Capillary electrophoresis separation of peptide diastereomers that contain methionine sulfoxide by dual cyclodextrin‐crown ether systems†

Abstract

A dual-selector system employing achiral crown ethers in combination with cyclodextrins has been developed for the separation of peptide diastereomers that contain methionine sulfoxide. The combinations of the crown ethers 15-crown-5, 18-crown-6, Kryptofix® 21 and Kryptofix® 22 and β-cyclodextrin, carboxymethyl-β-cyclodextrin, and sulfated β-cyclodextrin were screened at pH 2.5 and pH 8.0 using a 40/50.2 cm, 50 μm id fused-silica capillary and a separation voltage of 25 kV. No diastereomer separation was observed in the sole presence of crown ethers, while only sulfated β-cyclodextrin was able to resolve some peptide diastereomers at pH 8.0. Depending on the amino acid sequence of the peptide and the applied cyclodextrin, the addition of crown ethers, especially the Krpytofix® diaza-crown ethers, resulted in significantly enhanced chiral recognition. Keeping one selector of the dual system constant, increasing concentrations of the second selector resulted in increased peak resolution and analyte migration time for peptide-crown ether-cyclodextrin combinations. The simultaneous diastereomer separation of three structurally related peptides was achieved using the dual selector system.

Screening and identification of DPP-IV inhibitory peptides from deer skin hydrolysates by an integrated approach of LC–MS/MS and in silico analysis

“…Methionine (Met) is a target of oxidizing free radicals in peptides and proteins. Met residues of proteins are susceptible to ROS and are oxidized to methionine sulphoxide under mild oxidizing conditions (Bachi et al, 2013;Stadtman, Moskovitz, & Levine, 2003;Zhu, Heinemann, Schonherr, & Scriba, 2014). Methionine oxidation converts a hydrophobic residue (Met) into a hydrophilic one (Met(O)).…”

Stereospecific capillary electrophoresis assays using pentapeptide substrates for the study of Aspergillus nidulans methionine sulfoxide reductase A and mutant enzymes

“…The separation of the diastereomers of ac‐Lys‐Phe‐Met(O)‐Lys‐Lys‐Dnp could be achieved using the dual selector system developed for peptides in , i.e., 10 mg/mL sulfated β‐CD and 10 mM 18‐crown‐6 in 50 mM Tris buffer, pH 8.0, but at a migration time of more than 30 min. Resolution could also be achieved in the absence of the crown ether.…”

“…The separations of the diastereomers of ac‐Lys‐Asp‐Met(O)‐Asn‐Lys‐Dnp and ac‐Lys‐Asn‐Met(O)‐Asp‐Lys‐Dnp had been achieved in 50 mM Tris buffer, pH 8.0, containing 12 mg/mL sulfated β‐CD and 6 mM Kryptofix ® 22 . However, long migration times of more than 30 min were observed for the internal standard Fmoc‐β‐Ala.…”

Recent developments in capillary and microchip electroseparations of peptides (2013–middle 2015)

“…The enantiomeric and diastereomeric CE separations of peptides were systematically investigated in Scriba's group. In addition to the above (Section ) mentioned CZE separations of isomers of degradation products of aspartyl tetrapeptides, Gly‐Phe‐Asp‐Gly , and Ala‐Phe‐Asp‐Gly , the separations of peptide diastereomers containing methionine sulfoxide were investigated by CZE with a dual‐selector system composed of achiral crown ethers (15‐crown‐5, 18‐crown‐6, Kryptofix 21®, Kryptofix 22® and CDs (β‐CD, carboxymethyl‐β‐CD, and sulfated β‐CD) at pH 2.5 and 8.0 . No separation was observed in the sole presence of crown ethers but the addition of some of them, especially the Kryptofix® diaza‐crown ethers, resulted in significantly enhaced chiral recognition, see Fig.…”

“…Buffer additives are (A) 12 mg/mL sulfated β‐CD, (B) 12 mg/mL sulfated β‐CD plus 25 mM 18C6, (C) 12 mg/mL sulfated β‐CD plus 6 mM Kryptofix ® 22, (D) 12 mg/mL sulfated β‐CD, 25 mM 18‐crown‐6, plus 15% acetonitrile. Reprinted with permission from .…”