B
I O
P
R O C E S S
TECHNICAL
Lipids in Bioprocess Fluids
William G. Whitford
L
ipids are becoming a factor
in a growing number of
processes and products in
biotechnology. Their
functions include excipient
or indirect activity (e.g., binding- or
presentation-dependent activation of
immunological adjuvants or
membrane proteins), stabilization,
delivery, and pharmacokinetic
control of poorly soluble active
ingredients (such as in parenteral
liposome-stabilized amphotericin B).
Lipids can also serve in biochemical
or immunologic (e.g., diagnostic
assays), pharmaceutic (e.g., PUFAs
treat some mental illness), and
nutritional (e.g., ⍀-3 fatty acids)
capacities. Here I’m specifically
addressing lipids as factors in
bioreactor- and fermentor-based
operations — from culture media
formulation to secondary
purification.
Once, all naturally occurring
compounds soluble in nonpolar
solvents (such as benzene) were
termed lipids. Later they were
described as water-insoluble
biomolecules biosynthetically or
functionally related to fatty acids and
PRODUCT FOCUS: BIOLOGICS
PROCESS FOCUS: PRODUCTION AND
DOWNSTREAM PROCESSING
WHO SHOULD READ: ANALYTICAL
LABORATORIES, MANUFACTURING,
PROCESS DEVELOPMENT
KEYWORDS: LIPIDS, DISPERSION,
EMULSIONS, LIPOSOMES, MEDIA
SUPPLEMENTATION
LEVEL: ADVANCED
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JANUARY 2005
ABBREVIATIONS USED
ACP: Acyl carrier proteins
ADCF: Animal-derived component free
BSA: Bovine serum albumin
CD: Chemically defined
DMSO: Dimethyl sulfoxide
EtOH: Ethanol
FDA: Food and Drug Administration
This computer simulation of a hydrated
(blue) phosphatidylcholine bilayer (orange
and pink) containing 25 mol% cholesterol
(white) is from MAURICIO C. TRIPP AND SCOTT
FELLER AT THE WABASH COLLEGE DEPARTMENT OF
CHEMISTRY (WWW.LIPID.WABASH.EDU).
their derivatives. The word lipid,
though derived from ancient Greek,
was coined surprisingly recently
(1925), and its definition is not
entirely clear. Some people stress the
solubility aspect, under which even
some proteins would be included.
Other definitions stress structure or
biochemical derivation, thus
excluding steroids and many oils.
So there’s some confusion when
designing means for addition or
removal of lipids in aqueous
solutions. Less-polar materials to be
dispersed may be commonly
referred to as lipids (or not), and
many nonlipid amphiphiles (such as
fat-soluble vitamins and even some
proteins) can participate in their
recovery or delivery. No matter the
definition, bioprocess engineers are
universally concerned about the use
of lipids: Their limited solubility in
aqueous media and the propensity
of some to form various and
changeable structures in dispersion
makes their use a challenge.
HMW: High–molecular-weight
IgG: Immunoglobulin G
LMW: Low–molecular-weight
PEG: Polyethylene glycol
PF: Protein free
PIT: Phase inversion temperature
PUFA: Polyunsaturated fatty acids
SFM: Serum-free medium
STF: Surfactant-free
UIC: Urea-inclusion compound
UPSTREAM PROCESSES
Dispersions: Lipids can be dispersed in
aqueous media by a number of
techniques. Requirements of various
fluids used in production can be met
in three ways: by adsorbing lipids to
soluble carrier molecules, devising a
formula that drives lipid self-assembly
to a required particle size, or
dispersing (and stabilize) a lipid
mixture to particles of sufficient
transient size and stability (1, 2). Each
approach is a science in its own right
and has been used in developing
upstream bioprocess fluids (Table 1).
Lipids in an organism can be
carried by bile salts, albumin,
lipoproteins, and ACPs. Animal
serum, the original supplement
providing lipids to cells in culture,
contains various carrier proteins
bearing high levels of all lipids
required by animal cell systems. For
example, FBS contains about
300 µg/mL cholesterol and
30 µg/mL oleic acid. Such high
concentrations and diversity are
sometimes detrimental. Cells
cultured in serum are constantly
exposed to a number of steroids,
making it difficult to specify the
effects of any particular one. And the
bioactivity of some lipids can impair
the performance of cells in culture.
Serum extracts and lipid-rich
fractions are popular means of
providing high serum-lipid
concentrations. Commercially
available fractions of animal serum
contain performance-relevant lipids.
BSA is a commonly used vehicle for
lipids. You can make or purchase a
reduced-lipid BSA that adsorbs
desired lipids added to it (3).
Organic polar solvents such as
EtOH and DMSO are other
common carriers. Use of such
“solvents” to act as carriers is
limited mostly by their inherent
activity on cells. Toxicity comes
primarily from two actions: cytosolic
catalysis and phospholipid extraction
from cell membranes.
Cyclodextrins (naturally occurring
circular polymers of glucopyranose)
can increase lipid solubility (Figure
1). Their function is similar to other
adsorptive systems in that they encase
or chelate lipids with more watersoluble molecules. They are well
tolerated by biological systems and
considered a pharmaceutical
excipient by the FDA. Many
products, even parenteral drugs, have
cyclodextrins in their formulations
(e.g., Pfizer markets ziprasidone
stabilized with a cyclodextrin). Some
medium supplements use
cyclodextrin to solubilize cholesterol
and fatty acids (Figure 2) and deliver
high concentrations to mammalian
cells in culture.
Emulsions and microemulsions
are forms of lipid dispersions used in
bioprocess fluid supplementation.
Microemulsions, the more common,
take advantage of the propensity of
some amphiphiles and lipids to
predictably self-assemble and form
multimeric or aggregate structures
Table 1: Various lipid dispersion technologies; their potential and limitations
Features of Lipid Supplementation Approaches
Physical
Stability
ADCF
Potential
PF
Potential
Serum
High
No
No
Serum Extracts
High
No
Solvents
High
Yes
Albumins
High
rAlbumin
Emulsions
Low
Yes
Micelles
Liposomes
Cyclodextrin
CD
Active Lipid Formulation
Potential
Capacity
Adjustable
No
High
Minimally
No
No
High
No
Yes
Yes
Low
Yes
No
Nearly
Medium
Somewhat
Yes
Yes
High
Significant
High
Yes
Yes
Yes
Medium
Somewhat
Medium
Yes
Yes
Yes
Medium
Somewhat
High
Yes
Yes
Yes
High
Yes
based on certain physical properties
that each unit molecule presents in
an aqueous environment (4). Soap
bubbles are a common example of
such phenomena.
Emulsions are kinetically stable
(nonequilibrium) dispersions. In
simple emulsions, the lipid particle
size is reduced by introducing
hydrodynamic force (e.g., mixing).
Particles are then stabilized through
the surface activity/charge of such
added amphiphiles as polar lipids,
peptides, or synthetic polymers that
present their hydrophobic regions to
each particle’s lipid core and their
hydrophilic regions to the aqueous
media. Concentrated supplements are
sometimes further stabilized through
such means as increasing the viscosity
of the continuous polar phase.
A more complex emulsion
involves spheres of lipids in a
lamellar phase with aqueous particle
cores. These are termed lipid bilayer
vesicles or liposomes. Regardless of
the number of concentric parallel
bilayers within a liposome, all
resident nutrient lipids must be
components of the lamellae
(bilayers, as pictured on the first
page of this article). Sometimes
lamella-forming polar lipids can be
combined with nutrient lipids such
that the mixture generates
liposomes of suitable size and
stability. For example, cholesterol or
fatty acids can intercalate between
the acyl chains of phosphatidyl
choline. Although functional
mixtures of polar and nutrient lipids
so formulated have been
Figure 1: Molecular model of a
ß-cyclodextrin inclusion complex. Guest
molecule is 1,4-butanediol, a vitamin B6
precursor with both hydrophilic (blue)
and hydrophobic (yellow) regions.
REPRINTED WITH PERMISSION OF STEFAN IMMEL
AT THE TECHNICAL UNIVERSITY DARMSTADT,
HTTP://CARAMEL.OC.CHEMIE.TU-DARMSTADT.DE/
IMMEL
demonstrated, attempts at using
such technology commercially have
failed due to unacceptable shelf life
or lipid delivery kinetics.
Microemulsions of lipids in the
micellular lyotropic phase may be
formed using detergents or
surfactants to generate disperse
particles containing the active lipid at
their core. Further considerations are
important beyond primary
development of these microscale and
thermodynamically stable dispersions.
Animal cell culture requires a type
and concentration of surfactant
neither too toxic nor too disruptive
to cell membranes. That makes a
practical limit about 0.3 mg/L of
lipid dispersed by such means.
Aside from understanding the
basics of lyotrophic phase behavior in
pure surfactant–cargo–water mixtures
(as is often depicted in ternary-phase
diagrams), implementation of such
pure chemistries to real applications
can be a study in itself. Often other
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Figure 2: Cholesterol-dependent cells
cultured in basal SFM with no lipids
(black squares); SFM supplemented with
HyQ® LS250 lipid supplement (red
triangles); and SFM supplemented with
HyQ® LS250 and fed (arrow) with HyQ®
LS1000 lipid supplement (blue circles)
constituents of the continuous phase
(e.g., culture media) in which a
dispersion is built can strongly
influence the phase-forming
ingredients. Such constituents range
from HMW amphipathic molecules
to LMW hydrotropes and
kosmotropes (5).
Fermentation: Generally, lipids
exist (if at all) only incidentally and
in trace amounts in the media used
for the fermentation of plant, fungi,
bacteria, or yeast cells because such
cells synthesize what lipids they
require from simple precursors. But
a growing number of applications
call for the addition of fats, oils, or
fatty acids to media formulations
(6). Oils are sometimes added as
antifoam agents because they are
innocuous and can directly interfere
with bubble film formation. Many
cells used in fermentor-based
production possess no lipases
capable of hydrolyzing complex
lipids, so the oils are insignificant as
a nutritional substrate.
Some natural and recombinant
strains, however, do express cellular
membrane-bound or soluble lipases
that allow them to use fats and oils
as either carbon sources or
substrates for desired secondarymetabolite products. For example,
in the production of polyketides and
PUFAs it has become practical to
develop strains selected for desired
elongase or desaturase activity — or
to insert genes for exogenous lipases
or desaturases. Thus fermentation
can produce a desired product using
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relatively high levels (up to 3% w/v)
of added lipid as a precursor. For
example, lipase-deficient
Pseudomonas can be transformed
with exogenous lipase genes to
allow its use of lard or coconut oil
in production of polyhydroxyalkanoates (7). A number of lipids
— from individual fatty acids to
vegetable or animal oils and fats —
have been shown to significantly
increase product yield depending on
the strain used, its lipid metabolic
capability, and the desired end
product.
Because cells used for such
applications are generally more
resistant to hydrodynamic and
nonpolar phase stress than are higher
animal cells, those fats and oils can
be added to fermentors and allowed
to form crude emulsions simply
through the shear forces created by
an impeller and spargation apparatus
(8). However, as use of higherperformance, fermentation-based
lipid production grows, methods
such as PIT are being developed to
facilitate dispersion of nutrient oils in
large amounts (9).
Animal Cell Culture: Requirements
for better performance and avoidance
of serum, along with improved
understanding of culture systems, are
inspiring fresh interest in
supplementing cell-culture media
with certain lipids. Despite the
multiple roles lipids play for whole
organisms (e.g., energy stores and cell
signaling), in culture they function
primarily as structural components of
cell and organelle membranes.
In any living organism an essential
nutrient is a compound required for
growth and reproduction that it
cannot synthesize itself. Many
organisms, such as most prokaryotes,
can produce most of the dozens of
lipids they require from simple
precursors. In mammals, though, at
least two fatty acids (linoleic and
␣-linolenic acid) have proven
essential. With the more widespread
use of serum-free and CD media,
researchers are looking closely at the
requirements of mammalian cells
maintained with minimal
complements of identified lipids.
Early work performed with cells
JANUARY 2005
adapted to media with serumdetermined high lipid levels seems to
have provided an inaccurate picture of
their actual requirements. Many SFM
formulations now lack linoleic and/or
␣-linolenic acid but sustain indefinite
cell growth and full function. So it
appears that those “essential” fatty
acids are, in fact, not essential for
most animal cell culture (10).
Providing cells with appropriate
preformed lipids (e.g., certain fatty
acids, sterols, and phospholipids),
even though not essential, reduces
the need for their biosynthesis by
cells. The resulting more efficient
metabolism is especially evident
when the rate of cell division is
important or where cells produce
high levels of a transgenic product
(11). Some derived clones are truly
auxotrophs for particular lipids,
meaning that those lipids are
essential to them. For example, the
NS0 myeloma cell line requires large
amounts of exogenous cholesterol.
That unusual phenotype is caused
by the silencing of an enzyme
involved in cholesterol synthesis.
All the basic dispersion approaches
described above apply to the design
of full-complement animal cell
culture media, with the constraints
that dispersed lipids must be
physically and chemically stable for
over six months at 4–8° C, at high
dilution in a complex solution of up
to 40 disparate components, and at a
fixed pH and tonicity. The latter may
be a disadvantage in the application
of some dispersion technologies, but
it can favor others.
Media Supplements and Feeds:
Whatever dispersion technology is
used, the goal in designing a lipid
supplement for animal cell culture is
to produce a concentrated
dispersion of select lipids that upon
dilution are nontoxic, disperse to a
biocompatible form, are taken up by
cells in a controlled fashion, can be
microfiltered, and remain stable in
storage for up to a year.
Adjustable pH, tonicity, and
minor component levels are
advantages of using concentrated
supplements instead of buying
complete media. A concentrate’s
formulation is not constrained by
the stability requirements or
dispersion chemistry effects of the
many other components in fullcomplement media. For practical
reasons in application, supplement
feeds must be concentrated in the
range of 50–5000⫻. Several media
supplements are commercially
available, each possessing a unique
set of features (Table 2).
Materials and Applications:
Although thousands of distinct lipids
populate natural systems, relatively
few are used in serum-free media
formulations. There has been an
evolution of thought regarding
which lipids are beneficial, but
because most successful formulations
are proprietary, exact recipes cannot
be obtained. Lipids currently
mentioned in the literature include
cholesterol; cod-liver oil; soybean oil;
and oleic, linoleic, and palmitic acids.
Applications include clone-specific
essential and performance-enhancing
lipids in SFM formulations, special
requirements such as the need for
high levels of cholesterol in NS0based transgenic producers, and fedbatch procedures in bioreactors (12).
Beyond the vendors’ concerns for
chemical and physical stability, users
must exercise care to maintain
product integrity. Each supplement is
subject to its own set of
destabilization conditions. Factors
such as pH, temperature,
hydrodynamic force, and interaction
with introduced amphiphiles (e.g.,
antifoam agents) can destabilize even
the most carefully designed product.
Furthermore, many plastics — for
example, those used in flexible tubing
— can adsorb lipids from most
formulations in a matter of minutes.
Lipids for cell culture are
available from a number of sources
and vendors. Commercially available
enriched fractions are extracted from
such diverse starting materials as
animal serum, sheep’s wool, fish oil,
and soybeans. Individual lipids are
also purified or derived from those
naturally rich sources. Chemical
synthesis or derivatization is used to
make those that are found rarely in
nature or are abundant only in
unacceptable sources.
The most popular materials for
dispersing nonpolar lipids in SFM
include the Tween 80 nonionic
detergent, pluronic acids (block
copolymer surfactants), phosphatidyl
choline (lecithin), albumin (both
natural and recombinant), and
cyclodextrin. Lipid particle surface
and interfacial energies must be
overcome both in generating
metastable emulsions and
accelerating the equilibrium of
microemulsions. Equipment for
such purposes (e.g., the EmulsiFlexC50 high-pressure homogenizer
from Avestin) generates extreme
hydrodynamic force while
minimizing heat production.
Addition of chemical antioxidants
(e.g., ␣-lipoic acid and
␣-tocopherol) can reduce the
peroxidation of polyunsaturates, and
procedures that limit the
introduction of free oxygen can also
help in this regard. Interestingly,
many lipids seem to be exquisitely
protected from oxidation when
complexed with cyclodextrin (13).
New Directions: Only 10 years ago
the principles determining the
structures formed by amphiphilic
lipids were mostly a mystery to
application specialists. Increased
understanding of the chemistry and
physics involved — and greater
dissemination of this knowledge —
are providing theoretical bases for
new technological applications (14).
Most people now responsible for
formulation/manipulation of
bioprocess fluids have been exposed
to some principles involved in
lipid–water systems. But translation
of particular chemistries or
knowledge of physical structure to a
technology that is robust in handling
a particular lipid or amphiphile
application can be an inefficient and
frustrating experience.
New approaches are becoming
available for the dispersion of lipids
in aqueous solutions, which
determines a gap between even the
Table 2: Ready-to-use lipid supplements for animal cell culture, in alphabetical order by supplier; publicly available composition
status abbreviations are PF = protein free; CD = chemically defined; ADCF = animal derived component free; STF = surfactant free,
NA = not available
Supplier
Trade Name
Active Ingredients
Published Status
Dispersion Techology
HyClone
LS 1000
Cholesterol
PF, CD, ADCF, STF
Adsorption to cyclodextrin
HyClone
LS 250
Cholesterol and fatty acids
PF, CD, ADCF, STF
Adsorption to cyclodextrin
HyClone
LipiMate
Serum Lipids
NA
Protein stabilized emulsion
Invitrogen
1000⫻ Cholesterol Lipid Conc.
Cholesterol and fatty acids
PF, CD, ADCF, STF
Adsorption to cyclodextrin
Invitrogen
250⫻ Cholesterol Lipid Conc.
Cholesterol and fatty acids
PF, CD, ADCF, STF
Adsorption to cyclodextrin
JRH
Lipid Conc. (500x)
Cholesterol
PF, CD, ADCF, STF
Proprietary, unavailable
Prolient
LiPro
Serum lipids
NA
Protein stabilized emulsion
Serologicals
ExCyte
Serum lipids
NA
Protein stabilized emulsion
Sigma
SyntheChol NS0 Supplement
Cholesterol
ADCF
Proprietary, unavailable
Sigma
Lipid Mixture 1 CD
Cholesterol and fatty acids
CD
Pluronic stabilized emulsion
Thermo
Xten DLS
Steroid, fatty acids, vitamin E
PF, STF
Soluble steroid, patent pending
JANUARY 2005
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chemistries currently available and
those technologies now in practice.
When lipid dispersion formula
development is addressed, a
distinction should be made between
established practice, existing but
recent developments, and
interesting future directions. Basic
research is producing a variety of
technologies that will undoubtedly
provide exciting improvements to
those approaches currently in use.
Most of us are familiar with lipids
in the micellar or lamellar phase.
However, polar lipids can form
other lyotropic phases that are being
exploited in production of stable
dispersions with new and useful
properties. For example, some lipids
mixed in excess water will
spontaneously form structures in a
number of hexagonal, cubic, or gel
phases. New technologies allow
them (and some more familiar
phases) to be dispersed as small,
stable, submicron particles such as
hexosomes, cubosomes, bicelles, and
cochleates (Photo 1). As with
liposomes, such dispersed particles
can be composed entirely of the
structural lipid or used as a vehicle
for either polar or nonpolar cargo
(15). Furthermore, they can be
polymerized to provide very stable
and filterable particles (16).
Pullulan (a linear polymer of
maltotriose) can be derivatized with
cholesterol or acetate residues to form
a more amphipathic molecule capable
of forming dispersible nanoparticles
containing the cargo lipid (17).
Polymeric micro- or nanospheres can
be composed of natural or synthetic
compounds having quite varied and
dynamic properties (18). “Intelligent”
(environmentally responsive) targetspecific and biodegradable
nanoparticles are now becoming
practical. For example, hydrogel
nanospheres could provide sterile,
stable, targeted, and controlled
delivery of cargo lipids (19).
Nonpolar molecules can be
covalently linked to polar ones either
to make them water-soluble or
facilitate the formation of dispersible
particles. A familiar example is
ethoxylation (PEGylation) of poorly
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JANUARY 2005
Photo 1: A cryotransmission electron
micrograph of a dispersion of lipids in
the cubic phase (displaying ordered
internal lattices) amid ordinary lamellar
vesicles. REPRINTED WITH PERMISSION FROM
SPICER PT, ET AL. NOVEL PROCESS FOR
PRODUCING CUBIC LIQUID CRYSTALLINE
NANOPARTICLES (CUBOSOMES). LANGMUIR
17(19) 2001:
5748–5756.
soluble drugs; and lipids can be
similarly derivatized. PEG can be
bound either to the desired lipid
itself or to a vehicle lipid derivatized
to alter its mesomorphic
characteristics and therefore its
cargo-bearing ability (20).
DOWNSTREAM PROCESSES
Biotechnology products display a
range of properties from highlypolar soluble substances (such as
DNA and most proteins) to
nonpolar solubles such as lipids and
some polypeptides. Most of these
high-value products are relatively
fragile and require special mild
processing conditions. Such
products are often relatively dilute
and need to be separated from
complex mixtures including closely
related molecules and/or crude cell
debris. These factors make
economical separation difficult.
Until recently, most lipid separation
schemes were ad hoc, representing
scaled-up laboratory procedures.
More selective, biocompatible,
and scalable extraction and
precipitation schemes — along with
new resins and separation
media/sorbants — have recently
become available. Combinations of
new and conventional methods are
being explored to increase
scalability, efficiency, economy, and
specificity. The scale-down approach
enables large-scale systems to be
efficiently estimated in small-scale
models, helpful in commercializing
bioprocesses where speed and
economy are critical to success.
Downstream bioprocessors
concerned about handling lipids fall
into two camps: those who want the
lipids (as a product) and those who
don’t (considering lipids an
interfering contaminant).
As a Product: Lipids of various
types can be produced through
in vitro culture of bacteria, yeast,
fungi, and plant or animal cells.
Lipid-based products (and those
having significant nonpolar regions
or being otherwise related) range
from fatty acids to polyketides to
complex lipids such as fatty acid
glycosides or lipopolysaccharides.
Whether recovering them from
fermentors or bioreactors, producers
need to economically separate lipids
from culture supernatant and isolate
or purify desired products from the
total lipid fraction. That must be
accomplished while both
maintaining product integrity and
preventing introduction of
contaminants. Steps can in some
cases be efficiently combined, at
least in small-scale methods.
An extensive body of reports
describing innovative small-scale
extraction and purification has
appeared since Floch (21) described
the first significant development
(outline and review at
www.cyberlipid.org). Large-scale
techniques have also been developed
in the food and cosmetics industries,
often beginning with plant and fish
biomass. Methods include
distillation, solvent extraction/
partitioning, pressurized fluid
extraction, supercritical fluid
extraction, multistage or continuous
centrifugation, column
chromatography, and particularly, a
number of solid-phase or dry
downstream extraction approaches.
Large-scale bioprocessing
applications have, surprisingly, not
yet been comprehensively researched
and optimized — or at least not as I
have found in published literature.
Nevertheless, some large-scale
lipid extraction techniques developed
for other commercial applications are
beginning to be applied — with
various levels of success — to
fermentation and bioreactor product
purification. For example, very largescale and continuous or multistep
centrifugation techniques (as in the
dairy industry) can be used to collect
emulsions of lipids and gently,
efficiently remove dispersed particles
of any basic form. However, such
collected product commonly contains
many undesired components based
on coincident sedimentation, and
supernatant can retain lipid particles
larger than ~5 µm. So up to 25% of
the product can be lost.
Small lipid particles — such as
those remaining after most advanced
large-scale centrifugation approaches
(e.g., tubular-bowls) or are
dispersed by adsorption to carriers
or as mixed micelles — have been
recovered at large scale by
partitioning onto such immobilized
substrates as hydroxypropyl dextrin
mediated by zinc salts and organic
solvents (22). Another established
technique becoming popular
involves formation of UICs (23).
That approach is based on the
propensity of inclusion compounds
to selectively isolate longer chainlength fatty acids that have multiple
(preferably trans) double bonds.
UIC is inexpensive, biocompatible,
scalable, and currently applied to
the large-scale purification of
PUFAs from fish and plant oils.
As a Contaminant: Many of the
above methods cannot be used in
conjunction with the large-scale
harvest of proteins because they
damage or remove the product —
or they require incompatible
chemicals. However, because here
the lipid need not be recovered,
other techniques become available.
Most commercial media
formulations are screened to some
degree for compatibility with the
most fundamental purification
processes. However, some lipid
vehicles can be stable throughout
culture but, when depleted of their
original cargo, harbor new (often
cell-line dependent) hydrophobic
materials. Scale-up processes, culture
or primary harvest material
additives, and application of less
common approaches can cause
coalescence, precipitation, and phase
changes in dispersed lipids. Such
FOR FURTHER READING
Books
Vance DE, Vance JE. Biochemistry of
Lipids, Lipoproteins and Membranes.
Elsevier Science: 2002.
www.cem.msu.edu/~reusch/
VirtualText/lipids.htm
Morrison ID, Ross S. Colloidal
Dispersions: Suspensions, Emulsions,
and Foams. Wiley-Interscience: 2002.
Solutions, Surfaces, and Colloids:
Tutorial in Medicine (A. Kabanov,
Nebraska Medical Center)
www.unmc.edu/pharmacy/
wwwcourse/graduate/g_syllabus.html
Caffrey M. LipiDat: A Database of
Thermodynamic Data and Associated
Information on Lipid Mesomorphic and
Polymorphic Transitions. CRC Press,
Inc.: Boca Raton, FL, 1993.
Lipid Phase Tutorial (Nanoscale
Chemistry Research Group, University
of Birmingham)
www.nanochem.bham.ac.uk/
liquid_crystals/lc_index.htm
Journal Articles
Patravale VB, Date AA, Kulkarni RM
Nanosuspensions: A Promising Drug
Delivery Strategy. J. Pharm Pharmacol.
56(7) July 2004: 827–40.
Membrane Biophysics Tutorial
www-biology.ucsd.edu/classes/
bibc110.SU99/part1/reader.html
Strickley RG. Solubilizing Excipients in
Oral and Injectable Formulations
Pharm Res. 21(2) February 2004:
201–30.
Lipid/Water Systems (D. Weitz,
Harvard University)
www.deas.harvard.edu/projects/
weitzlab
Websites and Online Tutorials
American Oil Chemists Society
Educational Offerings
www.aocs.org/meetings/education
Lipid/Water Systems (Soft Condensed
Matter Group, University of Edinburgh)
www.ph.ed.ac.uk/cmatter/soft.html
Interfacial and Colloid Science Group,
(John Berg, University of Washington)
http://faculty.washington.edu/spc
Lipid Metabolism (M. King and S.
Marchesini, University of Brescia)
www.med.unibs.it/~marchesi/
lipsynth.html
Lipids Tutorial (W. Reusch, Michagen
State University)
activity brings up new process issues
due to consequent alterations in
rheology, filterability, and/or lipid
deposition characteristics of the inprocess harvest.
Primary clarification is often
accomplished by a disk-stack type
centrifuge that removes whole cells
and particulates down to ~1.0 µm.
But some harvests and downstream
steps require more work. Passing
supernatant through pad or depth
filters (such as Millipore’s Millistak+
A1HC, or Pall’s SUPRAdisc) made
of such materials as cellulose fibers
and diatomaceous earth can provide a
second level of clarification (down to
~0.2 µm). Although they remove
some lipids (mostly through
nonspecific particle entrapment), their
behavior in specific and qualitative
lipid removal has not been
Lipid/Water Systems (P. Spicer)
www.nonequilibrium.com
Surfactants and Microemulsions tutorial
www.fisica.unam.mx/liquids/tutorials/
microemulsions.htm
What Is Liquid Crystal Fluid? tutorial
www.lxdinc.com/AppNotes/fluid.htm
General Lipid Resource: Cyberlipid
Center (C. Leray) www.cyberlipid.org
Lipid Types Tutorial
www.cyberlipid.org/cyberlip/
desc0004.htm
systematically addressed. Products
purporting to specifically adsorb lipids
include Cuno’s Zeta Plus filters.
LIPID SCIENCE
People have been devising simple and
practical formulas for working with
oil dispersions for millennia.
Churning butter and adding egg
yolks to béchamel sauce are examples
of applying simple techniques
without necessarily understanding
the chemistry behind them. At the
other extreme, the modern
pharmaceutical, food processing,
agrochemical, and cosmetics
industries approach oil-and-water
dispersion issues with sophisticated
technologies. Interfacial and colloidal
chemistry, including such disciplines
as hydrocarbon chain packing and
JANUARY 2005
BioProcess International
51
lipid mesophase behavior, have been
brought to bear against lipid
dispersion issues (24, 25).
Currently, much of the lipidhandling technology applied in
bioprocessing is relatively crude. But
as biotechnologists apply even
existing chemistries and large-scale
commercial techniques from other
industries to scaled-down
approaches, much more specific,
efficient, and economical procedures
are sure to be developed.
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HyClone, 925 West 1800 South, Logan,
UT 84321; fax 1-435-792-8018;
[email protected].