Monday, February 25, 2008


It is not surprising that sooner or later some one or some fermentation technologists will come up with the idea of inventing 'disposable bioreactors' in this age of convenience and take aways and disposables. The idea is not new and have been in the minds of most fermentation technologists. It is only the question of:

1 What sort of fermentation is suitable for the kind of fermentation that can be carried out using disposable bioreactors?
2 What advantages do we get by using the disposable fermentors compared to the traditional and widely accepted stainless steel CSTRs?

At present the use of disposable fermentor bags are used only in small scale cultivation of mammalian cells and biologics in pharmaceutical and biotechnology companies.

There are several advantages and disadvantages in using these disposable bioreactor bags.
In terms of advantages these disposable bags are manufactured under sterile materials and under clean room environment. There is no need for cleaning processes and thus will shorten downtime and turn around time. As new bags are always used for each fermentation run there is lower risks of cross contaminations

The main disadvantage faced in using these disposable fermentors are scaling up of the production process. Comparing to CSTR with well established hydrodynamic characteristics which makes it easy for scaling up, disposable bioreactors hydrodynamics are unchartered territories

Maybe in the future this problem of scaling up the disposable bioreactors will be overcome. Since these disposable bioreactors are for single use and made of expensive plastics, there is the need to buy a large number of these bioreactors.

In essence the disposable bioreactor bags are just specialised plastic bags that are used to carry out fermentations. It fulfill the basic criteria of the fermentor but it does face limitations such as:

1 The type of mixing that can be carried outt in the disposable bioreactor bags. The bag will only only have to depend on simple mixing such as provided by the ' rocking mixing' and is not capable of a more powerful shearing mixing as seen in conventional bioreactors. Maybe this kind of mixings are more suitable for the fermentation of animal or plant cells? Less shear forces

Then there is the question of sensors. Sensors are expensive and could not be regarded as disposable item thrown away after every fermentation. These sensors could be re used again after sterilizations in the disposable plastic bags

Thus for the moment we can safely say that these disposable bioreactor bags are suitable for low volume high value fermentation
Read more!

Sunday, February 24, 2008


As I have been and will be very busy for the next two weeks with major consultancies. I would like to inform to all that I might not be free enough to blog on this blog for the stated period. Blogging will resume actively after the stated period. My apologies to all Read more!

Thursday, February 21, 2008


Cooling towers are common sights in the vicinity of buildings,factories and even power plants. These cooling towers function as heat transfers units that will throw away the heat to the environment so as to maintain ideal temperatures within the building environments or production processes.

One of the most popular cooling tower model is the water cooled cooling tower. Heat is transferred to the coolant water through heat exchangers and the heated water will be pumped to the cooling towers where the excess heat is loss to the surrounding environment through evaporation of the water

Most of these cooling towers are exposed to the environment and in fact act like bioreactors which can support high growth of various types of microorganisms. These build up of microorganisms are the source of many problems in various industrial processes such as microbial corrosion, loss of heat transfers and clogging of pipes and are even hazardous to our health as linked to the famous Legionella's disease.


Looking at the typical cooling tower show all the requirements of a septic fermentor

to be continued...breakfast now
Read more!


Throughout my tenure teaching Fermentation Technology to Microbiology and Biotechnology students at the University Malaya I have always stressed the importance of 'hands - on' practicals to the various students taking the course. For fermentation practicals each group of six students are assigned one unit of laboratory scale fermentor of 2 litres capacity throughout their course

Students taking on the course will cover the following practicals before that are deemed to have completed the fermentation technology course practicals. These practicals dominate many hours of continuous work over days and nights and even holidays. Students who do not complete each practical to the expectation of the lecturer will have to repeat the experiments until the practicals are properly executed

Anatomy of fermentor whereby the students are required to dismantle and identify the various components of the fermentor and study the various systems making up the fermentor

Students are required to learn the importance of cleaning the fermentor properly and to carry out COP cleaning

Students are required to assemble the fermentor and to check that everything is in order prior to autoclaving or sterilization of fermentor

Students also learn the anatomy and function of the various standard electrodes in the fermentor such as ph, Dissolved oxygen, foam probe, temperature probe. The students will learn the correct methods of calibrating the ph and DO probes

Students built their own cartridge filters using glass wool

Students carry out studies on rheology of the fermentation broth,
Students will carry out simple experiments to monitor change in viscosity of the fermentation broth

Types of primary, secondary and tertiary mixings using dyes as tracers. Newtonian and Non Newtonian broth will be used for comparative studies

Students will learn how to set up the fermentation console

Students learn how to carry out post sterilization procedures

Students carry out water loss studies from fermentor due to sterilizations

Students learn to carry out blank runs to test the integrity of the fermentors

Students learn aseptic methods of inoculating the fermentor

Students learn how to carry out aseptic samplings from the fermentor

Students carry out various tests for detecting microbial contaminations

Students carry out microbial isolation and screening studies

Students carry out growth curve studies at different temperature and ph

Students carry out microbial cell culture amplification

Students carry out complete fermentation monitoring experiments using various parameters such as substrate, ph, biomass products etc using various chemical, instrumental techniques

Students carry out OUR and OTR studies using the fermentors

Students carry out some down stream processing equipments

TROUBLE SHOOTING AND DIAGNOSTICS ( Continuous clinical observations)
Trouble shooting and diagnostics of the fermentor and fermentation process are continually carried out through out the whole fermentation practicals as part of their fermentation pathology exercise. Rapid physical, chemical, biochemical and microbiological techniques will be exposed

Where after learning all the above, the students will carry out detailed fermentation study of their choice incorporating all the lessons learned

It is sad to note ever since my retirement from the university those tight and demanding practicals might not be executed as I would like the students to do Read more!

Wednesday, February 20, 2008


This is a true story of a food microbiology practical being conducted in one of the outstanding universities in the country which has gone awry, not due to the lack of instructions how to prepare sauerkraut but due to ignorance and poor understanding of fermentation technology

Although the steps are clear and all the components needed to produce the sauerkraut fermentation are followed, the practical failed.

Why the practical failed is easily understood if one understand the concept of fermentation technology. The lecturer who instructed the class in preparing the sauerkraut fermentation made two key mistakes:

1 Using a large Kilner jar to hold the fermentation process but it only contain very little food substrate and a large volume of head space
2 The kilner jars were regularly opened up and stirred like an open pot cooking

In such situations the fermentation will fail because of too much exposure to air and oxygen which will prevent proper fermentation of sauerkraut from taking place

It is hoped that lecturers or teachers intending to carry out such fermentation studies learn from their mistakes :)). It is very embarrassing to note lecturers with PhD still failed to carry out fermentation when their uneducated ancestors can do the fermentation properly Read more!


Recently, by chance I came to read a chapter entitled"Tapai processing in Malaysia: A technology in Transition" by Zahara Marican and Yeoh Quee Lan in a book edited by K H Steinkraus entitled "Industrialization of indigenous fermented foods". The authors discussed in general about making tapai in Malaysia.

There were a few interesting points made by the authoress such as on page 252 which I quote "tapai fermentation is not an anaerobic process and this partly accounts for the low alcohol content. Furthermore the fermentation is normally arrested before all of the sugars are fermented, as the desired end product should not be alcoholic if it is to be consumed by those opposed to alcohol consumption for religious or other reasons"

In the above statement I only want to direct to the points that:
1 Tapai fermentation is not anaerobic
2 Partly account for the low alcohol content
3 fermentation arrested before all the sugars are fermented

From the above statements I seem to see that even within the same sentence or paragraph the two authors contradict themselves. While in the early statement they deny tapai is a fermentation process but in the later sentences they admit that the fermentation process is arrested

I would like to make a point here that in the production of tapai it it involves yeasts and fungi which are capable of living aerobically or by fermentation. In the beginning stage of tapai production, enzymes are released to release the free sugars. This explains in the process of the enzyme hydrolysis why the tapai is sweet. At this stage it may or may not be aerobic.
However, when alcohol are produced by the yeasts by metabolizing the sugars in the absence of oxygen, it is definitely fermentation. In fact it is a good example of the classic traditional fermentation

One must be aware that tapai production is slightly different compared to wine or beer fermentation, even though sugars are used and alcohol are produced. In tapai fermentation it involves the concept of solid substrate fermentation. Each of the cooked pulut or ubi is coated with a thin layer of viscous liquid which makes oxygen diffusion and penetration difficult and thus setting the environment for alcohol fermentation. The action of the sugar molecules with the liquid caused rheological changes making it sticky or viscous. This is attributed to the interaction of the hydrogen bondings between the sugar molecules and water molecules

The low alcohol content achieved by some makers of tapai is attributed to the slightly open or not tight packing and high room temperature which would result in the vaporization ot volatilization of alcohol to the environment. After all the alcohol as fermentation products are short chain and gave a lower evaporation or boiling point

The sad point I want to make here is that the article has not changed much despite advanced understanding of the biochemistry and biotechnology of the fermentation process. The chapter would have been more interesting rather than traditionally detailing the usual methods of making tapai and listing the various microorganisms.

It is in this aspect why I feel that until the understanding of fermentation technology is fully carried out, there is really no transformations in the technology of tapai making and the industry will still lag far behind

For those interested too see how to produce tapai or interested in making money out of tapai, please go to the following website:

The site is intersting as not only it gives tips to making tapai but valuable information in trouble shooting tapai making process Read more!

Monday, February 18, 2008


In times of energy shortage and increasing pollution by wastes, many companies have toyed with the idea of extracting methane as a fuel from the tons and tons of waste filling the landfills. In theory this idea seems to be very very attractive. However in reality, this idea might not be technically or economically viable. There are many problems that will be faced in attempting to produce and extract methane from landfill compared to doing anaerobic digestion in proper digesters

In normal anaerobic digesters the type and composition of wastes fed into the digestors can roughly be controlled, in terms of BOD loadings and HRT. This is not so in landfills. In landfills the wastes filling the landfill anaerobic bioreactor are of unknown composition. Most of these wastes are not amenable to anaerobic digestion as they consist of solid wastes made up of yard wastes, plastics, polystyrenes. Their loading rate and organic load could not be ascertained. As reported in previous blog here, some of the wastes in landfills are still not degraded after several decades

Compared to conventional anaerobic digesters, landfills are basically SSF or exhibit Solid Substrate fermentation. This will mean that that there will be not much liquid in SSF fermentation.

Anaerobic digestion usually perform well in high liquid environment where not only will it prevent oxygen diffusion but water is a necessary requirement for the nutrient supply and the growth of microorganisms. In fact a few studies in landfills show better decomposition if the landfill leachate is recycled.

In a normal operating anaerobic digesters treating sewage sludge it is shown that a long retention time of around 10 to 15 days is good for anaerobic digestion. In landfills this is much more longer as the landfill digester is not optimally designed as an anaerobic digester to produce methane. Thus in terms of methane production landfills are not technologically efficient process.
There will be a long slow rate of methane production stretching for years which economically not viable

Most anaerobic digesters are operated fed batch wise with HRT about 10 to 15. Landfill bioreactors are more effective as batch reactors functioning more as tombs continually taking the solid wastes until filled to the brim or even above the level of the earth

If we consider landfills as the biggest bioreactor compared to conventional anaerobic digesters, it will be difficult to mix or improve the mass transfers due to its sheer size. There are many points to consider where to collect the gases or prevent loss of methane gases. It is generally accepted that the loss of methane gas through landfills account to about 30 to 40%. This is economically unacceptable in any methane generating bentures. Technology of methane recovery in landfills has much to be desired.

To be commercially successful, anaerobic digesters need to be operated as High Rate digesters. This require stirring and mixing and letting the anaerobic digestion occurring in the thermophilic range is this possible in landfills?

We all know that anaerobic digestion is a very sensitive and instable process and require close monitoring and control. Is this possible in landfills anaerobic digesters? Read more!

Sunday, February 17, 2008


One of the weaknesses in carrying out fermentation technology is due to poor understanding of the behaviour and characteristics of microbial populations especially in the fermentor. Understanding the role and function of the microorganisms is more than just knowing what type of microorganisms we are dealing with or knowing the physical structure and dimension of microbial cells. There is more to that, and this lack of knowledge about the behaviour and composition of microbial populations in the fermentors are often regarded as the " Achilles Heel" of the fermentation process.

Whether we want to admit it that although the unit of microorganisms is the single microscopic cell but their behaviour are often characterised in a "multicellular fashion" in terms of the microbial population. Even microbiologists expressed the activity of the unicellular organisms in terms of the microbial population. The growth curve of the microorganisms is often expressed as microbial population growth against time :))

The concept of microbial population is complex in terms of its structure and composition. The properties of the microbial population is always dynamic and changing with time. We cannot and must not regard the microbial particles as innate or static particles in the fermentor. Everything during the fermentation process is in a state of dynamic flux with microorganisms affecting the fermentor functions and vice versa

In the study of microbiology there are two states of growth.
1First the real growth of the single cell which increase in size and biomass before dividing into two new daughter cells and
2 Second the population growth where a number of cells will divide to form high number of cells.

In the fermentor we are more concerned with the growth behaviour of the population of microorganisms or the microbial inocula introduced into the fermentor

The growth of the initial microbial population (inocula) is affected by the following factors:
1 Supply of nutrients
2 Diffusion of waste products
3 Mass transfers between the inocula and the broth environment

All though within the same inocula the initial composition of microbial cells within the inocula show different composition temporally and spatially. This to a degree depend on the time period of the inocula and the conditions of the inocula. Even though it is deemed that the inocula is taken at log phase that does not mean all the cells in the inocula are uniform occurring as young active cells

At the onset of the fermentation period after the microorganisms have adapted to the environment, there will be higher proportion of new young cells compared to old and dying cells. However on reaching the stationary phase there will be a shift in the composition of the cells with higher percentage of the population consisting of old and dying cells

To maintain a high proportion of young cells or for it to be continually in the log phase for most of the population either fresh substrate need to be added or a wastage of cells need to be carried periodically


While it is true microorganisms are unicellular and capable of existing independently, but in a population of microbes in the fermentor these microbial cells do not occur in a uniform unicellular suspension. In reacting to the stress of the fermentor environment such as intense aeration and shearing forces, these microorganisms tend to occur in the form of microbial aggregates such as a microbial floc, pellets, mats, granules or as biofilms.

These microbial aggregates will not only complicate the rheology and mass transfer processes but also the fermentation process on the whole. New considerations must be taken in operating the fermentor under such conditions such as stirring and supply of oxygen Read more!

Friday, February 15, 2008


In carrying out any fermentation studies using any type of fermentor, it is of utmost importance that we carry out the studies properly so that there will be no unwanted errors and that the results obtained are proper and truthful. We do not want at the end of our fermentation studies ending up with wrong and erroneous data which in their true value are useless.

In the process of carrying out the fermentation we need to look closely at the following points

In carrying out fermentation studies we are dealing with fermentors which often come with various sensors and electrodes to monitor the fermentation process. The electrodes need to be examined closely in terms of its structure and functionality. It is important to calibrate the electrodes before every use in the fermentation studies

Most forget that every electrode have its life time expectancies especially in the fermentation studies where the conditions are harsh and demanding and the lifespan of the electrodes are short

It is ad to note in certain incidence fouled, broken and dried electrodes are used to monitor the fermentation process

Before the use of the fermentors close examinations must be carried out on the various components of the fermentors from tubings to air filters. Blank run using rich medium should be used to detect if the fermentor is fit for operation.

The autoclaves used to sterilize the fermentors should be validify in terms of their efficacy in sterilizing the fermentor. The inner chamber temperature of the autoclave should be calibrated to ensure that the temperature shown outside is the same as shown inside. This is similarly applied for the pressure check.

There is the need that the autoclave is not over loaded with materials to be sterilized, if not the sterilization cycle need to be adapted

It is bad to assume that the microbial cultures used for the fermentation to be pure. Microbiological checks are need to be carried out up stream before the culture is used. Failure to do this would be a disaster to the fermentation studies as amplification of contaminants could occur with time length and temperature

Proper aseptic techniques during inoculation and sampling should be effectively carried out and confirmed or validated. The sampling line is one of the most common portals for the introduction of contaminants into the fermentor.


The volume of fermentation broth should be calculated to take into account of possible loss through samplings and dehydration. It is important that the volume filled must not enter or use up the headspace to avoid contaminations

Wrong volume of broth used could affect the results of the fermentation studies. We must be efficient and economical in determining the right volume for the fermentor

In most of the procedures above we must not assume or live in the belief that everything is proper and assuming that the fermentors are effectively sterilized, or that the cultures are not contaminated or the ph probe is working well as specified. There is the need to validate that all the procedures are correct and the probes are in 100% tip top conditions.

Studies or validation procedures must be carried out to confirm all the above steps. Validation of equipments or methods constitute part of HAACP procedures and cGMP mechanisms. It is sad to say that these very important and fundamental objectives are often not carried out especially if the fermentation process involves engineers.

This condition would not be acceptable in stringent and demanding industries such as in pharmaceutical fermentation industries

I have expressed my concern to a number of fermentator users in the local colleges and universities and inquire of the status or procedures carried out in their respective plants or laboratories. Sad to say the response have been very very poor from no response to a one word answers. This attitude does reflect the sad attitude of not willing to learn or scared of being criticised Read more!


Mixing or the stirring of fermentation broth in a CSTR is an important operating parameter in the fermentation process. The process of mixing is primarily carried out by the action of the stirrers and impellers, however secondary mixings of the fermentation broth are achieved by other modes such as sparging of the broth.

There are many factors which affect the mixing properties of a particular fermentation process such as broth rheology, power, geometry of vessel and design and configuration of impellers.

Why do we need to mix the fermentation broth? The main answer is to achieve homogenization of the broth so that the broth will show uniform composition, physically, chemically and even microbiologically throughout the active volume of the fermentor. This will avoid the formation of physical and chemical gradients which may lead to toxic pockets being formed in the fermentor

Mixing is accepted as the method to increase the efficiency of mass transfer between the microorganisms and the environment. This is achieved by reducing the boundary layer surrounding the cell and enhancing diffusion process through a thinner boundary layer

Mixings are proven to aid solubility and dispersion of gases as well as nutrients in the fermentor. This is especially important where the gas or solids show limited solubility

But what do we really see in the mixing process? In reality in the process of mixings carried out in a fermentor we can see two types of phenomena:
1 Macromixings
2 Micromixings

Macromixings are more involved at the larger scale seen at the level of the impeller and directly involved in the homogenization of the broth and solubility and dispersion of nutrients. Micromixings occur more at the molecular and nano level at the intimate proximity of the microorganisms. This is important especially at the level of the boundary layer. Although the two phenomena occur simultaneously during the mixing process but the efficiency and the operating regimes differ. The choice of the right mixing systems and operating conditions can help shift the the effect of micro or macromixing

One of the most common mistakes in operating the fermentor is fixing the mixing speed to a fixed value such as 200 rpm or whatever values without properly understanding the demand and impact of such mixings. Each type of fermentation requirements for mixing should be determined experimentally as no two fermentations are the same. The amount of mixing should be a balance between the requirements of:
1 Providing oxygen
2 Need to keep cultures in suspension
3 Reducing the damages to the microorganisms by the shear forces generated by mixing

In simple statement, mixings should be 'tailor made' for the specific type of fermentation Read more!


The heart of the fermentation system is the fermentor, where the crucial biotransformations occur. It is within this vessel where substrates, microorganisms and the fermentation products intermixed. Everything, be it gases, liquids and solids enter and leave the fermentor vessel at one time or other. Thus we see under a steady state conditions solids, gases and liquids will be spending a period of time in the fermentor. There is always a dynamic flux of the various phases in the fermentor

This flux of solids, liquids and gases in the fermentor occur under full regulation or control and efficiently executed by the use of pipes, tubings, pumps and valves. We could picture this controlled flux by the four components as the ' circulatory system' of the fermentor

The circulatory system ensures:
1 Heating and cooling of the fermentor
2 Supply input of nutrients
3 Supply of inocula
4 Removal of samples
5 Removal of broth
6 Controlling input of micronutrients, acids, alkalis, antifoams
7 Removal of exhaust air and other gases

When we talk about the valves it must be discussed in terms of pipes and tubings and the pumps. The three components form the complete system of flow control for gases and fluids.
In such situations we can see that valves will control:

1 Volume of fluids going through the pipes
2 Rate of flow
3 Direction of flow for the fluids

In most cases when the valves are involved in controlling the flow of fluids through the fermentor system, the driving force of the fluid through the valves will be carried out by the pumps that will provide the pressure or drive to move the fluid. In pressure or safety valve, the pressure is attributed to the positive pressure created in the fermentor by the air pumped into the fermentor and through the exhaist gas outlet.

There are many types of valves to choose from. A right valve is needed for a particular function in the fermentor. The choice of valves used depends on factors such as:

1 Do you need a fine control of the flow?
2 Do you need a rapid large volume flow of fluid?
3 Do you need a one way flow and avoid backflow of gases and liquids?
4 Do you need to divert flows of fluids?
5 Is the valves to be used needed in aseptic conditions?
6 Will the valve be of sanitary quality requirements?
7 Will it be easily cleaned by CIP or SIP?
8 Is the valve for safety pressure release?

The characteristics of commonly used valves in the fermentation plant, industries or in the fermentor system are as shown.

1 On and off control Ball valve,Gate valve
2 Large pipes Butterfly valve
3 One way flow Check valve
4 Sanitary flow Diaphragm valve
5 Flow regulation Globe valve
6 Slow pressure release Needle valve

Try to look at your whole fermentor system and note where all the valves are. (Note: Some of the valves are "hidden" by not being in the fermentor itself!!!) Read more!

Wednesday, February 13, 2008


The use of anaerobic digestion process in biological wastewater treatment has often been hailed as the ultimate answer in the treatment of various types of wastewaters. There are many advantages cited in using this fermentation technology such as wastewater stability, sludge reduction, dewaterability, energy and fertilizers. Yet such attractive technology that is in the dreams of environmental engineers are often the 'nightmare' of wastewater operations,

The sad truth is the anaerobic digestion process is very sensitive and instable process and easily prone to failure..

The failure of control of the anaerobic digestion is often attributed to the failure of understanding the microbiology and biochemistry of the process

The anaerobic digestion is a very complex fermentation process because:

1 It is a mix culture microbial populations all involving in the process
2 It is made up of two basic groups of microorganisms; acidogens and methanogens with their own physiological attributes requiring their own specific growth requirements but still dependent on each other in the anaerobic digestion process
3 Methanogens are obligate anaerobic bacteria very fragile, slow growers and easily killed by exposure to oxygen and other extreme conditions
4 Complex substrate loading to the anaerobic digestors
5 Very high solid content and complex rheology of the substrate
6 Poor mixing conditions pf most anaerobic digestors
7 Washouts and slug dosings or shock loadings affecting the bacteria
8 Effect of temperature and ph upon the two groups of microorganisms

Since one of the hall,arks of anaerobic digestion process is its sensitivity and stability, the success of its operation depend on close monitoring of the anaerobic digestion process. The optimu operating characteristics of an optimally run anaerobic digestor are marked by:

1 High volume of gas production
2 High percentage of CH4 to CO2 ratio
3 Neutral pH
4 Very low concentration of TVFA of less than 200mg/litre
5 Presence of short chain fatty acids such as acetic and propionic acids with little or no trace of butyric and valeric acids

The optimal range occurs in a very narrow range thus close monitoring are required to detect any changes and to take detection before the impending disaster

It should be noted that the TVFA profile is not the cause of a digestor failure but symptoms of the inefficient and uncoupling of the anaerobic digestion process between the acidogens and methanogens Read more!

Tuesday, February 12, 2008


The fermentation broth is a very complex soup or solution. Fundamentally, the fermentation broth is the sea of nutrients in which the microorganisms grow, reproduce and 'swim' . The fermentation broth supply the microorganisms with all the nutrients the microorganisms need to grow and produce the various fermentation products.

The fermentation broth too act as the medium for various physical, biochemical and physical reactions to take place. The fermentation broth will be implicated in all the mass and heat transfers that occur within the fermentor, and it will be the medium that holds the fermentation products formed.

The nature and composition of the fermentation broth temporally and spatially will affect the efficiency of the fermentation process. The interactions between the fermentation broth and the various components is complex and affect both directions

At any time the composition of the fermentation broth is complex consisting of anything that ends up in the fermentation broth. This includes:

1 Raw substrates
2 Fermentation products
3 Microorganisms and its derivative components
4 Chemical additives added to the fermentor
5 Gases such as oxygen and other metabolic gases

All three main phases; solid, liquid and gases are present in the fermentation broth and their possible interactions

One of the most important singular properties of the fermentation broth will be its rheological or viscosity characteristics. Let us not stick to strictly to the definition and quantification of rheology but rather try to appreciate it from its behaviour and its impact.

We always 'picture' fermentation broth as a thick gooey sticky mixture that is thick and viscous compounded by rising bubbles of gas exploding at the broth surface. Maybe this picture is too dramatic but in a way it is true!

The viscous nature or the rheological properties will affect the mixing regimes of the fermentor.
Viscosity is not a simple but a complex phenomena that is always changing and responding to various parameters. Very rarely can we describe a fermentation broth as following a Newtonian behaviour. In most cases it is a complex combinations of various Non Newtonian behaviour.

This poor understanding of the fluid behaviour of the fermentation broth will affect the efficiency of mixing and liquid circulations resulting in poorly controlled or less economical fermentation process

The viscousness of the fermentation broth is caused by the interactions of the various components in the fermentation broth. The interactions may occur between the components of the broth and the water or it could result from the interactions between the components themselves. Such interactions in the viscosity of the broth could occur at the level of the ions and molecules which involve the various ionic forces or it could involve at the macrolevel such as between the various biopolymers tangling and sliding with each other. The overall result will be that the fermentation broth will be viscous.

Now let us look at one of the components which make the fermentation broth viscous, that is the contribution of sugars to the viscosity. Sugar or the carbohydrates are the main carbon source in any fermentation media and supply the carbon needed for energy and skeleton structures of the cells and organic compounds

Experience have shown to us that sugar in solution is sticky, but dry sugar is not sticky. The stickiness or viscousness of the sugar in solution is caused by hydrogen bondings which develop between the sugar molecules and water. During the interactions of sugar and water the hydrogens in the water molecules and the hydrogen in the sugar molecules have an attraction for each other. Thus it is the hydrogen bondings that make the sugar sticky!

Thus we see that most of the viscosity in the fermentation broth is caused by the various hydrogen and other ionic bondings

The most crucial effect of viscosity is that it makes the situation very difficult to achieve proper and complete mixings. This will affect the various mass transfer processes that occur in the fermentor. poor mixings due to high viscosity will also result in the formation of various physical and chemical gradients

Viscosity makes scaling up studies difficult due to the change in behaviour of the fermentation broth such as difficulty in mass heat transfers, solubility of components and gases and mixings at the upper scale of fermentation process Read more!

Monday, February 11, 2008


In my years of being involved in fermentation and fermentors, I have regularly noticed that in most cases:

1 Fermentors are underused
2 Fermentors are improperly used
3 Fermentors are rarely serviced until they are broken down or unworkable

It is sad to note that in many laboratories and fermentation plants I have visited a significant numbers of the fermentors are left standing in one corner and being treated more as white elephants and occupying spaces. Fermentors are capital equipments and represents high investments

In mosr cases these fermentors should not have a premature retirement. They are still good and workable. All it needs is a minor repair and replacements. These fermentors should be retrofitted or refurbished as it is capable of giving many many more years of faithful service.

Often the 'failure' of these fermentors are not the failure of the equipment but the failure or poor understanding of the person who bought and operate these fermentors. Some of these failures could be easily repaired.

It is very important in any fermentation laboratory or plants that they have a dedicated workshop which can support in doing repairs and renovations to existing fermentors. And having an engineering workshop or department such as mechanical and electronic departments can be a real asset

If we looked at an old or dying fermentor we can see that in most cases:
1 The fermentor vessel or stainless steel body is really in good conditions
2 Most of these old fermentors are the 'old generation' fermentors which are mostly analogue driven and not digital or electronically or computer controlled

Still these old fermentors can function and they still fulfill the requirements of a good fermentor as mentioned in earlier blogs here.

Retrofitting or refurbishing is possible at fraction the cost of new fermentors. Valves and sensors could be replaced. Stirrers and motors could be changed. In fact modern computer control loops could be incorporated.

The question is whether there is the earnest desire to refurbish the old fermentors. In my experience these old analogue fermentors are more reliable and trustworthy than the new modern highly electronic controlled fermentors which easily break down! These old fermentors are indeed workhorses of the fermentation industries Read more!


One of the most stringent requirements in the design of a fermentor is the ability of the fermentor to maintain strict aseptic integrity throughout the fermentation process. Of course this is applicable in monoseptic fermentation involving a single pure culture fermentation. For such requirements, theretically there should be complete isolation or barrier between the fermentor content and the surrounding environment. This would literalloy mean that the fermentor is completely closed by wall or structure to the environment with no openings or holes that expose the content of the fermentor to the environment.

In reality this is difficult to achieve as fermentors need holes or ports that allow important connections for the insertion of impellers, electrodes, inlet gas and outlet gas and even sampling ports. If this is so then the design of the fermentor must reach a compromise where while on one hand it maintains its aseptic integrity and on the other hand it allows port holes for the necessary components to enter or leave the fermentor but by not compromising its aseptic integrity

The solution to this dilemma is by:
1 Providing seals
2 Maintaining the seals continually aseptic

The seal is especially crucial at the stirrer or agitator entrance to the fermentor. The stirrer is the essential component in any fermentor as it is involves in the mixing and homogennization of the contents of the fermentor. Any seal that is used in the agitator system will have to fulfil various requirements such as:

1 It should be able to maintain the state of aseptic integrity of the fermentor while functioning
2 It should be able to withstand the stress of repeated sterilizations of the fermentor such as SIPs
3 It should be able to withstand the various CIP procedures
4 It should be able to sustain the pressure developed within the fermentor
5 If the fermentor is involved in food and pharmaceutical fermentations, it should be built of sanitary materials
6 The seal components will not contaminate the fermentation process
7 It should be able to isolate the contents inside the fermentor from the external environment

In any seal there is usually a need for:
1 Packing structure within the seal
2 Lubricant to smoothen the rotation of the shaft within the seal
3 Steam sterilization of the seal to ensure no contaminations occurring through the seal

There are other seals within the fermentor which is made up of the O - rings that help in the close air tight sealing between two surfaces. We have discussed this section in previous blogs Read more!


If you go through the internet, you will be seeing a lot of advertisements trying to sell their cultures which are supposed to the ultimate solution to solving almost anything from pollution to bioremediation and even to composting among others. The vendors claimed to have discovered or concocted mixtures of species of microorganisms that seems to do the trick.

The question is does it really do the work as promised by defying the laws of microbiology itself.

Let us look at a simple example whereby a mixture of microorganisms are effective in reducing pollution of a heavily polluted water body

Questions that should be asked are;

1 Have these microbial cultures been successfully adapted to using the wastes in the polluted wastewaters
2 How are these cultures adapted to degrading the wastewaters?
3 Are there proven laboratory or feasibility studies supporting this claim?
4 Are the findings reported in well known peer reviewed journals

Throughout my experience in microbial degradation, it has always been the principles of microbiology that it is the substrate that chooses the microorganisms and not vice versa.
Adaptation involves various stages of not only adapting to toxic conditions but also developing the necessary enzymes to utilize the complex substrate. Microbes are not stupid! Given the choice they will use the easily assimilated carbon in the wastewaters before going after the difficult carbon.

Wastewaters especially the polluted and complicated wastewaters are not only toxic but contain a variety of carbon which are difficult to degrade. I cannot see how by just pouring hundreds of buckets of these 'special microorganisms' into the moving wastewaters such as rivers and streams will solve the pollution problem. Up to now I have only heard claims upon claims of these magical cultures doing wonders...Where are the proof in properly carried out experiments published in eminent scientific journals. I don't think it is wise or safe to take the opinion from salesmen selling the cultures, laymen or farmers

Often when they carry out the 'experiments' they monitor it months and years after the event

One thing I do agree the media which they used for building up the cultures are mainly mplass based which are rich in sugars and vitamins which support the growth of many microorganisms and are good fertilizers. So is the so called success in the use of these microorganisms more because of the 'magical bacteria' or because of the rich nutrients in the molasses? These molasses solution are often viscous and acidic which resulted in lowering the ph of the water and precipitating the colloids and chemicals. But will it be lasting in the long run and on a bigger scale? We need more research and experiments properly carried out before validating these products. If not a lot of people will get tricked and lose a lot of their money over the magical property of these microbes Read more!

Tuesday, February 5, 2008


Most, if not all, biological wastewater treatment plants are either not working or are not functioning efficiently to their expectations. This sad state of affairs gets worst with time due to changes in the operating parameters of the WWTP or due to poor monitoring and servicing of the WWTP.

The main contributing factor to this poor state of WWTP probably arises out of the attitude of the companies regarding the WWTP as a liability in that it does not contribute to the profits of the company. It is not surprising therefore that they will not invest more money in operating and maintaining the WWTP efficiently. In the first place they build the WWTP more as an excuse to get the necessary operating license to carry out their manufacturing activities

The WWTP has always been situated at the back of their manufacturing premise and not in front of their minds. The full realization and impact of poor housekeeping in maintaining an operating WWTP only comes too late when Department Environment officers slap them with compounds and threatening imprisonments and forced transfer of their manufacturing facilities


The failure of WWTP could be attributed to many factors such as design oversights, poor operation, poor maintenance and servicing of the WWTP. These various failures will ultimately affect the performance of the microbes in the WWTP which will ultimately affect the performance of the WWTP. After all a biological WWTP is nothing more than a bioreactor that support the growth and function of high concentration of microorganisms


WWTP failures reflect both the state of the pathology of the WWTP and give us vital clues to the where and why the WWTP fails. We should try to study the nature of these failures scientifically so that it will aid us in our decision whether we can repair, replace or improved the WWTP process. Failing to do this we might end up in the future building or renovating the WWTP which will end up in similar predicament years later


There are generally failure two main types of failures:

1 Sudden failure

2 Gradual failure

Sudden failure of WWTP is a failure that is not anticipated and usually occurs immediately beyond our control. Examples of sudden failure are electrical outage, blower burnt out, toxic chemicals or sudden hydraulic loading due to heavy rain. The impact of sudden failure is often very serious which can throw out the WWTP out of its function

Gradual failure of a WWTP occurs slowly and show cumulative deterioration with time. It is usually reflective of slow breakdown of system. Example of gradual failure of WWTP is exemplified by gradual clogging of the porous filters used in supplying air to the WWTP. Such failures could have been averted as the signals of impending failure are there. Such failures could have been avoided if remedial actions such as repair taken before turning to disaster

Previous records if kept could have indicated clues of impending failure, when the performance of the WWTP is slowly spreading out of optimum band state of operations. We should be able to see the loss in treatment efficiency of the WWTP over time in terms of BOD, COD and TSS reduction


In reality most failures go undetected because there are no staffs that are trained to monitor the performance of the WWTP. By the time the WWTP completely broke down, it is already too late to take preventive actions

Even if monitoring of the WWTP is carried out, it usually involves ‘end of pipe’ analyses and not ‘up pipe’ or ‘along pipe’ analyses. In most cases the analyses just involved the basic regulatory requirement parameters such as BOD, COD and TSS. These parameters are insufficient to show the state of health of the WWTP and you do not really know which unit process is facing the problem?


There are other parameters besides BOD, COD and TSS to give us the clues on the exact state of health of the WWTP. Any changes in the operating parameters of the WWTP will be reflected by the gradual change in the composition and properties of the microorganisms of the WWTP. There will also be changes in the chemical; parameters in the mixed liquour due to the changes in the microbial consortium in the WWTP.

The failure for nitrification could be indicated by very poor DO and high NH3 content of the wastewaters. High biomass washouts could be attributed presence of pin flocs within the WWTP are some of the examples of how additional parameters monitoring help to determine WWTP failures


To confirm the observations there might be the need to carry out scale down studies to pin point the cause of the WWTP failure. Samples of the mixed liquour from the WWTP are removed and transferred to various smaller vessels to study the effect of mixing, aeration or oxygen uptake. From the studies we can conclude if enough mixing or aeration are the culprit of the WWTP failure.

Additional respirometric experiments may be carried out to determine if the microorganisms are viable or if the wastewater is biodegradable.


Wastewater treatment in a WWTP usually involves the flow of the wastewater through a series of unit processes which include physical and biological treatment

Analyses of various parameters as the wastewater enter and leave the various unit processes from the influent inlet to the treated effluent outlet will be able to tell us:

1 Which unit processes are most effective in reducing the pollution load?

2 Which unit processes are the rate limiting steps and controlling the bottleneck of the process?

3 Which unit process are the ones having problems?

This information would be useful to allow us to improve the efficiency of the unit process in mind or even to increase the optimum rate to the limit for existing functioning unit processes

Simple measurements can be conducted by using ph and DO probe with long cables, together with simple tracer studies to see the problems of flow and circulations within the WWTP. Any short circuiting or improper mixings of the WWTP will be easily noticed

The success in determining the cause of plant failure and inefficiency will allow rapid action to remedy the situation or even to optimize the rate limiting steps in the wastewater treatment process. This will allow the factory to save time and costs.

If the failure is not detected immediately and the WWTP is allowed to deteriorate until whole system backfire then it will be difficult to pinpoint source of error and to repair it early.

Read more!

Monday, February 4, 2008


The common thing shared by both chemical synthetic industries and the microbial fermentation industries is that both are involved in the synthesis and production of chemical compounds. That is where the similarity ends.

Chemical industries produced both organic and inorganic chemicals in more varieties by carrying out the chemical reactions which utilize more energy and pressure compared to the fermentation industries. Microbial fermentation industries carry out the synthetic reactions under normal ambient temperature and pressure using microorganisms and enzymes to catalyze the changes. Their biological reactions do not require high energy or extreme conditions.

Fermentation products formed by the microorganisms are however more labile and occur in lower concentrations or in very dilute solutions. These characteristics have created problems not only in the production of the chemicals but also in the downstream isolation, concentration and purification of the fermentation products compared to the chemical industries.

It is interesting to note that most of the chemical downstream processing units for product recovery are also used in the fermentation industry downstream processing of chemical oroducts. However, in the case of downstream processing of fermentation products additional downstream processing units are added in the purification and concentration of fermentation products. Read more!

Saturday, February 2, 2008


In the past few decades there have been great interests in the cultivation of mammalian cells, including human cells in fermentors for the production of biologics for diagnostic and therapeutics used in human health care.
This is not a new direction in the progression of fermentation technology but a natural anticipated progress in the field of fermentation technology.

Historically speaking the field of fermentation technology has been based on the cultivation of microorganisms such as bacteria to carry out the transformation and production of fermentation products of great economic and industrial importance. It seems then that the power of the microbes are limitless in producing any kind of metabolites. Great strides in the development in microbial fermentation technology were made with parallel development in the study of microbiology, physiology, biochemistry and genetics with similar development in the design and construction of fermentors by bioprocess engineers.

The culmination of this progress is in a nutshell the fourth definition of fermentation by the engineers which defined fermentation being the study of the cultivation of high concentration of microorganisms in bioreactors microorganisms. The fermentor is seen as a special vessel which provides all the optimum conditions necessary to support the growth of high concentration of microorganisms and allow the transformations to occur.

With the development in plant biotechnology especially in the generation of tissue cultures, it then becomes a natural extension or progression to culture single plant cells in fermentors as there is really not much difference between microorganisms and plant cells when they occur as single plant cells.

Much knowledge, experience and breakthroughs in microbial cultivation in fermentors could be similarly applied to single plant cells. Well, at least superficially they seems similar, but in reality there are many problems faced in the cultivation of single plant and mammalian cells compared with the cultivation of microbial cells


As we have said earlier the common factor shared between microorganisms and single plant and mammalian cells is that all the cells for the cultivation in the fermentors are SINGLE cells. Beyond it are more differences than similarities.

We can see the main differences between the cultivation of microbial cells and mammalian cells in the following:

1 Larger size
2 Longer time to grow
3 Fragility of cells
4 Lower oxygen consumption
5 Tendency to form multicellular aggregates
5 Complex medium requirements
6 The cells might end up not producing the desired products efficiently

The above factors are crucial when one consider cultivating the mammalian cells in fermentor.

The fragility of mammalian cells means that these cells are easily broken up by the various shear forces generated in the fermention broth such as effect of mixing by impellers, liquid shears and bubbles cavitation

Due to the long growth time means that it takes very long period for the mammalian cells to reach optimal concentration in the fermentors. This situation would make the fermentation process more susceptible to microbial contaminations. There is a stringent requirement to prepare clean room standards for mammalian cell cultivation.

The cultivation of mammalian cells require complex media which is more costly adding to the costs of the fermentation

In mammalian cell cultivation the issue is not so much about using cell substrate for the highest yield or productivity but more of:
1 instability protein expressions which might deteriorate with time
2 protein produced require extensive post translational modifications especially glycosylation of protein such as in production of monoclonal antibodies

Bacterial systems are poor choice in the production of the biologics because bacteria cannot carry out complex post translational modifications of proteins.They can only produce simple proteins such as insulins.

These proteins formed too are not easily secreted by the bacterial system and tend to accumulate inside the bacterial cells as crystalline deposits. Their release from the cells would enticed complicated downstream processing which are costly and there is also the danger of endotoxins being produced if gram negative bacteria are involved

The released proteins might require resolubilization and refolding to bring it to its effective state. This does not consider percentage losses of products during downstream activities

In view of the above considerations, any fermentor to be used for mammalian cell cultivation must be:

1 Exhibit minimum shearing forces either liquid to liquid shearing, liquid to impeller shearing or
bubbles shearing
2 Provide durable aseptic integrity throughout the fermentation duration

As we already know the CSTR is the most widely used fermentor in industrial fermentations and its engineering principles and reliability well established. Modifications can be made to the CSTR to make it able to cultivate mammalian cells

The fermentation vessel should be rounded at the bottom and equipped with baffles to prevent vortex which could damage the cells.

Strong air sparging is avoided and laminar hydrodynamic flow is encouraged. Micro bubbles are generated in a laminar flow mixing for oxygenation. Since the mammalian cells require low amount of oxygen supply of oxygen minimum aeration to maintain DOT values of about 20% saturation

Strong aseptic adaptations might be included with increasing the number of seals to prevent contamination. The sampling port should be equipped with direct steam sterilization to prevent contaminations during samplings

to be continued.... Read more!

Friday, February 1, 2008


This may be considered the most easy, convenient way to start a fermentation by obtaining the cultures directly from culture collection centres. Superficially it seems that you save time and money by not trying to search and screen microbial isolates from various habitats to provide microorganisms for the fermentation process. But is it so?

While it is true certain groups of microorganisms can produce antibiotics, and obtaining these microorganisms should be economical and we are still assured of its antibiotic producing capabilities. It must be noted however that these microorganisms in the Culture Collections are just kept for reference and taxonomic purposes and studies. They are not industrial strains or high yielder. Logically if the strain is a super producer it would be kept under tight security under lock and key:))

The microorganisms maintained at the Culture Collections are products of isolation and identification years ago which are perpetually maintained by periodic sub culturings. Maybe these microorganisms have "lost" some of its important genetic traits?

It is better to search for new strains from exotic habitats in the hope of finding a super producing strain or a new metabolite Read more!


One of the most important aspects before and while running a fermentation study using a fermentor is to monitor whether that fermentor is effectively sterilized and/ or that during the whole fermentation run that the fermentor maintain its aseptic integrity. It is indeed very sad in my personal experience meeting with various fermentation technologists especially those trained from an engineering background that the question of sterility and contamination have never been seriously considered. What is important to them that the fermentor is 'functioning mechanically' and is yielding growth of microorganisms or producing tons and tons of data. Did they ever think for all the efforts, time, energy and costs that their data could be wrong or useless? In fermentation studies, wrong experiments can still yield data....wrong and error filled data!!?? And to make things worst they send their students on a wild goose chase analyzing these wrong data and reaching definitive conclusions? This must be the case of abuse of statistics and computer! After all if its rubbish data coming in you will get rubbish data or conclusions coming out

Most fermentation technologists take lightly to the concept of microbial contamination. This is not so in the stringent pharmaceutical fermentation industries where cGMP are always applied and validation of sterilizations and aseptic conditions are sustained as routine procedures. Any contamination would be a very heavy consequence in such industries.

Most fermentation technologists in universities, colleges and research institutes I met seemed to be in the state of denial thinking that:

1 Their fermentors are 100% sterile or can maintain aseptic integrity
2 They dont bother doing blank sterility test runs periodically to check if their fermentors are up to the sterilty validation
3 They do not periodically remove samples during fermentation runs to check for microbial contaminants

It is true that those fermentors before they are delivered are tested for sterility check for about 5 days equivalent of fermentation run. But these fermentors must be continually checked by conducting blank runs using rich broth and operated without any organism inoculated

It is impirtant to know if contamination occurs to determine whether it is due to:
1 Fermentor structural weakness
2 Operator failure
3 Feed or what ever Read more!