Frans_M_Everaerts_Isotachophoresis_378342.pdf

268 PRACTICAL ASPECTS
It can be seen from Table 9.3 that the step heights of the Li’ and Tris’ zones are
nearly constant (at this pH they are both strong ions), whereas the step heights of the
terminating zone ‘water’ increase considerably with increasing pH. At pH values below
about 4.25 (depending on the type of leading electrolyte used), ‘water’ cannot be used as
a terminator because disturbances will occur. At these pH values, the step height of the
pure ‘water’ zone is lower than that of, e.g., Tris’ and zone electrophoretic phenomena
can be expected. Also, above a pH of about 5-5.5, ‘water’ cannot be used as a terminator
because its effective mobility is too low. Therefore water can act as a terminator in the
approximate pH range 4-5.2. In a similar way, water can act as a terminator in the
approximate pH range 9-10 for anionic separations.
Van Hout [ 11 used water as a terminator* in the separation of, e.g., amino acids at pH
9.2. An example is shown in Fig.9.10 for the separation of glutamic acid, taunne, serine,
&cine, tryptophan and sarcosine. The terminator was water and the leading electrolyte
was 0.01 N hydrochloric acid-ethanolamine at pH 9.2. Barium hydroxide was added to
the terminator ‘water’ in order to suppress the amount of hydrogen carbonate present in
the solution and to raise the pH in the terminating reservoir. In Fig.9.10, however, a small
zone of hydrogen carbonate can be seen (see also Chapter 13).
9.6. PURIFICATION OF THE TERMINATOR
As already described in section 5.4, terminating electrolytes have to be very pure,
because if small amounts of impurities are present in the terminating zone (a large voltage
gradient), with higher mobilities than that of the terminating ions, they will be pushed
forwards through all preceding zones until they reach a zone boundary in accordance
with their effective mobilities. This procedure is a type of moving-boundary procedure
and can cause disturbances. In addition to chemical methods for purifying terminators,
such as recrystallization and distillation, an isotachophoretic method can also be used.
TABLE 9.4
SOME STEP HEIGHTS MEASURED WITH TWO DIFFERENT THERMOCOUPLES
The metals were measured in the operational system WKCAC: and the anionic species in the system
MTris/HCI*i(A). The step heights are given in millimetres, and relate to 0 PA.
Species
Formate
Acetate
Caprylate
Stearate
Cacodyla te
Salicylate
Butyrate
Palmitate
Thermocouple 1
162
186
223
26 0
298
172
202
253
Thermocouple 2
170
197
234
27 6
312
183
216
272
Species
+ Li
Tris‘
Ni*+
cu2+
Pb’+
BaZ*
Na’
(CJ N
Cd’+
*WKCAC is listed in Table 11.4 and MTris/HCI in Table 12.4.
Thermocouple 1
233
305
201.5
22 1
24 3
169
184
270
223
Thermocouple 2
*Later experiments show that often a terminating ion can better be added to the water (section 13.1.3).
24 I
321
214
24 1
262
180
195
285
236