Abstract
The idiopathic inflammatory bowel diseases, which include Crohn’s disease and ulcerative colitis, are multifactorial chronic conditions that result in numerous perturbations of metabolism in the gastrointestinal mucosa. Thus, methodologies for the qualitative and quantitative analysis of small molecule metabolites in mucosal tissues are important for further elucidation of mechanisms driving inflammation and the metabolic consequences of inflammation. High-performance liquid chromatography (HPLC) is a ubiquitous analytical technique that can be adapted for both targeted and non-targeted metabolomic analysis. Here, protocols for reversed-phase (RP) HPLC-based methods using two different detection modalities are presented. Ultraviolet detection is used for the analysis of adenine nucleotide metabolites, whereas electrochemical detection is used for the analysis of multiple amino acid metabolites. These methodologies provide platforms for further characterization of the metabolic changes that occur during gastrointestinal inflammation.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Hagan RL (1994) High-performance liquid chromatography for small-scale studies of drug stability. Am J Hosp Pharm 51:2162–2175
Colgan SP, Fennimore B, Ehrentraut SF (2013) Adenosine and gastrointestinal inflammation. J Mol Med 91:157–164
Bhatt DP, Chen X, Geiger JD, Rosenberger TA (2012) A sensitive HPLC-based method to quantify adenine nucleotides in primary astrocyte cell cultures. J Chromatogr B Analyt Technol Biomed Life Sci 889–890:110–115
Chunn JL, Young HWJ, Banerjee SK, Colasurdo GN, Blackburn MR (2001) Adenosine-dependent airway inflammation and hyperresponsiveness in partially adenosine deaminase-deficient mice. J Immunol 167:4676–4685
Knudsen TB, Winters RS, Otey SK, Blackburn MR, Airhart MJ, Church JK, Skalko RG (1992) Effects of (R)-deoxycoformycin (pentostatin) on intrauterine nucleoside catabolism and embryo viability in the pregnant mouse. Teratology 45:91–103
Lennon PF, Taylor CT, Stahl GL, Colgan SP (1998) Neutrophil-derived 5′-adenosine monophosphate promotes endothelial barrier function via CD73-mediated conversion to adenosine and endothelial A2B receptor activation. J Exp Med 188:1433–1443
Synnestvedt K, Furuta GT, Comerford KM, Louis N et al (2002) Ecto-5′-nucleotidase (CD73) regulation by hypoxia-inducible factor-1 mediates permeability changes in intestinal epithelia. J Clin Invest 110:993–1002
Ward JL, Tse CM (1999) Nucleoside transport in human colonic epithelial cell lines: evidence for two Na+-independent transport systems in T84 and Caco-2 cells. Biochim Biophys Acta 1419:15–22
Barrio JR, Secrist JA III, Leonard NJ (1972) Fluorescent adenosine and cytidine derivatives. Biochem Biophys Res Commun 46:597–604
Tsunoda M (2006) Recent advances in methods for the analysis of catecholamines and their metabolites. Anal Bioanal Chem 386:506–514
Rozet E, Morello R, Lecomte F, Martin GB, Chiap P, Crommen J, Boos KS, Hubert P (2006) Performances of a multidimensional on-line SPE-LC-ECD method for the determination of three major catecholamines in native human urine: validation, risk and uncertainty assessments. J Chromatogr B Analyt Technol Biomed Life Sci 844:251–260
Glover LE, Bowers BE, Saeedi B, Ehrentraut SF et al (2013) Control of creatine metabolism by HIF is an endogenous mechanism of barrier regulation in colitis. Proc Natl Acad Sci U S A 110:19820–19825
Green BT, Lyte M, Kulkarni-Narla A, Brown DR (2003) Neuromodulation of enteropathogen internalization in Peyer’s patches from porcine jejunum. J Neuroimmunol 141:74–82
Brown DR, Price LD (2008) Catecholamines and sympathomimetic drugs decrease early Salmonella Typhimurium uptake into porcine Peyer’s patches. FEMS Immunol Med Microbiol 52:29–35
Collins CB, Aherne CM, Kominsky D, McNamee EN et al (2011) Retinoic acid attenuates ileitis by restoring the balance between T-helper 17 and T regulatory cells. Gastroenterology 141:1821–1831
Monteleone I, Rizzo A, Sarra M, Sica G et al (2011) Aryl hydrocarbon receptor-induced signals up-regulate IL-22 production and inhibit inflammation in the gastrointestinal tract. Gastroenterology 141:237–248
Zelante T, Iannitti Rossana G, Cunha C, De Luca A et al (2013) Tryptophan catabolites from microbiota engage aryl hydrocarbon receptor and balance mucosal reactivity via interleukin-22. Immunity 39:372–385
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2016 Springer Science+Business Media New York
About this protocol
Cite this protocol
Kao, D.J., Lanis, J.M., Alexeev, E., Kominsky, D.J. (2016). HPLC-Based Metabolomic Analysis of Normal and Inflamed Gut. In: Ivanov, A. (eds) Gastrointestinal Physiology and Diseases. Methods in Molecular Biology, vol 1422. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3603-8_7
Download citation
DOI: https://doi.org/10.1007/978-1-4939-3603-8_7
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3601-4
Online ISBN: 978-1-4939-3603-8
eBook Packages: Springer Protocols