BS ISO 16632:2021 pdf download – Tobacco and tobacco products — Determination of water content — Gas-chromatographic method
Weigh (5,0 ± 0,25) g of the sample (8.1) into the dry extraction vessel (6.1). Record the weight to the nearest 0,000 1 g.
It is recommended that a minimum of two test portions be prepared and analysed for each test sample. The recommended procedure for portioned products such as snus is to analyse the entire portion by cutting the pouch in half and adding the tobacco and pouch material to the extraction vessel. Pipette 100,0 ml of extraction solution (5.5) into the extraction vessel and immediately seal the vessel. Place the extraction vessel in the shaker (6.2) and shake for 3 h. Remove the extraction vessel from the shaker and set it aside overnight.
The test portions should be gently swirled or mixed mechanically prior to removal of the analysis aliquot. Assemble a disposable syringe (6.3) with a 0,45 µm filter (6.3). Carefully transfer about 5 ml of the supernatant liquid into the disposable filtration assembly. Purge the filter of adsorbed water by disposing of a small volume of the extract. Filter the extract into a 2 ml GC injection vial and cap the vial. Store the filtered extract in a refrigerator below 4 °C until GC analysis, making certain of tight seals. If the extract is not analysed on the same day, store in a refrigerator.
The sample shall be allowed to equilibrate to ambient conditions prior to analysis. 8.3 Setting up the apparatus Set up the apparatus and operate the gas chromatograph (6.5) in accordance with the manufacturer’s instructions. Ensure that the peaks for water, internal standard and solvent are well resolved. Condition the system just prior to use by injecting two 0,5 µl aliquots of the extraction solution as a primer.
Suitable operating conditions are as follows:
— carrier gas: helium;
— linear velocity: 30 cm/s at 50 °C;
— injection temperature: 250 °C;
— injection liner: appropriate liner packed with glass wool;
— injection mode: splitless (split valve closed during injection, to be opened after about 1 min);
— injection volume: 0,5 µl;
— initial temperature: 60 °C;
— initial hold time: 0 min;
— temperature ramp A: 5 °C/min;
— final temperature A: 130 °C;
— final hold time A: 0 min;
— temperature ramp B: 10 °C/min;
— final temperature B: 170 °C;
— final hold time B: 5 min;
— total analysis time: 23,00 min;
— detector: 250 °C.
Optimize the GC conditions for analyte separation and sensitivity. Once optimized, the same GC conditions shall be used for the analysis of all standards and samples, including the same injection volume of 0,5 µl. An adjustment to the chromatographic conditions may be required depending on the instrument configuration and columns chosen for separation. NOTE High boiling point components can accumulate in the column. Typically, increasing the oven temperature to 220 °C for 20 min has been found to be sufficient to avoid carry over to the next analysis.
8.4 Calibration of the gas chromatograph
8.4.1 Procedure Inject an aliquot (0,5 µl) of each of the calibration solutions (5.7) into the gas chromatograph. Record the peak areas (or heights) of the water and internal standard (5.3). Calculate the ratio of the water peak to the internal standard peak (Y i = A H2O /A IS ) from the peak area (or height) data for each of the calibration solutions including the solvent blanks. Plot the graph of the concentrations of added water (X-axis) in accordance with the area ratios (Y-axis). Calculate a linear regression equation (y = a + bx) from these data and use both the slope and the intercept of the linear regression equation. If the correlation coefficient R 2 is less than 0,99, the calibration should be repeated. The signal (peak area or height) obtained for all test portions must fall within the working range of the calibration curve.
Perform this full calibration procedure daily. In addition, inject an aliquot of an intermediate concentration standard after every 20 sample determinations. If the calculated concentration for this solution differs by more than 3 % from the original value, repeat the full calibration procedure. Perform the full calibration if a new extraction solution is made. The calibration curve should include the solvent blank (i.e. extraction solution) (see 5.7).
NOTE The regression line does not pass through zero due to water present in the extraction solution. If the water content of the extraction solution exceeds 1,0 mg/ml, the batch should be rejected.
8.4.2 Blank test Because of the absorption of water by the extraction solution, create duplicate blanks per set of test samples exactly as the test portions, including shaking, filtering and transferring to injection vials.
8.4.3 Determination Inject aliquots (0,5 µl) of the test portion (see 8.2) from the sample extracts. Calculate the ratio of the water peak/internal standard peak (Y t ) from the peak area (or height) data. Calculate the mass concentration for each test portion aliquot using the coefficients of the linear regression [r t = (Y t − a)/b]. Calculate the mean value of the ratio from the replicate determinations. Injection volume may be optimised depending upon the column specification and chromatographic conditions.