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BS EN 17270:2019 Animal feeding stuffs: Methods of sampling and analysis. Determination of theobromine in feed materials and compound feed, including cocoa derived ingredients, by liquid chromatography, 2019
- undefined
- 1 Scope
- 2 Normative references
- 3 Terms and definitions
- 4 Principle
- 5 Reagents
- 6 Apparatus
- 7 Procedure [Go to Page]
- 7.1 Sample preparation
- 7.2 Extraction
- 8 Chromatographic analysis
- 9 Calculation of results
- 10 Accuracy
- 11 Test report
- Annex A (informative)Example HPLC-UV conditions [Go to Page]
- A.1 HPLC-UV analysis [Go to Page]
- A.1.1 LC system
- A.1.2 UV conditions
- A.1.3 Batch composition
- A.1.4 Peak identification
- A.2 Reagents [Go to Page]
- 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
- A.3 Apparatus [Go to Page]
- A.3.1 HPLC with UV detector [Go to Page]
- 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
- A.5 Example chromatogram
- Annex B (informative)Example LC-MS/MS conditions [Go to Page]
- B.1 General
- B.2 LC-MS/MS analysis [Go to Page]
- B.2.1 LC system
- B.2.2 MS conditions
- B.2.3 Batch composition
- B.2.4 Peak identification
- B.3 Reagents [Go to Page]
- 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
- B.5 HPLC conditions
- B.6 MS conditions
- B.7 Example chromatograms
- Annex C (informative)Method performance data obtained during single laboratory validation
- Annex D (informative)Method performance data obtained during collaborative trial [Go to Page]