Metabolite Yeast Extracts

Metabolite Yeast Extracts

The yeast extracts – U-¹³C (ISO1) and unlabeled (ISO1-UNL) – are designed for internal standardization (i.e., matrix spike-in) quantitation experiments and for quality control evaluations in untargeted and targeted metabolomics. The compounds in these extracts span broad metabolic classes (e.g., amino and organic acids, sugar phosphates, coenzymes) that are linked to various biochemical pathways (e.g., citrate and glyoxylate cycle, amino acid and nucleotide metabolism, pentose phosphate) and cellular/molecular processes (e.g., immune system, blood coagulation, DNA metabolism). These metabolites have been rigorously characterized by several LC-/GC-MS methodologies and are amenable to a variety of research uses after simple reconstitution.

Kit contents:

One dried-down yeast extract (U-¹³C, ISO1 or unlabeled, ISO1-UNL).
Document package (supplied via QR code). Package includes user manual, which contains example procedures and LC-MS methods for user reference.

Resources

Metabolite Yeast Extracts

Sets, Mixes, and Kits for MS 'Omics and MS/MS Screening

Reconstituting ISO1 or ISO1-UNL with Water Video

Labeled Metabolites: Fight Against Error Video

Stable Isotope Standards for Mass Spectrometry

For further information on the metabolite yeast extracts and its applications, please visit ISOtopic Solutions.

Frequently Asked Questions

What part of the life cycle is the yeast extract from?

The yeast (Pichia pastoris, strain CBS 7435) is in the exponential growth phase. In this phase, the cell is using most of the available substrate to reproduce. The doubling time for the yeast strain employed is a little less than 3 hours. Exponential growth is ensured through verification of the OD 600 measurements during fermentation. The extract preparation is detailed in PMID: 23086617.

What does the "U" in U-¹³C (98%) refer to?

The "U" denotes a uniformly labeled compound. For example, NADP⁺ with a formula C₂₁H₂₉N₇O₁₇P₃ has 21 C in its ¹³C-labeled form.

What is the recommended procedure for dissolving the dried metabolite yeast extracts?

The recommended procedure for solubilizing ISO1 and ISO1-UNL is as follows:

Reconstitute the extract in 2 mL solvent (e.g.; water, 50% methanol).
Vigorously shake by hand with intermittent high-speed vortexing (for a minimum of 2 min).
Centrifuge at 20°C for 5 min at 4000 rcf.
The clear standard solution can then be diluted (1/10 v/v) for direct use or prepared further for calibration and matrix addition.

A video demonstration of this procedure is shown in the resources section above.

Are there alternate reconstitution procedures that can be applied to obtain lipids?

Yes. Reconstituting the dried-down extracts in 2 mL of 50% methanol (instead of 2 mL water) enables lipid detection. Glycerophospholipids (e.g., PCs, PEs, PSs) and lysophospholipids (e.g., LPCs) have been reproducibly identified by RPLC-MS/MS (Q Exactive HF), while other classes have been observed at low concentration.

What sample types have these yeast extracts been measured in?

The extracts have been applied to human tissues (e.g., plasma) and cells (e.g., colon cancer) for QC or quantitative analysis by a variety of LC- and GC-MS methods. Outlined in the user manual (supplied with the kit shipment via QR code) of ISO1, for example, are three application examples that utilize isotope ratio analysis for absolute or relative metabolite quantification.

What are the commonly identified metabolites in the yeast extracts?

A tabulated list of metabolites observed across routine, batch-to-batch measurements is indicated in the metabolite yeast section of the S table Isotope-Labeled Products for Metabolic Research catalog. This list is not finite, as other metabolites have been identified with alternate protocols (e.g., coenzyme As – acetyl, malonyl, propionyl; glucose-1-phosphate; fructose-1-phosphate) or are subject to degradation (e.g., reduced nicotinamide adenine dinucleotide phosphate – NADPH). Please inquire if other metabolites are of interest and we will investigate.

What are examples of other cofactors observed in the yeast extracts?

Additionally detected by HILIC-MS (positive ESI, Q Exactive HF) are NADPH (nicotinamide adenine dinucleotide phosphate), NMN (nicotinamide mononucleotide), and ADPR (adenosine diphosphate ribose).

What classes of metabolites are commonly observed?

These extracts enable the routine identification of a breadth of metabolites across a panel of classes. This minimally includes amino acids and derivatives, organic acids and conjugates, sugar and sugar phosphates, vitamins, and coenzymes. Additionally characterized are a collection of nucleobases, nucleosides, and nucleotides, as well as lipids.

References

Castoldi, A.; Monteiro, L.B.; van Teijlingen Bakker, N.; et al. 2020. Triacylglycerol synthesis enhances macrophage inflammatory function. Nat Commun, 11(1), 4107. PMID: 32796836

Mairinger, T.; Weiner, M.; Hann, S.; et al. 2020. Selective and accurate quantification of N-acetylglucosamine in biotechnological cell samples via GC-MS/MS and GC-TOFMS. Anal Chem, 92(7), 4875-4883. PMID: 32096989

Rusz, M.; Rampler, E.; Keppler, B.K.; et al. 2019. Single spheroid metabolomics: optimizing sample preparation of three-dimensional multicellular tumor spheroids. Metabolites, 9(12). 304. PMID: 31847430

Zhang, Y.; Vera, J.M.; Xie, D.; et al. 2019. Multiomic fermentation using chemically defined synthetic hydrolyzates revealed multiple effects of lignocellulose-derived inhibitors on cell physiology and xylose utilization in Zymomonas mobilis. Front Microbiol, 10, 2596. P MID: 31787963

Galvez, L.; Rusz, M.; Schwaiger-Haber, M.; et al. 2019. Preclinical studies on metal based anticancer drugs as enabled by integrated metallomics and metabolomics. Metallomics, 11(10), 1716-1728. PMID: 31497817

Si-Hung, L.; Troyer, C.; Causon, T.; et al. 2019. Sensitive quantitative analysis of phosphorylated primary metabolites using selective metal oxide enrichment and GC- and IC-MS/MS. Talanta, 205, 120147. PMID: 31450417

van Tol, W.; van Scherpenzeel, M.; Alsady, M.; et al. 2019. Cytidine diphosphate-ribitol analysis for diagnostics and treatment monitoring of cytidine diphosphate-l-ribitol pyrophosphorylase A muscular dystrophy. Clin Chem, 65(10), 1295-1306. PMID: 31375477

Causon, T.J.; Si-Hung, L.; Newton, K.; et al. 2019. Fundamental study of ion trapping and multiplexing using drift tube-ion mobility time-of-flight mass spectrometry for non-targeted metabolomics. Anal Bioanal Chem, 411(24), 6265-6274. PMID: 31302708

Sullivan, M.R; Danai, L.V.; Lewis, C.A.; et al. 2019. Quantification of microenvironmental metabolites in murine cancers reveals determinants of tumor nutrient availability. Elife, 8, e44235. PMID: 30990168

Demarest, T.G.; Truong, G.T.D.; Lovett, J.; et al. 2019. Assessment of NAD⁺ metabolism in human cell cultures, erythrocytes, cerebrospinal fluid and primate skeletal muscle. Anal Biochem, 572, 1-8. PMID: 30822397

Hermann, G.; Schwaiger, M.; Volejnik, P.; et al. 2018. ¹³C-labelled yeast as internal standard for LC-MS/MS and LC high resolution MS-based amino acid quantification in human plasma. J Pharm Biomed Anal, 155, 329-334. PMID: 29704823

Guijas, C.; Montenegro-Burke, J.R.; Domingo-Almenara, X.; et al. 2018. METLIN: A technology platform for identifying knowns and unknowns. Anal Chem, 90(5), 3156-3164. PMID: 29381867

Si-Hung, L.; Causon, T.J.; Hann, S. 2017. Comparison of fully wettable RPLC stationary phases for LC-MS-based cellular metabolomics. Electrophoresis, 38(18), 2287-2295. PMID: 28691762

Schwaiger, M.; Rampler, E.; Hermann, G., et al. 2017. Anion-exchange chromatography coupled to high-resolution mass spectrometry: a powerful tool for merging targeted and non-targeted metabolomics. Anal Chem, 89(14), 7667-7674. PMID: 28581703

Ortmayr, K.; Hann, S.; Koellensperger, G. 2015. Complementing reversed-phase selectivity with porous graphitized carbon to increase the metabolome coverage in an on-line two-dimensional LC-MS setup for metabolomics. Analyst, 140(10), 3465-3473. PMID: 25824707

Krista Backiel

Marketing Manager and Metabolomics Manager

Krista Backiel is responsible for managing and promoting products that are utilized in metabolomics and clinical/diagnostic MS. She spends a lot of her time developing new products to assist customers in their diverse research efforts.

Andrew Percy, PhD

Senior Applications Chemist – Mass Spectrometry

Dr. Andrew Percy is the Senior Applications Chemist for Mass Spectrometry and the MS ‘Omics Product Manager at CIL. His responsibilities minimally involve providing technical support, overseeing product development, identifying new product market opportunities, assisting in the analysis of product-related applications, and writing/reviewing marketing literature.

Metabolic Research

Metabolite Yeast Extracts