Aalto University

The University of the chemical technology

CHEM-E3140 - Bioprocess technology II

Centrifugation
Downstream processing assignment

Iida Mäkeläinen
Henna Penttinen
Jenni Heikkinen
Vera Kutiavina
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Abstract

Centrifugation is widely utilized and important separation technique in many industrial fields
such as food, pharmaceutical, biotechnical and chemical industries. The operating principle
of centrifuges is based on centrifugal force that facilitates the separation of different
components in the mixture based on their sedimentation features. The applied force on
particles depends on the rotation speed and the radius of centrifuge rotor. Thus, centrifugal
force causes denser and larger components to migrate outward from the axis of rotation.
Generally industrial centrifuges are distributed into two different classes according to their
feeding type and operational principle. Filtration centrifuges differ from sedimentation
centrifuges by utilizing filtration material e.g. slots, holes, filtration cloth to filtrate liquid
material. The type of centrifuge that is chosen for industrial process is dependent on the
process and separable components.

Background

Centrifugation technique has been widely used in industrial and laboratory settings for over
100 years. The first studies utilizing centrifugation technique are from the beginning of the
1800s [1]. The first centrifuge for industrial use was presented by Antonin Prandtl in 1864 for
milk processing [2]. Later in 1878 Gustaf de Laval introduced continuous centrifuge for
industrial applications, enabling usage of centrifuges in wider range [1]. Currently centrifuges
are regularly utilized in different fields in laboratory-scale settings as well as in larger-scale
industrial processes.

Centrifugation is a separation technique where different components of mixture are

separated based on their density or particle size. The separation of different substances is
based on centrifugal force that is produced by high speed rotation. Centrifugal force causes
denser components to be directed away from the axis of rotation whereas lighter
components migrate towards it. [3]

Centrifugation is generally utilized with separation of insoluble particles from liquids, but it is
utilized as well in the separation of two immiscible liquids. For example, in industrial milk
processing the centrifugal separation of the cream from milk is based on the density
differences of the components. Centrifuges that are utilized in industrial setting, can be
classified into two different categories: sedimentation and filter centrifuges. The classification
is based on the general operation principles of the centrifuges, but the continuous and batch
feeding type of centrifuge is also taken into consideration in classification. [5]

Sedimentation centrifuges separate insoluble particles from liquids based on different

sedimentation features, such as size, shape, density and centrifugal force [6]. Sedimentation
is a process where molecules or particles sediment downward due to gravitational force.
Larger and denser particles sediment faster. Centrifugation facilitates sedimentation with
centrifugal force, separating the particles into solid pellet (precipitate) and liquid solvent
(supernatant). [7] In comparison to sedimentation centrifuges, centrifugal filtration utilizes
filtration material to separate the components from the mixture [5].
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Theory and mechanism

The working principle is based on centrifugal force that makes particles move toward bowl
wall of the centrifuge. This force is usually expressed with number of earth gravities (g). The
process of particles sedimentation depends on several factors. There is a direct correlation
between rate of sedimentation and size of particles, speed of centrifugation, density
difference between liquid and solid phase of solution and an inverse relation with fluid
viscosity. [5]

Figure 1. Particle in rotation.

Figure 1 shows how the particles move in

rotation. The centrifugal acceleration of the
particles means that the force initiates
particle movement outward from the axis of
rotation. Centrifugal acceleration occurs the
radial position (r) of the particles and the
square of the angular velocity (⍵). In figure 1
shows that angular velocity is calculated
using the tangential velocity (uθ) and radial
position. The force acting on a particle
increases, when the acceleration increasing.
That cause a sedimentation of the particles
at the axis of the rotation. [8, 9, 10]

Figure 2. The measurements of the

centrifuge.

Radius is depending of the shape and

type of the rotor of the centrifuge.
Basically, if centrifuges rotor is fixed angle
rotor as is shown in figure 2, there are two
different values for radius. Fixed angle
rotor means that tubes are in same
position during centrifugation and the
rotational speed cannot affect them. The other options are a swinging bucket centrifuge,
where tubes hanging in the rotor. In that kind of centrifuge radius is depending on rotational
speed as is shown in figure 3. When rotational speed higher, also the radius is increasing.
Centrifuges with tubes are generally used in laboratory scale, not in downstream processing.
[9]
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Figure 3. Radius varies with rotation

speed.

Calculations

Relatively centrifugal force (RFC) is the

unit, which is used when is question of
centrifugal force. Generally used
revolutions per minute (RPM) is not a
valid unit, because the force depends on
the radius of the centrifuge. If the RPM is
known and also the radius of the rotor, is
calculation of the RFC easy using the
equation:

!"#
𝑅𝐹𝐶   =   1,118 ∗ 𝑟  (!""")! ,

where the unit of the radius of the rotor (r) is millimetres. The unit of RFC is g as an Earth’s
gravitational force. [6, 9]

As has been mentioned before, sedimentation rate is directly related to the features of
particles, namely size and mass. Particles that are bigger in size than 5 µm accumulate at
the bottom due to gravitational force, whereas if the size is smaller the process can be
described with Brownian motion. That is why the smaller particles are, the stronger
centrifugal force is required for sedimentation. In that case, the required force can be
described with equation:

!∗! !
𝐹 =     ,
!

where m - mass of the particle,

v - velocity of the particle,
r - rotation circle radius.

The ratio between centrifugal force and gravitational force represents the centrifugal effect:

! !!
𝐶 = !     = !",

where G - gravitational force

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Classification

All modifications of centrifuges can be distinguished into two main types: sedimentation and
filtering centrifuges. In sedimentation centrifuges the centrifugal force is used to separate
solids from liquids or two liquids with different densities. Sedimentation centrifuges include
decanter, disk-stack, solid-bowl basket and tubular bowl centrifuges, which will be described
below. Filtering centrifuges use centrifugal force to pass a liquid through a filtration media,
such as a screen or cloth while solids are captured by the filtering media. [5]

Sedimentations centrifuges

There are many different types of sedimentation centrifuges and the best centrifuge type can
be chosen on the basis of application and specificity of the process.

Disc bowl centrifuge or disc stack centrifuge was first invented in order to separate cream in
milk production industry. These centrifuges usually used for separation of two liquids or
liquid and relatively low suspended solid phase. This centrifuge is composed by a vertical
rotor with several conical discs on it. These systems of conical spacers allow to increase the
sedimentation area. During rotation, centrifugal forces make denser solids move towards the
bowl of the centrifuge where they then can be collected. [5, 13]

Decanter centrifuge is sedimentation centrifuge, which consists out of a horizontally oriented

bowl in a shape of a cone with a conveying scroll inside. The main working principle is
related to the differences of densities of the liquids as denser liquids will drop to the bottom
wall of the centrifuge. Conveying scroll creates a liquid pool, solids settle to the bowl wall
and then transported by a conveying scroll to the end of a bowl where they are collected,
whereas less denser liquid returns back to the pool. To prevent accumulation of solid
particles on the scroll and reach optimal retention time for the separation, difference in
rotational speeds of bowl and scroll is used in decanter centrifuges. [5]

Hydrocyclone is one type of sedimentation centrifuges. It differs from other sedimentation

centrifuges so that it does not have any rotation parts. The centrifugal force form, when
suspension is pumped to colon with high speed. Because of the shape of the colon the
suspension start to swirl, which occur the separation of the solid particles to the walls of the
colon. The liquid material stays in the centre of the colon. The solid particles are removed of
the bottom of the colon and clarified liquid material in the top. Hydrocyclones is generally
used in continuous processes. [5]

Solid-bowl centrifuge is a good example about the batch-type centrifuge. It consists of the
one basket and it is generally used of collection particles of low concentration suspensions.
In addition, it is also a good method to separate fine particles from liquid material, because
of large diameter and high g forces. [5]

Filtration centrifuges

Filtration centrifuges similar as sedimentation centrifuges, but they are based on the filtration
of the liquid material. Mechanism of the filtration centrifuge is the same as in other
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centrifuges thus the separation is based on the centrifugal force. The difference is that
filtration centrifuges has the filtration material, for example slots, holes, a porous membrane
or filtrate cloth depending of the application and the purpose of the process. Filtration
centrifuges are in different types as sedimentation centrifuges. [5]

Filtration centrifuges can work as in continuous process as in batch process depending of

the type of the centrifuge. Generally, filtration centrifuges are used in continuous processes,
because the higher throughput capacity is often needed in industrial processes. Filtration
centrifuges can be used for separation of the suspension with high solid material
concentration, because of the filtration material. Solid particles form a cake on the filtration
material. The solid material can be washed in centrifuge with water if centrifuge is equipped
with rinsing system. That help for example to get all residual material out of solid material. [5]

In addition to centrifugal force, many other things affect to the centrifugation of the material
with filtration centrifuge, for example cake thickness, viscosity, screen area and the filtration
material. [1, 5] If the cake growth too big on the filtration material, it can be removed during
process. Batch process is more flexible in that case, but in some filtration centrifuges is
possible to cut the cake decreasing rotation speed during continuous process when solid
material drop to the bottom of centrifuge. [5]

Importance

Centrifugation is one of the most important and widely applied research techniques in
biochemistry, cellular and molecular biology, and in medicine. Current research applications
rely on isolation of cells, subcellular organelles, and macromolecules, often in high yields.
[11]

Today, centrifuges are routinely used in a variety of disciplines ranging from large-scale
commercial applications to laboratory-scale scientific research. The number of centrifuge
designs and configurations used in the mineral, petrochemical, chemical, medical,
pharmaceutical, municipal/industrial waste, dairy, food, polymer, energy and agricultural
industries (to name a few) seem almost as numerous as the applications themselves. [12]

Industrially, centrifuges are used for a variety of purposes related to separation of materials
on the basis of density. Generally, centrifuges are used throughout many manufacturing
industries e.g. food and agri business, pharmaceutical/biotechnology, environmental
industries and chemical industries. It appears in heavy industrial uses such as sugar and oil
refining and in light industrial separations such as dewatering of vegetables. [5]

Centrifugation and membrane filtration are the only techniques used for large-scale cell
harvesting. Centrifugation has advantages for dense (density > 1.03 g/cm3) and large
(diameter > 2 µm) microorganisms. For example, centrifugation is very efficient for
harvesting yeast. [14]
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Case study: Pickering Emulsion as an Efficient Platform for Enzymatic Reactions

without Stirring

A Pickering emulsion is an interesting new method to process enzymatic reactions without

stirring and immobilization. Stirring and shaking can cause the damage to enzymes’
structures. Because of high price enzymes should be efficiently recyclable. Immobilization of
enzymes is one possible solution for these problems, but sometimes it leads to a decrease
in enzyme activity. [15]

Figure 5 shows how a conventional biphasic enzymatic reaction can be converted into a
water-in-oil Pickering emulsion system by adding a small amount of solid particle emulsifier.
Pickering emulsions that are stabilized by solid particles have advantages over the
conventional surfactant-stabilized emulsions such as the ease of product−particle separation
and low toxicity of particles. The large oil−water interfacial area allows biphasic reaction
systems to access high efficiency, which has been extensively demonstrated in
hydrogenation, oxidation, as well as enzymatic reactions. Pickering emulsion systems were
found to enable biphasic reactions to proceed efficiently through autodiffusion of molecules
without need for stirring due to large interface area and short molecule diffusion distance.
[15]

Figure 5. Schematic figure of the Pickering emulsion method for stirring-free biphasic
enzymatic reactions. [15]

At the end of reaction, the emulsion is returned to a biphasic system by centrifugation, where
the solid particle emulsifier is picked for recycling. After the upper layer of organic product is
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removed through liquid transfer, the enzyme and emulsifier in the water phase can be
directly used for the next emulsion reaction. [15]
Conclusion

Although centrifugation technique is over a hundred years old it is still an important method
in the broad range of scientific, medical and industrial applications. Centrifuges are widely
used in a variety of disciplines ranging from laboratory-scale scientific research to large-
scale commercial applications. A centrifuge is used to separate particles or even macro-
molecules based on size, shape or density.

Centrifugal separation is used when gravity separation (settling, sedimentation or flotation) is

too slow and the particles do not settle readily or at all. A centrifuge can generate centrifugal
acceleration which is much greater than the Earth’s gravity. The centrifuges in industrial use
can be divided into two categories sedimentation and filter centrifuges which of each can be
subdivide into continuous or batch feeding type.

After fermentation, the biomass is commonly harvested by a centrifugation process to

separate insoluble particles, which are usually cells, subcellular organelles, viruses and large
molecules.
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References

[1] Wilson, Ian D. Poole, Colin F.. (2009). Handbook of Methods and Instrumentation in
Separation Science, Volume 1 - Centrifugation to Covalent Chromatography. Elsevier.
Online version available at: http://app.knovel.com/hotlink/pdf/id:kt00BYXIS1/handbook-
methods-instrumentation/centrifugation-covalent

[2] Holtbrügge, J., Kunze, A. K., Niesbach, A., Schmidt, P., Schulz, R., Sudhoff, D., &
Skiborowski, M. (2016). Reactive and Membrane-Assisted Separations. P. Lutze, & A. Górak
(Eds.). Walter de Gruyter GmbH & Co KG.

[3]JoVE Science Education Database. General Laboratory Techniques. An Introduction to the

Centrifuge. JoVE, Cambridge, MA, doi: 10.3791/5019 (2016), http://www.jove.com/science-
education/5019/an-introduction-to-the-centrifuge, 27.11.2016

[4] Anonymous, RPM vs. RCF, http://www.sorbio.com/index.php/rpm-vs-rcf, 27.11.2016

[5] Beveridge, T. (2000), Large-Scale Centrifugation, Minister of Public Works and

Government Services Canada

[6] Rickwood, D.,Centrifugation Techniques, (2001)

https://www.researchgate.net/publication/227982944_Centrifugation_Techniques,
27.11.2016

[7] Creighton, Thomas E.. (2010). Physical and Chemical Basis of Molecular Biology - 16.
Sedimentation by Centrifugation. Helvetian Press. Online version available at:
http://app.knovel.com/hotlink/pdf/id:kt0091X9N1/physical-chemical-basis/sedimentation-by-
centrifugation

[8] Centrifugation: Theory, Sedimentation Rate, Coefficient and Other Details, webpage,
http://www.biologydiscussion.com/cell-biology/centrifugation-theory-sedimentation-rate-
coefficient-and-other-details/3558 28.11.2016

[9] Clarke, P. 2009, pdf, Theory of sedimentation and centrifugation,

http://www.bbka.org.uk/local/iceni/bm~doc/pollensuspension-2.pdf

[10] Fundamental of particle technology, pdf, Chapter 8: Centrifugal separation,

http://www.particles.org.uk/particle_technology_book/chapter_8.pdf

[11] Cole-Parmer: Basics of Centrifugation. 2006.

http://www.coleparmer.com/TechLibraryArticle/30

[12] Handbook of Methods and Instrumentation in Separation Science page 49

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[13] Conical Plate centrifuge, Wikipedia, webpage,

https://en.wikipedia.org/wiki/Conical_plate_centrifuge 2.12.16

[14] Harrison R., Todd P., Rudge S., Petrides D. (2015). Bioseparations Science and
Engineering. Oxford University Press, 547 s.

[15] Pickering Emulsion as an Efficient Platform for Enzymatic Reactions without Stirring
Lijuan Wei, Ming Zhang, Xiaoming Zhang, Hongchuan Xin, Hengquan Yang,
School of Chemistry and Chemical Engineering, Shanxi University, Taiyuan 030006, China
American Chemical Society, September 28, 2016,
http://pubs.acs.org/doi/abs/10.1021/acssuschemeng.6b01776