Merck
  • Home
  • Search Results
  • Development of an NMR microprobe procedure for high-throughput environmental metabolomics of Daphnia magna.

Development of an NMR microprobe procedure for high-throughput environmental metabolomics of Daphnia magna.

Magnetic resonance in chemistry : MRC (2015-04-22)
Edward G Nagato, Brian P Lankadurai, Ronald Soong, André J Simpson, Myrna J Simpson
ABSTRACT

Nuclear magnetic resonance (NMR) is the primary platform used in high-throughput environmental metabolomics studies because its non-selectivity is well suited for non-targeted approaches. However, standard NMR probes may limit the use of NMR-based metabolomics for tiny organisms because of the sample volumes required for routine metabolic profiling. Because of this, keystone ecological species, such as the water flea Daphnia magna, are not commonly studied because of the analytical challenges associated with NMR-based approaches. Here, the use of a 1.7-mm NMR microprobe in analyzing tissue extracts from D. magna is tested. Three different extraction procedures (D2O-based buffer, Bligh and Dyer, and acetonitrile : methanol : water) were compared in terms of the yields and breadth of polar metabolites. The D2O buffer extraction yielded the most metabolites and resulted in the best reproducibility. Varying amounts of D. magna dry mass were extracted to optimize metabolite isolation from D. magna tissues. A ratio of 1-1.5-mg dry mass to 40 µl of extraction solvent provided excellent signal-to-noise and spectral resolution using (1)H NMR. The metabolite profile of a single daphnid was also investigated (approximately 0.2 mg). However, the signal-to-noise of the (1)H NMR was considerably lower, and while feasible for select applications would likely not be appropriate for high-throughput NMR-based metabolomics. Two-dimensional NMR experiments on D. magna extracts were also performed using the 1.7-mm NMR probe to confirm (1)H NMR metabolite assignments. This study provides an NMR-based analytical framework for future metabolomics studies that use D. magna in ecological and ecotoxicity studies.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Methanol, anhydrous, 99.8%
Sigma-Aldrich
Sodium azide, BioUltra, ≥99.5% (T)
Sigma-Aldrich
Sodium azide, purum p.a., ≥99.0% (T)
Sigma-Aldrich
Sodium azide, BioXtra
Sigma-Aldrich
Sodium azide, ReagentPlus®, ≥99.5%
Sigma-Aldrich
Methanol, HPLC Plus, ≥99.9%, poly-coated bottles
Sigma-Aldrich
Acetonitrile solution, contains 0.05 % (v/v) trifluoroacetic acid
Sigma-Aldrich
Acetonitrile solution, contains 0.1 % (v/v) trifluoroacetic acid, suitable for HPLC
Millipore
Acetonitrile solution, suitable for HPLC, acetonitrile:water 5:95% (v/v), 10 mM Ammoniumbicarbonate, pH 10,0
Sigma-Aldrich
Methanol, NMR reference standard
Sigma-Aldrich
Deuterium oxide, "100%", 99.990 atom % D
Sigma-Aldrich
Acetonitrile, anhydrous, 99.8%
Sigma-Aldrich
Deuterium oxide, standard, 99.98 atom %±0.01 atom % D
Sigma-Aldrich
Deuterium oxide, 99.9 atom % D, contains 1 % (w/w) 3-(trimethylsilyl)-1-propanesulfonic acid, sodium salt (DSS)
Sigma-Aldrich
Deuterium oxide, 99.9 atom % D, contains 0.75 wt. % 3-(trimethylsilyl)propionic-2,2,3,3-d4 acid, sodium salt
Sigma-Aldrich
Deuterium oxide, 99.9 atom % D
Sigma-Aldrich
Deuterium oxide, "100%", ≥99.96 atom % D
Sigma-Aldrich
Deuterium oxide, extra, 99.994 atom % D
Sigma-Aldrich
Deuterium oxide, 99.9 atom % D, glass distilled
Supelco
Methanol solution, NMR reference standard, 4% in methanol-d4 (99.8 atom % D), NMR tube size 5 mm × 8 in.
Sigma-Aldrich
Methanol solution, NMR reference standard, 4% in methanol-d4 (99.8 atom % D), NMR tube size 3 mm × 8 in.
Sigma-Aldrich
Methanol-12C, 99.95 atom % 12C
Sigma-Aldrich
Deuterium oxide, 99.8 atom % D
Sigma-Aldrich
Deuterium oxide, 60 atom % D
Sigma-Aldrich
Deuterium oxide, 70 atom % D
Supelco
Methanol solution, contains 0.10 % (v/v) formic acid, UHPLC, suitable for mass spectrometry (MS), ≥99.5%
Sigma-Aldrich
Acetonitrile, Preparateur, ≥99.9% (GC), One-time steel-plastic (SP) drum
Sigma-Aldrich
Deuterium oxide, filtered, 99.8 atom % D
Sigma-Aldrich
Acetonitrile, for DNA synthesis
Sigma-Aldrich
Deuterium oxide, 99.9 atom % D, contains 0.05 wt. % 3-(trimethylsilyl)propionic-2,2,3,3-d4 acid, sodium salt