The innoculum sizes for starter culture (controlled fermentation) seem to vary. How is it dependent on the organism being used or is there a minimum quantity that will ensure that the desired organism is responsible for most metabolic transformations.
Since you have not mentioned the fermentaion system and there is no generalize rule but its logic goes in the following way.
Inoculum % depends on time at which reaches log/exponential phase and shaking
1) Aerobic microbes uses oxygen for growth/final electron acceptor producing high amount of ATP hence growth rate is high. Hence you need a lower number of cell at start point (inoculum) to reached a desired number at a particular time (remember time at which you want desired growth is also imp,) .
So around 1% to 2% of inoculum is generally good enough for aerobic microbes. for e.g E.coli for 16 hrs in LB broth for 16 hr gives O.D of 5-7
2) Obligate Anerobic fermentative microbes produces less ATP, so require higher number of microbes in starting inoculum to reached a particular number in particular time,
So around 10% to 15% used. E.g Clostridium acetobutylicum for 24-36 hr in TYA medium give O.D 5-7.
3) Facultative anerobic microbes can grow at intermediate oxygen level and mid % of inoculum is needed
So around 5% to 10% is good enough. E.g yeast for 24 hrs at 70 rpm in MGYP medium gives OD of 7-8
4) mixed consorita of anerobes requires more inoculum % because of different growth of different microbes present.
So around 10 to 15%. E.g Methanogenic consortia takes 20 days in Msp medium to reach O.D of 2-3.
5) Anerobic respiration. Depending on final electron exceptor number of ATP produce varies. I have no persional experience so cant help you with that but initial inoculum % should be around 5 to 10%.
Usually the standard inoculation size is 1 % v/v of overnight culture in logarithmic phase. However, the inoculation size can be raised or lowered depending on the growth characteristics of the microorganism. Each microorganism is different when it come to its growth capabilities so its important to determine the generation/doubling time. This information would allow you to alter the inoculation size to obtain optimal fermentation conditions.
Hy, I agree with Deon with the following addition: one should keep in mind the characteristics of each fermentation system (microbial consortia, fermentation substrate, desired end-products, etc). Thus, for example when you are dealing with mixed microbial consortia as well as a complex fermentation substrate it is best to use a 5 to 10 % v/v starting inoculum.
Thus, I recommend a set of preliminary experiments through which you can specifically determine the optimal required inoculum size before designing the main experimental setup.
The fermenter seed size there is no thumb rule. As per lab practices we can use from 0.1 ( if culture growing very fast) and up-to 30 % v/v (Some time seed culture reusing for just bio-transformation activity ) . As it dependence upon what kind of microbes we are using.
So to develop optimum seed size we need to optimize the on bench scale by considering following points
1) Minimum seed size and also minimum lag phase during fermentation.
2) During scale up we should consider what capacity seed reactor is available.
Since you have not mentioned the fermentaion system and there is no generalize rule but its logic goes in the following way.
Inoculum % depends on time at which reaches log/exponential phase and shaking
1) Aerobic microbes uses oxygen for growth/final electron acceptor producing high amount of ATP hence growth rate is high. Hence you need a lower number of cell at start point (inoculum) to reached a desired number at a particular time (remember time at which you want desired growth is also imp,) .
So around 1% to 2% of inoculum is generally good enough for aerobic microbes. for e.g E.coli for 16 hrs in LB broth for 16 hr gives O.D of 5-7
2) Obligate Anerobic fermentative microbes produces less ATP, so require higher number of microbes in starting inoculum to reached a particular number in particular time,
So around 10% to 15% used. E.g Clostridium acetobutylicum for 24-36 hr in TYA medium give O.D 5-7.
3) Facultative anerobic microbes can grow at intermediate oxygen level and mid % of inoculum is needed
So around 5% to 10% is good enough. E.g yeast for 24 hrs at 70 rpm in MGYP medium gives OD of 7-8
4) mixed consorita of anerobes requires more inoculum % because of different growth of different microbes present.
So around 10 to 15%. E.g Methanogenic consortia takes 20 days in Msp medium to reach O.D of 2-3.
5) Anerobic respiration. Depending on final electron exceptor number of ATP produce varies. I have no persional experience so cant help you with that but initial inoculum % should be around 5 to 10%.
Dear Philippa Ojimelukwe ,
What kind of fermentation? In the case of research on alcoholic fermentation (beer, wine, cider) recommended for evaluation of yeast medium is used in 1-2 x 10E6 cfu / ml of initial inoculum. Thus, you can analyze the yeast growth curve and thus perform several kinetics experiments modifying the must or yeast.
Like Jai Ghosh said usually in any fermentation the inoculum size constitutes 10% of the entire volume of fermentation medium. However, depending on the organism and nature of fermentation it can be altered. It is good to inoculate with an inoculum with high number of the desired organism(s). This will hasten or drive the fermentation faster with better results. The number you stated (1-2 x 10E6) is good enough for fermentation.
Agreed with Mustafa Vohra.
But still u will need to optimize the amount of inoculum required for inoculation.
You will need to consider the number of cells in the inoculum.
For the purpose, try to use overnight culture/log phase cultures.
As for example, for 5% inoculum the ratio to be used could as follows:
5% means, 5 ml inoculum with OD@600nm = 1 (using same un-inoculated medium as blank) to be added into 100 ml of sterile nutrient medium.
So, you can go for preliminary set with 1 - 10% inoculum, to find which one is better for you and then finalize.
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