ISO 8128-2:1993

Overview

ISO 8128-2:1993 specifies a standardized method for determining the patulin content in apple juice, apple juice concentrates, and drinks containing apple juice using thin-layer chromatography (TLC). Patulin is a mycotoxin commonly found in fruit products, particularly apple-derived beverages, which poses health risks if consumed in excessive amounts. This internationally recognized method ensures consistent, reliable detection and quantification of patulin, supporting food safety and regulatory compliance. The method involves extraction of patulin with a mixture of ethyl acetate and chloroform, purification via silica-gel column chromatography, and qualitative and quantitative analysis by two-directional TLC with MBTH reagent development.

Key Topics

• Scope and Purpose
  Designed for routine patulin analysis in apple juices and related products, the method enables detection down to 25 µg/l based on a 50 ml sample, suitable for screening purposes.

• Analytical Principle

  • Extraction of patulin using a 3:2 volume mixture of ethyl acetate and chloroform.
  • Purification through silica-gel column chromatography.
  • Separation using bidirectional thin-layer chromatography.
  • Visualization by spraying with 3-methyl-2-benzothiazoline hydrazone (MBTH) hydrochloride solution, yielding a detectable fluorescent spot under UV light.

• Reagents and Equipment

  • High-purity solvents including ethyl acetate, chloroform, benzene, methanol, toluene, and acetic/formic acids for mobile phases.
  • Silica gel for column chromatography and TLC plates (20 cm x 20 cm) with a 0.25 mm silica layer.
  • Specific apparatus such as chromatographic columns, UV lamps, and a thermostatically controlled drying oven.

• Performance Characteristics

  • Limit of detection: 25 µg/l patulin in ready-to-drink apple juice.
  • Repeatability and reproducibility are quantified with statistical values to ensure reliability of results.
  • The method provides good separation of patulin from interfering substances like hydroxymethylfurfural (HMF).

• Operational Procedure
  Includes detailed steps for sample preparation, extraction, column purification, TLC development, staining, and result interpretation using fluorescence intensity comparison with patulin standards.

Applications

• Food Safety Testing
  Ensures compliance with regulatory limits for patulin in apple juice products, helping manufacturers prevent contaminated batches from reaching consumers.

• Quality Control in Juice Production
  Facilitates routine monitoring of patulin levels during processing and formulation to maintain product safety and quality.

• Research and Analytical Laboratories
  Provides a validated reference method for laboratories conducting patulin analysis in line with international standards.

• Regulatory and Certification Bodies
  Supports enforcement of food safety regulations by providing a standardized, internationally accepted analytical approach.

Related Standards

• ISO 8128-1:1993
  Specifies an alternative, more precise method for patulin determination using high-performance liquid chromatography (HPLC), suitable for confirmatory analysis or legal disputes.

• ISO 5725
  Details general standards for the accuracy (repeatability and reproducibility) of measurement methods, applicable to validating patulin testing procedures.

• AOAC Official Methods of Analysis
  Includes reference procedures for patulin determination supporting standardized laboratory practices.

Keywords: ISO 8128-2, patulin detection, apple juice safety, thin-layer chromatography, TLC method, mycotoxin analysis, apple juice concentrates, beverage testing, food contaminant detection, chromatographic patulin analysis.