1 Rf

Figure 8 Plot of HETP versus development distance showing the effect of diffusion coefficient. Curves 1 -4 dp = 5 ^m.

Figure 8 Plot of HETP versus development distance showing the effect of diffusion coefficient. Curves 1 -4 dp = 5 ^m.

Figure 9 Variation of the apparent HETP with development length. Effect of retention. Particle size 5 |im. 1, RF = 0.9; 2, Rf = 0.8; 3, Rf = 0.7; 4, RF = 0.5; 5, RF = 0.3; 6, RF = 0.2. (Reprinted with permission from Guiochon G and Siouffi A (1978) Journal of Chromatographic Science 16: 470-481.)

As p, P, r are kept constant, a plot of H versus (1 β€” Rf)/Rf or retention factor would be linear. Such

Figure 9 Variation of the apparent HETP with development length. Effect of retention. Particle size 5 |im. 1, RF = 0.9; 2, Rf = 0.8; 3, Rf = 0.7; 4, RF = 0.5; 5, RF = 0.3; 6, RF = 0.2. (Reprinted with permission from Guiochon G and Siouffi A (1978) Journal of Chromatographic Science 16: 470-481.)

plots can be drawn from data already published and are shown in Figure 9. The fit of a straight line is excellent as long as solutes are not strongly retained, i.e. when (1 β€” RF)/RF is not too large.

Influence of particle size Examination of Dm has shown that different performances may be attained depending on whether fine or coarse particles are considered. Tables 6 and 7 together with Figure 10 allow the comparison of the different terms A', B' and C with dp.

Best efficiencies are obtained with small particles if Dm is small and development length is short. H will increase very rapidly with increasing length of plate if very fine particles are used. If migration is long the coarser particles are more efficient, which is particularly evident from the plots. We have an explanation for the different claims of efficiencies: most demonstrations of HPTLC are made using mixtures of dyes that have rather large molecular weights and low Dms. Although the nanoplate size prevents long development, some early chromatographers performed development for too long a distance so that efficiency was less than with the older TLC plates. Moreover, the optimal velocity of solvent (v = 3.8) is kept constant for quite a long time with coarse particles but only for a short time with fine particles. As was

Table 6 Effect of change in particle size on the coefficients of the HETP equation

dp (m

A x 104

B' x 104a

C x 106

3

3.38

7.06

1.27

4

5.46

5.30

3.02

5

7.93

4.24

5.90

6

10.74

3.53

10.19

7

13.9

3.03

16.2

8

17.3

2.65

24.2

10

25.2

2.12

47.2

12

34.1

1.77

81.6

15

49.5

1.41

153.3

20

79.9

1.06

378.0

25

115.9

0.848

737.0

30

157.0

0.706

1.27 x103

40

254.0

0.530

3.02 x103

50

368.0

0.424

5.90 x103

Chromatographic conditions: A = 1; C = 0.01; 6 = 47.2 cm s"1; z0 = 0.5 cm.

ay = 0.7; RF = 0.7; Dm = 5 x 10"10 m2s"1. B' is assumed to be

2yDm/6dpRF.

Chromatographic conditions: A = 1; C = 0.01; 6 = 47.2 cm s"1; z0 = 0.5 cm.

ay = 0.7; RF = 0.7; Dm = 5 x 10"10 m2s"1. B' is assumed to be

2yDm/6dpRF.

Figure 10 Plot of HETP versus development distance showing the effect of particle diameter. Diffusion coefficient, 5x10~6cm2s~1. Numbers on curves represent particle size (| m).

demonstrated in HPLC, the race towards finer and finer particles looks meaningless in TLC for the development ought to be so short that the variance of sample deposition would be too large by far. On the other hand 15 |im particles appear to be a good compromise between efficiency and development length. From these results it is also obvious that one requirement of high performance TLC is the use of particles with a narrow size distribution. The mobile phase velocity is determined by the fine particles and efficiency by the coarse ones, so that one loses on both counts.

Table 7 Effect of change in particle size on coefficients of the HETP equation

dp (m

A x 104

B'x104

C'x105

3

2.69

14.12

0.0635

4

4.34

10.6

0.151

5

6.29

8.48

0.295

6

8.53

7.06

0.510

7

11.03

6.06

0.811

8

13.8

5.30

1.21

10

20.0

4.24

2.36

12

27.1

3.54

4.08

15

39.3

2.82

7.66

20

63.5

2.12

18.9

25

92.1

1.696

36.8

30

125.0

1.412

63.5

40

202.0

1.06

151.0

50

292.0

0.848

295.0

Chromatographic conditions as for Table 6, except Dm 1 x10"9m2s"1.

Chromatographic conditions as for Table 6, except Dm 1 x10"9m2s"1.

Influence of velocity coefficient 6 It is difficult to select a convenient liquid that keeps the critical solute within the RF range of 0.30-0.60. From the previous conclusions 6 must be as large as possible, but 6 is imposed by the choice of the chromatographic system and little can be done to change it.

Relative role of the different contributions to band broadening From the above calculations it is clear that the mass transfer C' only plays a minor role unless the plate is poorly prepared or made with very coarse particles, which are not used in current practice. It thus may be neglected in almost all cases, as shown in Figure 11, which gives variation of A', B' and C when dp = 5 |im orβ€” dp = 20 |im for a given Dm.

As expected from previous discussions, the B' term is often the only one which must be taken into account. For dp = 5 |im the packing term is only 0.25 H at L = 1 cm and 0.03 H at L = 10 cm. In contrast, it accounts for 0.9 H when L = 1 cm and dp = 20 |im. When development distance increases the contribution of molecular diffusion increases as the contribution of packing decreases. Molecular diffusion and packing irregularities are equal when the following condition is satisfied:

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