ISO 18252:2006

Overview

ISO 18252:2006 / IDF 200:2006 defines a routine gas–liquid chromatographic (GLC/GC) method for determining the sterol composition of anhydrous milk fat. The method analyzes sterols directly from the unsaponifiable fraction of extracted milk fat without purification or derivatization. The procedure is optimized for quantitative evaluation of cholesterol (≈98% of milk-fat sterols) and qualitative/quantitative assessment of major phytosterols when vegetable fats are present.

Key topics and technical requirements

• Scope: Routine determination of sterol composition in anhydrous milk fat; validated for samples containing ~28–32% vegetable fat.
• Principle: Addition of 5α‑cholestane as internal standard → saponification with methanolic KOH (2 mol/L) → extraction of the unsaponifiable matter with diethyl ether → analysis by capillary gas chromatography (GLC/GC) with flame ionization detection (FID).
• Standards/reagents: Reference sterols (cholesterol, campesterol, stigmasterol, β‑sitosterol) and prepared standard solutions; use of analytical-grade solvents (n‑hexane, ethanol, methanol) and anhydrous Na2SO4.
• Instrumentation: Fused silica capillary columns, split or on‑column injectors, temperature-programmable oven (up to ~310 °C in examples), carrier gases (He, H2 or N2) and computerized integration.
• Performance and limitations: The method provides repeatability and reproducibility data (interlaboratory precision reported in the standard). Because there is no purification step, the method requires caution for milk‑fat purity verification; suspicious samples should be analyzed using the reference method ISO 12078/IDF 159.
• Safety: Handling flammable solvents and heated GC equipment - comply with relevant laboratory safety rules.

Applications and users

• Who uses it: Food chemistry laboratories, dairy quality control teams, regulatory laboratories, and lipid analysts involved in milk and dairy product testing.
• Practical applications:
  • Quantifying cholesterol in anhydrous milk fat for nutritional labeling and research.
  • Detecting and profiling phytosterols to assess vegetable‑fat adulteration in milk fat blends.
  • Routine quality control of butter, milk powders, ghee and other anhydrous-dairy-fat products.
  • Supporting authenticity and compositional surveillance in dairy supply chains.

Related standards

• ISO 12078 / IDF 159 - reference method for sterol analysis (recommended if purity is in question).
• ISO 14156 / IDF 172 - extraction methods for lipids and liposoluble compounds.
• ISO 3696 - water for analytical laboratory use.
• ISO 707 / IDF 50 - recommended sampling for milk products.

Keywords: ISO 18252, anhydrous milk fat, sterol composition, gas liquid chromatography, GLC, GC, cholesterol, phytosterols, IDF 200, dairy analysis, unsaponifiable matter.