Journal of Membrane Science 352 (2010) 116125

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Journal of Membrane Science
j our nal homepage: www. el sevi er . com/ l ocat e/ memsci
Analysis of the mass transfers in an articial kidney microchip
Aissa Ould-Dris
b
, Patrick Paullier
a
, Laurent Griscom
c
, Ccile Legallais
a
, Eric Leclerc
a,
a
CNRS-UMR 6600, Laboratoire de biomcanique et de bio ingnierie, Universit de Technologie de Compigne, BP 20529, 60205 Compigne Cedex, France
b
EA: 4297, Laboratoire de transformation intgre de la matire renouvelable, Universit de Technologie de Compigne, BP 20529, 60205 Compigne Cedex, France
c
SATIE CNRS UMR 8029, ENS Cachan - Bretagne, Campus de Ker Lann, 35170 Bruz, France
a r t i c l e i n f o
Article history:
Received 23 October 2009
Received in revised form 26 January 2010
Accepted 1 February 2010
Available online 6 February 2010
Keywords:
Articial kidney
Polydimethylsiloxane (PDMS)
Polyethersulfone membrane (PES)
Mass transfer
Clearances
Microuidic
Microchip
a b s t r a c t
In this communication we demonstrate a conception of an articial microkidney using pertinent
microtools that more accurately mimic organ functions in vitro. We present a technique to integrate
polyethersulfone (PES) membranes usually used in hemodialysis inside a polydimethylsiloxane (PDMS)
microchip. The purpose of the microchip is to model glomerular ltration on-chip. Mass transfer of
urea (60Da), vitamin B12 (1355Da) and albumin (70,000Da) are investigated by using two types of
membranes (cut-off at 500,000Da and 40,000Da) in co-current and counter-current ow conditions.
The time of urea, vitamin B12 and albumin removal, and the mechanisms of mass transfer, are controlled
either by controlling the pore size of the membranes or by controlling the pressure proles along the
membrane via the owconditions. An analytical model, which is supported by our data, is put forth. The
model allows the extractionof the diffusioncoefcients of eachmolecule throughthe various membranes
studied. Due to the downscaling, the model and the experiments demonstrate that the dialysance in the
microchip is expressed by the sumof the diffusion and convection mass transfer components. The results
of this work support an analytical model which describes the mass transfer in a microchip modelling a
glomerular unit. Coupled with the advantages of the microuidic biochip (high surface/volume ratio,
reduction of the uid volumes), our data will complete the integration of further cellular functions for
the utilisation of the present microchip as a future in vitro model of a miniaturized bio articial kidney.
2010 Elsevier B.V. All rights reserved.
1. Introduction
In toxicity studies, the effects of a molecule or drug are often
tested directly with animals (in vivo studies) or by using the spe-
cic cells targeted by the molecule (in vitro studies). However,
most in vitro studies cannot reproduce in vivo studies due to the
lack of the systemic interferences and interactions between tissues
and organs of a live organism [1,2]. Advances in bioengineer-
ing are allowing the development of various complex bioreactors
that can reproduce the interactions between the organs such
as the liver, kidney, intestine or the lung [35]. These bioreac-
tors consist of cell culture chambers containing engineered tissue
and cell cultures interconnected and irrigated by a uidic net-
work that control the ows of culture medium. The irrigation
using a microuidic architecture permits continuous cell feeding,
and waste removal in addition to chemical loading and regula-
tion. Using these types of bioreactors, the toxicity of metabolites
of naphthalene or of benzo[a]pyrene, both metabolized by the
liver (in a liver cell culture chamber) and then transported to
a targeted organ (to the lung or to the intestine respectively

Corresponding author. Tel.: +33 03 44 23 79 43; fax: +33 03 44 23 79 42.

E-mail address: eric.leclerc@utc.fr (E. Leclerc).
for naphthalene or of benzo[a]pyrene) has been investigated
[35].
In order to rene cell culturing models and methods for toxico-
logical studies, bioreactors are nowbeing miniaturized frommacro
to micro scales [68]. The miniaturisation of the bioreactors allows
the reduction of the quantity of uids involved in cell culture and in
turn the reduction of chemicals used [9]. In addition miniaturisa-
tion technologies allowworking with physiological time of contact
between the investigated molecules and targeted tissues.
As the main organ of detoxication is the liver, there is a large
panel of studies working on the miniaturisation of liver bioreac-
tor [1012]. However, only a few works deal with articial kidney
bioreactors. The kidney is an organ of prime interest in toxicity
studies since it is involved in the process of ltration and excretion
of toxic metabolites.
Kaazempur-Mofrad et al. investigated the integration of a mem-
brane (with a 0.4m porosity) and the mass transfers of urea and
creatinine inside a microchip [13]. Combined with the progress
in bioarticial kidney technology [14], nano porous ultraltration
membranes [15], and Micro Electro Mechanical Systems (MEMS)-
based microchips [16], the works on renal microchips using a
dialysis membrane are hoped to be extended to miniaturized
portable renal replacement systems. However, to achieve this
goal, the microchips must provide a greatly increased surface area
0376-7388/$ see front matter 2010 Elsevier B.V. All rights reserved.
doi:10.1016/j.memsci.2010.02.007
A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125 117
Fig. 1. SEM images of the PES membranes: (A) 1FPH; (B) 8F. The scale bar is 10m.
and a signicant improvement in the clearance efciency of ure-
mic wastes over current hemodialysis technologies. Ideally, the
miniaturisation would enable the microchip to be portable and
potentially wearable [15].
In a recent study, Baudoin et al. demonstrated the feasibility of
culturing Madin Darby Canine Kidney cells (MDCK cells) inside a
polydimethylsiloxane (PDMS) microchip [17]. The microchip, used
as a miniaturized bioreactor, has several advantages compared to
traditional in vitro methods including a higher surface to volume
ratio, and dynamic chemical loadings. The results of the cultures
in the miniaturized bioreactor coupled with the effect of dynamic
ammoniumchloride loading onMDCKcell cultures have illustrated
the potential of using microchips for wider in vitro chronic toxicity
investigations. In the present paper, we investigate a glomerular
like ltration microchip based on the integration of a polyether-
sulfone membrane (PES) inside a PDMS microsystem that includes
microchannel networks and ltration chambers. The glomerular
ltration and the mass transfers in the microchip are characterized
by investigating experimentally and theoretically the behaviours
of urea, vitamin B12 and albumin molecules.
2. Materials and methods
2.1. Membranes
We used two types of at polyethersulfone (PES) membranes
providedby Membrana GmbH(reference numbers: MicroPES 1FPH
and MicroPES 8F). The polyethersulfone material is chosen because
it is widely used in the hemodialysers. The manufacturer gives the
following properties of the membranes. The 1FPH and 8F nomi-
nal pore sizes are 0.04m and 0.8m respectively, corresponding
to a ltration cut-off of molecular weights near 40,000Da and
500,000Da. SEM views of the membrane are shown in Fig. 1A
and B. The pure water transmembrane ow rates are respec-
tively 4mL/(mincm
2
bar) and245mL/(mincm
2
bar) at 25

Cfor the
1FPH and 8F membranes according to the manufacturer. The 1FPH
membrane is classied at the transition between the microiltra-
tion and the ultraltration processes, whereas the 8F membrane
is a microltration membrane. The thickness of the membranes
is 100m.
2.2. Microchip
The microchip is composed of three layers, including two PDMS
layers in which a PES membrane is sandwiched. Fig. 2A shows the
layered assembly of the parts composing the ltration microchip.
Each PDMS layer includes an inlet and outlet network and a l-
tration chamber. The PDMS ltration chamber and inlet/outlet
networks are fabricated by a photo lithographically dened replica
mouldingas describedintheliterature[18]. Theltrationchambers
are 200m deep, 11mm large and 13mm long. A microchannel
array of 100m in depth is located at the bottom of each cham-
ber, as shown by Fig. 2B. This array is added in order to create a
sieve, which is designed to avoid blockage of the ltration by the
deformationof the membrane onthe PDMS surface during the uid
perfusion. In each PDMS layer, inlet and outlet ports are drilled,
resulting in two separate microuidic circuits. An inlet and out-
let microchannel network (100m deep) is used to connect the
ltrationchamber andthe inlet andoutlet ports (Fig. 2A). The mem-
brane is sandwiched between the two PDMS layers. The microchip
integrity and the prevention of leaks are guaranteed by a sample
holder with 8 screws. The total surface available for ltration was
1.3cm
2
. The microchip and the sample holder are shown in Fig. 2C.
Fig. 2. (A and B) Principle of the microchip; (C) photography of the microchip.
118 A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125
Fig. 3. Experimental setup: (A) co-current (full arrow) and counter-current (dashed
arrow) owcircuits used during the mass transfer experiments; (B) dead end mode
for the transmembrane pure water ow rate determination.
For comparative purposes, a microchip without a sieve structure
has been also built.
2.3. Experimental setup
In order to analyse the mass transfer in the microchip, two per-
fusion circuits were used as shown in Fig. 3A. The rst circuit was
used to ow the retentate solution in which the molecule to lter
was loaded. The second circuit corresponded to the permeate solu-
tion in which the molecule to be ltered was extracted. Each circuit
included a 10mL uid reservoir and a peristaltic pump. In order to
control the pressure, four manometers were connected at the inlet
and outlet ports of the microchip (ports dened in Fig. 2A).
The experimental value of the transmembrane pure water ux
(P
H
2
O
) was measuredinthe microchip. For that purpose, ports 2and
3 were closed (Figs. 2A and 3B). The water was fed through port 1
and exited at port 4. The transmembrane pure water ux through
the membrane was measured in dead end ltration mode. A pre-
cision scale weighing the water collected at the port 4 was used
to calculate the ow rate. The membrane water resistance (R
m
)
was extracted from this ux by the relation: R
m
= PTM/Jf
H
2
O
(in
which is the viscosity, Jf
H
2
O
the transmembrane pure water ux
and PTM the transmembrane pressure P
1
P
2
dened in Table 1).
2.4. Filtration experiments
Co-current ow conditions were achieved by a ow stream in
the retentate and in the permeate owing in the same direction
(and therefore coming from respectively the ports 1 and 3). We
dened the counter-current ow conditions when the retentate
Table 1
Nomenclature, parameters and experimental extracted values used in the model.
1 Retentate footnote
2 Permeate footnote
f Filtrate footnote
i Inlet footnote
R Outlet footnote
L Length of the chamber 1cm
b Width of the chamber 1cm
e Height of each compartment 150m
Q Flow rate m
3
/s
Q
f
Filtration ow rate m
3
/s
Fluid velocity m/s
P Pressure Pa
C Concentration g/L
DL Dialysance m
3
/s
x
*
Abscise where Q
f
=0 m
Viscosity Pa s
V Volume of each compartment 1.510
9
m
3
Rm Membrane water resistance of the 1FPH
membrane
7.510
10
m
1
Rm Membrane water resistance of the 8F
membrane
7.510
9
m
1
C
10
Solute initial concentration in the retentate Section 2.5
C Equilibrium concentration Section 2.5
P
H
2
O
Experimental transmembrane water ux
(1FPH)
8mL/(mincm
2
bar)
P
H
2
O
Experimental transmembrane water ux
(8F)
80mL/(mincm
2
bar)
Dm Coefcient of diffusion of urea (1FPH) 210
12
m
2
/s
Dm Coefcient of diffusion of urea (8F) 210
11
m
2
/s
Dm Coefcient of diffusion of vitamin B12
(1FPH)
410
13
m
2
/s
Dm Coefcient of diffusion of vitamin B12 (8F) 610
12
m
2
/s
Dm Coefcient of diffusion of albumin (1FPH) 910
14
m
2
/s
Dm Coefcient of diffusion of albumin (8F) 210
13
m
2
/s
Thickness of the membranes 100m
and permeate arrived in the microchip from opposite directions.
This meant that the retentate arrived from port 1 and the perme-
ate from port 4. The ow rates used in the study were 150L/min
and 300L/min in co-current or counter-current conditions. Each
experiment for each membranes and owconditions was repeated
at least 2 times independently.
2.5. Solutes and measurements
The mass transfer was tested with a urea solution which has a
molecular mass of 60Da. This molecule was usedas a lowmolecular
weight marker in our experiments. The retentate solution was con-
centrated at 36mg/mL whereas initially the permeate solution was
pure water. In order to sample the solution, the perfusion pumps
were stopped and 50L was sampled from the retentate and per-
meate reservoirs. The urea concentration was measured using the
Bioassay Quantichrom
TM
Urea Assay Kit. The equilibrium concen-
tration was theoretically 18mg/mL.
The mass transfer was also tested with a vitamin B12 solution
which has a molecular mass of 1355Da. This molecule was used as
a middle molecular weight marker. The retentate solutionwas con-
centrated at 120mg/L whereas initially the permeate solution was
pure water. The vitamin B12 concentration was measured in the
retentate and permeate reservoirs using a spectrophotometer at
360nm. The equilibrium concentration was theoretically 60mg/L.
The mass transfer was nally testedwitha bovine albuminsolu-
tion which has a molecular mass of 70,000Da. This molecule was
used as a large molecular weight marker. The retentate solution
was concentrated at 1000mg/L in PBS solution (phosphate buffer
saline) whereas initially the permeate solution was PBS solution
(phosphate buffer saline). The albumin concentration was mea-
sured in the retentate and in the permeate reservoirs using a
spectrophotometer at 280nm. The equilibrium concentration was
theoretically 500mg/L.
A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125 119
Fig. 4. Schematic of the modelled ltration chamber and the associated pressure proles in (A) counter-current and (B) co-current ow conditions.
2.6. Theory
We propose a model to describe the transport of the molecule
in the microchip and through the membrane. The nomenclature is
summarized in Table 1. The model takes into account the following
assumptions:

the osmotic pressure is neglected (H1);

the inlet andoutlet effects are neglectedinbothcompartments of

the ltration chambers meaning that the ows were fully devel-
oped (H2);

hypothesis H2 leads that there is no boundary layer of uidveloc-

ity occurring along the membrane (H3);

themainresistancetothediffusivetransfer is locatedinthemem-
brane (there is no diffusion boundary layer in the compartments)
(H4);

the volume of the uid in each compartment is assumed to be

that of the reservoir (volume of the connections was neglected)
(H5);

the concentrations ineachcompartment were constant along the

membrane due to the very short length of the membrane (H6);

the viscosity in each compartment are equal due to high dilution

of the solutes (H7).
The objective of the model is to determine the concentrations
of the solutes vs. time in the retentate and permeate solution in
order to obtain a predictive method of the mass transfers. The pur-
pose of the model is also to establish an analytical method to nd
the dialysance and to characterize the efciency of the microchips.
The model is based on the solution of the mass balance equations,
the convective and diffusive equations of mass transfer across the
membrane and the transmembrane pressure equations along the
membrane. In the model, the retentate (that can be understood as
the plasma circuit in conventional hemodialyser) is denoted by the
uid 1, whereas the permeate is denoted as the uid 2 as shown
in Fig. 4.
The mass balance in the retentate and permeate is given by
dV
1
dt
+
dV
2
dt
=
dV
1
dt
=
dV
2
dt
= 0 (1)
This equation indicates that the initial volumes in both reser-
voirs were completely lled and remained constant during the
experiments. V
1
is the initial volume of the retentate witha solution
having an initial concentration of solute C
10
.
This leads to the following mass conservation:
d(V
1
C
1
)
dt
+
d(V
2
C
2
)
dt
= 0 V
1
dC
1
dt
+V
2
dC
2
dt
= 0

_
C
1
C
10
dC
1
=
V
2
V
1
_
C
2
0
dC
2
C
10
C
1
C
2
=
V
2
V
1
C
2
= (C
10
C
1
)
V
1
V
2
(2)
If the volume of uid in each compartment remains constant
then the average transmembrane pressure is equal to zero. Thus,
the mass balance in the retentate compartment can be written as:
dV
1
dt
=
_
L
0
J
f
bdx =
1
R
m
_
L
0
(P
1
P
2
)bdx = 0

_
L
0
P
1
dx =
_
L
0
P
2
dx (3)
in which J
f
is the transmembrane ux dened by J
f
= (P
1

P
2
)/
1
R
m
. In these equations we suppose that is no boundary layer
of uid velocity occurring along the membrane (H2 and H3).
We denoted x
*
the membrane abscissa in which the pressure
in each compartment P
1
(x

) = P
2
(x

), which leads to the following

mass ux in the retentate:
V
1
dC
1
dt
= b
_
x

0
_
J
f
C
1
+
D
m

(C
1
C
2
)
_
dx
b
_
L
x

_
J
f
C
2
+
D
m

(C
1
C
2
)
_
dx (4)
In this equation, it is assumed that there is no boundary layer of
the solute concentration along the membrane (H4). It is assumed
that the concentrations in each compartment were constant along
the membrane due to the very short length of the membrane (H6).
This leads to the following equation:
V
1
dC
1
dt
= b
_
C
1
_
x

0
J
f
(x) dx +C
2
_
L
x

J
f
(x) dx +
(C
1
C
2
)D
m

_
L
0
dx
_
(5)
120 A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125
Table 2
Experimental dialysance in L/min for the urea, the vitamin B12 and albumin for the 1FPH and 8F membranes.
Molecules
Urea Vitamin B12 Albumin
Co-current
a
Counter-current
a
Co-current
a
Counter-current
a
Co-current
a
Counter-current
a
150/150-1FPH 0.12L/min 0.3L/min 0.03L/min 0.174L/min 0.0054L/min
300/300-1FPH 0.12L/min 0.48L/min 0.03L/min 0.34L/min 0.0054L/min
150/150-8F 1.2L/min 0.36L/min 0.012L/min
300/300-8F 1.2L/min 0.36L/min 0.012L/min
a
Flow conditions.
Tosolve Eq. (5), we must calculate the pressure ineachcompart-
ment. Inthe model, we have neglected the sieve and have modelled
each compartment of the ltration chambers as a Hele Shawcham-
ber (be, parameters dened in Table 2 and Fig. 4). To determine
the pressure prole, we assume a fully developed ow and neglect
any lateral effects (H2). Indeed Beavers and Joseph [19] proposed
a model of the slip velocity along the membrane in laminar and
Stokes ow conditions. This model has been extended in the case
of a slowvelocityof ltrationwhencomparedtothe upper free ow
velocity [2022]. It is shown in these particular conditions that the
porous media can be consider as an impermeable media leading
to a velocity of ltration in the order of (1/R
m
)
0.5
. Thus, in a rst
approximation we assumed that the slip velocity is 0 in our ow
conguration. In our case, this led to consider an adherence condi-
tion at the membrane surface. The hypothesis will be validated and
discussed in the result section. Therefore, the pressure gradient in
each compartment was given by the HagenPoiseuille law:
dP
1
dx
= 12

1
v
1
e
2
1
(6)
dP
2
dx
= 12

2
v
2
e
2
2
To solve the system of the set of equations (6) we suppose that
the uids have the same viscosity (H7). We can also suppose that
the pressure prole is linear over the length of each compartment.
If the ltrate ow rate Q
f
does not exceed 1% of the ow rates Q
1
and Q
2
in the two compartments then it can be considered that the
pressure prole in each compartment is linear, meaning that the
difference of the transmembrane pressure is too weak to modify
the velocity in each compartment.
The ltrate ow rate, Q
f
is then dened as the internal ltration
ow rate calculated with the following equation:
Q
f
=
_
x

0
J
f
bdx =
b
R
m
_
L/2
0
(P
1
P
2
) dx (7)
In this equation, J
f
is equal to 0 at the abscissa x
*
=L/2. In the case
of the co-current ow conditions, Eq. (7) becomes:
Q
f
=
b
R
m
_
L/2
0
(P
1
P
2
) dx = 6
b
R
m
_
v
1
e
2

v
2
e
2
__
L/2
0
(L 2x) dx
=
3L
2
2R
m
e
3
(Q
1
Q
2
) (8)
in which Q
1
= v
1
be
1
and Q
2
= v
2
be
2
with e
1
=e
2
=e in our case.
we dene
Q
2
Q
1
= k then Q
f
=
3L
2
Q
1
LR
m
e
3
(1 k) (9)
FromEq. (9) we may notice that if Q
1
andQ
2
are equal thenQ
f
=0
Q
f
Q
1
=
3L
2
2R
m
e
3
(1 k)
Q
f
Q
2
=
3L
2
2R
m
e
3
1 k
k
(10)
The following equations are satised if the pressure proles are
linear and therefore if Q
f
/Q
2
<1%. This meant that we must check
that the following relation between the geometrical and physical
parameters is valid:
Q
f
Q
2
0.01 k
1
(1 +0.02R
m
e
3
)/3L
2
(11)
A similar analysis for the counter-current ow conditions from
Eq. (7) lead to an equation of Q
f
given by:
Q
f
=
b
R
m
_
L/2
0
(P
1
P
2
) dx = 6
b
R
m
_
v
1
e
2
+
v
2
e
2
__
L/2
0
(L 2x) dx
=
3L
2
2R
m
e
3
(Q
1
+Q
2
) (12)
This leads to write
Q
f
=
3L
2
Q
1
2R
m
e
3
(1 +k);
Q
f
Q
1
=
3L
2
2R
m
e
3
(1 +k);
Q
f
Q
2
=
3L
2
2R
m
e
3
(1 +k)
k
(13)
The linear pressure prole assumption is valid in counter-
current ow condition only if
Q
f
Q
2
0.01 k
1
0.02R
m
e
3
/3L
2
+1 (14)
Taking into account Eqs. (8) and (12), Eq. (5) can be rewritten as
follows:
V
1
dC
1
dt
=
_
Q
f
(C
1
C
2
) +
LbD
m

(C
1
C
2
)
_
=
__
Q
f
+
LbD
m

_
(C
1
C
2
)
_
(15)
Using Eq. (2), and expression of equilibrium concentration C

,
C

= C
10
(V
1
/(V
1
+V
2
)), the differential equation above can be
rewritten as follows:
dC
1
dt
=
__
Q
f
+
LbD
m

__
V
1
+V
2
V
1
V
2
_
(C
1
C

)
_
(16)
The solution of Eq. (16) is:
C
1
(t) = C
1
+(C
10
C

) exp
_

_
Q
f
+
LbD
m

__
V
1
+V
2
V
1
V
2
_
t
_
(17)
The average ux is given by the following equation:

J
f
=
Q
f
bL/2
(18)
Then the Peclet number across the membrane comparing con-
vective and diffusive mass transfer across the membrane can be
A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125 121
expressed by:
Pe =

J
f

D
m
=
Q
f
bL/2

D
m
(19)
Therefore, the differential equation (17) can be expressed using
the Pe number:
C
1
(t) = C
1
+(C
10
C

) exp
_
Q
f
_
1 +
2
Pe
__
V
1
+V
2
V
1
V
2
_
t
_
(20)
In co-current ow with Q
1
=Q
2
, Q
f
becomes equal to zero and
only diffusive transport takes place across the membrane. In this
case, Eq. (17) reduces to the following Eq. (21)
C
1
(t) = C
1
+(C
10
C

) exp
_

LbD
m

_
V
1
+V
2
V
1
V
2
_
t
_
(21)
The linearized formof Eq. (20) can be used to validate the model
with experimental results
ln
C
1
(t) C

C
10
C

= Q
f
_
1 +
2
Pe
__
V
1
+V
2
V
1
V
2
_
t (22)
Thus, experimentally the Pe number can be determined from
the slope and D
m
deduced.
The dialysance, DL, expresses the mass transfer (between the
inlet and outlet of the retentate compartment, via the relation:
Q
1i
Q
1r
C
1r
, the scales are dened in Table 1) per unit of time nor-
malized by the concentration difference between the retentate and
the permeate. The dialysance, that takes into account the multi
pass ows via the concentration C
2
, is expressed by the relation
according to Winger and Weiner [23]:
DL =
Q
1i
C
1i
Q
1r
C
1r
C
1i
C
2i
(23)
In the present model, we assume that the mass transfer in the
retentate can be expressed simply by the termV
1
(dC
1
/dt) leading
to rewrite Eq. (23) as follows:
DL = V
1
dC
1
dt
_
1
C
1
C
2
_
(24)
DL = V
2
dC
2
dt
_
1
C
1
C
2
_
(25)
The solution of Eqs. (24) and (25) lead to
ln[C
1
(t) C
2
(t)] = DL
_
1
V
1
+
1
V
2
_
t (26)
Thus, the dialysance can be deduced experimentally from the
slope of Eq. (26). Following the present theory, the modelled dialy-
sance is given by Eq. (15), Eqs. (24), (25) and (26):
DL =
_
Q
f
+
LbD
m

_
(27)
3. Results and Discussion
3.1. Microchip design and fabrication
In conventional PDMS microchip fabrication processes, the
microuidic network can be sealed to a PDMS or glass surface
by exposing the surfaces to a RF generated oxygen plasma that
allows a strong bonding of the PDMS to other PDMS or glass sub-
strates. This effect is assumed to be due to the oxidation of the
surface by the plasma creating OH bonds. However, as is the case
with PES membranes, we were unable to bond PDMS to it using
the plasma bonding technique. To achieve hermetic seal between
the PES membrane and the PDMS structures, we have devised a
microchip holder that allowed us to clamp and maintain pressure
on the PDMS parts. The holder allowed working with pressures up
to 1m of water (corresponding to 100mbar). In terms of ow rate,
this led to a maximal working ow rate of 700L/min. At higher
ow rates (and pressures) there was a radial leakage of the uid
through the membrane (data not shown).
3.2. Determination of the membrane water resistance
The values of the membrane water resistance (R
m
) are shown
in Table 1. In the case of the 1FPH membrane, the calcu-
lated resistance is equal to 7.510
10
m
1
resulting from the
experimentally measured transmembrane water ux equal to
81mL/(mincm
2
bar) (n10). This value is twice as high as
claimed in the manufacturers data (4mL/(mincm
2
bar)). In the
case of the measurements using the 8F membrane, the value of
the transmembrane water ux decreased with the time due to the
membrane clogging. Therefore the value of the transmembrane
water ux was 160mL/(mincm
2
bar) at the rst measurement
(compared to 245mL/(mincm
2
bar) value given by the manu-
facturer) and decreased to 80mL/(mincm
2
bar) at the second
measurement (n10). A limit value down to 20mL/(mincm
2
bar)
was reached after repeating the measurements consecutively. For
that reason, the transmembrane water ux was characterized
before and after each ltration experiment for all membrane. In
addition, the membrane was changedfor every ltrationtest. Using
this experimental procedure, a membrane water resistance (R
m
) of
7.510
9
m
1
was calculated, corresponding to a transmembrane
water ux of 80mL/(mincm
2
bar).
For comparative purposes, in the microchip without sieve, we
were unable to ow water through the membrane. This led to
an articially low value of the transmembrane ow rate (0.1 and
0.004mL/(mincm
2
bar) for the 8F and 1FPH membranes respec-
tively). This result can be attributed to the deformation and the
contact of the membrane with the bottom surface of the ltration
chamber. In addition, with this type of microchip, we observed a
corrugation of the membrane that locally increased the pressure
prole on the membrane. In this situation, we cannot assume a lin-
ear pressure prole along the membrane. The use of the sieve was a
key component in the design to achieve reproducible mass transfer
experiments.
3.3. Dialysis mode
In the case of the 150/150L/min and 300/300L/min owrate
conditions in co-current situations, mass transfers of the urea, of
the vitamin B12 and of the albumin were not affected by the vari-
ation of the ow rate as shown in Fig. 5AC. As similar pressure
proles were obtained in both uids along the membrane, it could
be expected that the local transmembrane pressure was always
close to zero leading to small convective effects. This suggested
the absence of diffusion boundary layer that could inuence the
mass transfers and to consider that all the resistance to the dif-
fusion was located in the membrane itself. Should the blood or
dialysate resistance be signicant, they would be different accord-
ing to the perfusion ow rate (i.e. the higher ow rate, the lower
dialysate resistance), which was not observed. This result was con-
sistent with the assumption in the model where the expression of
the diffusive ux(Eq. (4)) was not functionof these additional resis-
tances observed in conventional hemodialysers [24]. In the present
experimental cases, the diffusionwas theoreticallythe onlyprocess
of mass transfer.
For the three molecules and all ow rate conditions, we found
that the mass transfer kinetics depended on membranes pore size.
The equilibrium with urea was reached after 20min with the 8F
membranes whereas 150min were needed with the 1FPH ones
(Fig. 5A). This was attributedtothelarger poresizeleadingtohigher
coefcient of diffusion through the membrane. The data obtained
with the vitamin B12 showed the same tendencies as presented
122 A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125
Fig. 5. Superimposition of the modelled and experimental values of the concentra-
tion of the urea (A), of the vitamin B12 (B) and of the albumin (C) in the retentate
in co-current ow conditions with the 1FPH membranes (Exp: 150/150; Exp:
300/300, Model: black line) and with the 8F membranes (Exp: * 150/150; Exp:
300/300, Model: dashed line).
in Fig. 5B. In addition, the vitamin B12 mass transfer rate was
lower compared to urea transfer due to its larger molecular weight
(1355Da vs. 60Da). Finally, the equilibrium with albumin could
not be reached after 6h of perfusion using the 1FPH membranes
(Fig. 5C).
The diffusion coefcient, D
m
, in Eq. (21) was the only parame-
ter to adjust in the model in order to t the experimental results
obtained in the dialysis experiments in which there is no other
governing mass transfer mechanism. The other equation param-
eters were the geometrical parameters of the ltration chamber
(the discussion of the parameter choice will be performed in a
next section). The model led to a diffusion coefcient for urea of
210
12
m
2
/s for the 1FPH membranes and of 210
11
m
2
/s for
the 8F membrane (Table 2). In the case of the vitamin B12, the
model tted the experiences well if the diffusion coefcients were
set to410
13
m
2
/s for the1FPHmembranes andto610
12
m
2
/s
for the 8F membranes respectively. Finally, for the albumin, the
model tted the experiences if the coefcients of diffusion were
set to910
14
m
2
/s for the1FPHmembranes andto210
13
m
2
/s
Fig. 6. Superimposition of the modelled and experimental values of the concentra-
tion of the urea (A) and of the vitamin B12 (B) in the retentate in co-current (Exp:
150/150; Exp: 300/300; Model: black line) and counter-current (Exp: 300/300;
Model: dotted line; Exp: 150/150, Model: dashed line) owconditions used with
the 1FPH membranes.
for the 8F membranes respectively. These data were in agreement
withtheexpectedresults; namelytheobtainedcoefcients demon-
strated that urea diffusion was faster than vitamin B12 which was
faster than albumin diffusion. For both membranes and the three
molecules, the co-current ow conditions allowed working with a
Q
f
value equal to 0L/min as shown by Eq. (9). The model assump-
tions of noslipvelocity, Poiseuille prole andlinear pressure prole
(Q
f
/Q
1
<0.01) were therefore valid. In addition, this led to a Peclet
number equal to zero, representing a fully diffusion mode of mass
transfer. Finally, the dispersion between the theory and the exper-
iments in dialysis was found below 10%.
3.4. Diffusion-convection modes
In counter-current owconditions, the mass transfer was faster
due to an additional convection process through the membrane.
This was due to different transmembrane pressure. When com-
pared to the co-current ow conditions, the transfer time was
reduced as shown in Fig. 6A and B in the case of the 1FPH mem-
branes for the urea and vitamin B12. In counter-current ow
conditions, an increase in ow rates reduced the transfer time by
increasing the ow through the membrane (due to the local trans-
membrane pressure). For urea, this time was reduced from 60min
to 20min when the ow rates were increased from 150L/min to
300L/min (120min to 90 for the vitamin B12 respectively).
A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125 123
To model these experiments, we used Eq. (20), in which the D
m
values were those previously extracted from the dialysis experi-
ments. The model was dependent on the Peclet number, Pe, and the
ltrationowrate Q
f
. Inour owcongurations, we hadinEqs. (12)
and (13) k =1, that led to Q
f
0.18L/min and 0.36L/min when
Q
1
was set to 150L/min and 300L/min respectively with the 1
FPHmembrane. These values were in agreement with the assump-
tion of a linear pressure prole in each compartment, dened by
Q
f
/Q
1
<0.01 in the model. In addition these equations showed that
the augmentation of the ow rate (Q
1
and Q
2
) lead to an increase
in the value of Q
f
and therefore an increase of mass transfer via
convection (ultraltration process).
The Peclet numbers were 3 and 6 at 150L/min and 300L/min
respectively during the urea experiments. In counter-current ow,
both convection and diffusion appeared to play a role in the mass
transfer illustrated by a value of Pe equal to 3. This also explains
the faster transfer when compared to the co-current ow condi-
tions due to the addition of both phenomena (Fig. 6A). Then, in our
microchip, when the owrates Q
1
and Q
2
were increased, the con-
vectionof urea became more dominant (Pe =6). This was illustrated
by a faster mass transfer.
Due to the size of the molecules, the diffusion process was
weaker when we used the vitamin B12 when compared to the con-
vection process. This tendency was illustrated with Peclet numbers
equal to 15 and 30 at 150L/min and 300L/min respectively. In
those cases, the transfer of the molecules mainly resulted from
convection through the membrane. Thus for urea, both diffusion
and convection played an additional role in the mass transfer of
the urea while the mass transfer of the vitamin B12 was mainly
due to the convection. It should be noted that the difference
between the Peclet number between both experiments (urea and
vitamin B12 ltration with the 1FPH membranes) is dependent
Fig. 7. Sensibility of the model using the parameters of Table 2: (A) to the diffusion coefcient of the solute (: 410
12
cm
2
/s; : 410
13
cm
2
/s; : 410
14
cm
2
/s);
(B) to the ow rates (: Q
1
=Q
2
=300L/min; : Q
1
=Q
2
=75L/min; : Q
1
=Q
2
=150L/min); (C) to the height of the modelled chamber (: 300m; : 150m; :
75m); (D) to the ow rates if Dm =410
12
cm
2
/s (:Q
1
=Q
2
=150L/min; : Q
1
=Q
2
=300L/min; : Q
1
=Q
2
=75L/min); (E) to the transmembrane water ow rates at
Dm =410
13
cm
2
/s (: 80mL/(mincm
2
bar); : 1mL/(mincm
2
bar); : 8mL/(mincm
2
bar)).
124 A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125
Table 3
Clearances (CL) for the urea and the vitamin B12 reported in the literatures in microchip and in conventional hemodialysers compared to our microchip dialysance (DL) when
we used the 1FPH membranes in counter-current ow conditions.
CL or DL (mL/h) Q
1
/Q
2
Q
2
(mL/min) Surface (cm
2
) Q
2
/A (mL/(mincm
2
)) Membrane type
Present work
a
(urea) 0.018 1 0.15 1 0.15 PES (40kDa)
0.029 1 0.3 1 0.3 PES (40kDa)
Present work
a
(vit B12) 0.01 1 0.15 1 0.15 PES (40kDa)
0.02 1 0.3 1 0.3 PES (40kDa)
Kaazempur-Mofrad
a
(urea) [13] 15 1 0.033 0.0013 PC (200kDa)
40 1 0.033 0.0042 PC (200kDa)
70 1 0.033 0.007 PC (200kDa)
Brunet et al. (urea) (hollow bers) [27] 1380 1 16 6000 0.0027 AN69 (40kDa)
2640 1 32 6000 0.0053 AN69 (40kDa)
Jaffrin et al. (vit B12) (hollow bers) [24] 3300 1 500 11,500 0.043 AN69 (40kDa)
a
Data obtained in microchip.
on the coefcients of diffusion of the molecules through the
membrane.
Finally, for the urea and vitamin B12 using the 1FPHmembrane,
the dispersion between the theory in diffusion/convection mode
and the experiments was found at 10% without changing the value
of the coefcient of diffusion for each molecule in each membrane.
When the counter-current ow conditions were applied with
the 8F membrane, the retentate owed directly in the permeate.
The volume of uids in each circuit could not be kept constant
(and the retentate owing directly to the permeate, it did not per-
mit sampling of the solutions for measurements). There was no
more tangential owinside the chamber. These observations were
in opposition with the model assumption of Eq. (1) in which the
volumes in both reservoirs were initially completely lled and
remained constant during the experiments. Locally, this can be
explained by a pressure difference through the membrane so high
that the retentate penetrated directly in the permeate circuit at
the port locations. Thus, experimental and analytical results did
not match. In these situations, we found that Q
f
1.8L/min at
150L/min and the ratio Q
f
/Q
1
=0.012 meaning that the Poiseuille
prole assumption began to be incorrect. It probably resulted from
the apparitionof a slipvelocity showing the limitationof the model.
3.5. Model sensitivity to parameters
To superimpose the analytical and experimental data, we had
to introduce the geometrical and physical parameters in the
model. The transmembrane owrates used in the calculation were
extracted from the experimental data. The model did not take into
account the microchannels of the sieve and the microchannels of
the inlet and outlet networks. The ltration model represents an
ideal situation with two parallel at ow chambers separated by
a xed, inexible membrane. To reproduce the experimental data
we used a value of 150mthat corresponded to the mean value of
the height of each chamber (see Section 2.2). In Figs. 5 and 6, we
presented the data obtained with 150m. The coefcient of diffu-
sion for each membrane was determined by using the experiments
of each molecule in the diffusion modes.
InFig. 7, we have shownthe sensitivity of the model tothe trans-
membrane ow rate (ltration ow rate), the diffusion coefcient,
the chamber height and the ow rates. In the diffusion-convection
mode (Eq. (20)), the convection is an important phenomenon when
the microchip included the 1FPH membranes for the intermedi-
ate size molecules (such as vitamin B12). This was illustrated by
a small variation of the ltration time when the coefcient of dif-
fusion ranged from 410
13
m
2
/s to 410
14
m
2
/s (Fig. 7A). In
addition, theltrationtimewas stronglyinuencedbythevariation
of the ow rates or the chamber height at 410
13
m
2
/s (Fig. 7B
and C). On the contrary, at higher values of the coefcients of diffu-
sion (see at 410
12
m
2
/s in Fig. 7A), such as for the urea and the
small molecules, the diffusion in the microchip was not neglige-
able. In those cases both phenomena of convection and diffusion
acted on the ltration times (Fig. 7D). Finally, the sensitivity to the
transmembrane ow rate was presented. An augmentation of the
transmembrane owrate decreased the ltration time. This can be
understood by a higher porosity of the membrane (Fig. 7E).
3.6. Clearance and dialysance in the microchip
In Table 2, we have reported the dialysance of urea, vitamin
B12 and albumin studying the microchips. As it can be seen from
Table 3, the dialysance increasedwhenthe membrane porosity was
increased in the dialysis modes whereas the augmentation of the
size of the molecules contributed to reduce the dyalisance. Fur-
thermore, the augmentation of the ow rates did not increase the
dialysance. However, inconvectiondiffusionmodes, the dialysance
increased with the ow rate augmentation. In addition, the dialy-
sance decreased when the molecule size increased. This was due to
a predominance of the convectionterminthe equations incounter-
current ows when compared to the diffusion term(Pe 1). In our
microchip, the convection through the at membrane was there-
fore an important phenomenon of mass transfer even for the small
molecules. The model shows that the dialysance is the sum of the
diffusive term (via D
m
) and the convective term (via Q
f
). This ten-
dency is not usual inconventional hemodialysis inwhichthe global
mass transfer is mainly due to the diffusive mass transfer [24,25].
This is due to the asymptotic saturation of the mass transfer [24].
This leads to the expression of the dialysance and the clearance in
the conventional hemodialysis which is not the simple sum of the
convective and diffusive terms [2426]. We may notice that in the
microchip the membrane size is very small and we assumed that
there is no boundary layer to take into account during the mass
transfer. This results to a term Q
f
representing the convection that
is as much as important as the diffusion.
For comparative purposes, in Table 3, we have reported the
data described in the literature using conventional hemodyalisers
[24,27] and another type PDMS microchip previously developed
with a polycarbonate membrane (PC) [12]. The clearances of the
data found for the urea and vitamin B12 in the conventional
hemodialyser were higher than our dialysance. Those works were
done with an AN 69 membrane that have a typical cut-off about
30,00040,000Da (similar tp the 1FPH membrane in this study
[28]). Due to a smaller ow rate in our microchip (150L/min
and 300L/min compared to 16mL/min and 32mL/min [27], and
500mL/min[24]), anda short lengthanda small membrane surface
area (1cm
2
, when compared to 6000cm
2
[27] and to 11,500cm
2
[24]) the dialysance in our microchip was lower than in opti-
mized conventional hemodyaliser. In spite of the efciency of the
A. Ould-Dris et al. / Journal of Membrane Science 352 (2010) 116125 125
microchip appearing to be lower than those types of conventional
hemodialysers at their best working conditions, the comparison
must be balanced by the potential of the microchip to work with
smaller ow rates and smaller membrane surfaces. For the utili-
sation of the microchip as a miniaturized model of a glomerular
ltration, the obtained dialysances (up to 0.029mL/(hcm
2
)) illus-
trated the potential of our microchip as a miniaturized unit.
Finally, thecomparisonwiththestudyusingthemicrochips pre-
viously developed in the literature [13] must be balanced by the
type of membrane introduced in those microchips. Our microchip
seemed less efcient but those microchips using a polycarbonate
microitration membrane that have typical pore size larger than
0.2m corresponding to a cut-off above 10
6
Da. This explained
their high clearance compared to the present microchip data using
the 1FPH PES membranes (40,000Da).
4. Conclusions
We have introduced a microchip in which the mass trans-
fers of urea, vitamin B12 and albumin solutes were characterized.
The mass transfers with two types of membrane of polyethersul-
fone and for three types of molecules have been presented. The
microchip could be used either in dialysis mode or in diffusion-
convection mode. The introduction inside the microchip of a sieve
allowed controlling the mass transfer processes either by control-
ling the pressure inside in the ltration chamber or by controlling
the membrane porosity. This demonstrated that the mass trans-
fer could be discriminated with our microchip according to the
targeted applications. In addition, the proposed analytical model
allowed tting the experimental data with the same diffusion coef-
cient value for both co-current and counter-current ow. The
model adequacywiththeexperiments is demonstratedbytheinde-
pendence of the diffusion coefcient whatever the ow regimes
for a given couple of membrane and molecule. This is an encour-
aging result because it will permit to use the model as a predictive
analysis tool of our glomerular like microchip. Finally, in order
to propose a complete bio articial kidney, the proximal tubule re
absorption has to be included and investigated in the microchip.
Acknowledgments
This work was granted by the ANR Program PCV. We thank B.
Harten and Dr. H.D. Lemke fromMembrana GmbHfor the supply of
membranes used in this study and their fruitful discussions during
the microchip development. We also thank Pr. Michel Jaffrin for his
comments during the paper redaction.
References
[1] A. Guillouzo, Liver cell models in in vitro toxicology, Environ. Health Respect.
106 (1999) 511532.
[2] S.A. Roberts, Drug metabolism and pharmacokinetics in drug discovery, Curr.
Opin. Drug Disc. Dev. 6 (2003) 6680.
[3] S.H. Choi, O. Fukuda, A. Sakoda, Y. Sakai, Enhanced cytochrome P450 capac-
ities of Caco-2 and Hep G2 cells in new coculture system under static and
perfused conditions: evidence for possible organ-to-organ interactions against
exogenous stimuli, Mater. Sci. Eng. C 24 (2004) 333339.
[4] Y. Sakai, O. Fukuda, S.H. Choi, A. Sakoda, Development of a biohybrid simu-
lator for absorption and biotransformation processes in humans based on in
vitro models of small intestine and liver tissues, J. Artif. Organs 6 (4) (2003)
273281.
[5] L.M. Sweeney, M.L. Shuler, A cell culture analogue of rodent physiology: appli-
cation to naphthalene toxicology, Toxicol. In Vitro 9 (1995) 307316.
[6] L. Grifth, G. Naughton, Tissue engineeringcurrent challenges and expanding
opportunities, Science 295 (2002) 10091014.
[7] K. Viravaidya, M.L. Shuler, Incorporation of 3T3-L1 cells to mimic bioaccumu-
lation in a microscale cell culture analog device for toxicity studies, Biotechnol.
Prog. 20 (2004) 590597.
[8] K. Viravaidya, A. Sin, M.L. Shuler, Development of a microscale cell culture
analog to probe naphthalene toxcity, Biotechnol. Prog. 20 (2004) 316323.
[9] G.M. Walker, H.C. Zeringue, D.J. Beebe, Microenvironment design considera-
tions for cellular scale studies, Lab Chip 4 (2004) 9197.
[10] E. Leclerc, Y. Sakai, T. Fujii, Perfusion culture of fetal human hepatocytes in
microuidic-environments, Biochem. Eng. J. 20 (23) (2004) 143148.
[11] M.J. Powers, K. Domansky, A. Udapadhia, M.R. Kazempur-Mofrad, P. Kursawky,
D. Janigan, K.E. Wack, D.B. Stolz, R. Kamm, L. Grifth, A microarray perfusion
bioreactor for 3D liver culture, Biotechnol. Bioeng. 78 (2001) 257269.
[12] M.J. Powers, D. Janigan, K.E. Wack, Baker, D.B. Stolz, L. Grifth, Functional
behavior of primary Rat liver cells in a three-dimensional perfused microarray
bioreactor, Tissue Eng. 8 (3) (2002) 499513.
[13] M.R. Kaazempur-Mofrad, J.P. Vacanti, N.J. Krebs, J.T. Borenstein, A MEMS based
renal replacement system, in: Solid-State Sensor, Actuator and Microsystems
Workshop, South Carolina, June 610, 2004.
[14] E. Tziampazis, H.D. Humes, Extracorporeal kidney, in: A. Atala, R.P. Lanza (Eds.),
Methods of Tissue Engineering, Academic Press, San Diego, 2002, p. 987.
[15] W.H. Fissell, H.D. Humes, A.J. Fleischman, S. Roy, Dialysis and nanotechnology:
now, 10 years, or Never? Blood Purif. 25 (2007) 1217.
[16] Z. Yang, S. Matsumoto, R. Maeda, A prototype of ultrasonic micro-degassing
device for portable dialysis system, Sens. Actuators A 95 (2002) 274280.
[17] R. Baudoin, L. Griscom, M. Monge, C. Legallais, E. Leclerc, Development of a
renal microchipfor invitro distal tubule models, Biotechnol. Prog. 23 (5) (2007)
12451253.
[18] Y. Xia, G.M. Whitesides, Extendingmicrocontact printingas amicrolithographic
technique, Langmuir 13 (1997) 20592067.
[19] Beavers, D. Joseph, Boundary conditions at a naturally permeable wall, J. Fluid
Mech. 30 (1967) 197207.
[20] H. Ene, E.E. Sanchez-Palencia, Equations and surface phenomena for the ow
in a porous medium, J. Mech. 14 (1975).
[21] T. Levy, E. Sanchez-Palencia, On boundary conditions for uid ow in porous
media, Int. J. Eng. Sci. 13 (1975) 923940.
[22] A. Mikelic, Homogenization theory and applications to ltration through
porous media Filtration in Porous Media and Industrial Applications1998, Lec-
ture Notes Centro Internazionale Matematico Estivo (C.I.M.E.) Series, Lecture
Notes in Mathematics, vol. 1734, Springer, 2000, pp. 127214.
[23] K. Winger, M. Weiner, Acetate transfer across membranes of articial kidney
in vitro, Kidney Int. 15 (1979) 593600.
[24] M. Jaffrin, L. Ding, J.M. Laurent, Simultaneous convective and diffusivr mass
transfers in a hemodialyser, J. Biomech. Eng. 112 (1990) 212219.
[25] M. Jaffrin, B. Gupta, J. Malbrancq, A one dimensional model of silmutaneous
hemodialysis andultraltrationwithhighlypermeablemembranes, J. Biomech.
Eng. 103 (1981) 261266.
[26] C. Legallais, G. Catapano, B. von Harten, U. Baurmeister, A theorical model to
predict the in vitro performance of hemodialters, J. Membr. Sci. 168 (2000)
315.
[27] S. Brunet, M. Leblanc, G. David, D. Parent, S. Courteau, J. Cardinal, Diffusive and
convective solutes clearances during continuous renal replacement therapy at
various dialysate abd ultraltration ow rates, Am. J. Kidney Dis. 34 (3) (1999)
486492.
[28] A. De Vriese, F. Colyardin, J. Philipp, R. Vanholder, J. De Sutter, N. Lameire,
Cytokine removal during continuous hemoltration in septic patients, J. Am.
Soc. Nephrol. 10 (1999) 846853.