Journal of Analytical Sciences, Methods and Instrumentation, 2012, 2, 49-53
http://dx.doi.org/10.4236/jasmi.2012.22010 Published Online June 2012 (http://www.SciRP.org/journal/jasmi)
49
Microfluidic Behavior of Ternary Mixed Carrier Solvents
Based on the Tube Radial Distribution in Triple-Branched
Microchannels in a Microchip
Naoya Jinno, Masahiko Hashimoto, Kazuhiko Tsukagoshi*
Department of Chemical Engineering and Materials Science, Faculty of Science and Engineering, Doshisha University, Kyotanabe,
Japan.
Received March 1st, 2012; revised March 21st, 2012; accepted April 8th, 2012
ABSTRACT
Microfluidic behavior of ternary mixed carrier solvents of water-acetonitrile-ethyl acetate (2:3:1 volume ratio) was ex-
amined by use of a microchip incorporating microchannels in which one wide channel was separated into three narrow
channels, i.e., triple-branched microchannels. When the ternary carrier solution containing the fluorescent dyes, hydro-
phobic perylene (blue) and relatively hydrophilic Eosin Y (green), was fed into the wide channel under laminar flow
conditions, the carrier solvent molecules or fluorescence dyes were radially distributed in the channel, forming inner
(organic solvent-rich major; blue) and outer (water-rich minor; green) phases in the wide channel. And then, in the nar-
row channels, perylene molecules mostly appeared to flow through the center narrow channel and Eosin Y, which is
distributed in the outer phases in the wide channel, flowed through the both side narrow channels. A metal ion, Cu(II) as
a model, dissolved in the ternary mixed carrier solution was also examined. The Cu(II) showed fluidic behavior, trans-
ferring from the homogeneous carrier solution to the water-rich solution in the side narrow channels through the tri-
ple-branched microchannels.
Keywords: Triple-Branched Microchannels; Ternary Mixed Solvents; Fluorescence Dyes; Metal Ion; Tube Radial
Distribution Phenomenon (TRDP)
1. Introduction
The development of micro total analysis systems that
include a microchip or microfluidic device technology is
an interesting aspect of analytical chemistry [1,2]. Mi-
crofluidics exhibit various types of fluidic behavior of
solvents in a microchannel. The microfluidic behavior
has been examined by varying the channel configuration
and flow rate of the solvents, by using aqueous-organic
solvent mixtures, and by introducing specific obstacles
into the microchannel [3-5]. The fluidic behavior of the
solvents in the microchannel is related to the mixing,
separation, diffusion, and reaction of the solutes. Infor-
mation regarding the flow pattern of the solvents is im-
portant and useful to design microreactors or micro total
analysis systems [6,7].
Recently, we reported that when ternary mixed solvents
of water-hydrophilic/hydrophobic organic solvents are
fed into a microspace, such as microchannels in a micro-
chip or capillary tubes, the solvent molecules are radially
distributed under laminar flow conditions. This is called
the “tube radial distribution phenomenon” (TRDP) [8-
11]. In TRDP, in an organic solvent-rich carrier solution,
the organic solvent-rich major phase is formed around
the center of the microspace as an inner phase, while the
water-rich minor phase is formed near the inner wall as
an outer phase. On the other hand, in the water-rich car-
rier solution, the water-rich major phase is formed as an
inner phase and the organic solvent-rich minor phase as
an outer phase. TRDP forms a specific liquid-liquid
interface, which is not static but kinetic, in a micro-
space.
Until now, TRDP has been investigated by use of mainly
capillary tubes and straight single line microchannels
[8-11]. The triple-branched microchannels in which one
wide channel was separated into three narrow channels in
a microchip was designed and manufactured in our pre-
vious study to conform TRDP, where the ternary mixed
carrier solution was fed from the three narrow channels
to the one wide channels [8]. In this study microfluidic
behavior of the ternary mixed carrier solution in the tri-
ple-branched microchannels was examined with the re-
verse flow direction against the previous paper [8], i.e.,
*Corresponding author.

Microfluidic Behavior of Ternary Mixed Carrier Solvents Based on the Tube Radial Distribution
in Triple-Branched Microchannels in a Microchip
50
the flow direction form the one wide channel to the three
narrow channels. By the experiments, we could observe
unique fluidic behavior of the solvents and solutes in the
microchannels based on TRDP. The preliminary results
reported in a communication previously [12].
2. Experimental
2.1. Reagents and Materials
Water was purified with an Elix 3 UV system (Millipore
Co., Billerica, MA). All reagents used were obtained
commercially and were of analytical grade. Perylene,
Eosin Y, acetonitrile, ethyl acetate, and Cu(II) chloride
were purchased from Wako Pure Chemical Industries,
Ltd. (Osaka, Japan). Orange G was purchased from
Nacalai Tesque, Inc. (Kyoto, Japan). A microchip made
of quartz incorporating microchannels was manufactured
with Microchemical Technology (Kanagawa, Japan).
Figure 1 illustrates the setup of the microchip in the mi-
crochip holder (Figure 1(a)) and the enlarged view of the
microchip incorporating the triple-branched microchan-
nels (Figure 1(b)). As shown in Figure 1(b)), one wide
channel (300 μm wide × 40 μm deep) denoted as channel
W was separated into three narrow channels (each 100
μm wide × 40 μm deep) designated as channels N1-N3.
2.2. Fluorescence Microscope Equipped with a
CCD Camera System
A microchip incorporating the microchannels was set up
for the fluorescence microscope-CCD camera system.
Fluorescence in the microchannel was monitored using a
fluorescence microscope (BX51; Olympus, Tokyo, Japan)
equipped with an Hg lamp, a filter (U-MWU2, ex. 330 -
385 nm, em. >420 nm), and a CCD camera (JK-TU53H).
The ternary mixed solvents of the water-acetonitrile-
ethyl acetate mixture (3:8:4 volume ratio) contained 0.1
mM perylene and 1 mM Eosin Y. The carrier solution
was fed into the microchannels using a microsyringe
pump.
2.3. Cu(II) Analysis
The homogenous carrier solution of the water–acetone-
trile-ethyl acetate mixture (2:3:1 volume ratio) including
2.0 mM Cu(II) was fed into channel W at a definite flow
rate. 100 μL of the solution in channels N1-N3 was col-
lected through polytetrafluoroethylene (PTFE) tubes into
the corresponding vessel. The solution was dried under a
vacuum, and 0.5 M ammonia solution (100 μL) was
added to the residue for flow absorption measurement at
600 nm (modified SPD-6AV spectrophotometric detector,
Shimadzu Co., Kyoto, Japan).
3. Results and Discussion
3.1. Fundamental Experiments for TRDP in a
Microchannel
Microchips with single-branched microchannels having
widths of 100, 200, 300, and 400 μm and identical channel
depth of 40 μm were also manufactured. The influences
of channel width on TRDP in the microchannels were
examined with fluorescent dyes, perylene and Eosin Y,
dissolved in an organic solvent-rich carrier solution (wa-
ter-acetonitrile-ethyl acetate; 3:8:4 volume ratio). The
fluorescence images of the dyes in the solution fed into
the microchannels were observed with the fluorescence
microscope-CCD camera system (Figure 2). The flow
(a) (b)
Figure 1. Schematic representation of (a) setup of the microchip in the microchip holder and (b) the enlarged view of the mi-
crochip incorporating triple-branched microchannels.

Microfluidic Behavior of Ternary Mixed Carrier Solvents Based on the Tube Radial Distribution
in Triple-Branched Microchannels in a Microchip
51
rates in all microchannels (100 - 400 μm width) were
adjusted to achieve the same average linear velocity of
25 cm·min–1.
It is evident from Figure 2 that TRDP was observed in
all microchannels regardless of the channel width. Hy-
drophobic perylene molecule (blue) was distributed
around the center of the channel away from the side inner
walls of the channel, while relatively hydrophilic Eosin
Y molecule (green) was distributed near the side inner
walls of the channel. That is, the major inner phase (or-
ganic solvent-rich phase) and the minor outer phase (wa-
ter-rich phase) were formed in the microchannels of 100
- 400 μm width.
3.2. Effects of Flow Rates on TRDP in
Triple-Branched Microchannels
The effects of flow rate on TRDP in the triple-branched
microchannels were examined with an organic solvent-
rich carrier solution (water-acetonitrile-ethyl acetate,
3:8:4 volume ratio) containing Orange G by using the
bright-field microscope-CCD camera system. The Or-
ange G-containing carrier solution was fed at the flow
rates of 2.0 - 50.0 μL·min–1. The images are shown in
Figure 3. The hydrophilic Orange G molecule was dis-
tributed near the channel side inner walls, and the inter-
face between the water-rich outer phase and the organic
solvent-rich inner phase was distinguished because of the
color of Orange G (Figure 3). The TRDP was for the
first time observed with a bright-field microscope-CCD
camera system in our TRDP investigation. The system
was confirmed to be useful for TRDP observation in a
similar way to fluorescence system. At the low flow rates
of 2.0 and 5.0 μL·min–1, the liquid-liquid interface was
unstable, considerably changed, or appeared to instanta-
neously disappear, while the stability was observed at the
flow rates higher than 5 μL·min–1.
Thus, flow rates higher than 5 μL·min–1 created a sta-
ble liquid-liquid interface in the triple-branched micro-
channels under the present conditions.
Figure 2. Fluorescence images of fluorescent dyes dissolved
in a ternary mixed carrier solvent in microchannels with
channel widths of 100, 200, 300, and 400 μm and identical
channel depth of 40 μm. Conditions: Carrier, water-ace-
tonitrile-ethyl acetate (3:8:4 v/v/v) mixture including 0.1
mM perylene and 1 mM Eosin Y; and flow rates, 1, 2, 3,
and 4 μL·min–1 for microchannel widths of 100, 200, 300,
and 400 μm, respectively.
Figure 3. Effects of flow rates of the carrier solution on
TRDP. Conditions: Carrier, water-acetonitrile-ethyl acetate
(3:8:4 v/v/v) mixture containing 2 mM Orange G and flow
rate, 2.0 - 50 μL·min–1 for channel W.
(a) (b)
Figure 4. Fluorescence images of fluorescent dyes dissolved
in a mixed carrier solvent in triple-branched microchannels.
(a) Water-acetonitrile (1:4 v/v) and (b) water-acetonetrile-
ethyl acetate (3:8:4 v/v/v) mixtures. Conditions: Carrier,
aqueous-organic mixtures including 0.1 mM perylene and 1
mM Eosin Y; and flow rate, 6.3 μL·min–1 for channel W.
3.3. Fluorescence Images of the Dyes Based on
TRDP in Triple-Branched Microchannels
The microchip incorporating triple-branched microchannels
was shown in Figure 1(b) with their measurements. The
organic solvent-rich carrier solution (water-acetonitrile-
ethyl acetate, 3:8:4 volume ratio) containing Eosin Y and
perylene was fed into channel W of the microchip at the
flow rate of 6.3 μL·min–1. The mixture carrier solution
excluding ethyl acetate (water-acetonitrile, 1:4 volume
ratio) but containing the dyes was also fed into the
channel as a reference. Figure 4 shows the fluorescence
images of the dyes in the triple-branched microchannels
of the microchip. Figures 4(a) and (b) were obtained
with the water-acetonitrile mixture as a reference and the
water-acetonitrile-ethyl acetate mixture as ternary mixed
solvents, respectively.
The distribution behavior of the dyes, i.e., TRDP, was
not observed in the image of Figure 4(a) with the water-
acetonitrile mixture. On the contrary, the image of
Figure 4(b) with the ternary solvent mixture shows TRDP,
in which the hydrophobic perylene molecule (blue) was
distributed around the center of channel W away from
the channel side inner walls, while relatively hydrophilic
Eosin Y molecule (green) was distributed near the channel
side inner walls. The distribution phenomena of the ternary
mixed carrier solvents fed into the microchannels was
evident in the wide channel. And then, perylene molecules

Microfluidic Behavior of Ternary Mixed Carrier Solvents Based on the Tube Radial Distribution
in Triple-Branched Microchannels in a Microchip
52
mostly appeared to flow through the center channel
(channel N2) and Eosin Y, which is distributed in the
outer phases in channel W, flowed through channels N1
and N3, as shown in Figure 4(b).
3.4. Distribution of Eosin Y Based on TRDP in
Triple-Branched Microchannels
The homogenous carrier solution of the water–acetone-
trile-ethyl acetate mixture (3:8:4 volume ratio) including
0.5 mM Eosin Y was fed into channel W at a flow rate of
10 μL·min –1. 100 μL of the solution in channels N1-N3
was collected through PTFE tubes into the corresponding
vessel. The solution was subjected to flow absorption
measurement at 517 nm (modified SPD-6AV spectro-
photometric detector, Shimadzu Co., Kyoto, Japan). Eo-
sin Y concentrations in the center channel (channel N2)
and the two side channels (channels N1 and N3) were
0.36, 0.59, and 0.58 mM, respectively (Figure 5). The
concentrations were averages for five measurements. The
experimental data indicated that relatively hydrophilic
Eosin Y was distributed from the homogeneous carrier
solution to the water-rich solution in the side narrow
channels rather than the organic solvent-rich solution in
the central narrow channel. The distribution of the fluo-
rescent dyes in the triple-branched microchannels might
suggested the possibility of extraction or separation for
some solutes by utilizing the phase interface formed by
TRDP in these channels.
3.5. Distribution of Cu(II) Based on TRDP in
Triple-Branched Microchannels
Distribution of Cu(II) was examined in the triple-
branched microchannels by utilizing the kinetic liquid-
liquid or aqueous-organic interface formed by TRDP.
First, the calibration curve of Cu(II) was examined in a
manner similar to that described in the experimental sec-
tion (Sec 2.3). Cu(II) was determined over the concentra-
tion range of 0.5 - 10 mM with good linearity (R2 =
0.999).
Figure 5. Distribution of Eosin Y from the homogeneous
ternary mixed solution into the outer phases in the side
narrow channels by TRDP. Conditions: Carrier, water-
acetonitrile-ethyl acetate (3:8:4 v/v/v) mixture; flow rate, 10
μL·min–1 for channel W; Eosin Y concentration in the ho-
mogeneous solution, 0.50 mM.
The homogenous carrier solution of the water-ace-
tonitrile-ethyl acetate mixture (2:3:1 volume ratio) in-
cluding 2.0 mM Cu(II) was fed into channel W at the
flow rate of 40 μL·min–1. 100 μL of the solution in chan-
nels N1-N3 was collected with PTFE tubes into the cor-
responding vessel. Cu(II) concentrations in the center
wide channel (channel N2) and the two side channels
(channels N1 and N3) were 0.8, 2.5, and 2.7 mM, respec-
tively (Figure 6(a)). The concentrations were averages
for five measurements. As a reference, Cu(II) concentra-
tions were examined with the water-acetonitrile (2:3
volume ratio) carrier solution in a similar manner. Cu(II)
concentrations in channels N1-N3 were 2.1, 2.1, and 2.0
mM, respectively (Figure 6(b)). The experimental data
indicated that Cu(II) was extracted to the water-rich solu-
tion in the side narrow channels from the homogeneous
solution by the formation of a phase interface in the mi-
crochannel; we refer to this process as tube radial distri-
bution extraction (TRDE).
Moreover, we examined the Cu(II) distribution by
TRDE at other flow rates of 32 and 63 μL·min–1 in
channel W. At 32 μL·min–1, Cu(II) concentrations in the
center channel (channel N2) and the two side channels
(channels N1 and N3) were 0.8, 3.0, and 2.9 mM,
whereas, at 63 μL min–1, those were 1.1, 2.9, and 2.7 mM,
respectively. Comparing with the Cu(II) concentration,
2.0 mM, in the homogeneous carrier solution, the Cu(II)
concentration in the solution collected from the center
channel (channel N2) decreased, while that from the two
side channels (channels N1 and N3) increased at every
(a)
(b)
Figure 6. Distribution of Cu(II) in triple-branched micro-
channels. (a) Water-acetonitrile-ethyl acetate (2:3:1 v/v/v)
mixture and (b) water-acetonitrile (2:3 v/v) mixture carrier
solution. Conditions: Flow rate, 40 μL·min−1 for channel W
and Cu(II) concentration in the homogeneous solution, 2.0
mM.

Microfluidic Behavior of Ternary Mixed Carrier Solvents Based on the Tube Radial Distribution
in Triple-Branched Microchannels in a Microchip
53
flow rates. The obtained data showed that TRDE oc-
curred in the triple-branched microchannels of the mi-
crochip.
4. Conclusion
TRDP has been investigated by use of mainly capillary
tubes and straight single line microchannels. The TRDP
of the ternary mixed carrier solution in the triple-
branched microchannels was observed with fluorescence
microscope-CCD camera system. Consequently, pery-
lene molecules mostly appeared to flow through the cen-
ter narrow channel and Eosin Y flowed through channels
in the side narrow channels. The TRDP was also ob-
served through bright-light microscope-CCD camera
system. We tried to apply the phase interface formed by
TRDP, i.e., inner and outer phases, to the distribution or
extraction procedure of Cu(II) in the triple-branched mi-
crochannels. This process will be helpful for investigat-
ing separation and extraction in a microfluidic device.
5. Acknowledgements
This work was supported by a Grant-in-Aid for Scientific
Research (C) from the Ministry of Education, Culture,
Sports, Science, and Technology, Japan. This work was
also supported by “Advanced Study for Integrated Parti-
cle Science, and Technology,” Strategic Development of
Research Infrastructure for Private Universities, the Mini-
stry of Education, Culture, Sports, Science, and Techno-
logy, Japan.
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