4,4-Dimethyl-4-silapentane-1-ammonium trifluoroacetate (DSA), a promising universal internal standard for NMR-based metabolic profiling studies of biofluids, including blood plasma and serum |
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Authors: | Mohammed F Alum Paul A Shaw Brian C Sweatman Baljit K Ubhi John N Haselden Susan C Connor |
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Institution: | (1) Department of Investigative Preclinical Toxicology, Safety Assessment Division, GlaxoSmithKline R & D, Park Road, Ware, Hertfordshire, SG12 0DP, UK |
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Abstract: | Nuclear magnetic resonance (NMR)-based metabolic profiling of biofluids and tissues are of key interest to enhance biomarker
discovery for disease, drug efficacy and toxicity studies. Urine and blood plasma/serum are the biofluids of most interest
as they are the most accessible in both clinical and preclinical studies. However, proteinaceous fluids, such as blood serum
or plasma, represent the greatest technical challenge since the chemical shift (δ) and line-width (ν1/2) of internal standards currently used for aqueous NMR samples are greatly affected by protein binding. We have therefore
investigated the suitability of 4,4-dimethyl-4-silapentane-1-ammonium trifluoroacetate (DSA) as a universal internal standard
for biofluids. Proton (1H) NMR spectroscopy was used to determine the effect of serum pH (3, 7.4 and 10) and DSA concentration on the overall lineshape
and position of the trimethylsilyl resonance of DSA. The results were compared to that of 3-(trimethylsilyl)propionic acid
sodium salt (TSP). Both the chemical shift and line-width of the DSA peak were not significantly affected by pH or DSA concentration,
whereas these parameters for TSP showed large variations due to protein binding. Furthermore, the peak area of DSA correlated
linearly with its concentration under all pH conditions, whilst no linear correlation was observed with TSP. Overall, in contrast
to TSP, these results support the use of DSA as an accurate universal internal chemical shift reference and concentration/normalisation
standard for biofluids. In the case of proteinaceous biofluids such as serum, where no current standard is available, this
offers a considerable saving in both operator and spectrometer time. |
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Keywords: | 4 4-dimethyl-4-silapentane-1-ammonium trifluoroacetate (DSA) Metabolomics Metabonomics Proton NMR Blood plasma/serum Internal reference standard |
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