Simultaneous quantitation of isoprenoid pyrophosphates in plasma and cancer cells using LC-MS/MS

Yashpal S. Chhonker, Staci L. Haney, Veenu Bala, Sarah A Holstein, Daryl J Murry

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Isoprenoids (IsoP) are an important class of molecules involved in many different cellular processes including cholesterol synthesis. We have developed a sensitive and specific LC-MS/MS method for the quantitation of three key IsoPs in bio-matrices, geranyl pyrophosphate (GPP), farnesyl pyrophosphate (FPP), and geranylgeranyl pyrophosphate (GGPP). LC-MS/MS analysis was performed using a Nexera UPLC System connected to a LCMS-8060 (Shimadzu Scientific Instruments, Columbia, MD) with a dual ion source. The electrospray ionization source was operated in the negative MRM mode. The chromatographic separation and detection of analytes was achieved on a reversed phase ACCQ-TAG Ultra C18 (1.7 µm, 100 mm × 2.1 mm I.D.) column. The mobile phase consisted of (1) a 10 mM ammonium carbonate with 0.1% ammonium hydroxide in water, and (2) a 0.1% ammonium hydroxide in acetonitrile/methanol (75/25). The flow rate was set to 0.25 mL/min in a gradient condition. The limit of quantification was 0.04 ng/mL for all analytes with a correlation coefficient (r2) of 0.998 or better and a total run time of 12 min. The inter- and intra-day accuracy (85–115%) precision (<15%), and recovery (40–90%) values met the acceptance criteria. The validated method was successfully applied to quantitate basal concentrations of GPP, FPP and GGPP in human plasma and in cultured cancer cell lines. Our LC-MS/MS method may be used for IsoP quantification in different bio-fluids and to further investigate the role of these compounds in various physiological processes.

Original languageEnglish (US)
Article number3275
JournalMolecules
Volume23
Issue number12
DOIs
StatePublished - Dec 11 2018

Fingerprint

Ammonium Hydroxide
Diphosphates
Terpenes
Ion sources
Plasma Cells
hydroxides
cancer
Cells
Plasma (human)
Plasmas
Columbia (Orbiter)
Electrospray ionization
cholesterol
cultured cells
correlation coefficients
acceptability
ion sources
Physiological Phenomena
acetonitrile
Methanol

Keywords

  • Farnesyl pyrophosphate
  • Geranyl pyrophosphate
  • Geranylgeranyl pyrophosphate
  • LC-MS/MS

ASJC Scopus subject areas

  • Analytical Chemistry
  • Chemistry (miscellaneous)
  • Molecular Medicine
  • Pharmaceutical Science
  • Drug Discovery
  • Physical and Theoretical Chemistry
  • Organic Chemistry

Cite this

Simultaneous quantitation of isoprenoid pyrophosphates in plasma and cancer cells using LC-MS/MS. / Chhonker, Yashpal S.; Haney, Staci L.; Bala, Veenu; Holstein, Sarah A; Murry, Daryl J.

In: Molecules, Vol. 23, No. 12, 3275, 11.12.2018.

Research output: Contribution to journalArticle

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abstract = "Isoprenoids (IsoP) are an important class of molecules involved in many different cellular processes including cholesterol synthesis. We have developed a sensitive and specific LC-MS/MS method for the quantitation of three key IsoPs in bio-matrices, geranyl pyrophosphate (GPP), farnesyl pyrophosphate (FPP), and geranylgeranyl pyrophosphate (GGPP). LC-MS/MS analysis was performed using a Nexera UPLC System connected to a LCMS-8060 (Shimadzu Scientific Instruments, Columbia, MD) with a dual ion source. The electrospray ionization source was operated in the negative MRM mode. The chromatographic separation and detection of analytes was achieved on a reversed phase ACCQ-TAG Ultra C18 (1.7 µm, 100 mm × 2.1 mm I.D.) column. The mobile phase consisted of (1) a 10 mM ammonium carbonate with 0.1{\%} ammonium hydroxide in water, and (2) a 0.1{\%} ammonium hydroxide in acetonitrile/methanol (75/25). The flow rate was set to 0.25 mL/min in a gradient condition. The limit of quantification was 0.04 ng/mL for all analytes with a correlation coefficient (r2) of 0.998 or better and a total run time of 12 min. The inter- and intra-day accuracy (85–115{\%}) precision (<15{\%}), and recovery (40–90{\%}) values met the acceptance criteria. The validated method was successfully applied to quantitate basal concentrations of GPP, FPP and GGPP in human plasma and in cultured cancer cell lines. Our LC-MS/MS method may be used for IsoP quantification in different bio-fluids and to further investigate the role of these compounds in various physiological processes.",
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AU - Chhonker, Yashpal S.

AU - Haney, Staci L.

AU - Bala, Veenu

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AU - Murry, Daryl J

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KW - Geranyl pyrophosphate

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