Effect of citrulline and glutamine on nitric oxide production in RAW 264.7 cells in an arginine-depleted environment

JPEN J Parenter Enteral Nutr. Jul-Aug 2008;32(4):377-83. doi: 10.1177/0148607108319807.


Background: Nitric oxide (NO) is a highly reactive free radical essential for antimicrobial and tumor immunity as well as endothelial function. Arginine is a limiting factor in NO synthesis. Citrulline can be converted to arginine and might restore NO production when arginine availability is limited, while glutamine may competitively inhibit citrulline availability. We aimed to assess how these amino acids interact to generate NO using an in vitro model.

Methods: RAW 264.7 cells were exposed to various amino acid concentrations before and after lipopolysaccharide (LPS) stimulation, and NO production was assessed.

Results: NO production directly correlated up to 200 microM with arginine available after LPS stimulation (R(2) = 0.99). Provided the same arginine concentrations following LPS stimulation, low arginine precultured cells produced significantly less NO than high arginine precultured cells (P < .01). Citrulline added to low arginine preculture significantly increased NO production compared to cells in low arginine alone (P < .01). When glutamine was withdrawn before and after LPS stimulation, cells precultured in low arginine and citrulline produced NO equivalent to that of high arginine precultured cells. Additional citrulline provided after LPS stimulation additionally improved NO production beyond that observed in cells precultured in high arginine (P < .01), and NO production became less dependent on arginine availability (R(2) = 0.78).

Conclusion: Arginine availability is a limiting factor for NO production. Citrulline is a potential substitute to restore NO production when arginine availability is limited. Glutamine appears to be an important modulator that interferes with citrulline-mediated NO production.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Arginine / metabolism
  • Cell Line
  • Citrulline / antagonists & inhibitors
  • Citrulline / metabolism
  • Citrulline / pharmacology*
  • Dose-Response Relationship, Drug
  • Glutamine / pharmacology*
  • Lipopolysaccharides / pharmacology
  • Macrophages / drug effects*
  • Macrophages / metabolism*
  • Mice
  • Nitric Oxide / biosynthesis*


  • Lipopolysaccharides
  • Glutamine
  • Citrulline
  • Nitric Oxide
  • Arginine