Hi Reeta,

In reality it is much easier to control conditions at flask level than reactor level. The huge size and the growth of organism in a reactor is quite a trick to control. However, may be the huge quantatity of biomass and the efficient extarction of certain enzyme at reactor level make it seem that the reactor level has an enhanced result.

Many factors like nutrition ect plays a major role. At flask level the nutrition may be less, whereas in the reactor level the amount of nutrition is huge and the growth also enhances leading to huge production of required product.

However, conditions like temprature, pH, nutrition and the most important "innoculum growth phase" are the factors to be considered while going from a flask to a reactor level

cheers

Vivek Kempraj

Hi Reeta

Stirring is usually a key issue when growing filamentous fungi in a bioreactor.

impellers tend to destroy fungi´s hyphae, this is something you have to take into consideration.

moreover, fungi morphology in submerged growth is of paramount importance. Pellet or filamentous growth must be tackled, and from my experience, when extracellular enzyme production is the goal, filamentous growth will result in higher yields.

Best regards

Filipe

P.s. If you are getting the same enzyme production in bioreactor as in shaken flasks, i would consider it a successful scale up.

Dear Reeta: The most important limitation, which must be considered from passing from a flask to a reactor size is to provide the required amount of dissolved oxygen to the culture (for aerobic bioprocesses, which in most case are those used to produce extracellular enzymes). I disagree with the first comment - clearly culturing in a bioreactor vessel equipped with the necessary pH, DO and T probes and the regulatory circuits to control these parameters is much better than culturing in flasks. Flasks should be used only for preliminary screening and comparison not for process optimization. Hope this answers to your question.

I concur with Svetlozar. An additional advantage of bioreactors is

the option, in (industrial) fed-batch bioreactors, to programme feed profiles and, thereby, specific growth rate. In fed-batch cultures, both the identity of the growth-limiting nutrient and the specific growth rate (profile) can have a large impact on productivity, product yield on substrate and product titer. As Svetlozar rightly comments, rational, knowledge-based optimization of such (dynamic) processes cannot be based on shake-flask data.

Thankyou Filipe, Svetlozar, Jack and Vivek. Yes the process is for extracellular proteins by filamentous fungi, and I tried hard to maintain DO but as soon as the growth starts it goes down and am unable to attend it as increasing airflow beyond 5l/min increases pressure and increasing agitation might have harmful effects on filamentous fungi. So in this batch have planned to use oxygen rather than air to maintain DO to atleast 30%. In my present batch have attained a high growth by additional nutrition but still the protein level didn't increase.

The medium after after growth turned too viscous and now the mycelium started disitegrating as the medium viscosity is now decreasing. I am expecting protein secretion in this stage. Should I?

Dear Reeta,

With so little information this is a bit of a guessing game but:

1. do you use pH control in your fermenters? Use of ammonium-based media will cause strong acidification. This acidification can denature extracellularly produced heterologous proteins that are not acid tolerant. (Always check if the product of interes is stable in fermentation broth)

2. Don't be too careful with increased agitation of the cultures.

3. The standard way to prevent oxygen limitation in this type of processes is sugar-limited fed- batch cultivation, either with a preprogrammed feed profile that prevents oxygen limitation or with DOT/RQ-controlled feed.

Good luck,

Jack

Jack, thank you very much...Your comments are really useful. Yes pH is controlled at 4.0. Probably it could be a reason for not getting increased protein concentration. 2nd and 3rd point are too useful for me.