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Due to increasing developments in the biotechnological processing and bioreactor industry, there has also been an increase in the number of bioreactors currently available (See Fig. 1) (Oksman-Caldentey et al., 2002). The main function of a properly designed bioreactor is to provide a controlled environmental condition at which microorganisms can be able to perform the required reactions and transformations in the most optimum efficiency possible and achieve optimum growth and/or product formation (Chisti, 1989; Najafpour, 2007).
Figure 1: Bioreactor classification (Oksman-Caldentey, et al., 2002)
For aerobic bioprocesses the oxygen mass transfer is often the critical limiting factor in providing the optimal environment due to the low solubility of oxygen in the medium (Chisti, 1989, Garcia-Ochoa and Gomez, 2009). Agitation is widely used to provide for higher gas-liquid interfacial area due to the higher shear field and also better liquid-liquid/liquid-solid mass transfer. Under circumstances of agitation, sufficient transport rates need to be attained. Nevertheless, in general bioreactor design, other operational requirements such as sterile operation, power consumption and economics are considered (Chisti, 1989).
For aerobic bioprocesses the Stirred tank reactors (STR) has been the most commonly used type of biochemical reactor (Chisti, 1989, Garcia-Ochoa and Gomez, 2009). The STR has successfully been applied to processes such as aerobic fermentation and wastewater treatment, among others (Garcia-Ochoa and Gomez, 2009). Nevertheless, with STR still the most regular industrial bioreactor used for aerobic processes, supplementary reactors such as pneumatically driven reactors (e.g. the airlift reactors (ALR)) have been developed to counteract the limitations of the stirred tank and other mechanically agitated reactors (Chisti, 1989).
STIRRED TANK REACTOR
Stirred tanks, or conventional fermenters, have been widely used for culturing suspension cells since the 1960s. Mixing and bubble dispersion in a STR is achieved by mechanical agitation (Doran, 2009). Various different impeller shapes and sizes are used to produce different flow patterns within the vessel (Doran, 2009). The impeller is often mounted at the top of the bioreactor to drive the liquid downward in large-scale cell culture bioreactors. Baffles are used in the STR to reduce vortexing (Doran, 2009).
The aspect ratio of STRs (i.e. ratio of height to diameter) can be varied. The most economical STR shape would have an aspect ratio of 1 as it has the smallest surface to volume ratio (Doran, 2009). When aeration is required, the aspect ratio is increased, and thus provides for longer contact times between the gas and liquid as a greater hydrostatic pressure is produced at the bottom of the vessel (Doran, 2009). Temperature and heat transfer is controlled via internal cooling coils.
In these systems, a high number of variables influence the mass transfer and mixing, but the most important among them are stirrer speed, type and number of stirrers and gas flow rate used (Garcia-Ochoa, 2009).
Figure 2: Typical Stirred Tank Reactor for Aerobic Culture (Doran, 2009)
Airlift loop reactors have emerged as one of the most promising devices in chemical, biochemical and environmental engineering operations (Tongwang, 2006).
Airlift reactors are pneumatically driven reactors in which the energy input required for mixing is through a steam of gas or air, without any mechanical agitation (Oksman-Caldentey, et al., 2002; Najafpour, 2007). The air-driven bioreactor is equipped with a sparger at the bottom of the vessel (Vogel, 1997). It is widely used to culture fragile cells such as hybridoma cells, plant cell, tissue, and organ cultures (Flickinger, et al., 1999; Vogel, 1997).
The ALR consist of two tubes (Flickinger, et al., 1999). Air is introduced in the bottom of one tube through a sparger ring up to the part of the vessel cross section called the riser (Flickinger, et al., 1999; Najafpour, 2007). The gas and liquid travels upward as a two-phase co-current flow (Siegel et al., 1992). The riser is where most of the mass transfer takes place and has the higher gas holdup (Siegel et al., 1992). As the air bubbles rise to the top of the riser, gas is held up, fluid density decreases causing liquid in the riser to move upwards and the bubble-free liquid to circulate through the downcomer (Najafpour, 2007; Flickinger, et al., 1999). The resulting difference in the density of the liquid in the riser relative to the annular space within the bioreactor (the downcomer) causes variations in the mixing and dynamics of the fluid, which are greatly influenced by the viscosity, density difference, gas flow and gas bubble size (Oksman-Caldentey, et al., 2002). The density difference between riser and down-comer enables the liquid medium and cells to circulate with high turbulence (Najafpour, 2007; Oksman-Caldentey, et al., 2002).
The use of draught tube divides the flow in a riser and downcomer. (Oksman-Caldentey, et al., 2002). The flow passes up through the draught tube to the headspace of the bioreactor, where the excess air and the by-product disengage (Najafpour, 2007). The extent of gas disengagement will profoundly influence the gas holdup, liquid circulation velocity and mass transfer of ALRs (Siegel et al., 1992). Cooling can be provided by either making the draught tube an internal heat exchanger or with a heat exchanger in an external recirculation loop (Najafpour, 2007).
The gas is usually injected by static (diffuser stones, nozzles, perforated plates) or dynamic gas distributors (slot nozzles, venturi tubes, injectors or ejectors) (Oksman-Caldentey, et al., 2002).
There are three common airlift configurations, which are the internal loop airlift reactor, the external loop airlift reactor and the draught tube airlift reactor (Oksman-Caldentey, et al., 2002).
Figure 3: Pneumatically agitated reactors: (a) bubble column, (b) airlift reactor (internal loop), (c) airlift reactor (external loop), and (d) airlift reactor (draught tube). A, air inlet; G, gas exhaust. (Oksman-Caldentey, et al., 2002)
Figure 4: Gas and liquid flow pattern Figure 5: Airlift bioreactor with external with internal loop cycle recirculation pump
AIRLIFT BIOREACTORS VERSUS CONVENTIONAL STIRRED TANK REACTORS
Gas-Liquid Mass Transfer
Merchuk, 1990, states that the volumetric mass transfer coefficient, KLa, is 'the rate of gas transfer across the gas-liquid interface, per unit volume of the suspension and per unit of driving force.'
The volumetric mass transfer coefficient for external loop airlift Fermenter is estimated as (Najafpour, 2007):
KL = Mass Transfer coefficient
a = Gas-liquid interfacial area per unit volume of the liquid
ug = Gas velocity
The height of liquid in airlift reactors, hL, have an effect on KLa values as hL strongly influences the circulation velocity of the liquid and thus the overall gas holdup (Martens et al., 1993). The height of liquid can be as high as 60m in a large vessel causing the resulting pressure to increase the oxygen solubility and hence increasing the KLa value (Najafpour, 2007). The height of airlift reactors is typically about 10 times the diameter of the column (H = 10D) (Najafpour, 2007).
Due to the height and presence of a loop in the ALR, high efficiency mass transfer was shown as well as improvements in both the flow and mixing properties of the vessel (Vogel, 1997).
According to Malfait, 1981, an improved productivity of the ALR as compared to a STR was seen and this is mainly due to the higher overall mass transfer coefficient available in an ALR.
Of all nutrients supplied for aerobic fermentation, oxygen is the lowest solubility and often is the limiting nutrient (Chisti, 1989). Sufficient oxygen should be supplied to match the very rapid oxygen consumption by microorganisms in order to maintain the desired productivity (Chisti, 1989).
There is high gas-liquid contact area in ALRs as well as easy removal or replenishment of particles, and high heat and mass transfer rates (Tongwang, 2006). In an ALR, there is superior oxygen transfer performance (See Fig. 6) and provides great productivities with yeast, bacteria and filamentous fungi as stated by Oldshue, 1983, and Martens et al., 1993. Especially in low viscosity media, ALRs are capable of delivering oxygen at high transfer rates (Merchuk, 1990).
ALR aeration efficiency and performance are rather insensitive to changes in operating conditions, making ALR suitable for processes with changing oxygen requirements (Merchuk, 1990). However, a recent paper by Najafpour, 2007, claims that in an ALR however, as the microorganisms circulate through, the conditions change, and it is impossible to maintain consistent levels of carbon source, nutrients and oxygen through- out the vessel.
In an STR, better oxygen transfer is seen with the use of stirrers and baffles. It is possible to modify the rate of stirring to cater to greater flexibility in dealing with rheological changes within the medium that can influence oxygen transfer and mixing (Fontana et al., 1999).
Figure 6: Performance of ALRs and stirred tanks versus oxygen transfer rate. a, rectangular ALR; b, split-cylinder ALR; c, agitated tank; d, bubble column. (Orazem and Erickson, 1979)
Reactors must be able to have effective agitation and mixing capabilities so as to obtain a homogenous and uniform distribution of nutrients and transferred oxygen into the bulk medium. This is to prevent the presence of dead zones and possible anoxia (Chisti, 1989; Najafpour, 2007). The hydrodynamics within the reactor may also affect performance parameters such as mass and heat transfer as well as intensity of turbulence in the reactor (Chisti, 1989).
Properties of the fluids may complicate the mixing ability and oxygen transfer (Oolman and Blanch, 1986; Schugert, 1981). Some bacterial and yeast fermentation give rise to a highly viscous and non-Newtonian system (Chisti, 1989). With these non-newtonian media, sufficient mixing and mass transfer cannot be provided by ALRs (Doran, 2009). STRs would be more suitable for such viscous fluids, although, mass-transfer rates decline very rapidly with viscosities higher than 50-100 cP (Doran 2009). Multiple impellers can be installed in tall fermenters to improve mixing (Doran, 2009).
The degree of shear that may be tolerated to achieve sufficient oxygen mass transfer is an additional constraint on the mixing and mass transfer in biological systems. Excessive rate can cause damage to microorganisms and disrupt the cell wall, whereas, low shear can cause unwanted flocculation of cells, or even bacterial growth on the stirrer and reactor walls (Najafpour, 2007; Markl et al., 1987)
In STRs, there may be damage to the microorganisms passing through the high shear zone of the impeller (Vogel, 1997). Possible damage to fragile cells from the resulting shear forces and turbulence at the end of the stirrer blades can avoided by using an ALR (Flickinger et al., 1999). The possible shear damage caused to cells in STR may be a factor to its poor performance (Markl et al., 1987).
ALRs are known to have drastically lower levels of shear as compared to STRs (Oksman-Caldentey, et al., 2002). Due to its low shear, there is also low mixing; hence, the ALR can be used for growing fragile tissue, plant and animal cells (Najafpour, 2007). The flow regime in an ALR depends on the sparger used and the flow rate (Vogel, 1997). Various types of spargers can be used to provide different bubble size (Vogel, 1997).
The level of shear also affects the microbial morphology in other ways (e.g. whether a mold will grow as a mycelial mass or form pellets) (Chisti, 1989). This pelletized growth can further restrict oxygen transfer (Chisti, 1989).
The permissible levels of shear are predicted to be more stringent in the future due to increasing biotechnological advances with future uses on plant/animal cultures and genetically modified microorganisms which are less robust (Humphrey, 1984; Schuler et al., 1984)
The presence of foam causes reduced efficiency in gas-liquid separation (Najafpour, 2007). Variations in biomass concentration, viscosity, surface tension and ionic concentration can cause operational problems such as foaming, floatation and bubble coalescence (Oksman-Caldentey, et al., 2002).
Bubble coalescence often occurs in ALRs and should be minimized. If the feed enters the reactor at only one location, the microorganism will produce undesirable by-products due to it experiencing continuous cycles of high growth and starvation. This would result in low yields and high death rates and thus, multiple feed entries are used especially on a large scale (Najafpour, 2007).
In an STR, adequate headspace is provided by having the volume filled 70-80% with liquid so as to accommodate the presence of any foam, which may develop (Doran, 2009). To combat foaming further, a supplementary impeller called a foam breaker may be installed (Doran, 2009). Mechanical foam dispersal is generally preferred as chemical antifoam agents causes reduced rates of oxygen transfer (Doran, 2009).
Constant temperature and heat transfer is required to obtain the desired optimal microbial growth (Najafpour, 2007). More heat is generated from mechanical agitation than from sparging of compressed gas (Doran, 2009). This heat energy in the STR has to be removed to keep the temperature constant within the STR (Chisti, 1989) making temperature control for an STR more difficult.
When the heat released is high, such as the production of single cell proteins from methanol, it may be difficult to remove such high exothermic heat of reaction with a conventional stirred-tank design. Hence, ALRs may be the preferred choice (Najafpour, 2007; Doran, 2009)
A potential disadvantage of airlift technology is that it may not be possible to achieve cell growth as high as obtained in stirred reactors regularly (Oksman-Caldentey, et al., 2002).
A study done by Fontana, 2009, showed that the airlift had lower cellular growth as compared to an STR (See Fig below). However, due to the close results, this suggests that the airlift has potential for use in the production of the enzymes in large scale, with lower operating costs.
Figure 8: Time course of Aspergillus oryzae cultivation in WBEG medium in the airlift bioreactor. (A) Cell, reducing sugar, and dissolved oxygen concentration. (B) Endo- polygalacturonase (endo-PG) and exo-polygalacturonase (exo-PG) activities, and pH (Fontana, 2009)
Figure 7: Time course of Aspergillus oryzae cultivation in WBEG medium in the stirred tank reactor. (A) Cell, reducing sugar, and dissolved oxygen concentration. (B) Endo- polygalacturonase (endo-PG) and exo-polygalacturonase (exo-PG) activities, and pH (Fontana, 2009)
ALRs provide lower power economics as mechanical agitation would require a high energy input per unit volume so as to maintain the homogeneity of the system and aid oxygen transfer (Hess et al., 2002; Wu et al., 2002; Doran, 2009). Mechanically agitated reactors are unfeasible at volumes greater than about 500m3 as the power required to achieve adequate mixing becomes extremely high (Doran, 2009).
Not only ALRs are cheaper in terms of operating costs but also with capital costs. The reduced capital cost is mainly due to their simple mechanical design without the need for an agitator, gearbox, and structural steel (Chisti, 1989; Vogel, 1997).
STRs require complex mechanical seals to prevent contamination by unwanted microbial species (Chisti, 1989). Due to this complexity, STRs are thus more expensive and less robust than other bioreactors (Vogel, 1997).
According to Najafpour, 2007, however, though the ALRs may be cheaper due the fact that an agitator is not needed, but a larger air throughput is required and the air also has to be at a higher pressure, especially on large-scale. Vogel, 1997, suggests the use of air compressors to reduce power costs. These air compressors can be driven by steam and allows continued operation during power outages in large plants with minimal power generation control.
Further improvements can be made to get better energy economy by using a second sparger in the top part of the downcomer (Merchuk, 1990). By increasing the liquid velocity, then the free rising velocity of the bubbles is generated, causing the gas to be carried down, hence a longer gas-liquid contact time (Merchuk, 1990). This will then lessen the energy requirements as some of the gas is injected against a much lower hydrostatic pressure (Merchuk, 1990).
Sterility is difficult to maintain for long periods in an STR especially in continuous operations and where the impeller shaft must be inevitably pass into the vessel (Najafpour, 2007)
In an ALR, sterility is easily maintained, as there is less danger of contamination through seals of mechanical agitators (Siegel et al., 1992; Najafpour, 2007). Moreover, there is also a lower probability of mechanical failure in an ALR (Flickinger, et al., 1999). No maintenance of motors, gearboxes, seals and bearings are required in ALRs (Vogel, 1997). Due to the vertical orientation and lack of internals in ALRs, easier cleaning and sterilization can be maintained (Siegel et al., 1992).
Construction and Design Flexibility
ALRs are have very simple construction design allowing the possibility of constructing larger fermenters as the design is not limited by motor size, weight and shaft length as seen with STRs (Vogel, 1997; Siegel et al., 1992)
The ALR lack the flexibility as compared to STR as their design requires being more exact. Factors such as the geometry of the system has to be carefully defined as it can affect the circulation rates, the mixing time, and the volumetric mass transfer coefficient (Gavrilescu and Tudose, 1998).
According to Siegel, 1992, there has been very few data on ALR transport phenomena during the actual fermentation. A majority of the studies carried out on ALRs are on the growth kinetics during the fermentation process instead. These studies have been conducted on small bench-scale ALRs due to cost restrictions and complexities involved in large-scale studies. The bench-scale studies do not provide the basic hydrodynamic and mass transfer information essential for successful ALR scale-up.
Due to this lack of data, biochemical industries hesitate in using ALRs for commercial application. The STRs are still widely favored due to its 'off-the-shelf' convenience and well-defined performance and scale-up properties. Reports of improved performance by ALRs on the bench-scale is not enough to convince the biochemical industry in extensively using the ALRs due to doubts of its performance after scale-up (Siegel et al., 1992).
The limitations of STRs have led to the development of other bioreactors such as pneumatically driven bioreactors such as ALRs. The ALRs are seen to have the greatest potential with the possibility of providing the aeration and agitation with a low energy input. Particular interest is being focused on the use of ALR with tissue cultures due to its shear sensitivity, mildness and uniformity of turbulence.
The ALR has excellent contact among phases, nutrients are easily replenished and has high heat and mass transfer rates. There is easier temperature control in ALRs as compared to STRs.
While the ALR seems to be the better choice, biochemical industries still hesitate in using ALRs due to the lack of information and low maturity of the bioreactor.