Metabolites 2017, 7(4), 61; https://doi.org/10.3390/metabo7040061
NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver
Fariba Fathi 1, Antonio Brun 2
, Katherine H. Rott 2, Paulo Falco Cobra 1, Marco Tonelli 1, Hamid R. Eghbalnia 1, Enrique Caviedes-Vidal 3, William H. Karasov 2 and John L. Markley 1,*
, Katherine H. Rott 2, Paulo Falco Cobra 1, Marco Tonelli 1, Hamid R. Eghbalnia 1, Enrique Caviedes-Vidal 3, William H. Karasov 2 and John L. Markley 1,*
1
Biochemistry Department, University of Wisconsin-Madison, 433 Babcock Drive, Madison, WI 53706, USA
2
Department of Forest and Wildlife Ecology, University of Wisconsin-Madison, 1630 Linden Drive, Madison, WI 53706, USA
3
Instituto Multidisciplinario de Investigaciones Biológicas de San Luis, Consejo Nacional de Investigaciones CientÃficas y Técnicas, and Departamento de BioquÃmica y Ciencias Biológicas, Universidad Nacional de San Luis, San Luis, 5700, Argentina
*
Author to whom correspondence should be addressed.
Academic Editor: Peter Meikle
Received: 3 October 2017 / Revised: 7 November 2017 / Accepted: 8 November 2017 / Published: 16 November 2017
Abstract
Metabolites present in liver provide important clues regarding the physiological state of an organism. The aim of this work was to evaluate a protocol for high-throughput NMR-based analysis of polar and non-polar metabolites from a small quantity of liver tissue. We extracted the tissue with a methanol/chloroform/water mixture and isolated the polar metabolites from the methanol/water layer and the non-polar metabolites from the chloroform layer. Following drying, we re-solubilized the fractions for analysis with a 600 MHz NMR spectrometer equipped with a 1.7 mm cryogenic probe. In order to evaluate the feasibility of this protocol for metabolomics studies, we analyzed the metabolic profile of livers from house sparrow (Passer domesticus) nestlings raised on two different diets: livers from 10 nestlings raised on a high protein diet (HP) for 4 d and livers from 12 nestlings raised on the HP diet for 3 d and then switched to a high carbohydrate diet (HC) for 1 d. The protocol enabled the detection of 52 polar and nine non-polar metabolites in 1H NMR spectra of the extracts. We analyzed the lipophilic metabolites by one-way ANOVA to assess statistically significant concentration differences between the two groups. The results of our studies demonstrate that the protocol described here can be exploited for high-throughput screening of small quantities of liver tissue (approx. 100 mg wet mass) obtainable from small animals. View Full-TextKeywords:
NMR spectroscopy; liver tissue; extraction protocol; metabolite identification by NMR; diet, effect of on liver metabolites
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Fathi, F.; Brun, A.; Rott, K.H.; Falco Cobra, P.; Tonelli, M.; Eghbalnia, H.R.; Caviedes-Vidal, E.; Karasov, W.H.; Markley, J.L. NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver. Metabolites 2017, 7, 61.
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