NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver
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 metha...
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MDPI AG
2017-11-01
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| Series: | Metabolites |
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| Online Access: | https://www.mdpi.com/2218-1989/7/4/61 |
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| author | Fariba Fathi Antonio Brun Katherine H. Rott Paulo Falco Cobra Marco Tonelli Hamid R. Eghbalnia Enrique Caviedes-Vidal William H. Karasov John L. Markley |
| author_facet | Fariba Fathi Antonio Brun Katherine H. Rott Paulo Falco Cobra Marco Tonelli Hamid R. Eghbalnia Enrique Caviedes-Vidal William H. Karasov John L. Markley |
| author_sort | Fariba Fathi |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-12-17T15:20:09Z |
| format | Article |
| id | doaj.art-10ae90776d904005adf1c925ff89f333 |
| institution | Directory Open Access Journal |
| issn | 2218-1989 |
| language | English |
| last_indexed | 2024-12-17T15:20:09Z |
| publishDate | 2017-11-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Metabolites |
| spelling | doaj.art-10ae90776d904005adf1c925ff89f3332022-12-21T21:43:25ZengMDPI AGMetabolites2218-19892017-11-01746110.3390/metabo7040061metabo7040061NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian LiverFariba Fathi0Antonio Brun1Katherine H. Rott2Paulo Falco Cobra3Marco Tonelli4Hamid R. Eghbalnia5Enrique Caviedes-Vidal6William H. Karasov7John L. Markley8Biochemistry Department, University of Wisconsin-Madison, 433 Babcock Drive, Madison, WI 53706, USADepartment of Forest and Wildlife Ecology, University of Wisconsin-Madison, 1630 Linden Drive, Madison, WI 53706, USADepartment of Forest and Wildlife Ecology, University of Wisconsin-Madison, 1630 Linden Drive, Madison, WI 53706, USABiochemistry Department, University of Wisconsin-Madison, 433 Babcock Drive, Madison, WI 53706, USABiochemistry Department, University of Wisconsin-Madison, 433 Babcock Drive, Madison, WI 53706, USABiochemistry Department, University of Wisconsin-Madison, 433 Babcock Drive, Madison, WI 53706, USAInstituto Multidisciplinario de Investigaciones Biológicas de San Luis, Consejo Nacional de Investigaciones Científicas y Técnicas, and Departamento de Bioquímica y Ciencias Biológicas, Universidad Nacional de San Luis, San Luis, 5700, ArgentinaDepartment of Forest and Wildlife Ecology, University of Wisconsin-Madison, 1630 Linden Drive, Madison, WI 53706, USABiochemistry Department, University of Wisconsin-Madison, 433 Babcock Drive, Madison, WI 53706, USAMetabolites 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.https://www.mdpi.com/2218-1989/7/4/61NMR spectroscopyliver tissueextraction protocolmetabolite identification by NMRdiet, effect of on liver metabolites |
| spellingShingle | Fariba Fathi Antonio Brun Katherine H. Rott Paulo Falco Cobra Marco Tonelli Hamid R. Eghbalnia Enrique Caviedes-Vidal William H. Karasov John L. Markley NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver Metabolites NMR spectroscopy liver tissue extraction protocol metabolite identification by NMR diet, effect of on liver metabolites |
| title | NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver |
| title_full | NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver |
| title_fullStr | NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver |
| title_full_unstemmed | NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver |
| title_short | NMR-Based Identification of Metabolites in Polar and Non-Polar Extracts of Avian Liver |
| title_sort | nmr based identification of metabolites in polar and non polar extracts of avian liver |
| topic | NMR spectroscopy liver tissue extraction protocol metabolite identification by NMR diet, effect of on liver metabolites |
| url | https://www.mdpi.com/2218-1989/7/4/61 |
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