Scribd
Upload
4K views

Introduction To Chromatography Theory

This document provides an introduction to the theory of chromatography. It discusses plate theory, which views a column as divided into theoretical plates where analyte equilibration occurs. More plates lead to better separation. Rate theory takes a random walk approach and derives the Van Deemter equation relating plate height to velocity. Resolution measures the degree of separation between peaks and depends on narrow peak width (efficiency) and large spacing between peaks (selectivity).

Uploaded by

Akash Marathakam
Copyright
© Attribution Non-Commercial (BY-NC)
Available Formats
Download as PPT, PDF, TXT or read online on Scribd
4K views

Introduction To Chromatography Theory

This document provides an introduction to the theory of chromatography. It discusses plate theory, which views a column as divided into theoretical plates where analyte equilibration occurs. More plates lead to better separation. Rate theory takes a random walk approach and derives the Van Deemter equation relating plate height to velocity. Resolution measures the degree of separation between peaks and depends on narrow peak width (efficiency) and large spacing between peaks (selectivity).

Uploaded by

Akash Marathakam
Copyright
© Attribution Non-Commercial (BY-NC)
Available Formats
Download as PPT, PDF, TXT or read online on Scribd
You are on page 1/ 18

Introduction to Chromatography

Theory

[email protected]
The Theory of Chromatography
• Plate theory - older; developed by Martin &
Synge

• Rate theory - currently in use today


Plate Theory - Martin & Synge
1954 Nobel Laureates
• View column as divided into a number (N)
of adjacent imaginary segments called
theoretical plates

• within each theoretical plate complete


equilibration of analytes between stationary
and mobile phase occurs
Plate Theory - Martin & Synge
1954 Nobel Laureates
• Significance?
Greater separation occurs with:
– greater number of theoretical plates (N)
– as plate height (H or HETP) becomes smaller

• L = N H or H = L / N
where L is length of column, N is number
of plates, and H is height of plates
N can be Estimated
Experimentally from a
Chromatogram
• N = 5.55 tr2 / w1/22 = 16 tr2 / w2
where:
tr is retention time;
w1/2 is full width at maximum
w is width measured at baseline
Choice of Column Dimensions
• Nmax = 0.4 * L/dp

where:
N - maximum column efficiency
L - column length
dp - particle size

• So, the smaller the particle size the higher the


efficiency!
Efficiency Relative to Analysis
Time
today
90 mm L
3 um
today
150 mm L
N 5 um
1970’s
300 mm L
10 um

Analysis Time, min


10 100
First Important Prediction of
Plate Theory

Bandspreading - the width of bands


increases as their retention time
(volume) increases
Problem:
• A band exhibiting a width of 4 mL and a
retention volume of 49 mL is eluted from a
column. What width is expected for a band
with a retention volume of 127 mL eluting
from the same analyte mixture on the same
column?

• ANS: 10.4 mL
Second significant prediction of
plate theory

The smaller HETP, the narrower the


eluted peak
Plate Theory - Practical
Considerations
• Not unusual for a chromatography column
to have millions of theoretical plates

• Columns often behave as if they have


different numbers of plates for different
solutes present in same mixture
Rate Theory
• Based on a random walk mechanism for the
migration of molecules through a column

• takes into account:


– band broadening
– effect of rate of elution on band shape
– availability of different paths for different
solute molecules to follow
– diffusion of solute along length
Van Deemter Equation
• H=Aν 1/3
+ B/ν + C ν

where:

H is HETP (remember want a minimum!)


ν is mobile phase velocity
A, B, and C are constants
Van Deemter Equation
• H=Aν 1/3
+ B/ν + C ν
– first term - rate of mobile phase movement
through column (often just a constant)

– second term - longitudinal solute diffusion;


solute concentration always lower at edges of
column so solute diffuses longitudinally

– third term - equilibration is not instantaneous


Resolution
• Ideal chromatogram exhibits a distinct
separate peak for each solute

• reality: chromatographic peaks often


overlap

• we call the degree of separation of two


peaks:
• resolution = peak separation
average peak width
Resolution
• Resolution = ∆ tr / wavg

• let’s take a closer look at the significance of


the problem:
Resolution
• So, separation of mixtures depends on:

– width of solute peaks (want narrow)


efficiency

– spacing between peaks (want large spacing)


selectivity
Example
• What is the resolution of two Gaussian
peaks of identical width (3.27 s) and height
eluting at 67.3 s and 74.9 s, respectively?

• ANS: Resolution = 2.32

You might also like