Abstract
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.
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