Hi Tolga,

Perhaps some of the acids were present both within proteins and in free form. Eather that or it is a mistake.

Hi Polina,

Thx for your answer. I also talked to several scientists. They told me that some of the carbohydrate substances may bond to Nitrogen or reverse; thus the HPLC device may consider the substances as amino acids. The second accumption is the differences arise from the methods between a titrimetric simple protein anaylsis and an instrumental analysis. We all assume that the crude protein also includes the free form of amino acids, please correct me if I am wrong?

Dear Tolga Şahin ,

Interesting observation!

There are multiple reasons why this happens (apart from a mistake of course or a confusion in the way they quantify the AA content):

-First and most likely it has something to do with what you suggested yourself. The method for crude protein determination and for AA determination differ and are (unfortunately) not as absolute as one might wish.

-Second the crude protein determination is intrinsically an estimate. In a rather old paper it is already found that only about 15% of the crude protein in the herring silage was true protein and that ammonia N accounted for 8% of the crude protein is accounted for by the nitrogen from ammonia and the rest is mainly peptides and free amino acids. See:

Nicholson, J. W. G., & Johnson, D. A. (1991). Herring silage as a protein supplement for young cattle. Canadian Journal of Animal Science, 71(4), 1187-1196.

https://www.nrcresearchpress.com/doi/pdfplus/10.4141/cjas91-140

-Third in relation to the first two point read the following:

Mariotti, F., Tomé, D., & Mirand, P. P. (2008). Converting nitrogen into protein—beyond 6.25 and Jones' factors. Critical reviews in food science and nutrition, 48(2), 177-184.

https://hal.archives-ouvertes.fr/hal-02105858/document

Basically they state that the well-known 6.25 factor used in the crude protein determination is an oversimplfication. Multiple factors influence this rough estimate.

So in conclusion you’ve touched a very interesting point (not even mentioning the highly relevant issue of bio-availability and or apparent digestibility of amino acids). The findings are interesting by itself but use of both methods are still relevant when one compare different samples or circumstances.

Hope this shed some light on the matter.

Best regards.

Thank you for your kind response Dr. Keller,

When I sum up the crude ash, lipid and protein ratios at the figure I shared, the sum already exceeds 100% and there is even no place for NFE. So, there might be a mistake in this paper. However, I saw the same thing in some other articles that the ratios of total amino acids exceed the crude protein ratio. Therefore, as you mentioned above, this is a probability and it happens sometimes. I've also looked into the articles you shared. There is interesting information. It says that the total amino acids constitute ~60% of the crude protein in Herring silage. We are trying to make it clear that the crude protein can be less than total amino acids; however the info above makes it too difficult to understand. Do i miss something?

Dear Tolga Şahin ,

I think we are talking here about two different things. In the figure you mentioned the authors try to make clear the different fractions that contribute to the final crude protein amount. If you look at table 2 than the total amino acid amount is roughly 61% of the crude protein amount, which I can understand since the crude protein measurements are such a rough estimate.

However the other arguments (as I indicated too) can results in the other way around and the only reasonable (and main raison) is presumably simply the difference in analysis method (and the difference in what you really look at).

As said before I think the crude protein analysis is a measure by itself that is useful to compare different samples or differences between species. The total amino acid analysis is useful to compare between samples and/or species as well in terms of difference in amino acid composition.

The link between crude protein and amino acid is in the perfect world the same (at least the number should be) but in reality we have to do it with a weak relationship that sometimes make the total amino acid content lower and sometimes higher than the crude protein content (all depending the methods used to determine both).

Hope this brings some clarity in this indeed rather complicated and not fully understood phenomenon.

Best regards.

Dr. Keller,

Thanks for detailed information. I am fully satisfied with your answer.

Kind regards.