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