International Oenological CODEX

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Benzo[a]pyrene- Determination

COEI-2-HYDCAR Aromatic polycyclic hydrocarbons

Determination of benzo(a)pyrene in oenological charbons by HPLC

  1. Principle

Polycyclic aromatic hydrocarbons including benzoapyrene are extracted by hexane; the solvent is evaporated and the residue is taken up by the methanol-tetrahydrofuran for analysis by HPLC.

  1. Apparatus and reagents

2.1.    Reagents and calibrations

  • Acetonitrile for HPLC
  • Hexane for pesticide residues
  • Tetrahydrofuran for HPLC (THF)
  • Deionised and microfiltered water
  • Benzo[a]pyrene for HPLC.
    1.     Apparatus and chromatographic conditions
  • octadecyl type HPLC column
  • fluorimetric detector adjusted to the following detection conditions:
  • excitation wave length: 300 nm,
  • emission wave length: 416 nm.

Mobile phase:

  • solvent A: Deionised and microfiltered water
  • solvent B: acetonitrile

Variations in the composition of the solvent

 

TIME

in min

% solvent A

% solvent B

0

50

50

15

20

80

40

0

100

45

50

50

Flow 1.0 ml/mn

2.3.    Preparation of reference solutions

Benzoapyrene reference solution at about 100 mg/l in a methanol/THF mixture (50/50) stored for 3 years maximum in cold conditions.

Daughter solution at about 20 μg/l, prepared extemporaneous (0.5 ml of reference solution in 50 ml of methanol/THF then 1 ml of this intermediate solution in 50 ml de methanol/THF).

2.4.    Preparation of samples

2 g of oenological charbon are mixed in a 50 ml volumetric flask with 30 ml of hexane.

The polycyclic aromatic hydrocarbons are extracted for 5 min using a magnetic stirrer. The organic phase recovered by filtration is gathered in a evaporating flask and evaporated. The extract is taken up by 2 ml of a methanol/THF mixture (1/1, v/v) and injected.

  1. Results

 

The benzoapyrene content must not be higher than 1 μg/kg.

Remark: It is also possible to determine benzo[a]pyrene by chromatography in gaseous phase by an apolar capillary column with detection by mass spectrometry.