Analyzing Citramalic Acid Enantiomers in Apples and Commercial Fruit Juice by Liquid Chromatography-Tandem Mass Spectrometry with Pre-Column Derivatization

Molecules. 2023 Feb 6;28(4):1556. doi: 10.3390/molecules28041556.


Optically active citramalic acid (CMA) is naturally present as an acidic taste component in fruits, such as apples. The absolute configuration of CMA in such fruits was investigated by high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) following pre-column derivatization with a chiral reagent, benzyl 5-(2-aminoethyl)-3-methyl-4-oxoimidazolidine-1-carboxylate. The developed LC-MS/MS method successfully separated the enantiomers of CMA using an octadecylsilica column with a resolution and separation factor of 2.19 and 1.09, respectively. Consequently, the R-form of CMA was detected in the peel and fruit of three kinds of apple at concentrations in the 1.24-37.8 and 0.138-1.033 mg/wet 100 g ranges, respectively. In addition, R- CMA was present in commercial apple juice, whereas no quantity was detected in commercial blueberry, perilla, or Japanese apricot juice.

Keywords: benzyl 5-(2-aminoethyl)-3-methyl-4-oxoimidazolidine-1-carboxylate (CIM-C2-NH2); citramalic acid; high-performance liquid chromatography–tandem mass spectrometry (LC–MS/MS).

MeSH terms

  • Chromatography, High Pressure Liquid / methods
  • Chromatography, Liquid / methods
  • Fruit / chemistry
  • Fruit and Vegetable Juices
  • Malus* / chemistry
  • Stereoisomerism
  • Tandem Mass Spectrometry / methods


  • citramalate

Grants and funding

This research received no external funding.