Hello Cansu! The control should be made with both, acid and alkali solutions, by the nature of your process. In any case, its better to include both to be sure that the pH will be controlled. Don't forget to check how much alkali and acid solution is added by control. 

Hi Christian,

Thank you for Your answer. Do you start the pH Control at the same as you start the fermentation? Or do you wait until the end of the exponential Growth phase, for example?

Since the startting pH is high (~6.8), I thought it might be problematic to start the pH from the begining, then we will have to add so much acid solution to decreasse the pH to 5, and I am not sure how it effects the fermentation.

Hi, in our lab we have done lot of work with different Clostridium strains including beijerinckii 8052.  Regularly we adjust the starting pH at around 6.2-6.3 (usually P2 media).  During fermentation, we just adjust with base, because the main trouble during the ABE fermentation is the acid crash of the bacteria, after the acidogenesis.  We use a bioreactor that control the pH at 5. That's means, the pH starts at 6.2 and decrease with the acid production; then, the reactor do not let the pH become lower than 5, and the fermentation is run successfully, finishing in 3 days.  No acid addition is needed, although some people believe, if the pH increase a lot, you can add some acid to help in the reassimilation of acids.

As you mentioned, your fermentation stops when you reach some values of acid production (acetic and butyric), so I think you are suffering acid crash.  On the other hand you mentioned you control the pH at 5, and that should be enough to avoid the acid crash, so, I think maybe your pH probe can have some problem (or the calibration process or the buffers).  Sometimes the old pH probes just present several problems.  When your fermentation stops you can measure the pH with another method, just to try to confirm my hypothesis.  Other option is to try to control the pH at higher value, like 5.5.  But pH 5 should be good enough for 8052 strain.

In my experience, there are two key problems in ABE fermentation: 1. when the fermentation starts and then stops after some acid production, the problem is always acid crash.  2. If the fermentation is enable to start from the very begging or starts very slow, the problem is a high level of oxygen.

Dear Pablo,

Thank you for your reply. You are right, pH probes can give wrong results some times but I always check the pH of the broth with another pH probe. So, there is no problem on the probe. C.beijerinckii 8052 switches to solventogenesis at around pH 5, never below that, so that is why I don't think the problem is the acid crash, but inhibition by the acids. I will investigate that further. 

If you have any observations and experiences with the pH control with acids for ABE fermentation, I will be happy to hear.

Cansu,

Here is some info I could recall when I was working on this pH controlled process. Maybe helpful somehow.

Charles

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Biotechnology Letters

April 1993, Volume 15, Issue 4, pp 421–426

Controlled-ph batch butanol-acetone fermentation by low acid producing Clostridium acetobutylicum B18

Summary

C. acetobutylicum B18 produced a large amount of butanol over a wide range of pH (4.5–6.0). At pH 6.0 fermentation and cell growth were most active at pH 6.0, and the highest values of glucose consumption rate (4.37 g/L-h), butanol productivity (1.0 g/L-h), butyric acid recycle rate (0.31 g/L-h), and cell growth rate (0.2 h-1) were obtained. There existed a critical pH between 6.0 and 6.5 above which cells switched to organic acid producing mode. Clostridial stage appeared essential for solvent production by strain B18 but sporulation was not necessary for solvent formation.

Hi Cansu.

Ok, sorry, I just suggest 5.5 thinking in a problem with the probe, but I see you already check that.  We always use 5, and it works, just adding NaOH, and no acid addition.  

So, I am looking the data of a friend, which is co-worker in the lab and he has a mutant of 8052 strain able to produce 5.6 g/L of butyric acid, and this strain is also able to produce 9.6 g/L of butanol (less than wild type, but the acid doesn't stop the fermentation).  But the wild type does never yield that high butyrate concentration in our fermentations. Wild type produce around 2 g/L of butyrate, 14 g/L butanol and 10 g/L acetone, using P2 media with 60g/L of glucose (consuming around 50g/L).

Is your strain a mutant?  is there a problem with the acid re-assimilation?  what substrate are you using?

Regards!

Hello dear Cansu, it looks and interesting topic,  could you rephrase your question?

I hope to be helpful, regards