Theories of chromatography
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There are two theories that explain chromatography: plate theory and rate theory. Plate theory, developed in 1941, views the column as divided into theoretical plates where analytes completely equilibrate between the stationary and mobile phases. Rate theory, proposed in 1956, accounts for the dynamics of separation. Greater separation occurs with more theoretical plates and smaller plate height. The number of theoretical plates can be calculated using methods like half-height or USP, and depends on factors like column length, particle size, and retention time.
Theories of chromatography
- 1. 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
- 2. 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 phase and mobile phase Column Theoretical plate
- 3. Chromatographic principle Mobile phase Sample mixture Equilibrium establishes at each point (ideally) Stationary phase The molecules of the Retardation of rate mixture interact with the of movement of molecules of the Mobile molecules and Stationary Phase Each molecule Different distribution interacts differently coefficients and different with MP and SP net rates of migration
- 4. 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)
- 5. 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
- 6. N is a ratio of tR and σ of Wb which is 4σ •N = 5.55 tR2/ w1/22 = 16 tR2/ w2 where: tR is retention time w1/2 is width at h0.5 w is width measured at baseline
- 7. Where: N = Number of theoretical plates Ve = elution volume or retention time (mL, sec, or cm) h = peak height w1/2 = width of the peak at half peak height (mL, sec, or cm)
- 8. • Nmax = 0.4 * L/dp where: Nmax - maximum column efficiency L - column length dp - particle size • So, the smaller the particle size the higher the efficiency!
- 10. Band spreading - the width of bands increases as their retention time (tR) or retention volume (VR) increases
- 11. 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
- 12. The smaller HETP, the narrower the eluted peak
- 13. • 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