Evaluation of sample preparation protocols for quantitative NMR-based metabolomics

Snytnikova, Olga A.; Khlichkina, Anastasiya A.; Sagdeev, R. Z.; Tsentalovich, Yuri P.

doi:10.1007/s11306-019-1545-y

Cited by 33 publications

(53 citation statements)

References 39 publications

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“…Overall, these results indicate that for Blastocystis ST7, methanol is a better extraction solvent. This is in contradiction to one past publication, in which a comparison between methanol and ethanol both produced similar results [ 29 ].…”

Section: Discussioncontrasting

confidence: 99%

Establishing a Metabolite Extraction Method to Study the Metabolome of Blastocystis Using NMR

et al. 2021

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Blastocystis is an opportunistic parasite commonly found in the intestines of humans and other animals. Despite its high prevalence, knowledge regarding Blastocystis biology within and outside the host is limited. Analysis of the metabolites produced by this anaerobe could provide insights that can help map its metabolism and determine its role in both health and disease. Due to its controversial pathogenicity, these metabolites could define its deterministic role in microbiome’s “health” and/or subsequently resolve Blastocystis’ potential impact in gastrointestinal health. A common method for elucidating the presence of these metabolites is through 1H nuclear magnetic resonance (NMR). However, there are currently no described benchmarked methods available to extract metabolites from Blastocystis for 1H NMR analysis. Herein, several extraction solvents, lysis methods and incubation temperatures were compared for their usefulness as an extraction protocol for this protozoan. Following extraction, the samples were freeze-dried, re-solubilized and analysed with 1H NMR. The results demonstrate that carrying out the procedure at room temperature using methanol as an extraction solvent and bead bashing as a lysis technique provides a consistent, reproducible and efficient method to extract metabolites from Blastocystis for NMR.

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Section: Discussioncontrasting

confidence: 99%

Establishing a Metabolite Extraction Method to Study the Metabolome of Blastocystis Using NMR

et al. 2021

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“…Blood samples were collected by decapitation and kept at room temperature for 15 min, centrifuged at 3000 rpm for 15 min, and then serum samples were stored at −70 • C until analyzed. The extraction of metabolites from serum was performed by using a short sample preparation protocol earlier evaluated for quantitative NMR-based metabolomics [36]. Namely, 100 µL of ice-cold methanol (HPLC grade, Scharlau, Barcelona, Spain) and 100 µL of ice-cold chloroform (HPLC grade, Scharlau, Spain) were added to 100 µL of serum and vortexed for 30 s, kept on ice for 10 min, and incubated at −20 • C for 30 min.…”

Section: Metabolite Extraction and Nmr Spectroscopymentioning

confidence: 99%

Fucose Ameliorates Tryptophan Metabolism and Behavioral Abnormalities in a Mouse Model of Chronic Colitis

et al. 2020

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Growing evidence suggests that intestinal mucosa homeostasis impacts immunity, metabolism, the Central Nervous System (CNS), and behavior. Here, we investigated the effect of the monosaccharide fucose on inflammation, metabolism, intestinal microbiota, and social behavior in the Dextran Sulfate Sodium (DSS)-induced chronic colitis mouse model. Our data show that chronic colitis is accompanied by the decrease of the serum tryptophan level and the depletion of the intestinal microbiota, specifically tryptophan-producing E. coli and Bifidobacterium. These changes are associated with defects in the male mouse social behavior such as a lack of preference towards female bedding in an odor preference test. The addition of fucose to the test animals' diet altered the bacterial community, increased the abundance of tryptophan-producing E. coli, normalized blood tryptophan levels, and ameliorated social behavior deficits. At the same time, we observed no ameliorating effect of fucose on colon morphology and colitis. Our results suggest a possible mechanism by which intestinal inflammation affects social behavior in male mice. We propose fucose as a promising prebiotic, since it creates a favorable environment for the beneficial bacteria that promote normalization of serum tryptophan level and amelioration of the behavioral abnormalities in the odor preference test.

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“…Each sample of the frog tissue was weighted prior to homogenization: the typical heart sample weight was 70 mg and the typical liver sample weight was 175 mg. The tissue was placed in a glass vial and homogenized with a TissueRuptor II homogenizer (Qiagen, Netherlands) in 1,600 µL of cold (− 20 °C) MeOH, and then 800 µL of water and 1,600 µL of cold chloroform were added 42 . The mixture was shaken well in a shaker for 20 min and left at − 20 °C for 30 min.…”

Section: Animals R Amurensis Individuals Were Collected In Septembementioning

confidence: 99%

Metabolic response of the Siberian wood frog Rana amurensis to extreme hypoxia

Shekhovtsov

Булахова

Tsentalovich

et al. 2020

Sci Rep

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the Siberian wood frog Rana amurensis is a recently discovered example of extreme hypoxia tolerance that is able to survive several months without oxygen. We studied metabolomic profiles of heart and liver of R. amurensis exposed to 17 days of extreme hypoxia. Without oxygen, the studied tissues experience considerable stress with a drastic decrease of ATP, phosphocreatine, and NAD+ concentrations, and concomitant increase of AMP, creatine, and NADH. Heart and liver switch to different pathways of glycolysis with differential accumulation of lactate, alanine, succinate, as well as 2,3-butanediol (previously not reported for vertebrates as an end product of glycolysis) and depletion of aspartate. We also observed statistically significant changes in concentrations of certain osmolytes and choline-related compounds. Low succinate/fumarate ratio and high glutathione levels indicate adaptations to reoxygenation stress. Our data suggest that maintenance of the ATP/ADP pool is not required for survival of R. amurensis, in contrast to anoxia-tolerant turtles. Anoxia is a huge stress for the majority of vertebrates. Amphibians are considered to be relatively anoxia-intolerant, in contrast to a few turtle species and certain fish species 1-3 : the most resistant species are known to survive anoxia for a few days at most. However, a recent study 4 proved that the Siberian wood frog Rana amurensis can tolerate almost complete anoxia at 2-3 °C (below 0.2 mg/L oxygen, i.e., less than 1.5% of the normal concentration at this temperature) for several months. This is on par with the best vertebrate models from other classes. Moreover, while red-eared slider turtles, the most studied anoxia-tolerant tetrapods, are dormant under anoxia, the Siberian wood frog is able to react to stimuli, e.g., to flee when disturbed. R. amurensis is thus a unique model of extreme hypoxia tolerance among terrestrial vertebrates. The natural range of R. amurensis includes Northeastern China and Siberia from the Urals to the coast of the Okhotsk sea; its northern distribution is limited by about 71° N. It overwinters under ice in water bodies. Many of these lakes are shallow (< 3 m) due to thermokarst origin, i.e. resulting from local melting of permafrost. Experimental studies 4 demonstrated that during the winter oxygen is depleted in overwintering sites till almost complete anoxia. Siberian wood frogs overwinter for 6 to 7 months in different regions under hypoxia until they can leave the lakes in April-early May. The mechanisms of adaptation to the lack of oxygen are totally unknown for R. amurensis. Here we made an attempt to study this phenomenon using 1 H-NMR-based quantitative metabolomics. This method simultaneously yields concentrations of multiple metabolites 5. At the present, the detailed quantitative metabolomic composition is known for many human tissues, but for very few animals 6-8. To the best of our knowledge, this is the first report on the quantitative metabolomic analysis of amphibian tissues. We determined concentrations of ove...

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Evaluation of sample preparation protocols for quantitative NMR-based metabolomics

Cited by 33 publications

References 39 publications

Establishing a Metabolite Extraction Method to Study the Metabolome of Blastocystis Using NMR

Establishing a Metabolite Extraction Method to Study the Metabolome of Blastocystis Using NMR

Fucose Ameliorates Tryptophan Metabolism and Behavioral Abnormalities in a Mouse Model of Chronic Colitis

Metabolic response of the Siberian wood frog Rana amurensis to extreme hypoxia

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