Chemical detection of cysteine-rich circular petides in selected tropical Violaceae and Moringaceae families using modified G-250 and mass spectrometry

Abstract

Cysteine-rich circular peptides (CRCs) comprise a large family of gene encoded and low molecular weight polypeptides that has recently engaged the attention of scientists. This class of peptides exhibit a continuous circular configuration and a cystine knot backbone, which defines their resilient nature-directed structural design. Many CRCs have been reported in medicinal plants the first of which is kalata B1 cyclotide from the traditional African plant Oldenlandia affinis. Their detection and isolation can be very challenging and evasive. Only about 1% of plant species have so far been reportedly screened. A modified preliminary chemo-microscopic/macroscopic method involving the use of G-250 stain was applied followed by thin layer chromatographic protosite reaction for plant selection. This was followed by the Matrix Assisted Laser Desorption Time of Flight (MALDI-TOF) Mass spectrometry guided experiment for cyclotide discovery. The blue colouration produced upon sample reaction with modified G-250 led to the selection of potential circular peptide containing plant samples. A further MALDI-TOF MS-guided screening resulted in the detection of circular peptides and cyclotides in Moringa oleifera, Rinorea dentata, R. oblongifolia and R. brachypatela. Viola odorata and Viola tricolor (positive controls) indicated the presence of cyclotides. Results from this study can serve as proof-of-concept for plant selection based on preliminary cysteine-rich circular peptide detection in plants especially with the use of G-250 stain.

Keywords: Cysteine-rich Cyclotides, Violaceae, Moringaceae, TLC, Microscopy, MALDI-TOF MS