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BS EN 17270:2019

BS EN 17270:2019

$142.49

Animal feeding stuffs: Methods of sampling and analysis. Determination of theobromine in feed materials and compound feed, including cocoa derived ingredients, by liquid chromatography

Published By Publication Date Number of Pages
BSI 2019 26
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This document specifies a test method for the determination of theobromine in feed material or compound feed in the working range 27 mg/kg to 307 mg/kg using liquid chromatography coupled to a UV detector (HPLC-UV) or in the working range 49 mg/kg to 307 mg/kg using liquid chromatography with tandem mass spectrometry (LC-MS/MS).

This method has been fully validated using complementary compound feed for adult dogs and complementary compound feed for horses.

This method is also considered applicable for determining theobromine in baking chocolate using either HPLC-UV or LC-MS/MS systems.

The working range can be extended provided the extended range is validated.

PDF Catalog

PDF Pages PDF Title
2 undefined
7 1 Scope
2 Normative references
3 Terms and definitions
4 Principle
5 Reagents
9 6 Apparatus
10 7 Procedure
7.1 Sample preparation
7.2 Extraction
11 8 Chromatographic analysis
9 Calculation of results
12 10 Accuracy
11 Test report
13 Annex A (informative)Example HPLC-UV conditions
A.1 HPLC-UV analysis
A.1.1 LC system
A.1.2 UV conditions
A.1.3 Batch composition
A.1.4 Peak identification
A.2 Reagents
A.2.1 Orthophosphoric acid, Analytical reagent grade
A.2.2 Potassium dihydrogen phosphate, Analytical reagent grade
A.2.3 0,0125 mol/l potassium dihydrogen phosphate, pH 3,5
A.2.4 Acetonitrile, HPLC grade
A.2.5 90 % 0,0125 mol/l potassium dihydrogen phosphate, pH 3,5, 10 % acetonitrile
14 A.3 Apparatus
A.3.1 HPLC with UV detector
A.3.1.1 Solvent delivery system, capable of delivering a flow rate appropriate for the analytical column in use.
A.3.1.2 Degasser, optional, for in-line degassing LC mobile phases
A.3.1.3 Autosampler, capable of injecting an appropriate volume with sufficient accuracy
A.3.1.4 Column oven, capable of maintaining the column at a suitable temperature, for example 30 °C
A.3.1.5 Analytical column, capable of separating the analytes of interest
A.3.1.6 UV detector, capable of measuring at 278 nm
A.4 HPLC conditions
15 A.5 Example chromatogram
16 Annex B (informative)Example LC-MS/MS conditions
B.1 General
B.2 LC-MS/MS analysis
B.2.1 LC system
B.2.2 MS conditions
B.2.3 Batch composition
B.2.4 Peak identification
17 B.3 Reagents
B.3.1 Ammonium formate, ≥ 99,0 %
B.3.2 Methanol, high purity
B.3.3 Ammonium formate 12 mmol/l in water
B.3.4 Ammonium formate 12 mmol/l in methanol
B.3.5 Theobromine-d3 (7-methyl-d3), 99,9 % isotopic purity (Optional)
B.3.6 Theobromine –d3 stock solution, 200 µg/ml (Optional)
B.4 Apparatus
18 B.5 HPLC conditions
B.6 MS conditions
19 B.7 Example chromatograms
22 Annex C (informative)Method performance data obtained during single laboratory validation
23 Annex D (informative)Method performance data obtained during collaborative trial

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BS EN 17270:2019
$142.49