Computers and Chemical Engineering 30 (2006) 1464–1475

Bioprocess control: Advances and challenges

Joseph S. Alford ∗
Eli Lilly and Company, Indianapolis, IN 46285, United States
Received 15 February 2006; received in revised form 5 May 2006; accepted 16 May 2006
Available online 31 July 2006

Abstract
Most commercial process control tools and techniques used in the chemical process industries are applicable to bioprocesses (e.g. processes
that make certain medicines). While bioprocesses include several different unit operations, the focus of most of this paper will be the control of
bioreactors (i.e. fermentors) in which a near optimal environment is desired for microorganisms to grow, multiply, and produce a desired product.
Bioreactor control provides special challenges due to significant process variability, the complexity of biological systems, the need, in many cases, to
operate in a sterile environment, and the relatively few real-time direct measurements available that help define the state of the culture. This has led to
some creative solutions as well as the identification of topics where further research and development and vendor software functionality are needed.
© 2006 Elsevier Ltd. All rights reserved.

Keywords: Bioprocess control; Advanced process control; Data management; Bioprocess instrumentation; Alarm management

1. Introduction
In the early 1970s, when the author’s career in bioprocess
automation began, fermentation processes (e.g. making peni-
cillin), were perceived to be as much art as science. Making
antibiotics was not much more sophisticated than making wine.
Ingredients were put into a tank which was then closed and man-
ually sterilized, manually inoculated with living cells, and then
agitated and aerated for several days. The only automated feed-
back control was a pneumatic controller used for temperature.
Data recording consisted of operators walking the production
floor, periodically recording values from gauges onto a clipboard
sheet and then hand copying a few of these values onto the paper
manufacturing ticket. Most of the parameters of real value to the
fermentation culture [e.g. pH, dissolved oxygen (DO2 ), nutrient
concentrations] were not monitored or controlled on-line. Sci-
entists, in trying to evolve less variable and higher productivity
processes, were frustrated with the lack of appropriate sensors,
tools, and support systems. Symbolizing this “state of affairs,”
sometimes the best indicator as to how well a fermentation was
progressing was the subjective analysis of an experienced opera-
tor looking through a site glass located on the top of the fermentor
∗ Tel.: +1 317 276 7653; fax: +1 317 276 1403.
E-mail address: Alford Joseph S@Lilly.com.
and observing the color and texture of the foam layer riding on
top of the liquid broth.
Much progress has occurred during the past 30 years. Some
of this has resulted from a few companies, including Eli Lilly
and Company, using a combination of commercial technolo-
gies, complemented by significant customization of computer
systems (see Section 9), and the in-house development of novel
applications. Commercial solutions have been slowly catching
up to the needs of bioprocesses, although several gaps and oppor-
tunities still remain.
Regarding the current state of commercial computer control
systems used for bioprocesses, a variety of process control and
data analysis platforms and products currently exist. They gen-
erally represent variations of the generic system shown in Fig. 1.
With some exceptions, the low end of process control prod-
ucts involves benchtop bioreactors which are controlled via
personal computers (PCs) with Linux or Windows operating sys-
tems and which incorporate either commercial (e.g. LabView)
or bioreactor vendor proprietary application software packages.
Intermediate level systems may involve Programmable
Logic Controllers (e.g. Rockwell) + a Human Machine Interface
(HMI) + a historian.
High end systems (used in many fermentation pilot plants
and production facilities) utilize Distributed Control Systems
(e.g. Fox IA, Emerson DeltaV, Fisher Provox) which, in turn,
are usually linked to proprietary or commercial historians (e.g.

0098-1354/$ – see front matter © 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.compchemeng.2006.05.039
 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
Fig. 1. Generic bioprocess control and information system schematic.
Aspen Tech, OSI-PI). In some cases, DCS/Historian systems are
further interfaced with third party products (e.g. alarm loggers,
real-time expert systems, advanced data analysis software) to
provide additional functionality. They may also be interfaced
with other internal company systems such as laboratory assay
systems, manufacturing execution systems (MES), or plant site
data warehouses (e.g. Discoverant).
Improvements to bioprocess productivity generally come
from two sources: (1) cell lines and (2) process control.
In this paper, the focus is on bioprocess control. Biopro-
cess control is defined as providing a near optimal environment
for microorganisms to grow, multiply, and produce a desired
product. This includes providing the right concentration of nutri-
ents to the culture (e.g. carbon, nitrogen, oxygen, phosphorous,
sulfur, minerals), removing any toxic metabolic products (e.g.
CO2 ), and controlling important internal cellular parameters
(e.g. temperature, pH).
Note: While subtle differences exist in the definitions of the
terms bioreactor and fermentor, the terms are interchangeable for
many processes and will be used interchangeably in this paper.
The scope of bioprocess control theoretically includes not
only the sequence of steps leading up to and including the fer-
mentor (Fig. 2), but also, in many cases, to several of the product
recovery and purification steps (Fig. 3). For example, follow-
ing the fermentation, subsequent steps may involve rupturing
1465
Fig. 2. Culture growth stages.
the cells and eliminating cellular debris from the stream con-
taining desired product. Further, especially for mammalian cell
bioprocesses, there are a number of unit operations that exist
to provide several log reductions in viruses that may exist in
the product stream as a result of using certain mammalian cell
lines and/or due to any use of animal sourced (especially bovine)
raw materials. These viral reduction unit operations can include
detergent-based inactivation, column chromatography, low pH
(e.g. pH < 4) inactivation, and nanofiltration. With each of these
product recovery/purification operations (some of which are
shown in Fig. 3), the operation is either primarily manual, or con-
trolled with a local PC or programmable logic controller (PLC).
Availability of appropriate measurements and control algorithms
are generally satisfactory for post-fermentation unit operations.
Opportunities for improvement are primarily in the areas of data
analysis (described later) and automated batch sequencing.
Therefore, the most challenging unit operation from the per-
spective of process control is the bioreactor itself, and this will
be the focus for most of the rest of this paper.
The benefits of bioreactor automation have been the subject
of several conferences and articles in the literature, such that
automation is an expectation and no longer needs justification
for most manufacturing plants. Benefits have included:
A. process variability reduction;
B. productivity (yield) improvements;
C. increased on-line monitoring and troubleshooting capability.
1.1. Pre-inoculation operations: creating a consistent
optimum sterile medium
Bioprocess media composition has historically been deter-
mined by development scientists and engineers using statistical
1466 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
Fig. 3. Product recovery steps following fermentation.
“Design of Experiment” (DOE) methods in small-scale batch
equipment. A relatively small dimensional space is defined by
the scientist, based on experience and literature searches. This
space is typically explored in detail using three to five nutrients
of special interest (e.g. those nutrient concentrations expected to
highly influence culture growth and productivity, process eco-
nomics, etc.). This approach assumes that other factors (e.g.
sterilization operations and the control of earlier phases in grow-
ing the culture; see Fig. 2) are consistent. DOE methods typically
result in a broth nutrient “local optimum,” but not necessarily a
“global optimum.” Note: Broth nutrient optimization is a classi-
cal non-linear optimization problem.
Recent work with non-traditional techniques indicates greater
opportunity exists to (1) realize a “global optimum” with respect
to nutrient make-up and (2) understand and better control activi-
ties associated with pre-inoculation nutrient concentration vari-
ability (e.g. during sterilization). Specific to nutrient make-
up, “evolutionary computation” algorithms, such as particle
swarm optimization, have shown that, compared to traditional
approaches, a similar number of small-scale equipment exper-
iments can explore a higher dimensional space (i.e. number
of different nutrients) and larger ranges of individual nutri-
ents (Cockshott & Hartman, 2001; Cockshott & Sullivan, 2001;
Strobel & Hartman, 1999). Note that particle swarm optimiza-
tion, genetic algorithms, etc., are subsets of evolutionary com-
putation which, in turn, is a subset (along with neural nets and
fuzzy logic) of computational intelligence. Particle swarm opti-
mization (PSO) is a population-based stochastic optimization
technique, developed by Dr. Eberhart and Dr. Kennedy in 1995,
which was inspired by the social behavior of phenomena such
as bird flocking, i.e. a large number of birds eventually converg-
ing on a common roost. PSO shares several similarities with
evolutionary computation techniques such as genetic algorithms
(GA). The system is initialized with a population of random solu-
tions and searches for optima by updating generations. However,
unlike GA, PSO has no evolution operations such as crossover
and mutation. Reasons for choosing PSO versus other evolu-
tionary computation techniques are its ease of implementation,
relatively few parameters to adjust, reputation for finding global
optimum and fast convergence.
One of several differences between particle swarm and tra-
ditional methodologies is that, rather than defining fixed pre-
determined experiments using different nutrient concentrations;
an iterative approach is used in which the results from previous
iterations are used to define the next iteration of experiments.
To explain particle swarm further, using a global mountain
peak search analogy, consider the objective of finding the high-
est point in the Himalayan mountain range. A statistical design
approach might be to identify a “best guess” localized terrain,
encompassing a small number of mountains, which may or
may not include Mount Everest. “N” locations would be pre-
defined in this area, with many located on the boundaries of
the defined area. Climbers would journey to these locations and
measure the altitude (i.e. fermentation yield). The results, in
combination with response surface mapping algorithms, would
hopefully identify the approximate local point of maximum
altitude.
With particle swarm, a larger area (i.e. terrain) would be
defined, encompassing many mountains in the range. A team
of several climbers would be randomly distributed throughout
the range at which time they would note their altitude (i.e. ini-
tial broth composition and resulting fermentation yield—which
is iteration #1). Each of the climbers would then hike toward a
predefined new location (symbolic of a new chemical broth com-
position) and note their new altitude (i.e. fermentation yield),
which is iteration #2. The comparison of altitude changes (i.e.
yield improvement) for each of the climbers (from iteration 1 to
2) as well as where the highest climber (best yield) is located is
used to determine the direction (i.e. the next broth composition
to try) for each of the climbers for the following iteration.
 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
While not a guarantee, the particle swarm approach increases
the space that can be searched with a similar number of experi-
ments and increases the chance that a global optimum (i.e. Mount
Everest) will be found.
It has been suggested that the two techniques described above
can be used in complementary ways, with evolutionary algo-
rithms doing the initial broad search and a statistical design then
used for fine tuning.
The “particle swarm” evolutionary technique has been tried
on two existing fermentation processes at Lilly. In both fermen-
tation processes, broth medium composition providing >50%
yield improvement resulted, compared to the previous existing
production medium.
Regarding sterilization, it is known that large differences
can exist in the heat up and cool down times of sterilization
operations, depending primarily on the size of fermentors (e.g.
large production fermentors usually have longer total steriliza-
tion cycles than pilot scale fermentors). Since chemical reactions
of nutrient components are usually occurring during steriliza-
tion, differences in heat cycles can cause significantly different
post-sterilization broth composition of nutrients. For example,
glucose, when exposed to sterilizing temperatures, can form
polymers, can react with amino acids to generate Browning
reaction products, and can also react with inorganic phosphorus.
For some fermentations, the difference in heat cycles can cause
sufficient differences in post-sterilization nutrient composition
which can then significantly impact the subsequent extent that
the culture can grow and produce product. Work has been pub-
lished on characterizing the extent that chemical reactions occur
during sterilization, using a parameter defined as Ro (similar in
concept to the use of Fo in characterizing the degree to which
contaminating microorganisms are killed during sterilization).
Both Fo and Ro are integrating functions of time and tempera-
ture. It is suggested that Fo and Ro can be used collectively as a
basis of achieving both a desired kill of contaminating organisms
and a consistent post-sterilization nutrient composition (Boeck,
Wetzel, Burt, Huber, Fowler, & Alford, 1988; Boeck, Alford,
Pieper, & Huber, 1989).
2. Post-bioreactor inoculation operations
The remainder of the process control part of this article will
focus on the control of the post-inoculation culture environment,
typically by computer control systems. Inoculation normally
defines the beginning of seed and fermentor phases of the overall
bioprocess cycle. Topics considered include the current state of
sensors, control algorithms, and data analysis tools.
To provide the reader with a sense of process complexity,
the non-linear batch nature of a bioreactor operation is first
described.
A fermentation will begin with an inoculation step in which
a relatively small number of pure culture cells are transferred to
the bioreactor. The cells then grow exponentially until such time
that something (e.g. oxygen, carbon source) becomes limiting, or
something [e.g. substrate, dissolved carbon dioxide (dCO2 ), or
product] becomes inhibiting. The highest cell mass achieved will
normally be several orders of magnitude greater than the starting
1467
cell mass. This major load change is associated with correspond-
ing major changes during the time course of the fermentation
in oxygen and nutrient requirements, carbon dioxide removal,
“base” needed to maintain pH, etc. Many bioprocesses are addi-
tionally complicated in that an induction or product formation
de-repression activity occurs part way through the bioprocess in
which the culture is induced to change from a “growth” mode to
a “product synthesis” mode. This induction is often triggered by
a programmed shift in temperature or by a chemical addition.
Also, while this paper focuses primarily on computer-based
closed loop aspects of bioprocesses, there is significant work that
goes into the design of the bioreactor itself. As one example, an
agitator normally exists for most aerobic bioprocesses, the mis-
sion of which is to (1) break up sparging gas bubbles into smaller
bubbles (to increase bubble surface area to enhance oxygen and
carbon dioxide transfer) and (2) provide good mixing to achieve
a homogeneous broth. Homogeneity is needed, e.g. to provide a
uniform dissolved oxygen range throughout the fermentor such
that all cells have sufficient oxygen. Homogeneity is also needed
so that slug additions of caustic (for pH control) and antifoam
are quickly dispersed so as to, e.g. minimize local damage to
cells at the point of entry. Unfortunately, the agitator RPM and
types of agitator blades needed to achieve these goals often cre-
ates high shear conditions that are harmful to the cells. So, the
final reactor design plays an important role in the control of a
bioprocess and is usually a compromise with respect to shear
effects, mixing, gas mass transfer rates, and process economics
(i.e. utility costs).
3. Process measurements
“If you can’t measure it, you can’t manage it.”
3.1. Primary on-line sensors
A small number of different on-line sensors are commonly
used in bioprocesses. They include temperature, bioreactor back
pressure, gas flow rates, agitation rate, dissolved oxygen, and pH.
A few additional probes are used in some fermentations [e.g.
dissolved carbon dioxide, redox (oxidation reduction), and opti-
cal density]. Sometimes redundant sensors (e.g. electrochemical
probes such as pH and DO2 ) are used.
The challenge in the development of additional (especially
invasive) on-line sensors has historically been, and continues to
be:
• The need to operate dependably in a harsh, agitated, sterile,
and sometimes gritty environment.
• The need to maintain calibration over long time periods (e.g.
several weeks).
• Interpretation of output (e.g. for optical density in complex
media, redox).
3.2. Primary at-line sensors
Several additional at-line analytical measurements are avail-
able that can provide periodic values of important bioprocess
1468 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
Fig. 4. Process mass spectrometer (an example of Process Analytical Technol-
ogy).
parameters. Examples include:
• broth glucose concentration (e.g. YSI analyzer);
• broth glutamine concentration (e.g. Nova blood gas analyzer);
• cell size distribution (e.g. Lasentech).
While a few auto-sampling systems are known to exist, the
general limitations of these at-line sensors include (1) their use
of manual operations (e.g. sampling and sample preparation)
and (2) measurement frequency that may not practically allow
for closed loop feedback control.
3.3. Process analytical technologies
A few analytical systems have evolved over the past 20
years that provide powerful additional real-time or near real-
time information about bioprocesses. They include:
1. Gas analysis, using process mass spectrometry (replacing
earlier use of infrared carbon dioxide analyzers, paramag-
netic oxygen analyzers, and gas chromatographs).
These systems typically provide analysis of the concentra-
tions of oxygen, nitrogen, and carbon dioxide in gas streams.
They sometimes also provide information on other gases that
may be present (e.g. alcohols, hydrogen sulfide). One of the
most popular types of process mass spectrometers is the mag-
netic sector type (e.g. from Thermo-Electron & Hamilton
Sustrand) due to their very high accuracy and stability (see
Fig. 4).
Fig. 5. Culture oxygen uptake vs. time.
The information from these gas analyzers are typically
combined with other measurements or on-line estimates (e.g.
DO2 , gas flow rates, broth volume) to calculate and make
available in near real-time a large number of additional
parameters about a bioprocess. These include: culture oxy-
gen uptake (see Fig. 5), culture carbon dioxide evolution,
respiration quotient, kL a (oxygen mass transfer coefficient),
metabolic heat output, etc. This information has proven so
valuable to operators, scientists, and technical service per-
sonnel that Lilly has process mass spectrometers interfaced
to most of their development and production fermentors
(Alford, 2006).
2. Broth composition measurements, including use of near
infrared (NIR), mid infrared (MIR), ion chromatography,
and HPLC. These systems can provide analysis of nutrient
compositions, ammonia, product concentrations, and other
metabolites.
The Food and Drug Administration (FDA) is now actively
promoting the consideration of Process Analytical Technol-
ogy (PAT) in pharmaceutical manufacturing processes. They
define PAT as “a system for designing, analyzing, and con-
trolling manufacturing through timely measurements (i.e. dur-
ing processing) of critical quality and performance attributes
of raw and in-process materials and processes with the goal
of ensuring final product quality (U.S. Department of Health
and Human Services, Food and Drug Administration, 2004).”
It is expected that the adoption of the concepts and technolo-
gies behind PAT will help lead the pharmaceutical industry
in its evolution toward more advanced process control. This
is because several bioprocess measurements, historically per-
formed off-line with results obtained too late to be of pro-
cess control relevance, will now be performed more frequently
and in near real-time and, therefore, available as control loop
inputs.
 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
One of the issues in using PAT systems is whether these
are smart sensors or computer systems (PAT systems typi-
cally include computers that handle data processing, calibration,
analysis, etc., and which typically require software configur-
ing and/or programming). How these systems are defined will
impact the degree to which they must be validated.
3.4. Virtual sensors
Despite the range of measurements available with the above
listed technologies, some challenges still exist, e.g.
1. Estimating cell mass in fermentations using complex media
ingredients, i.e. ingredients that include solids (e.g. soy bean
grits).
2. Estimating broth primary carbon substrate (e.g. glucose) con-
centration on-line—for use in continuous control.
3. Estimating product concentration on-line.
Accurate estimates/measurements of these parameters are
important as most published bioprocess models include cell
mass, primary carbon substrate concentration and product con-
centration as primary “state” variables.
Some success has been reported in estimating parameters
such as these via use of virtual sensors. These sensors make use
of available on-line information and a model (via, e.g. statistical
regression, neural nets) to predict those parameters which are
impractical to continuously measure directly. Not surprisingly,
many of these virtual sensors make use of culture oxygen uptake
or carbon dioxide evolution (CER) calculated parameters (avail-
able from process mass spectrometry and other measurements)
as a key input, as these are strongly correlated to, e.g. glucose
consumption and cell growth. For example, some bioprocesses
(especially ones using complex media) have used the simple
equation:
dX
CER = k1 × + k2 × X (1)
dt
(where X is cell mass and t is time) to accurately estimate cell
mass on-line, especially for seed vessels and the growth phases
of fermentations (Alford, 1978). This equation assumes that cell
respiration activity (represented by CER) comes primarily from
two types of activities: (1) cell maintenance (proportional to
X) and (2) cell growth (proportional to dX/dt). Fitting Eq. (1)
to historical X and CER data results in the determination of
k1 and k2 . CER, already calculated on-line, then allows for the
additional on-line calculation of cell mass (X). As the batch
proceeds in time, this equation is eventually forgiving of any
error in the initial estimate of X (i.e. X at t = 0) and can be used
as a rational basis for certain process control decisions, such as
when to transfer the seed inoculum to the fermentor.
While virtual sensors can be quite accurate during growth
stages of cultures, they are often less accurate during product
synthesis portions of the process. This is because cellular activ-
ities involved with product synthesis are less well understood,
and significant cell death and product inhibition may be occur-
ring.
1469
3.5. Sensor advances/issues
In addition to advances in the measurements available online
and at-line, the technologies involved with sensors are also
rapidly progressing.
• Many sensors are becoming much smaller, leading to micro
sensor arrays.
• Sensors (and valves) are becoming smarter-providing mainte-
nance and diagnostic information as well as traditional mea-
surements and/or valve position indication. (Note: While this
information is valuable, it can also contribute to information
and alarm overload for operators.)
• Digital highways are available to communicate this addi-
tional information to computer control systems (e.g. Fieldbus,
Profibus).
• Some sensor-to-computer communications are becoming
“wireless.”
In general, the pharmaceutical industry moves slowly and
cautiously in adopting such new technologies—due in part to
the regulatory environment in which companies operate, the
need to prospectively validate (provide documented verifica-
tion) that they work, and the challenge of obtaining quality
control and management approval in a typically conservative
and bureaucratic change control environment. Therefore, bio-
process engineers in industry are watching these trends, but will
typically not be the first ones to try them out.
As an example of an opportunity for further research and
development, significant success has been achieved in using
neural nets as virtual sensors for applications such as on-line
substrate (e.g. glucose) concentration, cell mass, etc. However,
neural nets are known as a “black box” type of model meaning
they are not based on first principles (i.e. deterministic). One
of the consequences of this is that they are generally poor at
extrapolating. They are also weak in accommodating lags (usu-
ally requiring trial and error estimates of delay constants) which
are frequently encountered in bioprocesses. There is significant
appeal in developing “hybrid models” such that all available first
principle information is utilized in the model with neural nets
then used to estimate parameters (e.g. specific culture growth
rate constant) that is a complex non-linear function of several
parameters. Psichogios and Ungar (1992) successfully used this
approach in modeling a bioprocess. However, they separately fit
the model constants in the first principles model and the node
weights in the neural net. We would challenge vendors and uni-
versities to develop algorithms that could simultaneously fit all
the constants in both portions of the hybrid model, including
process delay parameters, which could then theoretically gener-
ate more meaningful values for the constants and more accurate
models.
4. Control techniques
A comprehensive review of fermentor control techniques was
published by Rani and Rao (1999). This review summarizes
and references over 100 other published articles, many deal-
1470 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
Fig. 6. Typical computer controlled fermentor.
ing with various aspects of advanced control. These include
dynamic programming, on-line adaptive control, on-line opti-
mization, non-linear control, and optimal control. Many of the
articles involve neural nets.
Bioprocesses employ most of the same types of control as
are used in other chemical industries. Over half of most bio-
process control loops can be handled by traditional single input
single output feedback PI (proportional + integral) controllers.
(See Fig. 6 for typical manipulated variables such as sterile air-
flow, nutrient feed, pressure, cooling water flow and agitation
rate, and controlled variables such as pH, temperature, and dis-
solved oxygen.)
The most frequently found more advanced type of control
are DO2 cascade controllers in which dissolved oxygen (i.e. the
master loop) is cascaded to the control loop for agitation, airflow,
substrate feed rate, or some other parameter (the slave loop).
Other examples of advanced control known to exist for indus-
trial bioprocesses are:
1. Multivariable control (e.g. to simultaneously control DO2
and dCO2 using independently controlled air and pure oxy-
gen gas feeds).
2. Model reference control (e.g. to control glucose based on
a virtual sensor). Model reference controllers are especially
effective in minimizing the problem of dead time (i.e. trans-
portation delay) associated with at-line instruments.
3. Adaptive control (e.g. adaptive gain).
While most of these techniques work satisfactorily, there are
reasons for caution and opportunities for improvement:
1. Process non-linearity
PID is a controller intended for processes that behave in a
linear fashion (or in a pseudo-linear region of a larger non-
linear space). Microorganism cultures are non-linear in many
respects. For example, the most common model for a limiting
substrate is the well-known Michaelis–Menten equation:
Max Rate × S
Reaction rate = (2)
Km + S
where Km is the Michaelis–Menten constant and is equal to
the substrate concentration at which the reaction rate is half
the Max Rate.
So, when a substrate (S) such as glucose becomes limiting
to the culture [when S in Eq. (2) becomes small compared to
Km ], then the relationship between metabolic activity (e.g.
glucose consumption) and glucose concentration changes
from zero order to first order. For some fermentations (e.g.
Bakers Yeast), there is an additional non-linear complication
in that glucose concentrations that are too high will cause the
culture to shift from a metabolism of making yeast to mak-
ing ethanol. The control of dissolved oxygen is another good
example where a brief power outage can cause the loss of
agitation causing the DO2 to go to near zero which, in turn,
causes the culture to slow and/or stop some metabolic activ-
ities. The dynamics of recovery will be much different than
if the DO2 had only dropped a small amount. The control
of dissolved CO2 for mammalian cell-based processes is yet
another example where concentrations too low or too high
are detrimental to the culture while a range in the middle is
OK.
2. Sub-optimal control
The pharmaceutical industry is not unlike many other
industries in which observers have noted that control loops
are often found in manual rather than automatic mode and
that loops are not well tuned. Some observers suggest a gap
exists between what is typically taught in university process
control courses and what is needed in industries such as the
batch oriented pharmaceutical industry.
More specifically, many university process control courses
focus on theoretical aspects of continuous “steady state”
types of processes and include such topics as Laplace trans-
forms, Nyquist Stability Criterion, Bode plots, Root Locus
diagrams, and Routh Stability Arrays. In contrast to this, a
large percentage of today’s industrial processes are batch
non-steady-state processes and the techniques mentioned
above are rarely used by practicing automation engineers.
There appears to be a need for more focus in educational
forums on dealing with process transients, process phase
transitions, and other non-linear control dynamics of batch
processing and more training on loop tuning techniques and
control valve characteristics and selection.
3. Advanced control algorithms (Fuzzy Logic, Model Predic-
tive Control, etc.)
As with new hardware technologies, the pharmaceutical
industry tends to move slowly and cautiously with regards to
new approaches in control algorithms.
Practicing engineers are monitoring the evolution of tech-
niques such as Fuzzy Logic Controllers and Model Predictive
Control (MPC)—and considering their potential application to
bioprocesses. For example, Nyttle and Chidambaram (1993)
 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
showed how a fuzzy logic controller could use glucose (sub-
strate) feed to force a fermentation to follow a predetermined
product (Penicillin) time varying concentration profile.
There is some reluctance to aggressively pursue such tech-
nologies for industrial bioprocesses due to:
A. Uncertainty as to how to prospectively validate them.
B. A desire to keep things simple for the supporting engineers
who are often instrument technicians or B.S. Chemical Engi-
neers with only one university course in process control (i.e.
do as much as possible with PID feedback and cascade tech-
niques).
C. Lack of compelling published evidence (so far) that such
algorithms significantly increase bioprocess productivity.
Specific to MPC, additional concerns include:
A. The technique was initially developed for continuous pro-
cesses. While also applicable to batch processes, the per-
ceived complexity is greater due to significant changes in
process gain and dynamics during the batch.
B. Reluctance to perturb the process to generate data to build
the model (especially regarding pH and temperature).
C. The perception (real or imagined) that outside consultants
are needed to help develop (and sometimes support) the pro-
cess model and overall optimization application.
D. Concerns of sustainability. A recent published survey by
Invensys (Rosenzweig, 2005) noted “The real issue for MPC
is keeping operators from turning it off. Over 50% of all pre-
dictive controllers are in manual.”
On the positive side, good progress has recently been made
in simplifying the implementation of MPC. For example, Inven-
sys, Emerson, Solutia, and Aspen Technology have developed
improved tools that reduce the effort, expertise, and expense
previously required with using MPC.
5. Process optimization
Much has been published regarding the creation of models
of bioprocesses as a means of better understanding the process
and in helping generate optimum control strategies. Tradition-
ally, control setpoints for industrial bioprocesses have been
determined by trial and error experimentation in pilot plants;
a sometimes long and labor intensive process. Note that some
setpoints are typically time varying, such as aeration, agitation,
and nutrient feed rates.
In teaming with process microbiologists, the author has had
experience with techniques in which a matrix of several biopro-
cess non-linear state equations is specified (a challenge in itself)
with parameter estimation (also non-trivial) then pursued using
experimental data. A similar size matrix of adjoint equations is
then specified and Hamiltonians, Pontryagin’s Maximum Princi-
ple, and Calculus of Variations then used to compute an optimal
time varying control variable setpoint (e.g. a nutrient feed rate).
These efforts have always been very time consuming and have
always resulted in better understanding of the bioprocess, but
1471
have only sometimes been successful in evolving an improved
process control strategy.
Of significant recent interest is the use of neural nets for such
purposes. This approach is less time consuming, is easier for
engineers to learn and use, and avoids the need to define deter-
ministic equations of the process. Vlassides, Ferrier, and Block
(2001) include a discussion and application of this approach for
wine fermentations.
However, as suggested before in Section 3.5, there is promise
in using hybrid models in defining the process and pursuing pro-
cess optimization in which neural nets would be complimented
with known first principle equations.
Another kind of process optimization involves minimizing
energy costs. Fermentors are often the highest energy con-
suming unit operation in a plant due to the high volumes of
compressed air and high agitation power required. Shields and
Kao (1994) developed and implemented a model in which
airflow and agitation RPM were managed so as to control
DO2 at a desired setpoint while minimizing the total energy
consumption.
6. Other automation system functionality
In addition to control algorithms, process control computers
are expected to provide additional utilities/functionality related
to process automation. These can include batch sequencing,
alarm management, interlocks, diagnostics, links to historians,
interfacing to PAT and at-line analytical systems, and pro-
viding effective easily configured color-graphic displays for
operators.
As a specific example of the opportunities and issues involved
with these other process control functions, alarm management
is discussed.
Alarm management is a significant problem area for most
companies in the chemical industries, including bioprocesses.
Nuisance alarms often exist which: (1) frustrates operators, (2)
trigger needless process deviation investigations, and (3) gives a
perception to others of a process NOT in control. Further, smart
alarms which: (1) can access all pertinent process information
within the computer, (2) provide insight to operators as to root
cause, (3) display the expected operator response, and (4) link
to paging systems are often difficult to configure. Opportunities
for improvement exist with both company implementation prac-
tices and with the functionality provided by vendor off-the-shelf
products.
The situation is gradually becoming worse as smart sensors,
smart valves, and communication protocols such as fieldbus
and high speed Ethernet are quickly expanding the amount of
information available to the process control system and plant
engineers are tempted to add more alarms in linking to some of
this information.
The companies doing a good job with alarms tend to be those
that adhere to the fundamental definition of an alarm—i.e. that an
alarm represents an abnormal condition that requires a response.
Lack of adherence to this is the single greatest source of nuisance
alarms and information overload to operators.
1472 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
From a vendor product perspective, many systems provide
little more than the ability to compare a current value with a
setpoint and generate an alarm should this gap exceed a user
specified value. If this alarm is then sent to a historian or alarm
logger, the alarm record contains little more than a calendar
time stamp, a cryptic description of the parameter in alarm, and
perhaps a value for the parameter in alarm. Alarm record sorting
capability is minimal as there is little alarm tag information to
sort on. Alarm operator acknowledge records may or may not
exist to help in understanding the history of and pursuing root
cause analysis of process deviation events.
Process control system historian vendors have had suffi-
ciently limited capability that many industrial companies have
resorted to use of third party alarm loggers. In the case of Eli
Lilly and Company, a real-time expert system was developed
and interfaced to the bioprocess historians (Alford et al., 1999a;
Alford et al., 1999b).
To illustrate with a bioprocess example: a DO2 alarm in a
typical commercial control system might normally be generated
by comparing a DO2 probe output (current value) to a prede-
termined number (e.g. setpoint). If this difference exceeds a
predetermined threshold, an output is triggered to control room
displays and devices and a record is generated to the histo-
rian containing the measured value, loop tag, and calendar time
stamp. However much more is possible. A smarter alarm might
be generated by using coded “if-then-else” rules to look at all
other relevant information within the automation system (e.g.
on-line oxygen uptake and kL a calculated parameters, redun-
dant DO2 probes, the relative phase time, etc.), in which case a
more accurate conclusion might be that the control DO2 probe is
drifting and is no longer valid. Such a “smarter” alarm might then
indicate this more “intelligent conclusion” in the console presen-
tation to operators, and also note the suggested “predetermined”
operator response to the alarm. If the facility is unmanned during
portions of the week, the system would “page” the appropriate
personnel. Alarm records in the historian would include fer-
mentor #, lot #, batch phase/step identifier, alarm class (safety,
product quality, environmental, etc.), and other tag informa-
tion to facilitate alarm record sorting and report generation. In
addition, the capability would exist to display the alarm record
information in “relative time” (i.e. time since the beginning of
the batch or batch phase). For batch processes, such as most
pharmaceutical processes, this “relative time” is far more valu-
able to analysts than “calendar time, since the importance of
an alarm (or any trend plot) is usually from the perspective of
when it occurred relative to the beginning of a batch or process
phase.
As value adding as the above described smarter alarm might
seem, the “end in mind” is suggestive of even more alarm
functionality—specifically the ability to compare a parameter’s
(e.g. DO2 ) value in a currently running batch process to the “time
varying” range of values established by the historical average ±1
or 2 standard deviations of recent satisfactory production lots.
The current value’s movement outside this time varying band
might then be the logic that triggers an alarm (see Fig. 7).
Lilly’s use of real-time expert systems in most of their bio-
process development and manufacturing facilities accomplishes
Fig. 7. Current process trend compared to historical average profile ±2S.D.
all of the above described “intelligent and remote alarming”
functionality. One bioprocess manufacturing plant, following
installation of this system, reported a 4% yield increase and
double digit reduction in variability, primarily due to ear-
lier identification and better management of poor perform-
ing fermentations. Also, as the name “expert system” implies,
such systems offer the opportunity to practically capture and
put on-line the expertise of experienced operators, engineers,
and technical service personnel in analyzing incoming data.
It is of interest that of several manufacturing plants report-
ing on productivity in the months following Lilly’s only early
retirement program (in 1993 in which all plants suffered key
personnel losses), the one using a real-time expert system
was the only one reporting no significant plant productivity
loss.
However, since real-time expert systems are third party
functionality that must be separately supported, validated, and
interfaced to other systems, the hope is that commercial pro-
cess control and historian vendors will evolve their systems to
accommodate more of this intelligent alarming and record anal-
ysis functionality. Alford, Kindervater, and Stankovich (2005)
describe alarm management best practices in greater detail and
notes some of the things process control and historian commer-
cial vendors can do to help.
7. Historians and data analysis
In general, process automation systems today are capturing
large volumes of bioprocess data and storing them in databases.
Further, computer control/historian utilities are available and
relatively easy to use so that data, in terms of trend plots, is
easy to visualize. In addition, tools are available to help mine
the information and knowledge content in the stored data. These
tools include model building software (e.g. neural nets), statisti-
cal analysis (e.g. JMP), principle component analysis, decision
tree generation, three dimensional graphics, rule-based systems
to analyze incoming data in real-time, etc.
 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
Companies providing effective data analysis tools include
SAS, Aegis, Spotfire, Gensym, Curvacious, and several oth-
ers. Unfortunately, most of these tools are not embedded within
commercial historians and therefore require interfacing the his-
torians to third party systems—or at least manually downloading
selected data sets into Excel spreadsheets and then further trans-
ferring the data to the analysis tool.
The problem is that, while several excellent commercial
data analysis and mining products exist, they are not used
very much in the bioprocess industry. This has frequently been
attributed to the significant manual effort required in prepar-
ing data sets for presentation to the applicable tool. A rule of
thumb is that it takes about 90% of the overall data mining
project time and effort to select and prepare data sets for analy-
sis and about 10% of the time to run the data analysis tool. This
90% prep time is a major barrier to greater use of data mining
tools.
To help develop and evaluate bioprocess data mining tech-
niques, Lilly joined a MIT sponsored consortium on fermen-
tation data analysis in the early 1990s, chaired by Prof. Greg
Stephanopoulos. As part of this effort, Lilly examined 2 years
of bioprocess data from one of their plants to find about 115
lots for which the data was perceived as valid. A large num-
ber of lots was necessary given the large number of vari-
ables affecting the process and the high natural variability
of the process. For example, in just considering the inocula-
tion step, a presumed set number of cells from a seed tank
(based on measured liquid volume and estimated cell den-
sity) must be transferred to the fermentor, during which time
the cells could become oxygen starved or come into contact
with insufficiently cooled transfer piping. Therefore, a fermen-
tor lot will not always start out with the same number of
viable cells—which can then significantly impact the subsequent
growth, oxygen uptake, dissolved oxygen, and other trends ver-
sus time.
Regarding whether fermentations were acceptable for anal-
ysis, lots that became contaminated or had sensors that failed
during the run were identified from batch records and thrown
out. Then, each of the process trends (about 30/lot) were called
up and visually inspected to look for data outliers (e.g. spuri-
ous points due to momentary power failures during electrical
storms). Outlier points were then deleted from the data set to be
used for analysis.
The next challenge was having to estimate each parameter at
specific time intervals (required for some analysis tools). This
required either averaging multiple data points or decompression
of compressed data in the historian for continuous variables (e.g.
pH). For off-line assays, this required taking once/day assays and
estimating (via interpolation) values at other times.
Once this work was done, the data was normalized and
presented in the format expected by the analysis tool. One
of the several mining techniques applied to the data set was
principle component analysis, shown in Fig. 8. One of the
benefits of this analysis was the elimination of most pro-
posed hypotheses regarding the difference between good and
bad runs that relate to the time period between inoculation
(time = 0) and induction; i.e. the cause of bad runs is probably
1473
Fig. 8. Principle component analysis.
not any aspect of bioreactor control between inoculation and
induction.
If this effort sounds daunting, this is just the effort required
in looking at a single unit operation, i.e. the fermentor. Consider
the more general case when trying to determine the root causes
of variability in a down-stream operation (see Fig. 3). The cause
could be in any of the preceding unit operations, such as the
fermentation, or, in fact, in a manually controlled operation early
in the life of the culture (see Fig. 2). In collecting data to analyze,
some of the data may need to be manually added to the database
(i.e. from manually monitored steps of the overall operation) and
portions of the data may come from different historians, stored
in different formats. Further, there is typically not a one to one
correlation of fermentor lots to purification lots (e.g. due to the
combining of lots for some operations) so lot tracking genealogy
is often a challenge.
So, the key to greater progress in data mining is in evolving
tools to facilitate data set preparation. Products such as Process
Insights (from Pavillion) and Discoverant (from Aegis) have
significant data collection and preparation functionality to help
with this.
8. Concluding comment
The state of bioprocess control is consistent with a recent
article (Spear, 2005) in which it is noted that the process control
industry clearly has a problem. There are two speeds in play:
(1) the rapid pace of digital communications development and
(2) the more sedate rate (measured in years, if not decades) of
many of its customers.
New biosensors, smarter field devices, and digital commu-
nications are about to give the bioprocess industry significantly
more data than it had before. The problem is that users are not
yet very good about efficiently consolidating, analyzing, and
mining the data that they already have. Plants already have large
1474 J.S. Alford / Computers and Chemical Engineering 30 (2006) 1464–1475
numbers of nuisance alarms and data stays as data rather than
becoming information and knowledge.
If technologists learn to be smarter in what they do with the
data they already have, they will be in much better position to
intelligently manage the increase in data as new and smarter
field devices and measurement techniques are developed.
While pharmaceutical companies and vendors are working
on these improvement opportunities, research institutions can
be working on:
1. Better understanding (including models) of the relationship
between intracellular metabolic pathways and extracellular
parameters. (Note that fermentor control is primarily about
trying to influence the cells internal environment by manip-
ulating the external environment.)
2. Improving hybrid neural net functionality.
3. Fast, efficient, and easy to configure plant scheduling tools.
4. More user friendly versions of advanced control techniques.
9. Additional information: the Lilly fermentation
computer system
In the early 1970s, Lilly scientists and engineers defined
a vision for the monitoring and control of bioprocesses that
could not be realized with commercial systems available at that
time. Therefore, a team was organized that developed Lilly’s
customized fermentation process control and historian system
(Alford, 1981, 1990).
During the years that followed, Lilly incorporated new com-
puter science technology as it became commercially available. A
major PAT addition was the use of process mass spectrometers.
This system was replicated into all of Lilly’s major fermenta-
tion research, development, and manufacturing facilities during
the late 1970s and early 1980s. Plant site historians were elec-
tronically linked together to facilitate sharing and comparing
of process data. Interface to real-time expert systems started in
1990. Many of these systems are still operating today, over 25
years later.
Several features of this system remain as advanced function-
ality as compared to today’s commercial systems. Automation
vendors should make note that these are practical to do and are
value adding to customers:
1. On-line changeability of all process control recipe logic.
This is especially valuable for research and development
operations when changes are frequent and cannot all be
prospectively anticipated prior to a run.
2. Automated audit trails for on-line changes.
3. Continuous and batch sequencing logic in a single configu-
ration environment (i.e. both paradigms implemented in the
same recipe).
4. Process control recipe creation that is simple enough that it
can be done by technicians.
5. Trend plots displayed in relative time (i.e. time since the
beginning of the batch—or batch step).
6. Process data from different fermentors at different plants
displayed on the same plot (especially valuable in scaling
up processes and in comparing different plants making the
same product).
7. The ability to display alarms, operator comments, off-line
generated discrete assay data and process control continu-
ous data on the same plot/screen.
8. Interface with rule-based expert systems for real-time pro-
cess diagnostics, data analysis, and intelligent remote alarm-
ing applications.
9. Use of relative input–output addressing. This enables the
same application recipe to be used for any similarly instru-
mented bioreactor.
10. Data records which can be sorted, queried, and displayed
by tank #, lot #, process step/phase, and any other tagged
attributes.
11. Ability to view data plots and receive alarms from remote
locations, including employee homes.
12. Ability to decompress historized data into interpolated esti-
mates at periodic time intervals (required for some data
analysis tools).
13. Ability to calculate bands (i.e. the time varying average
value ±2 standard deviations) based on analysis of selected
historical data, for use as a backdrop in process control data
trends.
Acknowledgments
The author wishes to acknowledge the commitment to excel-
lence and the many opportunities given to him by Eli Lilly &
Co. management during his career. He further acknowledges the
contributions of his mentors, peers, and associates for their great
help in projects and in his understanding of bioprocesses.
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