Download - Theories of chromatography
There are two theories to explain chromatography
Plate theory - older; developed by Martin & Synge in 1941
Rate theory - currently in use Proposed by van Deemter in 1956 Accounts for the dynamics of the separation
View column as divided into a number (N) of adjacent imaginary segments called theoretical plates Within each theoretical plate analyte(s)
completely equilibrate between stationary phaseand mobile phase
ColumnTheoretical plate
Chromatographic principle
The molecules of the mixture interact with the molecules of the Mobile
and Stationary Phase
Retardation of rate of movement of
molecules
Each molecule interacts differently
with MP and SP
Different distribution coefficients and different
net rates of migration
Stationary phase
Mobile phaseSamplemixture Equilibrium
establishes at each point (ideally)
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 or height equivalent to theoretical plate (HETP)
The number of theoretical plates that a real column possesses can be found by examining a chromatographic peak after elution by various methods like
- half-height method- USP method
•N = 5.55 tR2/ w1/2
2 = 16 tR2/ w2
where: tR is retention timew1/2 is width at h0.5 w is width measured at baseline
N is a ratio of tR and σ of Wb which is 4σ
Where: N = Number of theoretical platesVe = elution volume or retention time (mL, sec, or cm)h = peak heightw1/2 = width of the peak at half peak height (mL, sec, or cm)
• Nmax = 0.4 * L/dp where:Nmax - maximum column efficiencyL - column lengthdp - particle size
• So, the smaller the particle size the higher the
efficiency!
Band spreading - the width of bandsincreases as their retention time (tR) or retention volume (VR) increases
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
The smaller HETP, the narrower the eluted peak
• It is 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