Determination of selenosugars in crude human urine using high‐performance liquid chromatography/atmospheric pressure chemical ionization tandem mass spectrometry

Abstract

The narrow gap between essentiality and toxicity of selenium requires detailed investigations on selenium metabolism in order to find suitable indicators for the selenium status in the human body. Current methods for quantitative selenium speciation in human urine are based on separation by high-performance liquid chromatography (HPLC) coupled online with elemental mass spectrometry (MS), and the potential of molecular MS detection techniques for the reliable identification and quantification of selenosugars in crude human urine has not been utilized. Now we report the development of an HPLC tandem mass spectrometric (MS/MS) method for the reliable determination in crude human urine of three significant selenium urinary metabolites, collectively termed selenosugars, namely methyl 2-acetamido-2-deoxy-1-seleno-beta-D-galactopyranoside (SeGalNAc), methyl 2-acetamido-2-deoxy-1-seleno-beta-D-glucopyranoside (SeGluNAc) and methyl 2-amino-2-deoxy-1-seleno-beta-D-galactopyranoside (SeGalNH2). Reversed-phase HPLC, with and without cation-exchange guard columns, was applied for the separation of the selenosugars, and atmospheric pressure chemical ionization (APCI) and selected reaction monitoring (SRM) were used for selective and sensitive detection. The collision-induced dissociation behaviour of the selenosugars was studied in detail using APCI triple quadrupole MS/MS and electrospray ion trap MS. The developed method was applied to urine samples collected prior to and after selenium supplementation for the quantification of SeGalNAc using both external calibration and the method of standard additions. Additionally, SeGalNH2 was detected in urine samples after Se supplementation. Finally, neutral loss scanning was explored as a possible method for the detection of unknown methyl-selenosugars.

Selenium biochemistry and its role for human health

“…41 Minor selenosugars have been also detected in urine, including SeMethyl-N-acetyl-glucosamine (MSeGluNAc) 54 and the deacylated analogue of SeGalNAc, SeMethyl-N-amino-galactosamine (MSeGalNH 2 ). 55 It has been also hypothesized that in case of Se excess, HSe À can be metabolized entering into the stepwise methylation pathway (Fig. 1 path p).…”

A New Tool Based on the Use of Stable Isotopes and Isotope Pattern Deconvolution (IPD)‐ICP‐MS for Nutritional and Clinical Studies

“…To this end, the separation of seleno species was performed using a RP (C 18 ) column. A first peak eluting at a retention time of 2.7 min that could correspond to inorganic Se [most of it, about 95%, matched the retention time of Se(IV) and did not contain selenosugars] [41]. As can be seen, for natural Se, two main seleno species were detected.…”

Use of 3‐nitrobenzonitrile as an additive for improved sensitivity in sonic‐spray ionization mass spectrometry

“…The selenosugar 2‐acetamido‐2‐deoxy‐1‐seleno‐β‐D‐galactopyranoside (SeGalNAc), which is the main selenium metabolite found in human urine, showed substantial signal and S/N enhancement in 3‐NBN‐assisted V‐EASI‐MS. Almost an order of magnitude higher intensity, accompanied by a 7.4 higher S/N ratio, was observed for its protonated molecule ( m/z 300) in the presence of 500 µg mL −1 of 3‐NBN using an ion transfer capillary temperature of 200°C (Fig.…”