batch fed batch and continuous fermentation pdf

Batch Fed Batch And Continuous Fermentation Pdf

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An anaerobic fermentation process was developed for production of natural propionic, acetic and succinic acids from l -lactic acid using Selenomonas ruminantium. The l -lactic acid was quickly converted to a racemic mixture and there was no enantiomeric preference for further metabolism. Nutrients in corn steep liquor and yeast extract were necessary for optimal production of propionic acid.

An anaerobic fermentation process was developed for production of natural propionic, acetic and succinic acids from l -lactic acid using Selenomonas ruminantium. The l -lactic acid was quickly converted to a racemic mixture and there was no enantiomeric preference for further metabolism. Nutrients in corn steep liquor and yeast extract were necessary for optimal production of propionic acid.

batch fermentation pdf

In fact, human monoclonal antibodies hMAbs produced by CHO cells have played a major role in both the diagnostic and therapeutic markets for decades.

Since that approval in , scores of chimeric, humanized, and human MAbs have gained approval and entered clinical use. In development of all pharmaceutical production processes, including those involving hMAbs produced by CHO cells, decisions regarding the best process parameters and methods are made based on cost, time, and titer comparisons. Often, multiple scalable platforms are examined before a final process is transferred to pilot- or scale-up laboratories 2 — 8.

The objective of our research project was to compare the performance of batch, fed-batch, and perfusion processes — the three primary methods for hMAb production — using a single, laboratory-scale bioprocess controller. In the batch method, all nutrients are supplied in an initial base medium. The fed-batch method adds nutrients once they are depleted. The perfusion method circulates medium through a growing culture, allowing simultaneous removal of waste, supply of nutrients, and harvesting of product.

We compared two perfusion processes that differed in the way that cells are retained in a bioreactor. In one setup, we used an alternating tangential-flow ATF filtration device, which uses hollow-fiber filters to retain cells. In the second setup, we cultivated cells in a packed-bed bioreactor in which they were immobilized on a solid support matrix.

Table 1: Parameters common to the tested methods for production of a human monoclonal antibody hMAb. Materials and Methods We compared four methods for hMAb production, controlling all experimental runs with an Eppendorf BioFlo bioprocess control station. Table 1 overviews the four experimental setups. We inoculated cells from a cryopreserved stock vial at a density of 0. Using this method, we inoculated each bioreactor with cells that were at about the same passage and duration of culture after thawing.

Vessels: For the batch experiment, we used a 2-L Eppendorf water-jacketed, autoclavable glass vessel. It is easily programmable to give precise control of temperature, agitation, gas supply for sparging and overlay, pumping for addition and removal of liquids, and exhaust with heating or condensation.

And it can use sensors for pH, dissolved oxygen DO , reduction—oxidation, and CO 2 for measurement and control. Cultures were agitated at rpm. The glass vessel and BioBLU 5c single-use vessels were equipped with a pitched-blade impeller. For the packed-bed bioreactor, we used a basket impeller. We measured pH using a potentiometric sensor autoclavable vessel or optical pH probe single-use vessels and controlled it at 7.

All cultures were inoculated at a final density ranging from 0. In the batch run, we held that temperature constant. Table 2 summarizes important process parameters. Inoculation density, gassing control, DO control, and agitation were the same for all four experiments. To collect the highest-quality sample from growing cultures, we connected a sterile 5-mL syringe to the sample port Luer lock, then removed and discarded a dead volume of 5 mL.

Then we collected a second 5-mL sample in a new syringe to provide a fresh, viable sample for analytics and fresh sample storage. Later we accounted for that daily volume loss when calculating the fed-batch feed amounts.

Direct cell counting was not possible for cultures grown in the packed-bed bioreactor because cells were growing in or on Fibra-Cel disks. However, it is possible to correlate cell growth with glucose consumption trends. We analyzed the cell density and glucose consumption trend in the ATF perfusion run and used the results to estimate cell density in the BioBLU 5p bioreactor.

Triphasic glucose consumption represents lag phase, log phase, and a plateau for stationary phase. We used the trendline equations from each phase to calculate cell density. Photo 2: BioCommand block-logic automated feed program turns on the pump at a set point of 2. This allows a slow and steady feed each day so as not to shock growing cultures with rapid changes in osmolality or temperature. Set points such as pump speed which control volume delivered and duration of feed can be customized as the volume of a culture increases over time.

With a pump set point of 2. Photo 2 shows this automated program. Feed Program Line 2 uses the Counter loop to turn on Pump 1 at a set point of 2. The process ended when cell viability and density of the culture began to decline. To prevent filter clogging, we increased the ATF flow rate to 1. During cultivation, cell density reached a point at which the demand for DO was too high for the 1-SLPM maximum flow sparge drawer. So we used a 5-SLPM maximum flow overlay drawer using the same three-gas automatic gassing control algorithm.

Thus, a maximum flow of 5 SLPM could be achieved. For both perfusion experiments, we determined perfusion rates as follows: Based on culture health as indicated by growth rate and metabolite concentration, we steadily increased the perfusion rate in vessel volumes per day VVD.

Table 3 lists the adjustments we made over time. We decreased glutamine feeding concentration to reduce ammonia production. Results For each of the four cultivation conditions, we compared cell growth and viability, metabolic profile, and the total amount of product expressed. Figure 1: Batch culture results; left cell density and percent viability, right metabolic profile. Batch Culture: The batch process reached a peak viable cell density of 1.

On day 5, the cells had consumed all of their initially supplied glucose. Lactate concentration increased up to 2. Time-shifted to the decrease of glucose concentration, it then decreased — which suggests that those cells underwent a metabolic switch from glucose to lactate consumption. Within 9 days, the culture had produced a total amount of 0.

Figure 2: Fed-batch culture results; left cell density and percent viability, right metabolic profile. Feeding the culture also delayed the onset of lactate consumption, which began around day 5 and was not complete until day Figure 3: ATF perfusion results; left cell density and percent viability, right metabolic profile.

This phenomenon is attributed to metabolic changes that occur in response to high lactate concentrations, as has been documented 9. Within 16 days, the culture had produced a total amount of 4. Figure 4: Packed-bed perfusion results; left cell density calculated from glucose consumption, right metabolic profile.

Perfusion Cultures: We compared two perfusion processes that used different methods for cell retention. The former process reached a peak cell density of 7. We increased the perfusion rates over the course of these processes to a maximum of 1.

Table 4: Comparing peak cell densities and antibody titers obtained in the different processes. In the ATF process, glucose concentration varied. That can be attributed to the fact that the consumption rate was so much higher than in the fed-batch process that it was challenging to keep up with cellular demands. In the packed-bed perfusion, we expected a high consumption rate and therefore managed to keep glucose levels more stable. Ammonia levels increased and decreased based on our perfusion rate changes Figures 3 and 4.

Table 4 summarizes the peak cell densities and antibody titers obtained in all these different processes. Discussion The BioFlo system allows for seamless transition between bioprocess protocols, which enabled us to compare the performance of batch, fed-batch, and perfusion processes using a single bioprocess control station.

This was critical to our study because the ease of transitions among methods was part of our evaluation. With the ATF perfusion process, we achieved a noteworthy peak cell density of 7.

That represents fold and 2. The choice of which culture mode to use for production depends on budget, time available, and the quantity of hMAb required. Fed-batch culture provided a fivefold increase in hMAb yield, but over a substantially longer period, requiring much more human intervention and equipment that is more complex, although medium use remained low.

However, it required additional capital investment and laboratory space for the ATF filtration unit and controller, with operation of complex equipment, taking multiple daily samples, and monitoring of vessel volumes.

It also used a much larger quantity of cell culture medium. Setup of the packed-bed vessel involved simply connecting it to the control station for standard monitoring of vessel volume, and sampling was uncomplicated. Moreover, the method produced peak cell densities close to that of the ATF process but without additional capital equipment or complex operations.

Application of single-use equipment is on the rise in bioprocess development because it reduces turn-around times and cross-contamination risks. We used BioBLU single-use vessels successfully for our fed-batch process as well as both perfusion culture modes. Adding an ATF filtration device and using a solid growth-support matrix in a packed-bed vessel both allow for simple cell retention with higher cell and product yields.

Much greater productivity can be achieved through prolonged perfusion culture with up to two or three months of continuous antibody harvesting after a single inoculation. References 1 Jayapal KP, et al. BioProcess Int. BMC Proc. Stacey S. Willard is a senior bioprocess technical applications specialist, Amanda Suttle is an associate scientist, Xiaofeng Kevin Han is a research scientist, Michelet Dorceus is a bioprocess applications specialist, and corresponding author Ma Sha is director of technical applications at Eppendorf, Inc.

Fed-batch culture

Skip to Main Content. A not-for-profit organization, IEEE is the world's largest technical professional organization dedicated to advancing technology for the benefit of humanity. Use of this web site signifies your agreement to the terms and conditions. Model predictive control of a nonlinear fed-batch fermentation process Abstract: Bioprocesses, which are involved in producing different antibiotics and other pharmaceutical products, may be conveniently classified according to the mode chosen for the process: either batch, fed-batch or continuous. From the control engineer's viewpoint it is the fed-batch processes, however, which present the greatest challenge to get a pure product with a high concentration. To achieve this goal, control of the following parameters has significant importance dealing with these processes: temperature, pH, dissolved oxygen DO2.

Fed-batch culture is, in the broadest sense, defined as an operational technique in biotechnological processes where one or more nutrients substrates are fed supplied to the bioreactor during cultivation and in which the product s remain in the bioreactor until the end of the run. In some cases, all the nutrients are fed into the bioreactor. The advantage of the fed-batch culture is that one can control concentration of fed-substrate in the culture liquid at arbitrarily desired levels in many cases, at low levels. Generally speaking, fed-batch culture is superior to conventional batch culture when controlling concentrations of a nutrient or nutrients affects the yield or productivity of the desired metabolite. Nutrients such as methanol, ethanol, acetic acid, and aromatic compounds inhibit the growth of microorganisms even at relatively low concentrations. By adding such substrates properly lag-time can be shortened and the inhibition of the cell growth markedly reduced. In a batch culture, to achieve very high cell concentrations, e.

JavaScript seems to be disabled in your browser. You must have JavaScript enabled in your browser to utilize the functionality of this website. Supply of a substrate is a must for any bioprocess, even if it is just mineral salts, light and CO 2 for algae. Usually, it is in the form of a sugar, which is either provided just at the start of a process or added over time. The choice of which method to use will depend on the organism, application and final goal. Tony Allman.

Comparing Culture Methods in Monoclonal Antibody Production: Batch, Fed-Batch, and Perfusion

In fact, human monoclonal antibodies hMAbs produced by CHO cells have played a major role in both the diagnostic and therapeutic markets for decades. Since that approval in , scores of chimeric, humanized, and human MAbs have gained approval and entered clinical use. In development of all pharmaceutical production processes, including those involving hMAbs produced by CHO cells, decisions regarding the best process parameters and methods are made based on cost, time, and titer comparisons. Often, multiple scalable platforms are examined before a final process is transferred to pilot- or scale-up laboratories 2 — 8.

Data can be given on request. The IP rights of the cV yeast strain are exclusively owned by Terranol. Extending this phase in a fed-batch process increases the yield, rate and additionally reduces needed yeast amount for pitching. Extending this phase, as long as possible, would further enhance yield and economy of the process. To realise the concept a fermentation monitoring technique was developed and applied.

Cultivation technique of bacteria: Batch, Fed-batch and Continuous culture technique

The Difference Between Batch, Fed-batch and Continuous Processes

Moreover, unlike other model organisms, S. This type of fermentation is used for production of proteins from recombinant microorganisms. Antioxidant activities were determined by three methods comprising diphenyl picrylhydrazyl radical scavenging assay DPPH , radical cation decolorization assay ABTS and reducing power.

 - Но будем надеяться, что он этого не узнает. ГЛАВА 76 У подъезда севильского аэропорта стояло такси с работающим на холостом ходу двигателем и включенным счетчиком. Пассажир в очках в тонкой металлической оправе, вглядевшись сквозь стеклянную стену аэровокзала, понял, что прибыл вовремя. Он увидел светловолосую девушку, помогающую Дэвиду Беккеру найти стул и сесть. Беккера, по-видимому, мучила боль.

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References

Она показала на экран. Все глаза были устремлены на нее, на руку Танкадо, протянутую к людям, на три пальца, отчаянно двигающихся под севильским солнцем. Джабба замер. - О Боже! - Он внезапно понял, что искалеченный гений все это время давал им ответ. - Три - это простое число! - сказала Соши.  - Три - это простое число. Фонтейн пребывал в изумлении.

 - Халохот думал, что поблизости никого. Халохот какое-то время наблюдал за происходящим, потом скрылся за деревьями, по-видимому, выжидая. - Сейчас произойдет передача, - предупредил Смит.  - В первый раз мы этого не заметили. Сьюзан не отрываясь смотрела на эту малоприятную картину.

 Умно, - сказала Сьюзан. Стратмор продолжал: - Несколько раз Танкадо публично называл имя своего партнера. North Dakota. Северная Дакота. - Северная Дакота. Разумеется, это кличка. - Да, но я на всякий случай заглянул в Интернет, запустив поиск по этим словам.

А теперь выходи. Но Мидж эта ситуация явно доставляла удовольствие.

За ее спиной ТРАНСТЕКСТ издал предсмертный оглушающий стон. Когда распался последний силиконовый чип, громадная раскаленная лава вырвалась наружу, пробив верхнюю крышку и выбросив на двадцать метров вверх тучу керамических осколков, и в то же мгновение насыщенный кислородом воздух шифровалки втянуло в образовавшийся вакуум. Сьюзан едва успела взбежать на верхнюю площадку лестницы и вцепиться в перила, когда ее ударил мощный порыв горячего ветра.

Стратмора видно не. В ужасе от того, что ее ожидало, она направилась к кабинету шефа. Когда Сьюзан уже сделала несколько шагов, что-то вдруг показалось ей странным. Она остановилась и снова начала вглядываться в глубь помещения Третьего узла. В полумраке ей удалось различить руку Хейла.

 - Мидж посмотрела в монитор и постучала костяшками пальцев по столу.  - Он здесь, - сказала она как о чем-то само собой разумеющемся.  - Сейчас находится в шифровалке.

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