Facile and fast enrichment of phosphopeptides prior to matrix-assisted laser desorption/ionization time-of-flight mass spectrometric analysis using natural nanoparticle-bentonite

• Use nontoxic and inexpensive natural nanomaterial bentonite to the enrichment of phosphopeptides.
• The operation is facile, fast, and highly effective, without nanomaterial preparation, desalting and elution process. Phosphopeptides binded on bentonite can be detected directly.
• All the phosphorylation sites in a-casein and ß-casein can be detected unambiguously even at low fmol level.
• Open up new horizons for the application of bentonite and natural nanomaterials.

A novel strategy for facile, fast and highly effective enrichment of phosphopeptides by bentonite followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is presented. Bentonite is a Al(III)-containing, nontoxic and inexpensive natural nanomaterial with good biocompatibility. By employing bentonite into phosphopeptides enrichment, phosphopeptides binded on bentonite can be detected directly after simple wash, without nanomaterial preparation, desalting and elution process. The whole enrichment procedure can be easily completed within 10 min. Tryptic digest products from several standard proteins and nonfat milk are pretreated using bentonite to demonstrate the efficiency of this method, all the phosphorylation sites in a-casein and ß-casein can be detected unambiguously even at low fmol level. With all the advantages mentioned above, this method is of great potential for future studies of complex phosphoproteomes, and opens up new horizons for bentonite application.

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