Enantioseparation and quantitative determination of two homologous beta amino acids found in Fabaceae plants

Nóra Gampe; Lilla Ladocsi; Ida Fejős; Imre Boldizsár; András Darcsi; Szabolcs Béni

doi:10.1016/j.chroma.2022.463089

Enantioseparation and quantitative determination of two homologous beta amino acids found in Fabaceae plants

J Chromatogr A. 2022 Jul 19:1675:463089. doi: 10.1016/j.chroma.2022.463089. Epub 2022 May 7.

Authors

Nóra Gampe¹, Lilla Ladocsi¹, Ida Fejős¹, Imre Boldizsár², András Darcsi¹, Szabolcs Béni³

Affiliations

¹ Semmelweis University, Department of Pharmacognosy, Budapest, Hungary.
² Semmelweis University, Department of Pharmacognosy, Budapest, Hungary; Eötvös Loránd University, Department of Plant Anatomy, Institute of Biology, Bioactive Compounds Group, Institutional Excellence Program, Budapest, Hungary.
³ Semmelweis University, Department of Pharmacognosy, Budapest, Hungary. Electronic address: beni.szabolcs@pharma.semmelweis-univ.hu.

PMID: 35613505
DOI: 10.1016/j.chroma.2022.463089

Abstract

Non-protein amino acids are important metabolites of the Fabaceae family, possessing valuable biological effects in addition to their toxic properties. We have previously identified two non-protein amino acids homoproline and homopipecolic acid in Ononis species for the first time, and herein the study was extended to investigate further Fabaceae species (O. spinosa, O. arvensis, M. sativa, A. vulneraria) with medicinal, food or cosmetic uses. As the enantiomers of these beta amino acids can carry different activity or toxicity, our aim was to develop a chiral separation method for homoproline and homopipecolic acid enantiomers and apply it to plant samples. For this purpose, dansylated derivatives were prepared and a cyclodextrin-modified capillary electrophoresis in addition to a chiral HPLC method were developed. Although baseline separation was achieved by CE applying mono-(6-N-pyrrolidine-6-deoxy)-β-CD, mono-(6-N-piperidine-6-deoxy)-β-CD or sulfated-gamma-cyclodextrin at pH 6.0, the HPLC method was found to be more suitable for the analysis of the plant samples. Both homoproline and homopipecolic acid were confirmed in plant samples as racemates. The quantitative determination of homoproline and homopipecolic acid in several Fabaceae species were also aimed. Since these molecules can be found in the plants as esters, sample preparation was optimized to liberate the target molecules. Several SPE methods were tested for sample purification and a HPLC-MS/MS method using C8 stationary phase was developed and validated. The presence of homoproline and homopipecolic acid could be confirmed in all species ranging from 1 µg/g through 500 µg/g homopipecolic acid and 6 µg/g to 60 µg/g homoproline and significant changes could be observed between species, geographical origins, and botanical parts. Generally O. spinosa root samples were found to be the richest sources of the two amino acids, but a high variance could be observed between species.

Keywords: Capillary electrophoresis; Chiral HPLC; Derivatization; Homopipecolic acid; Homoproline.

MeSH terms

Amino Acids* / chemistry
Electrophoresis, Capillary / methods
Fabaceae*
Stereoisomerism
Tandem Mass Spectrometry

Substances

Amino Acids