Overview
ISO 15517:2003 - Tobacco - Determination of nitrate content - Continuous-flow analysis method specifies a validated laboratory procedure for measuring nitrate content in both manufactured and unmanufactured tobacco using a continuous-flow analyser. The method uses chemical reduction and colorimetric detection to report nitrate on a dry-weight basis and is suitable for routine quality control, regulatory testing and research.
Key topics and technical requirements
- Principle: Nitrate in an aqueous (or 5% acetic acid) tobacco extract is chemically reduced to nitrite (hydrazinium sulfate + copper catalyst) and reacted with sulfanilamide and N‑1‑naphthyl‑ethylenediamine dihydrochloride to form a coloured azo complex measured at 520 nm by a colorimeter.
- Sample preparation: Grind tobacco to pass a 1 mm sieve, determine moisture, weigh a test portion (~250 mg), extract (typically 25 ml water), shake 30 min and filter prior to analysis.
- Reagents and standards: Specifies hydrazinium sulfate/copper reagent, sulfanilamide/NED colour reagent, potassium nitrate primary standard and working standards (covering expected range, e.g., ~10–200 µg/ml nitrate). When extracting with 5% acetic acid, standards and wash cycles must use 5% acetic acid.
- Apparatus: Continuous-flow analyser with sampler, dialyser, heating bath, delay coils, 520 nm colorimeter (flow cell) and recorder. Annex B provides an example analyser layout.
- Optimization and quality control: Includes procedure to optimise hydrazinium sulfate concentration to ensure complete nitrate → nitrite reduction without over-reduction of nitrite to nitrogen. Calibration with at least five standards and routine priming/wash cycles are required.
- Calculation and reporting: Result expressed as percentage nitrate on a dry-weight basis using the measured concentration, extract volume, sample mass and moisture content; ISO gives the explicit calculation method. Results reported to two decimal places.
- Precision: Results from an international collaborative study (12 labs) give repeatability and reproducibility limits for common tobacco types (examples: flue-cured ~0.11% nitrate, r ≈ 0.03% and R ≈ 0.12% for water extraction).
Applications
- Routine tobacco nitrate determination in manufacturing QC laboratories.
- Regulatory compliance testing and inter-laboratory comparisons.
- Research on tobacco chemistry, agricultural practices and combustion/smoke formation studies where nitrate levels are relevant.
- Method integration where nitrate and reducing-substance or reducing-carbohydrate assays are performed simultaneously (5% acetic acid extraction recommended).
Who should use this standard
- Tobacco industry analytical laboratories
- Regulatory and public health testing labs
- Independent testing services and research institutions
- Manufacturers of continuous-flow analysers and analytical reagents
Related standards
- ISO 15152 - Nicotine by continuous-flow analysis
- ISO 15153 - Reducing substances by continuous-flow analysis
- ISO 15154 - Reducing carbohydrates by continuous-flow analysis
Keywords: ISO 15517:2003, tobacco nitrate determination, continuous-flow analysis, nitrate content in tobacco, continuous flow analyser, colorimetric detection, tobacco testing.