I am not specialize area, but came across such studies as attached, hope it is helpful.

Regards

Article New mechanistic model to simulate microalgae growth

Article Nutrient and media recycling in heterotrophic microalgae cultures

I  assume that you have already seen my paper on NIFTs that you might find useful. There are also references  within that will be of help. Regards, Elly

to answer directly to your question. there is no preset period of time  for the onset of nutrient limitation. This will depend on the species in culture, as well as on general culture conditions which will determine its physiological activity. two-three months seems an inordinately long time though, but remember the definition of nutrient limitation, which is fairly different from the ability to survive in a nutrient-low medium

Thank you very much for the reference, Mustafa Tekke. I'll have a look at them. 

Elly, thanks a lot for the paper and references. It's very helpful.  

A question, how do you determine a true NIFT response if we don't have a control? Sometimes the Ft of before and after nutrient addition has a slight changes but how do we know if it's a false or true response. 

Yes, I agree with you Domenico.

The cultures in which I used were approximately 2 months old.

What do you suggest to test the nutrient limitation in the culture?  I assume they were nutrrient-limited based on the culture age.

Thank you very much for your opinion.

 Most probably they were, but all I can suggest is that you use your original culture to inoculate new ones in a series of fresh media, each prepared omitting one nutrient source. In this case, seawater  without any enrichment will serve as your control, both to check if you get a NIFT response as well as for the classic growth experiment (I assume you use natural seawater, but you should try with artificial seawater, or at least with water from some oligotrophic area, if you have one handy).

good luck and keep us posted, domenico

Great All theBest