Overview
ISO 7358:2021 specifies an HPLC method for the determination of bergapten in essential oils of bergamot, lemon, bitter orange and lime that are fully or partially reduced in bergapten. The standard describes both normal‑phase and reversed‑phase HPLC approaches and permits either an internal standard (coumarin) or an external standard calibration for quantitative analysis, using diode‑array UV detection.
Key topics and technical requirements
- Scope: Quantitative determination of bergapten (5‑methoxypsoralen) in specified Citrus essential oils reduced in bergapten.
- Methods:
- Normal‑phase HPLC with silica column and non‑polar mobile phases (examples: hexane/ethyl acetate or hexane/chloroform).
- Reversed‑phase HPLC using a C18 column and a ternary mobile phase (water/acetonitrile/methanol) with a defined dilution solvent.
- Calibration options:
- Internal standard method using coumarin (recommended purity ≥98 %) and calculation of a response factor (K).
- External standard method following ISO 8432 procedures.
- Reagents and purity: Bergapten reference substance (≥95 % purity) and analytical/HPLC‑grade solvents (chloroform, hexane, ethyl acetate, acetonitrile, methanol, water).
- Instrumentation: Typical HPLC system with appropriate columns (silica or C18), ternary pump for gradients (reversed phase), 0.45 µm PTFE filtration, and diode‑array UV detector adjustable to 254 nm or 312 nm.
- Sample preparation: Follow ISO 356 for test sample preparation; dissolve any solid deposit by moderate heating and filter before injection.
- Performance and reporting: Procedure includes operating conditions, calculation formulas (internal/external), typical chromatogram and UV‑Vis spectrum in annexes, and statements on precision/repeatability.
Applications and users
ISO 7358:2021 is intended for:
- Quality control and routine testing laboratories in the flavour & fragrance and essential oil industries.
- Manufacturers of bergamot, lemon, bitter orange and lime essential oils for batch release and supplier verification.
- Analytical service labs conducting bergapten content assays for product safety and composition analysis (e.g., assessing phototoxic furanocoumarin residues).
- Regulatory and conformity assessment bodies requiring standardized, reproducible HPLC methods.
Practical uses include verifying reduction processes (fully or partially reduced bergapten), ensuring product consistency, and generating reliable quantitative data for safety assessments and labeling.
Related standards
- ISO 356 - Essential oils - Preparation of test samples (referenced for sample prep).
- ISO 8432 - Essential oils - Analysis by HPLC - General method (referenced for HPLC procedures).
- Prepared by ISO/TC 54 (Essential oils); second edition adds the reversed‑phase alternative.
Keywords: ISO 7358:2021, bergapten, HPLC, essential oils, bergamot, lemon, bitter orange, lime, coumarin, internal standard, reversed phase, normal phase, diode-array UV.