Characterization of Small Combinatorial Chemistry Libraries by ¹H NMR. Quantitation with a Convenient and Novel Internal Standard

Abstract

A novel silane standard, 1,4-bis(trimethylsilyl)benzene (BTMSB), is introduced for the generic quantitation of small organic molecules in DMSO-d(6) solution by (1)H NMR. This standard is an easily weighable solid and is stable for at least 1 month in DMSO solution, and its (1)H NMR spectrum contains a strong singlet in a region usually free of signals. With a set of certified standards, concentration determination with about 2% precision and accuracy is verified after solution preparation with fully automated procedures, thus making very effective the characterization of small combinatorial chemistry libraries for identity and purity when combined with other physicochemical or biochemical tests. As an example, for a set of about 400 compounds, results of (1)H NMR characterization are compared to the more customary LC-UV-MS method. NMR and MS data agree for identity on the vast majority of cases (84% positive and 5% negative), whereas the remaining cases (11%) are marked as highly impure only after NMR spectra analysis. Most importantly, determination of concentration rather than that of relative purity appears the right choice for a correct evaluation of biochemical potency.

Quantitative NMR in synthetic and combinatorial chemistry

“…In any case, the intensity and phase response should be checked using a sample with a single intense line and varying its position in the spectral window. If the response is found non-linear but reproducible in the range of interest, a correction function can be introduced [13]. This takes into account the off-resonance effect due to non-constant excitation of the rf pulse, as well.…”

Chemical-shift referencing and resolution stability in gradient LC–NMR (acetonitrile:water)

“…Both DSA (4,4-dimethyl-4-silapentane-1-ammonium trifluoroacetate) and dioxane have been used for protein samples because they were shown to not interact with proteins [25,29]. The internal standard N,N 0 ,N 00 -trimethyl(2-trimethyl) ethylammonium iodide (TMS-A) was proposed as a way to minimize some of the limitations of TSP and DSS [30], and other compounds have been proposed to minimize some of the limitations of TMS [31][32][33].…”

Design and Synthesis of Non-peptide RGD Mimics for Evaluation of Their Utility as Anti-platelet Agents

“…The purity of compounds 3a-e were measured by the internal standard method using 1,4-bis(trimethylsilyl) benzene. 38) Reagents and conditions: (a) NaBH 4 , MeOH, 0°C, 83%; (b) tert-butyl 4-(3-bromopropyl)piperidine-1-carboxylate, NaH, DMF, r.t., 29%; (c) 4 M HCl-AcOEt, r.t., 75%. 2 mmol).…”