Identification of Pigmented Substances in Black Honey from Leucosceptrum canum: Novel Quinonoids Contribute to Honey Color

Leucosceptrum canum (Lamiaceae) honey (LCH) is a rare and high-value honey with an unusual dark brown color. Its pigments may be related to the purported health benefits of the honey and could serve as indicators of authentic LCH. Here, we purified the pigmented substances contributing to LCH's color and identified them using high-resolution mass spectrometry and nuclear magnetic resonance spectroscopy. The major pigmented substances were 1-[4-(3-carboxy-propylamino)-3,6-dioxo-cyclohexa-1,4-dienyl]-pyrrolidine-2-carboxylic acid (GPBQ), 1-[4-(3-carboxy-propylamino)-3,6-dioxo-cyclohexa-1,4-dienyl]-pyrrolidine-2-carboxylic acid. (GAPBQ), and 2,5-di-(N-prolyl)-para-benzoquinone (DPBQ). According to UHPLC-DAD data recorded at 370 nm, GPBQ, GAPBQ, and DPBQ contents ranged from 2.58 to 5.13, 2.93 to 4.96, and 4.26 to 9.22 mg/kg, respectively. These amino acid-quinone conjugates, found here for the first time, are pigments that can serve as markers to control the quality of LCH.