The in-depth characterization of excipients is a prerequisite for their safe application in pharmaceutical products. In case of surfactants, this task can be a challenge, since many industrial products are mixtures of variable composition. In this work, mass spectrometric methods are applied to characterize some ethoxylated surfactants that are widely used by the pharmaceutical industry. Among them are ethoxylated fatty alcohols with ether structure (e.g., Brij®), ethoxylated fatty acids with ester structure (e.g., Myrj®), ethoxylated sorbitane fatty acid esters (e.g., Tween®), ethoxylated glycerides (e.g., Tagat®), and Triton® X-100. MALDI-TOF mass spectrometry is best suitable to obtain molecular mass distributions of polymeric products, namely those with higher molecular mass. Electrospray and nanoelectrospray molecular mass shows a greater tendency for multiple charges. However, it is best suitable for small MM products, and multiple charges have been de-convoluted successfully using the MaxEnt™ 3 algorithm. Tandem mass spectrometry helps to identify the chemical composition, e.g. for identification of acyl chains. The work is intended to serve as a reference for mass spectrometric characterization of surfactants in the course of R&D, validation or change control.