Determination of low levels of <sup>2</sup>H‐labeling using high‐resolution mass spectrometry: Application in studies of lipid flux and beyond

Herath, Kithsiri; Zhong, Wendy; Yang, Jinggang; Mahsut, Ablatt; Rohm, Rory J.; Shah, Vinit; Castro-Perez, José; Zhou, Haihong; Attygalle, Athula B.; Lee, Kang; Singh, Sheo B.; Johns, Douglas G.; Cleary, Michele A.; Hubbard, Brian K.; Previs, Stephen F.; Roddy, Thomas P.

doi:10.1002/rcm.6776

Cited by 16 publications

(18 citation statements)

References 19 publications

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“…The downside, however, is that it is not specific to particular fatty acids, and thus does not provide information on isotope enrichment per molecule [ 30 ]. To overcome this shortage, approaches with increased specificity have been developed by combining stable isotope tracers with mass spectrometry analysis [ 31 ]. In this updated approach, deuterated water (D 2 O) is often used in cultured cells or animals.…”

Section: De Novo Lipogenesis (Dnl)mentioning

confidence: 99%

Regulation and Metabolic Significance of De Novo Lipogenesis in Adipose Tissues

2018

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De novo lipogenesis (DNL) is a complex and highly regulated process in which carbohydrates from circulation are converted into fatty acids that are then used for synthesizing either triglycerides or other lipid molecules. Dysregulation of DNL contributes to human diseases such as obesity, type 2 diabetes, and cardiovascular diseases. Thus, the lipogenic pathway may provide a new therapeutic opportunity for combating various pathological conditions that are associated with dysregulated lipid metabolism. Hepatic DNL has been well documented, but lipogenesis in adipocytes and its contribution to energy homeostasis and insulin sensitivity are less studied. Recent reports have gained significant insights into the signaling pathways that regulate lipogenic transcription factors and the role of DNL in adipose tissues. In this review, we will update the current knowledge of DNL in white and brown adipose tissues with the focus on transcriptional, post-translational, and central regulation of DNL. We will also summarize the recent findings of adipocyte DNL as a source of some signaling molecules that critically regulate energy metabolism.

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Section: De Novo Lipogenesis (Dnl)mentioning

confidence: 99%

Regulation and Metabolic Significance of De Novo Lipogenesis in Adipose Tissues

2018

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“…Although our data suggest that one can blur the lines between the analytical methods, a head-to-head comparison would be necessary to identify true limits. We should also add that advances in high-resolution mass spectrometry can offer novel advantages in cases where one aims to quantify low levels of labeling [ 23 ], especially if multiple stable isotopes have been administered [ 24 ].…”

Section: Discussionmentioning

confidence: 99%

Isotope Fractionation during Gas Chromatography Can Enhance Mass Spectrometry-Based Measures of 2H-Labeling of Small Molecules

et al. 2020

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Stable isotope tracers can be used to quantify the activity of metabolic pathways. Specifically, 2H-water is quite versatile, and its incorporation into various products can enable measurements of carbohydrate, lipid, protein and nucleic acid kinetics. However, since there are limits on how much 2H-water can be administered and since some metabolic processes may be slow, it is possible that one may be challenged with measuring small changes in isotopic enrichment. We demonstrate an advantage of the isotope fractionation that occurs during gas chromatography, namely, setting tightly bounded integration regions yields a powerful approach for determining isotope ratios. We determined how the degree of isotope fractionation, chromatographic peak width and mass spectrometer dwell time can increase the apparent isotope labeling. Relatively simple changes in the logic surrounding data acquisition and processing can enhance gas chromatography-mass spectrometry measures of low levels of 2H-labeling, this is especially useful when asymmetrical peaks are recorded at low signal:background. Although we have largely focused attention on alanine (which is of interest in studies of protein synthesis), it should be possible to extend the concepts to other analytes and/or hardware configurations.

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“…Labelling with glucose or heavy water will lead to generalized incorporation of the label into a large number of different metabolites, while more pathway-specific tracers can be chosen, e.g., serine or palmitoyl-CoA for the sphingolipid pathway [25], or lipid head groups such choline can be used to investigate biosynthesis of choline-containing lipid classes. Commonly-used labelled substrates for tracing lipid flux are glucose or glycerol [26,27,28], fatty acids [19,29], amino acids [17,27,30], or heavy water (D 2 O) as an unspecific tracer [31,32]. In animal or human studies, the tracer can be a substantial cost factor, as rather large amounts may be required to achieve a sufficient degree of labelling, and because injectable tracers for human studies must be sterile.…”

Section: Analytical Considerationsmentioning

confidence: 99%

Analytical Considerations of Stable Isotope Labelling in Lipidomics

Triebl

Wenk

2018

Biomolecules

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Over the last two decades, lipids have come to be understood as far more than merely components of cellular membranes and forms of energy storage, and are now also being implicated to play important roles in a variety of diseases, with lipid biomarker research one of the most widespread applications of lipidomic techniques both in research and in clinical settings. Stable isotope labelling has become a staple technique in the analysis of small molecule metabolism and dynamics, as it is the only experimental setup by which biosynthesis, remodelling and degradation of biomolecules can be directly measured. Using state-of-the-art analytical technologies such as chromatography-coupled high resolution tandem mass spectrometry, the stable isotope label can be precisely localized and quantified within the biomolecules. The application of stable isotope labelling to lipidomics is however complicated by the diversity of lipids and the complexity of the necessary data analysis. This article discusses key experimental aspects of stable isotope labelling in the field of mass spectrometry-based lipidomics, summarizes current applications and provides an outlook on future developments and potential.

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Determination of low levels of ²H‐labeling using high‐resolution mass spectrometry: Application in studies of lipid flux and beyond

Cited by 16 publications

References 19 publications

Regulation and Metabolic Significance of De Novo Lipogenesis in Adipose Tissues

Regulation and Metabolic Significance of De Novo Lipogenesis in Adipose Tissues

Isotope Fractionation during Gas Chromatography Can Enhance Mass Spectrometry-Based Measures of 2H-Labeling of Small Molecules

Analytical Considerations of Stable Isotope Labelling in Lipidomics

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Determination of low levels of 2H‐labeling using high‐resolution mass spectrometry: Application in studies of lipid flux and beyond

Cited by 16 publications

References 19 publications

Regulation and Metabolic Significance of De Novo Lipogenesis in Adipose Tissues

Regulation and Metabolic Significance of De Novo Lipogenesis in Adipose Tissues

Isotope Fractionation during Gas Chromatography Can Enhance Mass Spectrometry-Based Measures of 2H-Labeling of Small Molecules

Analytical Considerations of Stable Isotope Labelling in Lipidomics

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Determination of low levels of ²H‐labeling using high‐resolution mass spectrometry: Application in studies of lipid flux and beyond