BS ISO 17780:2015 pdf download – Animal and vegetable fats and oils — Determination of aliphatic hydrocarbons in vegetable oils
8 Preparation of the test sample
Prepare the test sample in accordance with ISO 661.
9.1 Chromatography column preparation
9.1.1 Preparation of AgNO 3 impregnated silica gel Preparation of the silver nitrate silica gel column (for 3 columns): weigh 45 g of silica gel (5.1) in a 500 ml round-bottomed flask (6.3) protected by aluminium foil. With a Pasteur pipette (6.12), add drop by drop the silver nitrate solution (5.12) shaking continuously. Shake well for 30 min to homogenize. After completing, cover the flask with aluminium foil and allow to stand at room temperature for 12 h before use. In order to improve homogenization, it is recommended to use an automatic shaker. If no automatic shaker is available, it is possible to put the flask in a rotatory evaporator equipment and rotate for 30 min without vacuum.
NOTE 1 The impregnated silica gel can be stored one week at room temperature in a desiccator, provided the flask is protected with aluminium foil. NOTE 2 For screening purposes, AgNO 3 impregnated silica gel can be replaced by non-impregnated silica gel. The results will be similar or higher than those obtained using silvered silica gel. NOTE 3 For refined vegetable oils other than refined olive pomace oil, AgNO 3 impregnated silica gel can be replaced by non-impregnated silica gel.
9.1.2 Column packing In a beaker, suspend 18,5 g of silver nitrate impregnated silica gel (9.1.1) in n-hexane (5.4). The slurry is introduced onto the column (6.1) containing 40 ml of n-hexane and pack the column by tapping it gently using a glass rod (6.2). Add at least 0,5 cm to 1 cm of sodium sulfate (5.3) on top of the AgNO 3 – silica gel, and compress the AgNO 3 – silica gel bed with a stream of nitrogen. Rinse the AgNO 3 – silica gel with another 60 ml of n-hexane (5.4) to eliminate impurities in the AgNO 3 – silica gel. The column should be covered with a black paper cylinder or with aluminium foil to avoid oxidation of the silver nitrate. Elute the solvent until the level of the solvent in the column is about 0,5 cm higher than the AgNO 3 – silica gel bed. Put a 250 ml round-bottomed flask (6.3) under the chromatography column.
9.2 Elution of the hydrocarbon fraction Weigh to the nearest 1 mg, 1 g of the sample in a beaker and add 1 ml of the solution of the internal standard (5.6), transfer the solution to the chromatographic column (9.1.2) with the aid of a Pasteur pipette (6.12) and let the sample penetrate into the stationary phase. Wash the beaker with two portions of 1 ml of n-hexane (5.4) and introduce the solution into the column. Elute the hydrocarbon fraction with 55 ml of n-hexane (5.4) with a cadence of approximately 15 drops every 10 s, collecting the fraction in a 250 ml flask (6.3). Evaporate most of the solvent up to 1 ml or 2 ml with a rotary evaporator equipped with a water bath set to 35 °C (6.4). Transfer the concentrated solution to a 10 ml conical tube. Concentrate the solvent up to 0,5 ml from the conical tube under a stream of nitrogen, using either a water bath at 35 °C or an automatic evaporator (6.5). Take care that, in both evaporation steps, the residue is not evaporated to dryness to avoid loss of the volatile alkanes.
Adjust the elution volume of n-hexane by the analysis of 1 ml solution of paraffin (5.17) and collecting consecutive fractions of 50 ml, 10 ml, and 10 ml, and analysing each one by gas chromatography (GC).
9.3 Gas chromatography
9.3.1 Gas chromatography setup
Install the column (6.9) in the gas chromatograph (6.7) and check the working conditions by injecting the solvent, n-hexane (5.4). The baseline should be straight with a small positive drift. If the drift is high,proceed to condition the column, for a negative drift check the connections of the column.BS ISO 17780 pdf download.