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Method


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Updated 2019-12-03

 Method - expression of analytical results

The expression of analytical results and calculated values may depend on several issues. It is well known that food composition data usually are expressed per 100 g - contradictory to the Système International (SI system) where it would be more appropriate to express values per kilo (kg). There has been a few attempts to do so, e.g. the Swedish 1978 food composition table, but the nutrition community did not receive this well.
Likewise, the use of kilocalories (kcal) as energy unit is still widely appearing despite the fact that the unit was internationally discarded more that 40 years ago in preference of the Joule (kJ) in the SI system.

Other expressions that may cause confusion is the "equivalent" expressions, like the expressions used for for vitamin A activity (RE, RAE, etc.), niacin equivalents, expression of vitamin A and D in international units, fatty acids expressed per 100 g FA - is it really FA (fatty acids) or FAME (fatty acid methyl esters)? - amino acids expressed per 100 g, per g N or per 16 g N, etc.

In addition, there are methodologically different expressions for a range of components. Regulations or analytical method specifications/standards may even prescribe different expressions - even for the same component.

A few examples of different expressions for the same component are
 

Protein
 
The recommended method (FAO 2003) for protein determination is "the sum of amino acid residues" (see also Protein). The acceptable method of protein determination is the most commonly used calculation of the so-called "crude protein" from the amount of total nitrogen (N) analysed in the food by the Kjeldahl or comparable method (Dumas, Kjel-Foss (automated Kjeldahl using antimony-based catalyst), Kjeltec, etc).

By multiplying the total nitrogen content with a food matrix specific factor, the nitrogen-to-protein conversion factor (NCF) or Jones' factor:

    protein content = total nitrogen content x specific conversion factor  

the "crude protein" content is found.

However, the total nitrogen content measured with different analytical methods, give similar, but not same results. Furthermore, differences are dependent of the food matrices. As an example, the AOAC Official Method 992.15 Crude Protein in Meat and Meat Products Including Pet Foods, Combustion Method, First Action 1992, mentions in a foot note that "Results using this method average 1.01 * results using 928.08" (928.08 being the AOAC "standard" Kjeldahl method) - this means that the nitrogen content measured with the newer combustion method is higher than the nitrogen content measured with the traditional Kjeldahl method.
 

Total lipid/total fat

The recommended method (FAO 2003) for determination of (total) lipid/fat is as "fatty acids and expressed as triglycerides, as this approach excludes wax esters and the phosphate content of phospholipids"; this definition imply the use of gas chromatographic methods.
The acceptable methods are the traditional gravimetric methods involving extraction with one or more solvents, but the different methods yield different results (see also Lipids).
 

Fatty acids
 
Errors concerning fatty acid values often occur because it is not clear on which basis the fatty acid values are given. Values for fatty acids in profiles are usually given as % of total fatty acids (or g/100 g total FA). There is an ambiguity build into this expression, because "total fatty acids" can mean (at least) two things

Unidentified fatty acids are commonly designated as a value named "unknown" or "unidentified" when figures are given. The amount of unknown FAs can easily be 10-20% of the total of known and unknown fatty acids.
Therefore, the expression where only known FAs are included in the sum leads to a (gross) overestimation of the content of the fatty acid contents and this expression should be avoided.
 

Thiamin
 
In thiamin analysis, the standard analytical methods, e.g. AOAC 942.23, 953.17, 957.17, 986.27 and EN 14122, use thiamine chloride hydrochloride (AOAC. thiamine-HCl) as standard solution. The analytical results are expressed as thiamine chloride hydrochloride (thiamine-HCl), accordingly.
However, the EN 14122 standard opens up for recalculation of the analytical results as follows:

EN 14122

This means that thiamin can also be expressed as thiamin (1+) ion and thiamin chloride, which will evidently give different values for the nutrient, depending on the expression.

Newer HPLC methods using post-column derivatisation also detect content of 2-(1-hydroxyethyl)thiamine (HET) (mw: 381.33), which has the same thiamine activity as thiamine. HET gives a significant contribution to the total vitamin B1 activity.
Therefore, vitamin B1 should be expressed as

Vitamin B1 = thiamine + 2-(1-hydroxyethyl)thiamine

When vitamin B1 is expressed as thiamin chloride hydrochloride (mw: 337.28), HET is added to the determined thiamine content after multiplication with (337.28/381.33).
Investigations have shown that 7-24% of thiamin detected in samples of animal origin derives from HET. Similarly, HET contributed to 37% of the total amount of vitamin B1 in dried yeast (Jakobsen, 2008).
 

Pantothenic acid
 
Customarily, the reference standard in pantothenic acid analysis is calcium D-pantothenate, and the most common ways of expressing pantothenic acid are either as D-pantothenic acid (molecular mass 219.237) or as calcium D-pantothenate (476.536).  However, older official methods, e.g. AOAC 945.74, also indicate the possibility of expressing the pantothenic acid potency as sodium pantothenate (molecular mass 241.219).
Due to the difference in molecular mass of the three compounds, there will be a difference in the value determined depending on the chosen expression.
To calculate from D-pantothenic acid to calcium D-pantothenate use a factor 1.087. Similarly, to calculate from calcium D-pantothenate to D-pantothenic acid use a factor 0.920.
To calculate from D-pantothenic acid to sodium pantethonate use a factor 1.100. Similarly, to calculate from sodium pantothenate to D-pantothenic acid use factor 0.909.
 

Vitamin B6
 

The term vitamin B6 refers to a series of components, mainly pyridoxine (pyridoxol), pyridoxal and pyridoxamine, chemically slightly different forms of the vitamin. Vitamin B6 occurs in foods as pyridoxin, pyridoxal and pyridoxamine, and may be present in both the free for or a chemically bound state, e.g. as phosphates and glucosides. Hence, the extraction of the chemically bound forms is extremely important and usually involves enzymes or hydrolysation (heating with acids).
After releasing the bound state of the vitamin, its components are separated chromatographically and more recently by HPLC, followed by determination of the vitamin B6 fractions either microbiologically (microbiological assay) or by HPLC.
The standard solutions used in the microbiological assay are pyridoxine-HCl, pyridoxal-HCl, pyridoxamine-HCl. This gives rise to different ways of expressing values for vitamin B6, the hydrochloride expression and the free base expression. As the molecular weights of pyridoxine, pyridoxal and pyridoxamines are 82, 82 and 70 percent of the weight of the hydrochlorides, there is good reason to be alert. Failure to account for this difference leads to serious errors in the interpretation of the final vitamin B6 content.
It is therefore important to determine if values are designated as hydrochloride or free-base components.
 

Vitamin C

Be aware that older vitamin C methods determine acsorbic acid only. The preferred methhods shall determine both acsorbic acid and dehydroascorbic. Vitamin C is the sum of ascorbic acid and dehydroascorbic acid.
 

 

Phosphorus
 

Another example with different expressions for the same component is Phosphorus. Phosphorus can be expressed as the element P, the normal component used in food composition. However, phosphorus is in food inspection and legal documents often expressed as phosphorus pentoxide, P2O5 and analysed as such.
To recalculate results given as P2O5 to P, use a factor of 0.437.
 

 


 References

 


 



 News
NEVO-Online 2019

2019-11-19
New edition of Dutch food composition database NEVO-online: compositional data on more than 2150 food items - 2019. See NEVO website.
 
USDA ARS FoodData Central

2019-10-01
USDA's Beltsville Human Nutrition Research Center (BHNRC) has established FoodData Central. Available at fdc.nal.usda.gov.
 
UK Composition of Foods online

2019-05-23
Food Databanks National Capability extended dataset based on PHE’s McCance and Widdowson’s Composition of Foods Integrated Dataset.
 
Composition of foods integrated dataset (CoFID) 2019

2019-05-22
McCance and Widdowson’s 'composition of foods integrated dataset' on the nutrient content of the UK food supply. Available from Public Health England's website.