60 BioProcess International 9(8) SEPTEMBER 2011

B I O P R O C E S S TECHNICAL
An Approach to Design and
Performance Testing of an
Impeller-Driven Single-Use Mixer
Bojan Isailovic and Bruce Rawlings
PRODUCT FOCUS: ALL BIOLOGICS
PROCESS FOCUS: PRODUCTION,
MANUFACTURING
WHO SHOULD READ: MANUFACTURING
AND PROCESS ENGINEERS, ANALYTICAL
KEYWORDS: FLUID DYNAMICS, GOOD
ENGINEERING PRACTICE, DISPOSABLES
LEVEL: INTERMEDIATE
S
ingle-use technology is now
well established for many
bioprocessing unit operations,
including sterile filtration and
product storage, where it provides
specific benefits such as elimination of
cleaning and cleaning validation,
reduced risk of contamination, and
enclosed systems for safer handling.
Recently, single-use technology has
been introduced into more complex
unit operations such as purification by
chromatography (1), tangential-flow
filtration (2), and mixing systems.
More complex processing steps,
however, present a challenging task for
developers of single-use products.
Mixing systems fall into this category
because the range of applications can
be very wide, and mixing requirements
often differ substantially among those.
Developers of single-use mixers must
take into consideration several factors:
liquidliquid and solidliquid mixing,
suspensions, immiscible liquids, fluid
viscosity, shear-sensitive fluids,
susceptibility to foaming, container
shape and dimensions, process scale,
mixing times, fluid-contact materials,
capital and running costs, sampling
and monitoring, extended operation
and durability, particulate generation,
power transfer, footprint, hardware
requirements, and temperature control.
Here we report on an extensive
study of the design, development, and
performance testing of a single-use
mixing system for liquidliquid and
solidliquid mixing, incorporating
features that provide ease of use and
mixing capabilities that are comparable
with those of stainless steel systems.
PRINCIPLES OF MIXER DESIGN
Our initial task was to look at the
principles behind traditional mixer
design to determine how we might
apply them to a single-use engineered
solution that required ease of use for
multiple installations. In parallel, we
also reviewed the design and
performance attributes of currently
available single-use systems to identify
their strengths and limitations and
take those into account when
designing our new system.
Currently available single-use
mixer systems can be grouped into
four types: rocking mixers, pumped-
loop mixers, stir-bar mixers, and
mechanically driven impeller mixers.
Rocking Mixers: These systems are
based on wave generation inside a
flexible bag placed on a flat-bed
rocker (Figure 1). The noninvasive
nature of the mixer is a benefit, but
we were concerned about mixing
efficiency for poorly soluble solids as
well as scalability, power input, and
the general complexity of controls
necessary to ensure effective mixing
with different fluid types and fill
volumes.
Tote ready for use (TOP) and with
uninflated mixer in position (BOTTOM)
62 BioProcess International 9(8) SEPTEMBER 2011
Pumped-Loop Mixers: This type
relies on pumping fluid from a
container through a recirculating loop
to induce mixing within a container
(Figure 2). Although it provides a
simple and contained process, fluid
velocities in the container can be
severely limited by pumping capacity
of the loop. This limitation can restrict
those capabilities, especially regarding
solids dispersion and sedimentation of
low-solubility solids, mixing times,
and high-viscosity fluids. Power input
is generally low in these systems, but it
remains an acceptable approach for
liquidliquid applications.
Stir-Bar Mixers: Magnetically
driven stir bars or levitating impellers
linked to superconducting drives have
been incorporated into several single-
use mixing systems (Figure 3). Such
mixers have the benefit of simplicity
but can suffer from insufficient power
input for some applications, from
restrictions on flow dynamics within
containers to scalability.
Mechanically Driven Impeller
Mixers: A mechanically driven
impeller or paddle mixer (Figure 4)
has some significant benefits over
other types, especially regarding
power input and flow patterns.
However, this approach requires
careful consideration of engineering
principles for incorporation into a
single-use system.
In a recent evaluation of single-use
mixers (3), Sinclair and Monge noted
that each type currently available was
restricted by its design to specific
applications and that no system was
optimal for general-purpose mixing.
Similarly, Strahlendorf and Harper (4)
identified an unmet need for scalable
systems and also found that the
mixing systems were not currently
optimized for suspension products.
They believe this represents an
opportunity for innovation.
It was evident from our review that
none of the existing designs could
meet all of our criteria for an easy-to-
use, scalable mixer capable of handling
a very broad range of applications. We
concluded, however, that a
mechanically driven impeller design
offered potentially the best
opportunity, provided that some basic
design and engineering issues could be
addressed successfully.
CRITICAL REQUIREMENTS
FOR AN IMPELLER MIXER
In our studies on traditional mixer
design, we identified the basic
engineering principles and
requirements that would be critical in
development of a versatile, high-
performance, mechanically driven
impeller mixer for single-use
applications. The most important
properties (and their significance) can
be summarized as follows.
Power Input: Mixing efficiency is
strongly influenced by power input
(transfer of energy into the mixing
process). The benefit of an impeller
directly coupled to a motor is that
power input is maximized and can be
precisely controlled and adjusted.
Factors influencing power input
include fluid density, impeller power
number, speed, and diameter.
Flow Patterns: A flow pattern that
distributes fluid rapidly and efficiently
throughout a container is an important
requirement. This pattern must ensure
an absence of dead zones, especially in
the container extremities. Flow must
accommodate mixing of low-density
powders on a liquid surface and of
high-density solids in the base of the
container, as well as liquidliquid
mixing and high-viscosity fluids.
These are very different and
demanding requirements.
Flow regimes are defined by
turbulent and laminar-flow
components. Strong turbulent flow is
desirable when using turbine impellers
and can be advantageous for many
mixing applications. On the other hand,
some impellers operate well in a laminar
flow regime (e.g., close-clearance
impellers in high-viscosity fluids).
Shear forces carry out mixing
processes. In an impeller system, shear
forces represent shear stress and are
induced primarily by a combination of
two effects:
Fluid velocity and relative motion
of liquid layers within a container (the
dominant component for impeller
mixers in most, but not all, fluids)
Shear forces in the immediate
vicinity of impeller blades (where a
small amount of fluid experiences
high shear).
Container Geometry: The shape and
dimensions of a container are critical
properties that can substantially
influence flow patterns and mixing
Figure 1: Rocking mixer action Figure 2: Pumped-loop mixer action
Figure 3: Magnetic stir-bar mixing action

Figure 4: Mechanically driven impeller
mixing action
64 BioProcess International 9(8) SEPTEMBER 2011
performance. Most stainless steel,
stirred tanks are cylindrical with a
liquid height:diameter ratio greater
than one (H/T > 1), and they often
have dished bottoms to prevent
accumulation of solids in their corners
(Figure 5). This cylindrical shape can
induce undesirable tangential flow or
swirling of the liquid resulting in
reduced power draw by the impeller
and lower mixing efficiency, but such
effects can be minimized by the
introduction of wall baffles.
By contrast, many single-use
containers have a rectangular cross-
section (Figure 6), with a 3D profile
that is either cuboid (H/T = 1) or
rectangular (H/T < 1). This
noncylindrical profile can be
beneficial for two reasons: First, the
reduced height can improve top-to-
bottom, axial-flow distribution; and
second, the rectangular profile can
minimize undesirable tangential flow,
the corners acting like a baffle to
break up the regular flow pattern and
promote vertical flow.
We considered other properties and
features in our design approach, as well:
for example, dynamic sealing of an
impeller shaft, access for liquids and
solids, external hardware (tote), materials
of construction, and control systems.
Those aspects are described below.
APPLYING CRITICAL REQUIREMENTS
FOR DESIGN AND FUNCTION
First and foremost, we wanted to
ensure that our new design would be
based on proven engineering principles
for mixer performance. We first
approached this objective through a
detailed research study into the
scientific and engineering principles of
mixing processes and system design.
This study involved a substantial
amount of engineering expertise and
experience within our working
environment, together with the
application of good engineering
practice (GEP) during development
and testing (5).
Container Design: We based our
container design on an existing three-
dimensional (3D) single-use
biocontainer, using the same film
material as it used. This approach
enabled us to benefit from proven
performance and qualification studies
established for the existing product.
However, that containers geometry
a rectangular cross-section, a shorter
height than width, and a flat bottom
is different from that of a
conventional cylindrical mixing vessel.
Those characteristics required that
special features be incorporated to
ensure achievement of the required
mixing properties and performance.
Impeller Type: Designing an ideal
impeller for the wide range of
intended mixing duties was a critical
part of this project. To help explain
the basis for our final decision, we
summarize below the main types and
properties of general-purpose
impellers, which fall into three main
categories: radial flow, hydrofoil, and
axial flow.
Radial-Flow Impellers: An example
of a radial-flow impeller is a paddle
stirrer with rotating vertical flat
blades. This design discharges fluid
predominantly radially (horizontally)
toward the vessel wall. It can create
higher shear and turbulence at the
blade surface than some other types
will, but it also provides low pumping.
Radial action produces flow patterns
(Figure 7) with two circulating flow
loops, one above and one below the
impeller. Mixing occurs between those
loops. Such a flow pattern is not ideal
because it causes stratification or
compartmentalization in a tank,
resulting in poor homogeneity of the
bulk fluid. Although the flow pattern
can be converted to a strong top-to-
bottom flow using suitable baffles, we
did not consider this type of impeller to
be ideal for a general-purpose, single-
use system.
Hydrofoil impellers contain tapered,
twisted blades. Their flow pattern has a
greater vertical (top to bottom)
distribution than flat-blade impellers do,
and the flow is very uniform and
streamlined, with lower shear than
other types have. Although these
properties are favorable for many mixing
applications, the flow pattern cannot be
reversed by changing the rotation of the
hydrofoil. For reasons discussed below, a
reverse-flow feature was an important
requirement for our mixer, so we did not
select a hydrofoil impeller.
Axial-Flow Impellers: An example of
an axial flow impeller is a pitched-
blade impeller with angled blades
(Figure 8). This generates both axial
and radial flow in low- to medium-
viscosity fluids, as shown in Figure 9
Figure 5: Mechanically driven impeller
mixing action, stainless steel mixer
H
T
H / T > 1
Figure 6: Mechanically driven impeller
mixing action, single-use mixer
T
H
H / T 1
Figure 7: Radial-flow impeller flow pattern
66 BioProcess International 9(8) SEPTEMBER 2011
with a down-pumping flow. The
impeller produces slightly higher shear
at its blade surface than a hydrofoil
does, which gives a good balance
between pumping and shear action. So
this is considered to be a good general-
purpose impeller. Therefore, we chose
a pitched-blade design with four blades
for our development program.
Impeller Blade Pitch Angle: The
angle of a blades pitch influences
several mixing properties, from the
balance between axial and radial flow
to the pumping capability and shear
rate. These combined effects affect
both mixing performance and flow
patterns of a mixer.
One other key factor helped
determine our final selection of pitch
angle: namely the requirement to
operate a mixer in both down-flow
and up-flow pumping directions by
reversing the motor and shaft rotation.
This would provide greater flexibility
for use. To ensure the same pitch
angle for both pumping directions, we
incorporated a 45 angle. The benefits
of this feature are discussed below.
Blade Diameter: The ratio of blade
diameter to vessel diameter (D/T) is
an important parameter because it
affects flow pattern and power input
and consequently mixing efficiency.
D/T values of 0.330.50 have been
studied and used in design and
operation of conventional stirred
tanks. Larger blade diameters (higher
D/T ratios) provide higher power
input at lower speeds, which improves
mixing. However, it also increases
radial flow at the expense of axial
flow, which changes the flow pattern
and can negatively affect mixing
performance. With D/T ratios of 0.55
and higher, flow becomes
predominantly radial and not ideal for
general-purpose mixers.
As with many variables, it is a
matter of arriving at the optimum
balance for a given application. For
example, we sized the blade for a
200-L rectangular container to give a
D/T ratio of 0.41 and 0.55 for the
respective wider and narrower
container dimensions we offer. That
range ensures good power input and
sufficient axial flow.
Impeller clearance from the tank
bottom needs to be taken into
consideration in a mixer design. The
ratio of clearance to liquid height
(C/H) is an important parameter that
ranges typically from 1/3 to 1/6. For
down-flow pumping impellers, a very
small clearance can cause
predominantly radial flow with high
shear and reduced pumping. But that
may be beneficial for sweeping solids
from the tank bottom at the
expense of bulk homogeneity.
For our single-use system, we found
a C/H ratio in the region of 1/4 to be
optimal for mixing performance in the
wide range of duties defined by user
requirements. Nevertheless, we set the
ratio at 1/7. Lowering the impeller
nearer to the base of the container
allows for use of smaller working
volumes without exposing the impeller
above the liquid surface. Such exposure
can generate excessive turbulence and
undesirable effects on sensitive
products. In-house mixing tests at this
clearance showed satisfactory mixing
efficiency. This feature can also assist in
the drainage phase of a mixing process
to ensure homogeneity of the whole
solution and good product recovery.
Pumping Direction: We considered
the ability to operate an impeller in a
down-flow or up-flow pumping
direction to be beneficial for
extending the range of mixing duties
that our system could fulfill. By
reversing the motor rotation, our
pitched-blade impeller with its 45
angle could provide this function.
In many applications,
counterclockwise rotation is preferred
Figure 8: Pitched-blade impeller Figure 9: Pitched-blade impeller flow pattern Figure 10: Flanged drain port and outlet valve
Nonwetted
locating plate
Figure 11: Solidliquid mixing, 30 g/L TSB in 200 L water at 200 rpm

0.0
0.2
0.4
0.6
0.8
1.0
1.2
0 5 10 15 20 25 30 35
Time (minutes)
N
o
r
m
a
l
i
z
e
d

U
V
2
8
0

A
b
s
o
r
b
a
n
c
e
Draining Mixing
SEPTEMBER 2011 9(8) BioProcess International
67
for down-flow pumping. It has the
specific benefit of creating an area of
high turbulence below the impeller to
lift solid particles that may rest in the
container bottom.
Conversely, clockwise rotation for
up-flow pumping creates an area of
turbulence above the impeller. This
can be preferred when incorporating
light solids into bulk fluids and for
gentler mixing. It is particularly
beneficial when fluid is sensitive to air
entrainment or foaming, such as in
media preparation with low-density
powders.
Impeller Speed: It is important to
operate a pitched-blade impeller at a
speed that will ensure turbulent flow.
Our studies showed that the chosen
impeller design could achieve
turbulence in the range of 100200 rpm
for most applications. The exception is
with mixing of very viscous fluids, for
which higher speeds may be required
but can be limited by total power and
heat build-up due to viscous energy
dissipation. If that limitation
significantly restricts mixing efficiency,
a different and more appropriate
impeller may be needed (e.g. a close-
clearance impeller).
Wall Baffles: We considered the
effects of baffles on flow patterns and
mixing performance to be sufficiently
beneficial for some applications to
include this optional feature into our
single-use mixer. Baffles can reduce
swirl and help convert tangential flow
to vertical flow, which enhances top-
to-bottom mixing. Vortices created at
high impeller speeds are also
minimized by baffles to reduce air
entrainment and foaming. We made
the baffles optional using removable
vertical plates attached to three sides of
the tote, with the fourth side
containing its doors. Their width is
based on standard baffle configurations
used in conventional stirred tanks (8%
and 10% of the containers width and
depth, respectively).
Drain ports at the base of
containers potentially introduce dead
zones that can trap solids and
unmixed fluids. To eliminate that
possibility, we designed a specially
engineered drain port with an integral
flange flush at the bottom surface
(Figure 10). Actuation by opening and
closing the drain port happens
through a bayonet mechanism that
gives a positive indication of the port
position (either fully open or fully
closed). This function is critical for
solidliquid applications to ensure no
partial opening of the port or
entrapment of solids in operation
before the outlet valve is opened.
Tote Design: We based the tote for
supporting our flexible mixing system
on a design previously developed for
3D storage containers, after adapting it
to incorporate the various attachments
(motor drive unit for impeller shaft,
air/gas manifolds, baffles, and control
system). Full-width double doors were
incorporated to provide ease of access
for installation of the mixer assembly.
And we had to take other features into
consideration for our final design: e.g.,
a small footprint on weigh scales, low
height (<1.4 m, 4.5 ft), mobility, and
weight.
The user control interface (UCI) in
Photo 1 was designed to control the
impellers direction and speed, to
manage interlocks for security in
operation, and to provide for inflation
and a gas blanket. As part of normal
mixer operation, the mixing chamber
is first inflated before addition of
fluid and powder. That ensures a
consistent envelope for mixing a range
of volumes within the system and
provides an opportunity for gas
blanketing (using nitrogen for
example) when oxygen-sensitive
materials are mixed.
PERFORMANCE TESTING
We designed our single-use mixing
system for use in a wide range of
mixing operations. It was therefore
important to carry out performance
testing that simulated such operations,
in some cases representing worst-case
conditions. We evaluated different
solids and liquids under a range of
conditions. Our investigations can be
summarized as follows: liquidliquid
and liquidsolid mixing; low-density
and high-viscosity liquids; floating
and settling solids; high-foaming
PRELIMINARY STUDY RESULTS
AND CONCLUSIONS
Results confirmed that a mechanically
driven, pitched-blade impeller could
deliver adequate power and flow
distribution for these duties.
SolidLiquid Applications
Short mixing times of <1 minute to
three minutes
Efficient lifting of high-density solids
from bottom of container
Effective entrainment of a floating,
slightly hydrophobic powder
No foaming even at fast impeller speed.
LiquidLiquid Applications
Homogeneity of system in <30 seconds
Capable of mixing very high-viscosity
fluids (1,000 cP)
Photo 1: User control interface
Table 1: Mixing times for different applications
Fluid phase Type of Test Test Materials Mixing Time
Solidliquid Settling solids NaCl (2 kg) into water < 1 minute*
Solidliquid Floating solids Terrific broth (1 kg) into water 3 minutes***
Liquidliquid Homogeneity NaCl (300 g/L): NaOH (10M) and
red tracer dye
<30 seconds**
Liquid Homogeneity
(high viscosity)
Red tracer dye into glycerol
>1,000 cP (160 L)
20 minutes**
* Conductivity measurement ** Visual inspection of dye dispersion ***Turbidity measurement
68 BioProcess International 9(8) SEPTEMBER 2011
liquids; effects of impeller speed,
baffles, pumping direction, and liquid
volume; dead-zone tests; homogeneity
of mixing; and long-term use.
Preliminary Studies: To assess the
mixing properties and efficiency for a
solidliquid mixing system, we added
tryptic soy broth (TSB) powder at
30 g/L to 200 L of water using an
impeller speed of 200 rpm. We
monitored mixing efficiency by
pumping the liquid at 2 L/min through
a recirculating loop from outlet to inlet
and continuously measuring TSB
concentration with an inline ultraviolet
(UV) absorbance meter. We defined
the mixing time as the point at which a
steady UV absorbance reading at 280
nm (A
280
, corresponding to protein
concentration) was achieved. The
impeller was then stopped and the
solution drained with continuous
monitoring for any change in
absorbance that would indicate
nonuniform mixing and dead zones
within the container.
Figure 11 shows that the TSB was
fully mixed within three minutes. The
draining profile confirmed that
mixing was uniform, with no dead
zones. Similar studies (data not
shown) were performed using solid
liquid suspensions. Turbidity
measurement provided similar results.
We designed another series of tests
to establish typical mixing times and
fluid homogeneity under four widely
different test conditions (Table 1).
Variable parameters such as impeller
speed (200 rpm), pumping direction
(down), volume (200 L), and baffles
(not inserted) were kept constant
unless stated.
Extended Program: Having
confirmed good mixing performance
in preliminary studies, we put in place
a more extensive test program. Test
protocols were designed to study the
effect of parameters such as impeller
speed, liquid volume, and insertion of
baffles while testing a broader range
of solids and liquids. We divided this
program into two parts: liquidliquid
and solidliquid mixing. Table 2
summarizes the conditions for 23 tests
in the program.
Performance Test Conclusions: From
our complete set of performance test
results, we concluded the following:
Our mixer system met performance
targets set at project initiation.
Efficient mixing could be
achieved for a range of diverse and
challenging applications.
Studies highlighted the
importance of optimizing mixing
conditions to achieve peak
performance.
PUTTING KNOWLEDGE TO PRACTICE
Our approach to design and
performance testing of a new single-
use mixer system was based on three
basic and essential steps: first, a full
understanding of engineering
principles behind the performance of
mixing systems; second, application of
those principles to design of a single-
use mixer in accordance with GEP,
focusing on ease of use, safety, and the
widest possible range of applications;
and third, extensive performance
testing to confirm system capability.
Performance testing showed that
Good engineering and design
principles for mixer systems can be
applied successfully to single-use
technology
Design principles already
incorporated into existing stainless
steel mixers can be applied to single-
use systems
A system based on a mechanically
driven, pitched-blade impeller can
provide a practical alternative to
traditional stainless steel mixers for a
Table 2: Test parameters for a range of applications
Application
Solute
Liquid
Volume
Impeller
Speed Baffles
Mixing
Time Ingredients
Floating or
Settling
Liquid
Liquid
Conductivity adjustment Sodium chloride in water
Initial: 200 g/L
Final: 10 g/L
Settling 200 L
200 L
200 L
100 rpm
150 rpm
200 rpm
Yes
Yes
Yes
<1 min
<1 min
<1 min
Final formulation 1% v/v acetone in water Floating 50 L
200 L
200 L
100 rpm
100 rpm
*100 rpm
No
No
No
<1 min
<1 min
<1 min
200 mL dye into 250 cP corn syrup Settling 200 L 200 rpm No <1 min
1.25% v/v Polysorbate 80 in water Settling 200 L 200 rpm No 10 min
Solid
Liquid
Conductivity adjustment 0.17 M sodium chloride (10 g/L) in water Settling 200 L
200 L
200 L
100 rpm
150 rpm
200 rpm
Yes
Yes
Yes
< 1 min
< 1 min
< 1 min
1M sodium chloride (58.44 g/L) in water Settling 200 L 200 rpm No <1 min
Buffer preparation Dulbeccos PBS (9.6 g/L) in water Settling 200 L 200 rpm No <5 min
Citrate buffer (21 g/L) in water Settling 200 L 200 rpm No 6 min
Ammonium sulphate (132 g/L) in water Settling 200 L 200 rpm No **15 min
Media preparation Dulbeccos Modified Eagles Medium
(DMEM) 5.36 g/L in water
Floating 50 L
200 L
200 L
100 rpm
100 rpm
200 rpm
No
No
No
5 min
14 min
4 min
Terrific Broth (HD pellets) 5 g/L in water Settling 200 L
200 L
100 rpm
200 rpm
No
No
10 min
6 min
Terrific Broth (LD powder) 47.6 g/L in water Floating 200 L 200 rpm No 18 min
Tryptic Soy Broth 30g/L in water Floating 200 L 200 rpm No 4 min
Vaccines Aluminum hydroxide 1.4 g/L in water Settling 200 L 200 rpm No 8 min
* Up-flow pumping; all other tests with down-flow pumping. ** Includes seven minutes for addition of ammonium sulphate
SEPTEMBER 2011 9(8) BioProcess International
69
wide range of applications in
bioprocessing.
Scalability should further increase
the mixer designs capabilities. Work
is now in progress on scaling up this
single-use mixer as well as
incorporating other beneficial
features.
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1 Warner TN, Nochumson S.
Rethinking the Economics of
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5860.
2 Nicholson P, Storm E. Single-Use
Tangential Flow Filtration in Bioprocessing:
An Approach to Design and Development.
BioProcess Int. 9(1) 2011: 3847.
3 Sinclair A, Monge M. Evaluating
Disposable Mixing Systems. BioPharm Int.
9(2) 2009.
4 Strahlendorf K, Harper K. Mixing in
Small-Scale Single-Use Systems. BioProcess
Int. 8(10) 2010: 4249.
5 Botterill M, Rawlings B. Applying
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Bojan Isailovic is a senior R&D engineer,

and Bruce Rawlings is senior marketing
manager for Pall Life Sciences, Pall Life
Sciences, Europa House, Havant Street,
Portsmouth, Hampshire PO1 3PD, UK;
44-23-9230-3303, fax 44-23-9230-2506;
www.pall.com.
To order reprints of this article, contact
Rhonda Brown (rhondab@fosterprinting.com)
1-800-382-0808. Download a low-resolution
PDF online at www.bioprocessintl.com.