ISO/DIS 23776

DRAFT INTERNATIONAL STANDARD
ISO/DIS 23776
ISO/TC 34/SC 6 Secretariat: SAC
Voting begins on: Voting terminates on:
2020-12-02 2021-02-24
Meat and meat products — Determination of total
phosphorous content
Viandes et produits à base de viande — Détermination de la teneur en phosphore
ICS: 67.120.10
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ISO/DIS 23776:2020(E)
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ISO/DIS 23776:2020(E)
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Foreword ........................................................................................................................................................................................................................................iv

1 Scope ................................................................................................................................................................................................................................. 1

2 Normative references ...................................................................................................................................................................................... 1

3 Terms and definitions ..................................................................................................................................................................................... 1

4 Principle ........................................................................................................................................................................................................................ 1

4.1 Inductively coupled plasma optical emission spectrometry (ICP-OES) method ........................... 1

4.2 Spectrometric method ...................................................................................................................................................................... 1

4.3 Gravimetric method ............................................................................................................................................................................ 1

5 Sampling ........................................................................................................................................................................................................................ 2

6 Inductively coupled plasma optical emission spectrometry (ICP-OES) method ................................2

6.1 Reagents........................................................................................................................................................................................................ 2

6.2 Apparatus .................................................................................................................................................................................................... 2

6.3 Procedure .................................................................................................................................................................................................... 3

6.4 Determination ......................................................................................................................................................................................... 3

6.5 Calculation and expression of results.................................................................................................................................. 3

6.6 Limit of detection .................................................................................................................................................................................. 4

6.7 Precision ....................................................................................................................................................................................................... 4

6.8 Repeatability ............................................................................................................................................................................................. 4

6.9 Reproducibility ....................................................................................................................................................................................... 4

7 Spectrometric method .................................................................................................................................................................................... 4

7.1 Reagents........................................................................................................................................................................................................ 4

7.2 Apparatus .................................................................................................................................................................................................... 5

7.3 Preparation of test sample ............................................................................................................................................................ 6

7.4 Procedure .................................................................................................................................................................................................... 6

7.4.1 Test portion .......................................................................................................................................................................... 6

7.4.2 Determination .................................................................................................................................................................... 6

7.5 Calibration graph .................................................................................................................................................................................. 7

7.6 Calculation .................................................................................................................................................................................................. 7

7.7 Precision ....................................................................................................................................................................................................... 7

7.8 Repeatability ............................................................................................................................................................................................. 7

7.9 Reproducibility ....................................................................................................................................................................................... 7

8 Gravimetric method .......................................................................................................................................................................................... 8

8.1 Scope and field of application .................................................................................................................................................... 8

8.2 Definition ..................................................................................................................................................................................................... 8

8.3 Sample ............................................................................................................................................................................................................ 8

8.4 Reagents........................................................................................................................................................................................................ 8

8.5 Apparatus .................................................................................................................................................................................................... 8

8.6 Procedures .................................................................................................................................................................................................. 9

8.6.1 Preparation of test sample ...................................................................................................................................... 9

8.6.2 Test portion .......................................................................................................................................................................... 9

8.6.3 Mineralization .................................................................................................................................................................... 9

8.6.4 Determination .................................................................................................................................................................... 9

8.6.5 Blank test .............................................................................................................................................................................10

8.6.6 Expression of results .................................................................................................................................................10

8.6.7 Repeatability ....................................................................................................................................................................10

8.7 NOTES ON PROCEDURE ...............................................................................................................................................................10

9 TEST REPORT ........................................................................................................................................................................................................11

Annex A (informative) Interlaboratory testing ......................................................................................................................................12

Bibliography .............................................................................................................................................................................................................................18

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Barriers to Trade (TBT) see the following URL: Foreword - Supplementary information

The committee responsible for this document is ISO/TC 34, Food products, Subcommittee SC 6, Meat,

This secondedition cancels and replaces the firstedition (ISO 13730:1996), which has been technically

— A new test method - inductively coupled plasma optical emission spectrometry (ICP-OES) method

— The introductory texts for “Foreword”, “Normative references” and “Terms and definitions” have

This International Standard specifies three methods for the determination of total phosphorous content

- Inductively coupled plasma optical emission spectrometry (ICP-OES) method, Spectrometric method

and Gravimetric method of all kinds of meat and meat products, including poultry and livestock.

For the inductively coupled plasma optical emission spectrometry (ICP-OES) method, the limit of

detection (LOD) is 1.0mg/kg and limit of quantify (LOQ) is 3.0 mg/kg if the mass of test portion is 0.5g,

The following documents are referred to in the text in such a way that some or all of their content

constitutes requirements of this document. For dated references, only the edition cited applies. For

undated references, the latest edition of the referenced document (including any amendments) applies.

ISO 3696:1987, Water for analytical laboratory use — Specification and test methods

mass of phosphorous pentoxide determined by the procedure specified in this International Standard,

After digestion, the sample is determined by inductively coupled plasma emission spectrometer with

the characteristic spectral line wavelength of phosphorous. In a certain concentration range, the

spectral line signal intensity of phosphorous is proportional to its concentration, and is quantified by

Drying of the test portion and incineration of the residue. After cooling, hydrolysis of the ash with

nitric acid. Filtration and dilution followed by the formation of a yellow compound with a mixture of

ammonium monovanadate and ammonium heptamolybdate. Photometric measurement at a wavelength

Mineralization of a test portion with sulphuric and nitric acids. Precipitation of the phosphorous as

quinoline phosphomolybdate. Drying and weighing of the precipitate. An alternative method of

Sampling is not part of the method specified in this International Standard. A recommended sampling

It is important that the laboratory receive a sample which is truly representative and has not been

Start from a representative sample of at least 200 g. Store the sample in such a way that deterioration

6.1.3 Nitric acid (5 + 95): take 50mL nitric acid(6.1.1), slowly add 950mL water, mix.

6.1.4 phosphorous standard stock solution (1000mg/L) is certified by the state and is awarded the

standard solution of certified reference material. c (P) =1000 mg/l; c (P O ) =2294 mg/l.

6.1.5 Standard series solution of phosphorous: accurate extract standard reserve liquid, dilute

standard series solution with nitric acid solution (5+95). The mass concentration is 0mg/L, 20.0mg/L,

NOTE According to the sensitivity of the instrument and the actual content of phosphorous in the sample,

the concentration range of the standard solution should be adjusted appropriately.

6.2.3 Microwave digestion instrument: with polytetrafluoroethylene digestion internal tank

6.2.4 Electric hot plate with adjustable temperature control, or graphite digestion unit.

6.2.6 Homogenizer, high speed pulverizer: capable of samole pulverizing and homogenizing.

6.3.1 Sample pre-treatment: Samples with low water content are mixed together after removing

Microwave digestion: weigh, 0.2g~0.5g of the test portion (accurate to 0.001g, the sample with more

moisture content can be appropriately increased to 1g~2g) in the microwave digestion internal tank

(6.2.10) with Polytetrafluoroethylene digestion tube (6.2.10), add 5mL~10mL nitric acid(6.1.1), stand

for 1h or overnight, screw the tank cap, follow the standard operation steps of the microwave digestion

instrument to digest. Take out after cooling, slowly open the tank cap and vent, flush the inner cap with

a little water, put the digestion tank on electric hot plate (with adjustable temperature control)(6.2.4)

or in the ultrasonic water bath(6.2.5), heat 30min at 100°C or ultrasonic degassing for 2min~5min,

6.4.1 Instrument reference conditions: optimize the operating conditions of the instrument; make

the instrument sensitivity and other indicators to meet the requirements of the analysis. The reference

conditions for the instrument operation are observation mode: horizontal observation; power: 1150W;

plasma gas flow: 15L /min; auxiliary gas flow: 0.5L /min; atomized gas flow: 0.65L /min, measured line:

6.4.2 Standard curve drawing: the standard series working solution is injected into the inductively

coupled plasma emission spectrometer, and the intensity signal response of the analytical spectral line

is determined. When the element concentration is abscissa, the spectral line intensity response value is

Sample determination: the blank solution and sample solution are injected into the inductively coupled

plasma emission spectrometer respectively. The signal response values of the spectral line strength

are measured, and the concentration of phosphorous in the solution is obtained according to the

The results of the interlaboratory comparison in the test method will be attached soon

The limit of detection (LOD) is 1.0mg/kg and limit of quantify (LOQ) is 3.0 mg/kg if the mass of test

The precision of the method has been established by an interlaboratory test, carried out in accordance

The absolute difference between two independent single test results, obtained using the same method

on test material in the same laboratory by the same operator using the same equipment within a short

The absolute difference between two independent single test results, obtained using the same method

on identical test material in different laboratories with different operators using different equipment,

Use only reagents of recognized analytical grade and distilled or demineralised water or water of at

Mix 1 volume of nitric acid [65 %( m/m); ρ =1, 40g/ml (7.1.1) with two volumes of water.

Dissolve 2.5g of ammonium metavanadate in 500 ml of boiling water. Cool and add 20 ml of the nitric

7.1.4 Ammonium heptamolybdate solution, [(NH ) Mo O ·4H O], 50g/l (CAS 12027-67-7)

Dissolve 50g of ammonium heptamolybdate tetrahydrate in about 800 ml of warm water (at approx.

50°C). Cool and transfer quantitatively to a 1000ml volumetric flask. Dilute to the mark with water

Mix one volume of the nitric acid (7.1.2) with one volume of the ammonium metavanadate solution

(7.1.3). Subsequently add one volume of the ammonium heptamolybdate solution (7.1.4) and mix. Please

7.1.6 Potassium dihydrogen phosphate (KH PO ), (CAS: 7778-77-0, >99.99%) is certified by the state

Dissolve in water 479.4 mg of potassium dihydrogen phosphate (KH PO ), previously dried for 3 h at

Transfer quantitatively to a 1000 ml volumetric flask. Dilute to the mark with water and mix.

7.1.7 Phosphate standard solutions, containing between 0.05 mg and 0.30 mg of P O per millilitre.

Transfer by pipette or burette to 100 ml volumetric flasks 10 ml, 20 ml, 30 ml, 40 ml, 50 ml and 60 ml of

the phosphate stock solution (7.1.6). Add 10 ml of the nitric acid (7.1.2). Dilute to the mark with water

Note the range of standard curve can be adjusted according to the use of different concentrations of

Pipette 2 ml of the nitric acid (7.1.2) and 30 ml of the colour reagent (7.1.5) into a 100 ml volumetric

Important: all glassware shall be thoroughly cleaned using a phosphate-free detergent and then rinsed

7.2.1 Mechanical or electrical equipment capable of homogenizing the laboratory sample. This

includes a high speed rotational cutter, or a mincer fitted with a plate with holes not exceeding 4.5 mm in

7.2.4 Spectrometer, capable of being used at a wavelength of 430 nm±2 nm, or a photo-electric

7.2.8 Muffle furnace, with adjustable temperature control, temperature (550±25) °С

For details of this and other apparatus needed for the incineration procedure, see ISO 936.

7.2.12 Mechanical meat mincer, laboratory size, fitted with a plate with holes of diameter not

Homogenize the laboratory sample with the appropriate equipment (7.2.1). Take care that the

temperature of the sample material does not rise above 25°C. If a mincer is used, pass the sample at

Fill a suitable airtight container with the prepared test sample, close the container and store in such

a way that deterioration and change in composition are prevented. Analyse the test sample as soon as

NOTE If it is required to check whether the repeatability requirement (7.8) is met, carry out two single

7.4.1.1 Weigh, to the nearest 0.001 g, about 5 g of the prepared test sample. Carry out the mineralization

of the test portion (7.4.1) by using an incinerator (7.2.8) and the method described in ISO 936. Take up

the resulting ash in 10 ml of the nitric acid (7.1.2) using a stirring rod to aid dissolution. Cover the dish

with a watch glass and heat for 30 min on boiling water bath (7.2.2). Allow to cool and transfer the liquid

quantitatively to a 100 ml volumetric flask (7.2.7). Dilute to the mark with water, mix and filter through

Pipette 20 ml of the clear and colourless filtrate (7.4.1.1) into a 100ml volumetric flask (7.2.7) and add

30 ml of the colour reagent (7.1.5) by pipette. Dilute to the mark with water and mix. Allow to stand for

7.4.2.1 Measure the absorbance at a wavelength of 430 nm ±2 nm in a glass cell (7.2.5) against the

blank solution (7.1.8), using the spectrometer or the photo-electric colorimeter equipped with an

7.4.2.2 Read the phosphorous concentration of the sample solution from the calibration graph

7.5.1 Pipette 20 ml of each phosphate standard solution (7.1.7) into 100 ml volumetric flasks.

Add to these solutions 30 ml of the colour reagent. Dilute to the mark with water to obtain concentrations

of 10μg, 20μg, 30μg, 40μg, 50μg and 60μg of P O per millilitre, respectively. Mix and allow standing for

7.5.3 Plot the measured absorbance values, corrected for the blank value, against the concentrations

of the diluted phosphate standard solutions (7.5.1). Construct the best-fitting straight line through the

plotted points and the origin. The specified minimum correlation coefficient R ≥0.95.