Overview
ISO 15153:2003 specifies a laboratory method for determining the content of reducing substances in tobacco using a continuous-flow analysis technique. Applicable to both manufactured and unmanufactured tobacco, the method measures reducing substances expressed as glucose equivalents after extraction in 5% acetic acid and colorimetric detection at 420 nm. ISO 15153 is intended for routine tobacco analysis, quality control and comparative studies.
Key topics and technical requirements
- Scope: Applicable to all tobacco types (manufactured and unmanufactured).
- Analytical principle: Reduction of yellow hexacyanoiron(III) to colourless hexacyanoiron(II); decrease in color measured at 420 nm using a continuous-flow analyser.
- Sample preparation:
- Grind sample to pass a 1 mm sieve; determine moisture content.
- Typical test portion: ~250 mg tobacco extracted with 25 ml of 5% acetic acid, shaken 30 minutes and filtered.
- Reagents: Includes Brij 35 (polyoxyethylene lauryl ether), NaCl solution, 1 mol/L NaOH, alkaline potassium ferricyanide solution, and D-glucose for standards.
- Standards and calibration:
- Stock glucose solution (~10 mg/ml) and a series of working standards (typical range e.g. 0.2–2.5 mg/ml) for calibration.
- Plot peak height vs. glucose concentration to obtain sample concentration (c, mg/ml).
- Calculation: Convert measured concentration to percentage of reducing substances (expressed as glucose) on a dry-weight basis using the sample mass, extract volume and measured moisture content. Results are reported to one decimal place.
- Equipment: Continuous-flow analyser components (sampler, proportioning pump, dialyser, heating bath, delay coils, 420 nm colorimeter and recorder). Annex A provides an example layout.
- Performance: International collaborative data (12 labs) provide repeatability and reproducibility examples (e.g., Burley mean 2.3%: r = 0.7, R = 1.7; Oriental mean 15.1%: r = 1.2, R = 2.5).
Applications and users
- Who uses it: Tobacco manufacturers, analytical and QA laboratories, regulatory agencies, research institutions and standardization bodies.
- Practical uses:
- Routine quality control and batch verification of tobacco leaf and products.
- Product characterization and comparative analysis of tobacco types (Burley, Oriental, flue-cured).
- Research into tobacco composition, processing effects, and formulation decisions where reducing substances (sugars and other reducing compounds) are critical.
- Support for regulatory reporting and inter-laboratory studies.
Related standards
- ISO 15152 - Determination of total alkaloids (nicotine) by continuous-flow analysis
- ISO 15154 - Reducing carbohydrates - continuous-flow analysis method
- ISO 15517 - Determination of nitrate content - continuous-flow analysis method
Keywords: ISO 15153, reducing substances, tobacco analysis, continuous-flow analysis, continuous flow analyser, reducing sugars, glucose equivalents, tobacco quality control.