What is eddy and longitudinal diffusion?

The longer the path length of the molecule, the more be elution time and hence band broadening occurs. This effect is called Eddy's diffusion. Eddy's diffusion occurs in HPLC as the packed column is used in this technique. The longitudinal diffusion is zero/negligible.

What is eddy diffusion in chromatography?

If we have a distinction between the time it takes a set of molecules to move through the column based only on different path lengths, we have broadened the peak. This is known as eddy diffusion.

How can longitudinal diffusion be reduced?

The concentration of analyte is less at the edges of the band than at the center. Analyte diffuses out from the center to the edges. This causes band broadening. If the velocity of the mobile phase is high then the analyte spends less time on the column, which decreases the effects of longitudinal diffusion.

Why is longitudinal diffusion a more serious problem in gas chromatography?

Why is longitudinal diffusion a more serious problem in gas chromatography than in liquid chromatography? Diffusion coefficients of gases are 10^4 times greater than those of liquids. Therefore, longitudinal diffusion occurs much faster in gas chromatography than in liquid chromatography.

Why do bands spread in chromatography?

A band of solute invariably spreads as it travels through the column and emerges at the detector with a standard deviation, σ. Plate height (H) is proportional to the variance ( 2) of the chromatographic band: the smaller the plate height, the narrower the band.

Fundamentals of HPLC 30 - Describing Longitudinal Diffusion

How does particle size affect HPLC?

Particle size (dp), or the mean diameter of the spherical supports used to pack a column, is a physical dimension that has a significant impact on the performance of an HPLC column. Smaller particle sizes have been shown to offer higher peak efficiencies.

What is band broadening in HPLC?

Band broadening is a phenomenon that reduces the efficiency of the separation being carried out –leading to poor resolution and chromatographic performance. This is problematical in terms of both the quality of the separation obtained and the accuracy with which sample components can be quantified.

Why do smaller particles give better resolution in HPLC?

The particle size of a column packing affects the efficiency (theoretical plates) of a column. Smaller particle size improves efficiency of a separation without increasing run time, column length, or flow rate.

What is the difference between molecular and eddy diffusion?

If the convective motion is laminar (non-turbulent), you use the molecular diffusion coefficient. If the motion is turbulent, you use the eddy diffusivity, which is supposed to simulate the enhancing effect of turbulence.

How do you calculate eddy diffusion?

u θ = ( Γ / 2 π r ) exp ( − r 2 / 4 ν t ) . (12.128, 12.129) Equation (12.128) shows the interesting fact that the eddy diffusivity initially increases with time, a behavior different from that in molecular diffusion with constant diffusivity.

Is eddy diffusion independent of flow rate?

Eddy diffusion, the “A” term, a constant, does not change as the mobile linear flow rate changes.

What are the three terms that make up the van Deemter equation?

The Van Deemter equation is governed by three cumulative terms: (A) eddy diffusion, (B) longitudinal diffusion, and (C) mass transfer. A loss in peak efficiency can be observed as a wider analyte band, and therefore, these three terms can also be viewed as factors that contribute to band broadening.

Why is van Deemter equation important?

Understanding the van Deemter equation allows the determination of the optimum mobile phase velocity. The theoretical plate number N_th shows the relation between retention time and peak width and describes column quality and separation power.

What is Tracer diffusion?

Tracer diffusion and Self-diffusion, which is a spontaneous mixing of molecules taking place in the absence of concentration (or chemical potential) gradient. This type of diffusion can be followed using isotopic tracers, hence the name.

What is fronting and tailing?

There are many different causes to “fronting” or “tailing” peaks, but most can be easily remedied. For example, fronting peaks are often caused by column overload or overpacking. Similarly, tailing peaks can be caused by underpacking, or by having a sample that is too viscous.

What causes peak broadening?

Peak broadening is affected by the mobile phase flow rate and the particle size of the packing material in the columns. With the latter the smaller the particle size the better the peak resolution.

What is column efficiency?

Column efficiency, indicated as the number of theoretical plates per column, is calculated as N = 5.54 (t_R / w_0.5)² where t_R is the retention time of the analyte of interest and w_0.5 the width of the peak at half height.

What is the difference between C18 and C8 columns?

C18 has 18 carbon atoms while C8 has only 8 carbon atoms. C18 has a longer carbon chain, but C8 has a shorter one. C18 has higher retention while C8 has shorter retention. C18 has higher hydrophobicity, but C8 has a lower hydrophobicity.

What is pore size in HPLC?

As a general rule, a pore diameter of 10 nm or less should be used for analytes below 3,000 Da. A pore diameter of 10 - 13 Da is recommended for samples in the range of 3,000 - 10,000 Da. For samples above 10,000 Da, including peptides and proteins, a 30 nm material provides the best efficiency and peak shape.

Why are HPLC particles porous?

Theory and experiments have clearly established that particles used for the HPLC separation of molecules should have pores that are sufficiently large to allow free movement of such molecules within the pore structure and not restrict diffusion [1, 2].

What is the formula for Rf value?

The Rf value of a compound is equal to the distance traveled by the compound divided by the distance traveled by the solvent front (both measured from the origin).

What is the formula for retention factor?

f) The retention factor (k) is the ratio of the amount of analyte in the stationary phase to the amount in the mobile phase. It is generally calculated by k' = (tR - tM)/tM = tR'/tM.

Why is retention time important?

Retention time plays an important role in the H₂S removal efficiency as previously demonstrated by Chaiprapat et al. [37]. Longer reactant retention within the enzyme allows time for gas to absorb/react within/with the enzyme.