1H high-resolution magic-angle spinning (HR-MAS) NMR analysis of ligand density on resins using a resin internal standard.
We recently attempted to generate an affinity chromatography adsorbent to purify cytochrome P450 4A1 by coupling 11-(1'-imidazolyl)-3,6,9-trioxaundecanoic acid to Toyopearl AF-Amino 650 M resin. Variations in ligand density for several resin batches were quantified by high-resolution magic-angle spinning (HR-MAS) NMR spectroscopy using a novel resin internal standard. The uniquely designed ImQ internal resin standard yields its signature resonance in a transparent region of the analyte spectrum making suppression of the polymer background unnecessary. This method enabled us to target a reasonable ligand density for enzyme purification and provides an advantageous alternative to quantitation against soluble standards or protonated solvent.
Department of Chemistry, University of Kansas, 1251 Wescoe Hall Drive, 2010 Malott Hall, Lawrence, KS 66045-7582, USA.
Laura H
LH
Lucas
Matthew A
MA
Cerny
Yakov M
YM
Koen
Robert P
RP
Hanzlik
Cynthia K
CK
Larive
D000818:Animals; D002846:Chromatography, Affinity / Q000295:instrumentation* / Q000379:methods; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008745:Methylation; D015394:Molecular Structure; D051381:Rats; D012015:Reference Standards; D012117:Resins, Synthetic / Q000032:analysis* / Q000737:chemistry*; D014316:Tritium / Q000737:chemistry
D008024:Ligands; D012117:Resins, Synthetic; D014316:Tritium