Global centering-based normalization is a commonly used normalization approach in mass spectrometry-based label-free proteomics. It scales the peptide abundances to have the same median intensities, based on an assumption that the majority of abundances remain the same across the samples. However, especially in phosphoproteomics, this assumption can introduce bias, as the samples are enriched during sample preparation which can mask the underlying biological changes. To address this possible bias, phosphopeptides quantified in both enriched and non-enriched samples can be used to calculate factors that mitigate the bias.


We present an R package phosphonormalizer for normalizing enriched samples in label-free mass spectrometry-based phosphoproteomics.

Availability and implementation

The phosphonormalizer package is freely available under GPL ( > =2) license from Bioconductor (

Supplementary information

Supplementary data are available at Bioinformatics online.

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