Analysis of cocoa flavanols and procyanidins (DP 1-10) in cocoa-containing ingredients and products by rapid resolution liquid chromatography: single-laboratory validation

J AOAC Int. 2014 Jan-Feb;97(1):166-72. doi: 10.5740/jaoacint.13-034.


Recently, a multilaboratory validation (MLV) of AOAC Official Method 2012.24 for the determination of cocoa flavanols and procyanidins (CF-CP) in cocoa-based ingredients and products determined that the method was robust, reliable, and transferrable. Due to the complexity of the CF-CP molecules, this method required a run time exceeding 1 h to achieve acceptable separations. To address this issue, a rapid resolution normal phase LC method was developed, and a single-laboratory validation (SLV) study conducted. Flavanols and procyanidins with a degree of polymerization (DP) up to 10 were eluted in 15 min using a binary gradient applied to a diol stationary phase, detected using fluorescence detection, and reported as a total sum of DP 1-10. Quantification was achieved using (-)-epicatechin-based relative response factors for DP 2-10. Spike recovery samples and seven different types of cocoa-based samples were analyzed to evaluate the accuracy, precision, LOD, LOQ, and linearity of the method. The within-day precision of the reported content for the samples was 1.15-5.08%, and overall precision was 3.97-13.61%. Spike-recovery experiments demonstrated recoveries of over 98%. The results of this SLV were compared to those previously obtained in the MLV and found to be consistent. The translation to rapid resolution LC allowed for an 80% reduction in analysis time and solvent usage, while retaining the accuracy and reliability of the original method. The savings in both cost and time of this rapid method make it well-suited for routine laboratory use.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biflavonoids / chemistry*
  • Cacao / chemistry*
  • Catechin / chemistry*
  • Chromatography, Liquid / methods*
  • Molecular Structure
  • Proanthocyanidins / chemistry*
  • Reproducibility of Results
  • Time Factors


  • Biflavonoids
  • Proanthocyanidins
  • procyanidin
  • Catechin