166 Electrophoresis 2005, 26, 166–175

Xiangchun Xuan Band-broadening in capillary zone electrophoresis

Dongqing Li
Department of Mechanical
and Industrial Engineering,
University of Toronto,
Toronto, Ontario, Canada
with axial temperature gradients
It is widely accepted that Joule heating effects yield radial temperature gradients in
capillary zone electrophoresis (CZE). The resultant parabolic profile of electrophoretic
velocity of analyte molecules is believed to increase the band-broadening via Taylor-
Aris dispersion. This typically insignificant contribution, however, cannot explain the
decrease in separation efficiency at high electric fields. We show that the additional
band-broadening due to axial temperature gradients may provide the answer. These
axial temperature variations result from the change of heat transfer condition along the
capillary, which is often present in CZE with thermostating. In this case, the electric
field becomes nonuniform due to the temperature dependence of fluid conductivity,
and hence the induced pressure gradient is brought about to meet the mass continuity.
This modification of the electroosmotic flow pattern can cause significant band-
broadening. An analytical model is developed to predict the band-broadening in CZE
with axial temperature gradients in terms of the theoretical plate height. We find that
the resultant thermal plate height can be very high and even comparable to that due to
molecular diffusion. This thermal plate height is much higher than that due to radial
temperature gradients alone. The analytical model explains successfully the phenom-
ena observed in previous experiments.

Keywords: Axial temperature gradients / Band-broadening / Capillary zone electrophoresis /

Joule heating / Thermal plate height DOI 10.1002/elps.200406141

1 Introduction
Capillary zone electrophoresis (CZE) is a routine analytical
technique for fast and efficient separation of charged
species in aqueous solutions. Under the influence of an
externally applied electrical field, the sample introduced
as a plug (i.e., the sample zone) at one end migrates to the
other end of a fused-silica capillary. The migration velocity
of a particular ionic species is the vector sum of the bulk
electroosmotic (EO) velocity of the background electro-
lyte solution and the electrophoretic (EP) velocity of that
species. As a result, the analyte separates into bands
representing different components in the sample, which
will arrive at a fixed detector at different times.
The separation efficiency of capillary electrophoresis is
often expressed in terms of the theoretical plate height, H,
defined as H = s2 /L where s2 is the variance of the ana-
lyte concentration at the detector, and L is the distance
Correspondence: Prof. Dongqing Li, Department of Mechanical
and Industrial Engineering, University of Toronto, 5 King’s College
Road, Toronto, Ontario, Canada M5S 3G8
E-mail: dli@mie.utoronto.ca
Fax: 1416-978-7753
Abbreviations: EO, electroosmotic; EP, electrophoretic
between the injection point and the detector. In CZE, one
tries to reduce H, i.e., make s2 as low as possible. Under
ideal circumstances, where molecular diffusion is the only
source of band-broadening, s2 = 2Dt with D and t being
the diffusion coefficient and the elution time of the spe-
cies, respectively. Thus, the lower bound for the theoreti-
cal plate height is hideal = 2D/um, where um = L/t is the
migration velocity of the species. In practice, however,
there are various sources of band broadening that could
contribute to s2 in CZE [1–3]. One such source is the
Joule heating effect caused by an electric current passing
through the buffer solution [4–9].
Joule heating in capillary electrophoresis can lead to an
increase and a radial gradient in the buffer temperature
[10–16]. Since most fluid properties are temperature-
sensitive, both the EP velocity and the molecular diffu-
sion of analyte species become nonuniform over the
channel cross-section [16]. These radial nonuniformities
would cause band-broadening in CZE by the so-called
Taylor-Aris dispersion [17, 18]. The resultant thermal
plate height has been examined both theoretically and
experimentally [19–25]. It was found that this thermal
plate height is ordinarily small compared to that caused
by molecular diffusion except at some extreme condi-
tions [24, 25].

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Electrophoresis 2005, 26, 166–175 Band-broadening in CZE with axial temperature gradients 167

So far, however, no work has been done on the effects of axial

temperature gradients on the band-broadening in CZE.
These axial temperature gradients could arise from the var-
iation of heat transfer condition or the variation of buffer con-
ductivity along the capillary [26]. By inserting the middle part
of a capillary into a cooling water-bath while remaining the
rest parts in the air, Gaš [26] investigated the oscillation of
concentration profile and changes in concentration at sites of
jumps of the radial heat flux. Xuan et al. [27, 28] studied the
reservoir-based thermal end effects and the Joule heating
effects on EO capillary flow. They observed sharp tempera-
ture drops near the ends of the capillary and a high tempera-
ture plateau in the middle section. These axial temperature Figure 1. Illustration of CZE with axial temperature gra-
gradients make the electric field nonuniform along the chan- dients: (a) schematic diagram; (b) temperature distribu-
nel, which in turn induces pressure gradients in order to tion; (c) electric field distribution; (d) induced pressure
distribution.
maintain the fluid continuity during the flow. Indeed, these
authors observed concave-convex-concave fluid velocity
profiles from the inlet to the outlet of the capillary [28]. Porras
In this paper, we will develop an analytical model to cal-
et al. [14, 15] and Palonen et al. [24, 25] studied the influence
culate the theoretical plate height in CZE with axial tem-
of solvent on the fluid temperature and the thermal peak-
perature gradients. These axial temperature gradients
broadening in CZE at high electric fields. They obtained sig-
result from the variation of heat transfer condition along
nificantly low separation efficiency at high electric fields,
the capillary as shown in Fig. 1. We will show that the re-
which could not be explained by the radial temperature gra-
sultant thermal plate height can be very high and even

CE and CEC
dients induced thermal plate height. They argued that this
comparable to that due to molecular diffusion. The tem-
huge discrepancy might be attributed to the temperature
perature dependence of fluid properties is taken into
jumps at junctions between the thermostated (forced air-
consideration. This consideration should make the pres-
cooling) and unthermostated (still air-cooling) regions [25]. In
ent model applicable to most practical CZE.
practice, this variation of heat transfer condition along a
capillary often exists in CZE with thermostating. This is be-
cause a short length of the capillary at each end must be left
outside the thermostating cartridge for the sample injection 2 Theory
and detection, respectively.
In order to focus this study of the thermal plate height
dependence on the axial temperature gradients, we limit
Consider a homogeneous capillary with variable heat
ourselves to neutral analytes so that the contribution from
transfer conditions along its length direction. As shown
nonuniform EP flow profile is eliminated. In this section,
schematically in Fig. 1, the injection and detection regions
we first derive the temperature and flow profiles of the
are exposed to the stationary air (i.e., unthermostated)
fluid over the channel cross-section in each region, i.e.,
while the majority of the channel in the middle is thermo-
the injection region, the forced air-cooling region and the
stated by the forced air-cooling. As a result, the fluid
detection region as labeled in Fig. 1. Next, we use the
temperatures in the injection and detection regions are
continuity conditions of both the electrical current and the
higher than in the middle region (Fig. 1b). Since the fluid
mass flow rate along the capillary to determine the local
conductivity is increased with temperature, the electric
electric field and the local pressure gradient in each
field in the middle low-temperature (cooling) region
region. Finally, the theory of Taylor-Aris dispersion is
should be higher than the rest in order to meet the
applied to calculate the theoretical plate height in the
requirement of current continuity (Fig. 1c). Then, the
whole capillary.
requirement of mass continuity forces the appearance of
axial pressure gradients (Fig. 1d). The fluid flow profile is
no longer uniform, and the resultant Taylor-Aris dispersion
2.1 Notation
will increase the band-broadening in CZE. This thermal
plate height is different from that due to the parabolic-like a radius of ionic species, 1 nm
EP flow profile of the analyte species as described above.
A, B defined constants in fluid viscosity, A = 2.76161026
Essentially, the increased band-broadening due to the
kgm21s21 and B = 1713 K
effects of axial temperature gradients is identical to that
caused by the nonuniformity of zeta potential [29, 30]. D diffusion coefficient

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168 X. Xuan and D. Li Electrophoresis 2005, 26, 166–175

e charge of a proton, 1.602610219 C p polyimide coating

E applied electric field pd pressure-driven
h surface heat transfer coefficient, 130 and w wall
4000 Wm22K21 for the still-air cooling and the
forced-air cooling
2.2 Temperature and flow profiles of the fluid in
H plate height each region
k thermal conductivity, 0.61, 1.38, and 0.15 Wm21K21
for fluid (water), wall and coating As the capillary length is much larger than its internal di-
ameter (each region has several to tens of centimeters
kB Boltzmann’s constant, 1.381610223 JK21 while the capillary internal diameter is of 100 mm for the
L capillary length, 6 cm, 40 cm, and 4 cm for the present problem) and the Reynolds number is small (less
injection region, forced air-cooling region, and than 0.2 for the present problem), the transition regions in
detection region both temperature and flow fields near the capillary
N plate number entrance and at the junctions of unthermostated and
thermostated regions are negligibly small [30] and there-
P induced pressure
fore are not considered. This implies a discontinuity in the
DP ´induced pressure difference temperature and velocity at the above junctions. In each
r radial coordinate region itself, however, there are no axial variations.
R radius, 50 mm, 160 mm, and 180 mm for the internal In each region (the subscript i = 1,2,3 occurring later in
wall, outer wall, and coating this paper indicates the injection region, the thermostated
T temperature, 298 K (T0 for ambient) region and the detection region, respectively; see Fig. 1)
where the heat transfer condition is uniform along the
T cross-sectional average fluid temperature
length direction, Joule heating produces a parabolic-like
u axial velocity temperature profile across the capillary column [4–16].
uslip slip velocity The internal wall temperature Tiw and the temperature
U overall heat transfer coefficient difference between the center and the internal wall DTi
can be derived from the governing equations [13, 15],
V applied voltage between capillary ends
X0 the distance between the detector and the capillary E 2i k
i Rl ¼ 2Ui ðT iw
T 0 Þ (1)
outlet, 2 cm E 2i k
i R2l ¼ 4k l DT i (2)
y normalized radial coordinate 

i ¼ k0 1 þ a Ti
T 0
k (3)
zv valence of ions
Ti ¼ T iw þ DT i =2 (4)
a temperature coefficient of electrical conductivity,
0.02 K21 where Ei is the local electric field in region i, and k0 and k i
e permittivity, 305.7 exp(-T/219)68.854610 212 are the fluid electrical conductivity at the ambient tem-
CV21m21 perature T0 and the cross-sectional average fluid temper-
ature T i, Rl and kl are the capillary internal radius (i.e., the
Z dynamic viscosity
fluid domain) and the fluid thermal conductivity, respec-
2
s variance of the analyte concentration tively. Here, we employ a linear dependence of electrical
k0 electrical conductivity of fluid at T0 0.5 Sm21 conductivity on the fluid temperature with coefficient a.
The overall heat transfer coefficient Ui in Eq. (1) is given by
m mobility
1/Ri[ln(Rw/Rl)/kw 1 ln(Rp/Rw)/kp 1 1/hiRp], where hi is the
o defined coefficient describing the radial tempera- surface heat transfer coefficient resulting from both con-
ture gradient vection and radiation from the capillary outer surface. The
x zeta potential, 250 mV subscripts w and p represent the capillary wall and poly-
imide coating, respectively. Combining Eqs. (1) and (2)
Subscripts: gives the relationship between DTi and Tiw,
eo electroosmotic
Ui ðT iw
T 0 ÞRl
ep electrophoretic DT i ¼ (5)
2k l
i region in a capillary, i = 1,2,3 Then, by replacing T i in Eq. (3) with Eq. (4), and in turn
l fluid substituting the resultant Eq. (3) into Eq. (1), we have

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Electrophoresis 2005, 26, 166–175 Band-broadening in CZE with axial temperature gradients
1 Zi ¼ AexpðB=T i Þ
T iw ¼ T 0 þ  (6)
2Ui E 2i k0 Rl
að1 þ Ui Rl =4k l Þ
Whereas the denominator of the second term on the right
hand side of Eq. (6) can be mathematically negative, it is
physically impossible because Joule heating is supposed
to increase the fluid temperature. This phenomenon is
attributed to the positive-feedback auto-thermal effect,
where the fluid temperature cannot be held and will be
continuously increased until run-away [12, 20]. It is now
straightforward to derive the cross-sectional temperature
profile Ti(r) and the corresponding average value T i in
each region,
Ui Rl ðT iw
T 0 Þ  
T i ðr Þ ¼ T iw þ 1
r 2 R2l right (7)
2k l
Ti ¼ T iw ð1 þ Ui Rl =4k l Þ
T 0 Ui Rl =4k l (8)
where r denotes the radial position.
We have shown earlier that the axial temperature gra-
dients induce a pressure gradient in each region of the
capillary in order for the mass continuity to be met
(Fig. 1d). Therefore, we will determine the profile of a
combined EO and pressure-driven flow in each region in
the presence of radial temperature gradients as pre-
sented above. Since the Reynolds number in CZE is
generally small, we only need to solve the Stokes equa-
tion to get the velocity profile.
In the limit of a thin electrical double layer, the EO velocity,
ui, slip, calculated from the Helmholtz-Smoluchowski for-
mula acts as a “slip” boundary condition [5, 29–31],
ui;slip ¼
eiw zE i =Ziw (9)
where e and Z are the permittivity and the dynamic vis-
cosity of the fluid, respectively, and x the zeta potential.
The subscript iw denotes the fluid properties calculated at
the wall temperature Tiw. The temperature dependence of
zeta potential is not taken into consideration because this
problem has not been studied in details so far. We recog-
nize that the variations of zeta potential along the capillary
could substantially alter the results presented here. From
our previous experiments [28], however, we find that the
temperature dependence of zeta potential is essentially
small in the range of temperature change involved in this
work. With the use of slip velocity, the electrostatic force
drops from the Stokes equation, which thus reduces to
   
1d dui dP
rZi ¼ (10)
r dr dr dz i
where (dP/dz)i denotes the induced pressure gradient in
the axial direction in region i. The viscosity Zi depends on
the fluid temperature, and is empirically expressed by
 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
169
(11)
with A and B being constants. Thus, we have l/Zi =
exp(2B/Ti)/A. After expanding the exponential part of this
expression as a Taylor series around T = Tiw and retaining
only the first two terms, we have
1 1 
 
ffi 1 þ oi 1
r 2 R2l (12)
Zi Ziw
where Ziw = Aexp(2 B/Tiw) and oi = BDTi/T2iw, and Eq. (7)
has been used in the exponential expansion. Integrating
Eq. (10) with the “slip” boundary condition at the capillary
internal wall and the symmetric condition at the center
yields
eiw z
ui ðy Þ ¼
Ei

Ziw
  2 " ! #
qP Rl 1 1 r2 r2

1 þ oi
1 þ oi
oi 2 (13)
qz i 4Ziw 2 2 Rl R2l
The first term on the right hand side of Eq. (13) represents
the still plug-like EO velocity profile (identical to the slip
velocity in Eq. 9), and the second term describes the
pressure-driven flow profile. Note that the latter profile is a
little bit different from a parabola due to the small quartic
term caused by radial temperature gradients.
2.3 Electrical field and induced pressure
gradient in each region
We assume an incompressible flow in CZE (i.e., without
accounting for the density change with temperature).
Moreover, the capillary wall is a perfect insulator. Then,
we can use the current continuity and the mass continuity
to determine the local electrical field and the local pres-
sure gradient in each region [32]. Equivalently, the con-
tinuity of current density and the continuity of cross-sec-
tional average velocity can be used instead because of
the uniform fluid density and channel geometry.
The electric current in an electrokinetic flow is composed
of two parts: one is the streaming current due to the
motion of charged ions carried by the bulk fluid, the other
is the conduction current due to the EP motion. The
streaming current is generally much smaller than the
conduction current unless the channel size is comparable
to the thickness of electrical double layers (on the order of
nanometers) [31, 32]. Therefore, we account for only the
conduction current density Ji = k i Ei from the Ohm’s law.
Thus, the requirement of current continuity in each region
forces
1 E 1 ¼ k
k 2 E 2 ¼ k
3 E 3 (14)
170 X. Xuan and D. Li Electrophoresis 2005, 26, 166–175

For simplicity, we assume the heat transfer conditions 2.4 Theoretical plate height
around the injection (i = 1) and detection (i = 3) regions are
exactly the same. Then, E1 = E3 is found from Eq. (14) As demonstrated above, axial temperature gradients
because k1 = k
3. In addition, the electrical voltage across induce a pressure-driven fluid flow (see the second term
the whole capillary column is given by on the right hand side of Eq. 13), which is superimposed
upon a bulk plug-like EO flow (see the first term on the
E 1 L1 þ E 2 L2 þ E 3 L3 ¼ E 0 ðL1 þ L2 þ L3 Þ (15) right hand side of Eq. 13). This pressure-driven flow
causes Taylor-Aris dispersion, for which the variance of
the analyte concentration s2 in region i is given by [17, 18]
where E0 is the average or nominal electric field applied
!
externally, L1, L2 and L3 indicate the lengths of the injec- R2l u
2i;pd
2
si ¼ 2 D i þ
tion, forced air-cooling, and detection regions, respec- 48D  i ti (20)
tively. Since we already get the relation between the
average temperature T i (appearing in k i as shown in Eq. 3) i;pd ¼
Mi DPi =Li
u (21)
and the local electric field Ei in the preceding section, their
values are now available from Eqs. (14) and (15). These i (i = 1,2)
ti ¼ Li =u (22a)
results are omitted here for compactness.

As to the mass continuity, the cross-sectional average 3

t3 ¼ ðL3
X 0 Þ=u (22b)
 i in region i is first derived from
velocity displaystyleu
Eq. (13) as where D  i is the local diffusion coefficient extracted at the
average fluid temperature T i, and X0 is the distance be-
i ¼ miw E i
Mi DPi =Li
u (16) tween the detector and the outlet. We use the Stokes-
Einstein equation D = kBT/6pZa to calculate the diffusion
where miw = 2eiwx/Ziw is the EO mobility calculated with coefficient where kB is the Boltzmann’s constant and a is
fluid properties at the internal wall temperature Tiw, Mi = the radius of analyte molecules [31]. The effect of non-
1 uniform diffusion coefficient on the band-broadening is
(1þ oi) Rl2/8Ziw can be viewed as the hydrodynamic
3 neglected here [16]. As the variance is addible, the total
conductivity* in the presence of radial temperature gra- plate height Htot is expressed as
dients, and DPi is the pressure difference in region i. Then,
the mass continuity requires Htot ¼ Hdif þ HaxT (23a)
,
m1w E 1
M1 P1 =L1 ¼ m2w E 2
M2 ðP2
P1 Þ=L2 X
3
Hdif ¼ 2  i ti
D Ldet (23b)
¼ m1w E 1 þ M1 P2 =L3 (17) i¼1

where P1 amd P2 indicate the induced pressures at the
two ends of the forced cooling region as labeled in Fig. 1d.
The two reservoirs at capillary ends are assumed open to
air. Note that m1w = m3w, E1 = E3, and M1 = M3 have been
invoked in Eq. (17). As the local electrical field Ei has been
obtained above, P1 and P2 are easily determined from
Eq. (17) as
m2w E 2
m1w E 1
P1 ¼ (18)
M1 =L1 þ M2 ð1 þ L3 =L1 Þ=L2
P2 ¼
P1 L3 =L1 (19)
Now, the fluid flow profile in each region of the capillary is
known (see Eq. 13).
* This name is borrowed from the definition of thermal conduc-
tivity in heat flow density q = kDT/L where DT is the “driving
force” for the heat flow.
,
R2l X 2i;pd
3 u
HaxT ¼  i ti Ldet (23c)
24 i¼1 D
where Hdif and HaxT represent the theoretical plate height
due to molecular diffusion and the effects of axial tem-
perature gradients, respectively, and Ldet = L1 1 L2 1 L3 2
X0 is the distance between the inlet and the detector (not
shown in Fig. 1a).
Equation (20) assumes that the elution time ti in each
region of the capillary is much larger than the radial diffu-
sion time td , RI2/D, which is typically on the order of
seconds, so that the theory of Taylor-Aris dispersion may
be used. This condition might be problematic in the gen-
erally short injection and detection regions. However, we
can still use the Taylor-Aris dispersion formula to make a
preliminary prediction of the band-broadening. The theo-
retical plate height is a little underestimated in the present
analytical model.

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Electrophoresis 2005, 26, 166–175 Band-broadening in CZE with axial temperature gradients
3 Results and discussion
In this section, we apply the analytical model developed
above to an exampled CZE with axial temperature gra-
dients. The local electric fields, local fluid temperatures,
local flow profiles, and theoretical plate heights are first
examined quantitatively. Then, a comparison is made
about the thermal plate height due to the perturbed fluid
flow (axial temperature gradients) and the perturbed EP
velocity profile (radial temperature gradients), respectively.
After that, the combined effect of these two flow pertur-
bations is addressed. Finally, the present model is used to
explain the phenomena observed in previous experi-
ments. All parameters including fluid properties, channel
dimensions and properties are referred to the nomen-
clature unless otherwise indicated. The convective heat
transfer coefficients are: hi = 130 Wm22 K21 in both the
injection and detection regions, and h2 = 4000 Wm22K21 in
the forced air-cooling region [15, 24, 25].
Figure 2 shows the comparison of average fluid tem-
peratures T i in different regions of the capillary. One can
see that T i in either the injection region or the detection
region is always higher than T i in the forced air-cooling
region. This axial temperature variation becomes more
pronounced at higher electric fields. The equivalent
temperature T eq of the fluid in the whole capillary is
also shown in Fig. 2, which is converted from


keq ¼ k0 1 þ a T eq
T 0 where the equivalent fluid
conductivity is given by
L1 þ L2 þ L3
keq ¼ (24)
ðL1 þ L3 Þ=
k1 þ L2 =
k2
This equivalent temperature simulates the temperature
data extracted from the experimentally measured fluid
conductivity. It is shown that Teq is higher than the fluid
Figure 2. Cross-sectional average fluid temperatures in
different regions of the capillary and the equivalent fluid
temperature along the whole capillary. All other parame-
ters used in the calculation are referred to the notation
(Section 2.1).
 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
171
temperature in the forced air-cooling region where the
local electric field is actually larger than the nominal elec-
tric field E0 (Fig. 3). Therefore, if one uses E0 to calculate
the average fluid temperature in CZE supposed to be
uniformly thermostated, there would be an even larger
discrepancy between the calculated temperature and the
experimentally predicted fluid temperature, i.e., Teq. This
temperature deviation is exactly the phenomenon Porras
et al. [15] observed when they studied the influence of
solvent on the fluid temperature.
Figure 3 shows the local electric fields in different regions
of the capillary. As predicted, the electric field becomes
nonuniform along the capillary because of the tempera-
ture and hence fluid property variations in different
regions caused by Joule heating and different heat trans-
fer conditions. The forced air-cooling region undergoes a
higher electric field than the injection and detection
regions. It is interesting to note that there are two “self-
regulating” mechanisms in this system. Firstly, the re-
sponse of the electric field reduces Joule heat in the high
temperature region (lower electric field in the injection and
detection regions), and increases Joule heat in the low
temperature region (higher electric field in the forced air-
cooling region). This mechanism is in contrast to the
aforementioned auto-thermal effect resulting from the
temperature dependence of fluid conductivity (see Eq. 6).
Secondly, the decrease in EO velocity in the high temper-
ature region is partially offset by a local decrease in fluid
viscosity.
Figure 4 shows the profiles of bulk fluid velocity in differ-
ent regions of the capillary. The convex curvature of the
velocity profile in the injection and detection regions is
significant (Fig. 4a). On the contrary, the velocity profile in
Figure 3. Local electric fields in different regions of the
capillary. The dashed-point line shows the case in the
absence of axial temperature gradients. All parameters
used in the calculation are referred to the notation
(Section 2.1).
172 X. Xuan and D. Li
Figure 4. Profiles of bulk fluid velocity in different regions
of the capillary: (a) injection region and detection region;
(b) forced air-cooling region. All parameters used in the
calculation are referred to the notation (Section 2.1).
the forced air-cooling region (Fig. 4b) is only slightly con-
cave even at high electric fields. This difference is attrib-
uted to the much lower induced pressure gradient in the
thermostated region (Fig. 1d). In addition, the curvatures
of velocity profiles in all regions are not simply increased
with the rise of electric field. It is believed to be the
reflection of the aforementioned “self-regulating” mech-
anism.
Figure 5 shows the curves of theoretical plate heights with
respect to the electric field. At low electric fields (e.g., less
than 15 kV/m for the present problem), the effects of axial
temperature gradients are insignificant so that the result-
ant thermal plate height HaxT is negligible. The plate height
Hdif due to molecular diffusion predominates the total
plate height Htot. With the rise of electric field, HaxT is
increased significantly due to enhanced axial temperature
gradients (see solid line B in Fig. 6 for a zoom-in view). In
the meantime, however, Hdif is decreased because the
analyte migrates faster at higher electric fields. Conse-
quently, HaxT becomes comparable to Hdif at high electric
fields. For example, HaxT can be as high as 60% of Hdif at
around 45 kV/m in Fig. 5. The theoretical plate height
under ideal circumstances, Hideal, where molecular diffu-
sion is the only source of band-broadening, is close to Hdif
meaning that Joule heating induced temperature effects
only slightly influence molecular diffusion. Hideal is inver-
sely proportional to the electric field, while Htot varies
relatively
 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Electrophoresis 2005, 26, 166–175
Figure 5. Theoretical plate heights at different electric
fields. See Eqs. (23) and (27) for the definitions of different
plate heights illustrated in the figure. Hideal (defined in the
introduction) indicates the minimum plate height in CZE
where molecular diffusion is the only source of band-
broadening. Only HradT refers to the secondary Y-axis (i.e.,
right ordinate). All parameters used in the calculation are
referred to the notation (Section 2.1).
slowly in a range of electric fields. As demonstrated in
Fig. 5, Htot is decreased from 0.624 mm to 0.473 mm when
the electric field is doubled from 25 kV/m to 50 kV/m. The
reason we limit the discussion to field strengths up to
55 kV/m is the consideration of the high fluid temperature
attained in the free air-cooling region (around 907C as
shown in Fig. 2, i.e., the liquid is close to boiling). This
large temperature elevation is caused by the relatively
high electrical conductivity (0.5 S/m) we used in the cal-
culation. If we use a lower conductivity, say 0.2 S/m, the
maximum electric field without causing the liquid boiling
in the free air-cooling region can be at least doubled, i.e.,
over 100 kV/m. The decreasing of HaxT at electric fields
larger than 40 kV/m in Fig. 5 will still be seen in the case of
a low conductivity while the electric field may be as high
as 100 kV/m. We believe the self-regulating mechanism
partially cancels the effects of axial temperature gradients
at high electric fields.
Figure 5 also shows the thermal plate height HradT due to
radial temperature gradients alone (actually the induced
parabolic-like EP velocity profile of analyte molecules).
Note that the right ordinate (in nanometers) has different
units than the left one (in micrometers). In the absence of
axial temperature gradients, the electric field is uniform
along the whole capillary. The temperature formulae
derived in Section 2.2 are still effective, but the subscript i
now drops. The EP velocity is given by the Nernst-Ein-
stein equation uep = DzveE/kBT where zv is the valence of
ions and e is the charge of a proton [31]. Substituting the
diffusion coefficient D with the Stokes-Einstein equation
yields the EP velocity,
Electrophoresis 2005, 26, 166–175 Band-broadening in CZE with axial temperature gradients
uep ¼ zv eE=6pZa (25)
Combining Eqs. (12) and (25) gives

 
uep ¼ uep;w 1 þ o 1
r 2 R2l (26)
where uep,w = zveE/6pZwa denotes the EP velocity calcu-
lated with fluid properties at the internal wall temperature
Tw. Thus, the thermal plate height due to the perturbed EP
flow is given by
o2 R2l u2ep;w
HradT ¼ u (27a)
96D m


m ¼
ew zE=Zw þ uep;w 1 þ 12o
u (27b)
where D  is the diffusion coefficient at the average fluid
temperature, and u m indicates the migration velocity of
analyte species averaged over the capillary cross-sec-
tion. One can see in Fig. 5 that HradT (see the vertical axis
on the right) is much lower than HaxT, and is indeed insig-
nificant compared to Hdif even at high electric fields. This
fact may explain why the separation efficiency measured
in Palonen’s experiment deviated appreciably from the
prediction of Eq. (27) [25], which will be addressed after
we present the following formulations for charged ana-
lytes.
Since CZE is used to separate charged chemical species,
where the fluid flow perturbation due to axial temperature
gradients and the EP flow perturbation due to radial tem-
perature gradients should exist simultaneously, it is
important and necessary to investigate the combined
Figure 6. The combined thermal plate heights HT (solid
lines) and the ratio of HT to Hdif (dashed lines) for ions of
different charge: Case A, zv = 21; case B, zv = 0; case C, zv
= 11. All parameters used in the calculation are referred
to the notation (Section 2.1).
 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
173
temperature effects on the band-broadening. In this
regard, the observed velocity profile of an analyte ion in
region i is the sum of Eqs. (13) (fluid velocity) and (26) (EP
velocity),
" ! #
 
qP R2l 1 1 r2 r2
ui ðyÞ ¼
miw E i
1 þ oi
1 þ oi
oi 2 þ
qz i 4Ziw 2 2 Rl R2l
" !#
r2
þ mi;ep;w E i 1 þ oi 1
2 (28)
Rl
where mi,ep,w = zve/6pZiwa is the EP mobility corresponding
to the above uep,w in Eq. (26). Note that the subscript i is
inserted into the EP velocity part because it is also de-
pendent on the local electric field and fluid properties.
Then, the cross-sectional average value of the parabolic
part of the velocity profile in Eq. (28), which contributes to
the band-broadening, is expressed as
i;pd ¼
Mi DPi =Li þ 12oi mi;ep;w E i
u (29)
And the average migration velocity of analyte species
 i in Eq. (22) for calculating the elution time ti in each
(i.e., u
region of the capillary) is now changed to


i ¼ miw E i
Mi DPi =Li þ mi;ep;w E i 1 þ 12oi
u (30)
Different from the profile of induced pressure-driven fluid
velocity, where the curving direction is region-sensitive
(see Fig. 4), the EP velocity profile is curved in the same
direction in the whole capillary because the fluid temper-
ature is always higher at the capillary center. Therefore,
the resultant curvature of observed velocity profile for
charged analytes is increased in some regions while
decreased in other regions compared to that of neutral
analytes. Moreover, the elution time ti in Eq. (23) is asso-
ciated with the sign of charged analytes. These varieties
complicate the combined thermal plate height HT. Those
HT for negative, neutral and positive ions are shown in
Fig. 6. The ratios of HT to Hdif for these ions are also dis-
played. As expected, HT for charged ions follow the same
trend as neutral ions when the electric field is increased.
The difference in the magnitude of HT is mainly attributed
to the different migration speed for they are all much
higher than HradT (see Fig. 5).
Figure 7 shows the comparison of theoretical and experi-
mental plate numbers of 2,4-dinitrophenol in EtOH solu-
tion at different applied voltages, V = E0L. The experi-
mental plate numbers (symbols) were adapted from
Palonen et al.’s measurements (Fig. 2b in [25]). The theo-
retical plate numbers (solid line), N = Ldet/(HT 1 Hdif), were
calculated with the analytical model presented above
without accounting for the contributions to theoretical
plate height from injection, detection, and adsorption,
etc. [1–3]. The dashed line represents the calculated plate
174 X. Xuan and D. Li
Figure 7. Comparison of theoretically predicted plate
numbers N = Ldet/(HT 1 Hdif) (solid line) using the present
model with those measured in Palonen et al.’s experiment
(symbols with short straight lines in between, adapted
from Fig. 2b in [25]) at different applied voltages V = E0L.
The dashed line shows the theoretical plate numbers
calculated from N = 0.4 Ldet/(HT 1 Hdif) that accounts for
the contributions to band-broadening from injection,
detection and adsorption etc. Working parameters used
in the calculations include: L1 = 5 cm, L2 = 22.5 cm, L3 =
2.5 cm, Ldet = 28.5 cm, Rl = 37.5 mm, Rw = 167.5 mm, Rp =
187.5 mm, kl = 0.174 Wm21K21, kw = 1.5 Wm21K21, kp =
0.15 Wm21K21, EO mobility meo = 23.7610210 m2V21s21
at 237C, EP mobility mep = 21.2361028 m2V21s21 at 237C,
fluid viscosity Z = 1.0861023 Kgm21s21 at 237C, temper-
ature coefficient of viscosity 0.018 K21, electrical con-
ductivity K = 0.05 Sm21 at 237C, temperature coefficient
of conductivity a = 0.0207 K21 and molecular diffusivity D
= 5.34610210 m2s21 at 237C ([14, 15, 24, 25]; Palonen,
personal communication).
numbers by using N = 0.4 Ldet/(HT 1Hdif) because the real
plate numbers are only 30% – 50% of those illustrated in
the solid line according to Palonon et al.’s calculations
[24, 25]. One can see that our model predicts properly the
trend of plate numbers with respect to the applied volt-
ages, i.e., electric fields. Also, the theoretically predicted
plate numbers are close to the experimentally measured
values when we take into consideration the various
sources of band-broadening as Palonen et al. did (see the
dashed line in Fig. 7). These consistencies support the
analytical model developed in this work.
4 Concluding remarks
Many experimental factors in CZE can cause band-
broadening and thus decrease the separation efficiency.
Although Joule heating effects in CZE have been studied
for a long time, all previous efforts are made to the non-
uniformity of EP velocity arising from the radial tempera-
 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Electrophoresis 2005, 26, 166–175
ture gradients. In this paper, we have demonstrated that
axial temperature gradients can generate a much larger
thermal plate height than that due to radial temperature
gradients. These axial temperature gradients often exist
in CZE with thermostating, where the variation of heat
transfer condition along the capillary is generally present.
An analytical model has been developed to explicitly cal-
culate the theoretical plate height for CZE with axial tem-
perature gradients. The electric field, fluid temperature,
and flow profile in each region of the capillary are also
available from this analytical model. The results obtained
from this analytical model have been successfully used to
explain the phenomena observed in previous experi-
ments.
Financial support from the Natural Sciences and Engi-
neering Research Council (NSERC) of Canada, through a
research grant to D. Li is gratefully acknowledged.
Received May 2, 2004
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