peaks at their base.

(a) Findt1,t2andwawith units of minutes.

(b) The width of a peak at half-height isrole="math" localid="1663582749865" w1/2(Figure 23-9). If the plate number for cach peak is the same, findw1/2for each peak.

(c) The area of a Gaussian peak is1.064×peakheight×w1/2. Given that the areas under the two bands should be equal, find the relative peak heights(heightR//heightS).

  1. Use equation 25-1to estimate the length of a column required to achieve1.0×104plates if the stationary phase particles size is10.5,5.0,3.0,or1.5μm

  2. If the retention time was 20mins on the 10.0μm particle size column, what is the retention time on the 5.0,3.0,or1.5μmcolumns from part (a)? Assume that flow rate is constant for all columns.

  3. Use equation25-2to estimate the pressure of the column in (a) given that the pressure of the10.0μmcolumn was4.4Mpa

  4. If the flow rate was2.0mL/min , what is the baseline width for the peaks on 10.5,5.0,3.0,or1.5μmcolumns form part (a)?

  5. Which of these column configurations would require a UHPLC instrument?
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