Chromatography (Contd.

The concept of visualizing a packed column as a stack of discrete theoretical plates (each of which
represents an ideal equilibrium stage) common in chemical engineering applications, is also extended to
chromatography. Thus, for N number of theoretical plates in a column of length L, the Height
Equivalent to a Theoretical Plate (HETP), H, is given by

H L/N

The concept of theoretical plates generally does not give an accurate representation of the conditions in a
chromatographic column, since equilibrium is seldom attained in chromatography. However, the
pertinence of the HETP concept lies in the fact that H is proportional to band broadening lower the
value of H, lower the base width and narrower (and sharper) is the solute peak, resulting in higher
resolution. An empirical equation was obtained by van Deemter et al [xx] correlating H to the mobile
phase velocity, u, which explicitly considers the finite time taken for the solute to attain equilibrium
between the stationary and mobile phases (unlike the plate model, which assumes that equilibration is
infinitely fast):

B
H A Cu
u

where parameters A, B and C respectively incorporate band broadening effects due to eddy diffusion,
axial (longitudinal) diffusion and mass transfer resistance. The eddy diffusivity parameter, A, depends on
the variability of path length followed by the mobile phase and solute molecules through the stationary
phase they take different tortuous paths at random through the stationary phase. This results in base
widening of the solute peak as different paths are of different lengths. Smaller the packing particle size
and narrower the particle size distribution, smaller the value of A. Higher the mobile phase velocity, lesser
the time spent by solute inside the column, and correspondingly lower the effects of longitudinal
diffusion, quantified by the second term in the van Deemter equation. The third term in the van Deemter
equation incorporates the effects of diffusional mass transfer resistance, due to which the solute takes a
certain amount of time to equilibrate between the stationary and mobile phases. Now, if the velocity of the
mobile phase is high, and the solute has a strong affinity for the stationary phase, then the solute in the
mobile phase will move ahead of the solute in the stationary phase with resultant band broadening. Higher
the velocity of mobile phase, higher the broadening and lower the effectiveness of separation.

The fact that the base width of a chromatographic peak depends on the rate of elution, as well as on the
different alternative paths available to the solute molecules as they travel through the stationary phase, is
taken into account in the van Deemter model. From a practical point of view, the van Deemter model
emphasizes that the effectiveness of separation, of which H is a direct measure, is a function of the
process throughput, which directly depends on u.

The van Deemter equation may be nondimensionalized as

1
 b
H* a *
cu*
u

Where

H
H *
dp
ud p
u*
Dm

Here dp is the average particle diameter of the column packing and D m is the solute diffusivity. The
normalized mobile phase velocity, u* is basically the Peclet No. for chromatography.

A typical van Deemter plot is shown in Fig. xx.xx. With increasing u, H is seen to decrease sharply till a
minimum is reached, beyond which H increases monotonously. At the minimum value of H, base width of
the solute peak is minimum which corresponds to optimum chromatographic separation the
corresponding value of u is thus the optimum mobile phase velocity.

The process of elution chromatography can be mathematically modeled by either of two approaches, one as an
equilibrium stage separation process (akin to stagewise extraction) and the other as a diffusion-reaction process.

In the former approach, the continuous packed column is assumed to be a series of discrete stages of fixed volume,
in each of which the solution and adsorbent are in equilibrium. Solvent flows from one stage to the next (at a
constant vol. rate Q), but adsorbent of one stage is confined to that stage alone. Now consider the n th stage. Let yj
denote the solute concentration in liquid (defined as mass solute per volume solvent) exiting j th stage, and qj denote
solute concentration in solid adsorbent (defined as mass solute per volume of adsorbent) in the j th stage. Then rate of
net solute accumulation in the nth stage = Q(yn-1 yn). The total volume (solid+liquid) per stage, (assumed same for
all stages) VS = VB/N, where VB= bed volume and N = no.of stages. Let denote the fraction of stage volume V S that
is liquid. Then (1 -) is the fraction of stage volume that is solid. The solute accumulation in stage n takes place
both in the solid and liquid phases, hence

2
(Rate of solute accumulation in liquid) + (Rate of solute accumulation in solid) = Net rate of solute accumulation

VS dyn / dt (1 )VS dqn / dt H yn 1 yn

+ =

Although most actual adsorption equilibrium relations are nonlinear, it may be argued that chromatography dilutes
all solutes and at high dilution all equilibrium relations become linear. This permits the use of linear equilibrium
relation

qn Kyn

where K is the equilibrium constant (a function of temp.). This may be substituted in Eqn. (xx.xx) above to give:

[{ (1 ) K }VS ] dyn / dt H yn1 yn

The conditions for solving the above ODE are :

For t<0, yn=0, n = 1, 2, ., N; at t=0, y0=yF, n =1.

(where yF is the total solute concentration in feed solution charged into the column).

The solution is

yn n 1e

yF n 1 !

where is a dimensionless time given by

Ht

N [{ (1 )}K ]VB

The dimensionless solute concentration (y/yF) expressed as a function of , actually follows the well-known Poisson
distribution. As N increases, the profiles gradually approach the Normal or Gaussian distribution, which is that
observed experimentally:

y t / t0 1
2

exp
y0 2 2

Where y0 is the maximum solute concentration in an eluted solute peak, t0 the corresponding retention time, and 2
the variance (where w=4 ). For Gaussian peaks, the number of theoretical plates, N, may be related to both
retention time tR and base width w as

3
 t R2 tR2
N
2 ( w / 4)2

Expressions for yield and purity of eluted solute fractions:

The eluted solutes exiting from the column are collected as fractions over different time intervals.

Let yk = concn. of kth solute in the solution and y = yk = total solute concn. Also let Q = volumetric flow rate of
solvent. Hence total volume eluted between times t1 and t2 :

Q(t1-t2)

Total amount of solute in a particular fraction collected between times t1 and t2 :

t2

yQ.dt
t1

Total amount of solute charged into the column :

yQ.dt
0

Yield, in that particular fraction, of total solute:

t2

y.dt
t1

y.dt
0

Yield, in that particular fraction, of kth solute

t2

y .dt
t1
k

y .dt
0
k

Purity, in that particular fraction, of kth solute:

4
 t2

y .dt
t1
k

t2

y .dt k
t1

where the summation is over all solutes present. For a binary sample (consisting of solutes A and B, say) the purity
of A in eluate collected between times t1 and t2 is

t2

y
t1
A .dt
t2 t2

y
t1
A .dt yB .dt
t1