Intro to Chromatographic Separations Chap 26. Originally based on separation and identification by color Originally based on separation and identification.

Intro to Chromatographic SeparationsIntro to Chromatographic Separations

Chap 26Chap 26

• Originally based on separation and identification by Originally based on separation and identification by colorcolor

• All have in common:All have in common:

(a)(a) Stationary phaseStationary phase

(a)(a) Mobile phase (eluent)Mobile phase (eluent)

(a)(a) Separation based on differences in migration Separation based on differences in migration rates of components of a mixturerates of components of a mixture

i.e., The extent of which components are i.e., The extent of which components are partitionedpartitioned between mobile and stationary phasesbetween mobile and stationary phases

AAmobilemobile ⇌⇌ A Astationarystationary

CHROMATOGRAPHYCHROMATOGRAPHY

Quantitative Description of Column EfficiencyQuantitative Description of Column Efficiency

► Two related terms used:Two related terms used:

(1)(1) Plate height, HPlate height, H

(2)(2) Number of theoretical plates, NNumber of theoretical plates, N

(3)(3) Related by: Related by: (Eqn. 26-16) (Eqn. 26-16)

► Efficiency increases as NEfficiency increases as N ↑↑ and as Hand as H ↓↓

HL

N

LH

2

2

R

Wt

16N

(Eqn. 26-17)(Eqn. 26-17) (Eqn. 26-21)(Eqn. 26-21)

Effect of Particle Size on Plate HeightEffect of Particle Size on Plate HeightFig. 26-11Fig. 26-11

• Smaller particles reduce H and increase resolutionSmaller particles reduce H and increase resolution

Optimization of Column PerformanceOptimization of Column Performance

Objectives:Objectives:

• reduce zone broadeningreduce zone broadening

• alter relative migration rates of solutesalter relative migration rates of solutes

Column ResolutionColumn Resolution (R (RSS) ) ≡ ≡

(Eqn. 26-24)(Eqn. 26-24)

Recall thatRecall that efficiency increases as N and as Hefficiency increases as N and as H

• lengthening column increases Nlengthening column increases N

• lower H by altering flow rate of mobile phase orlower H by altering flow rate of mobile phase or

decreasing particle size of packingdecreasing particle size of packing

BA

ARBR

WWtt

])()[(2

RS

ttRR

WW

22σσ

LH

2

2

R

Wt

16N

(Eqn. 26-17)(Eqn. 26-17) (Eqn. 26-21)(Eqn. 26-21)

Desire RS > 1.50

The General Elution Problem in ChromatographyThe General Elution Problem in ChromatographyFig. 26-15Fig. 26-15

Effect of Solvent Variation on ChromatogramsEffect of Solvent Variation on ChromatogramsFig. 26-14Fig. 26-14

Fig. 26-12Fig. 26-12

The General Elution Problem in ChromatographyThe General Elution Problem in Chromatography

Solution to the General Elution ProblemSolution to the General Elution Problem

• For liquid chromatography (LC):For liquid chromatography (LC):

• vary kvary kAA by varying composition of mobile phase during by varying composition of mobile phase during elution (elution (gradient elutiongradient elution or or solvent programmingsolvent programming))

• For gas chromatography (GC):For gas chromatography (GC):

• use temperature programminguse temperature programming

Result of Temperature ProgrammingResult of Temperature ProgrammingD

ete

cto

r S

ign

al

Gas ChromatographyGas Chromatography

Chap 27Chap 27

Schematic of a Gas ChromatographSchematic of a Gas ChromatographFig. 27-1Fig. 27-1

A Soap-Bubble FlowmeterA Soap-Bubble FlowmeterFig. 27-2Fig. 27-2

Sample Port and InjectorSample Port and Injector

• Must flash volatize andMust flash volatize and introduce sample onto columnintroduce sample onto column

• Heated ~ 50°C above BP ofHeated ~ 50°C above BP of least volatile compoundleast volatile compound

• May be:May be:

• split (cap columns & GC-MS)split (cap columns & GC-MS)

• splitless (packed columns)splitless (packed columns)

Fig 27-4

Column ConfigurationsColumn Configurations

Two types:Two types:

• Packed columns (L ~ 1 – 5 m)Packed columns (L ~ 1 – 5 m)

• Open Tubular Column (L ~ 1 – 100 m)Open Tubular Column (L ~ 1 – 100 m)

Typical Fused Silica Capillary ColumnTypical Fused Silica Capillary Column

~20 – 100 m~20 – 100 m

Fig. 27-6Fig. 27-6