Commission Regulation (EEC) No 2568/91Show full title

[F1 6.3. Gas chromatography U.K.

6.3.1. Working conditions for split injection: U.K.

• injector temperature: 300 °C,

• detector temperature: 320 °C,

• integrator-recorder: the parameters for integration should be fixed so as to give a correct assessment of the areas. Valley-valley integration mode is recommended,

• sensitivity: about 16 times the minimum attenuation,

• amount of solution injected: 1μl,

• oven programming temperatures: initial 235 °C for six minutes and then rising at 2 °C/minute up to 285 °C,

• injector with 1: 15 flow divider,

• carrier: helium or hydrogen at about 120 kPa pressure.

These conditions may be adjusted in accordance with the characteristics of the chromatograph and the column to give chromatograms meeting the following requirements: internal standard peak within approximately five minutes of the time given in 6.3.2; the internal standard peak should be at least 80 % of the full scale.

The gas chromatographic system must be checked injecting a mixture of the stock solution of cholestadiene (5.6) and n-nonacosane solution (5.8). The cholesta-3,5-diene peak must appear before the n-nonacosane (Figure 1c); if it does not occur two actions can be undertaken: reduce the oven temperature and/or use a less polar column.

6.3.2. Peak identification U.K.

The internal standard peak appears at approximately 19 minutes and the 3,5-stigmastadiene at a relative retention time of approximately 1,29 (see Figure 1b). The 3,5-stigmastadiene occurs with small quantities of an isomer, and usually, both elute together as a single chromatographic peak. Nevertheless, if the column is too polar or shows a high resolving power, the isomer can appear as a small peak before and close to that of stigmasta-3,5-diene (Figure 2). In order to ensure that the stigmastadienes are eluted as one peak, it is advisable to replace the column by one which is either less polar or has a wider internal diameter.

Note 9: U.K.

Stigmastadienes for reference can be obtained from the analysis of a refined vegetable oil by using less amount of sample (1 to 2 g). Stigmastadienes originate a prominent and easily identifiable peak. U.K.

6.3.3. Quantitative analysis U.K.

The stigmastadienes content is determined according to the formula:

mg/kg of stigmastadienes =

where:

A s = area of stigmastadienes peak (if the peak is resolved into two isomers, sum of areas of the two peaks), A c = area of internal standard (cholestadiene), M c = mass of standard added, in micrograms, M o = mass of oil taken, in grams.

Detection limit: about 0,01 mg/kg.

[F2 Note 10: When stigmastadienes appear in concentrations of more than 4 mg/kg, if quantifying is required, the method of the International Olive Council for determination of sterenes in refined oil must be applied.] ]

Textual Amendments

F2 Inserted by Commission Implementing Regulation (EU) 2019/1604 of 27 September 2019 amending Regulation (EEC) No 2568/91 on the characteristics of olive oil and olive-residue oil and on the relevant methods of analysis.

Textual Amendments

F1 Inserted by Commission Regulation (EC) No 656/95 of 28 March 1995 amending Regulation (EEC) No 2568/91 on the characteristics of olive oil and olive-residue oil and on the relevant methods of analysis and Council Regulation (EEC) No 2658/87 on the tariff and statistical nomenclature and on the Common Customs Tariff.