Quantification of phenolic acid metabolites in humans by LC–MS: a structural and targeted metabolomics approach
Abstract
Aim: Co-metabolism between a human host and the gastrointestinal microbiota generates many small phenolic molecules such as 3-hydroxy-3-(3-hydroxyphenyl)propanoic acid (3,3-HPHPA), which are reported to be elevated in schizophrenia and autism. Characterization of these chemicals, however, has been limited by analytic challenges. Methodology/results: We applied HPLC to separate and quantify over 50 analytes, including multiple structural isomers of 3,3-HPHPA in human cerebrospinal fluid, serum and urine. Confirmation of identity was provided by NMR, by MS and other detection methods. The highly selective methods support rapid quantification of multiple metabolites and exhibit superior chromatographic behavior. Conclusion: An improved ultra-HPLC–MS/MS and structural approaches can accurately quantify 3,3-HPHPA and related analytes in human biological matrices.
Papers of special note have been highlighted as: • of interest; •• of considerable interest
References
- 1 Metabolomics analysis reveals large effects of gut microflora on mammalian blood metabolites. Proc. Natl Acad. Sci. USA 106(10), 3698–3703 (2009). • Overview of gut microbiota effects on metabolites in blood.
- 2 . The interplay between the intestinal microbiota and the brain. Nat. Rev. Microbiol. 10(11), 735–742 (2012). • Overview of gut microbiota-derived interactions with the brain.
- 3 . The co-metabolism within the gut–brain metabolic interaction: potential targets for drug treatment and design. CNS Neurol. Disord. Drug Targets 15(2), 127–134 (2016).
- 4 . The occurrence of (-)-β-m-hydroxyphenyl-hydracrylic acid in human urine. J. Biol. Chem. 225(1), 269–278 (1957).
- 5 . Investigation of the relation between anaerobic bacteria genus clostridium and late-onset autism etiology in children. J. Immunoassay Immunochem. 35(1), 101–109 (2014).
- 6 . Increased urinary excretion of a 3-(3-hydroxyphenyl)-3-hydroxypropionic acid (HPHPA), an abnormal phenylalanine metabolite of Clostridia spp. in the gastrointestinal tract, in urine samples from patients with autism and schizophrenia. Nutr. Neurosci. 13(3), 135–143 (2010). •• Data linking urinary phenolic acid metabolites, intestinal anaerobes and mental illness.
- 7 The human urine metabolome. PLoS ONE 8(9), e73076 (2013).
- 8 . LC–MS-based metabolomics: an update. Arch. Toxicol. 88(8), 1491–1502 (2014).
- 9 . Role of the small intestine, colon and microbiota in determining the metabolic fate of polyphenols. Biochem. Pharmacol. 139, 24–39 (2017). • Overview of intestinal microbiota-mediated metabolism of polyphenols.
- 10 . Sulfation and glucuronidation of phenols: implications in coenyzme Q metabolism. Methods Enzymol. 400, 342–359 (2005).
- 11 . Urinary excretion of phenolic acids in human subjects on a glucose diet. Clin. Chim. Acta 9, 224–227 (1964).
- 12 . Urinary 3-(3-hydroxyphenyl)-3-hydroxypropionic acid, 3-hydroxyphenylacetic acid, and 3-hydroxyhippuric acid are elevated in children with autism spectrum disorders. Biomed. Res. Int. 2016 9485412 (2016). •• Data linking urinary phenolic acid metabolites, intestinal anaerobes and mental illness.
- 13 . Use of deuterated tyrosine and phenylalanine in the study of catecholamine and aromatic amino acid metabolism. In: Proceedings of the Second International Conference on Stable Isotopes, October 20-23, 1975. Oakbrook, IL. Klein ER, Klein PD (Eds). Energy Research and Development Administration, WA, DC, USA, 385–391 (1976).
- 14 . The relative contribution of the small and large intestine to the absorption and metabolism of rutin in man. Free Radic. Res. 40(10), 1035–1046 (2006).
- 15 . Chlorogenic acid, quercetin-3-rutinoside and black tea phenols are extensively metabolized in humans. J. Nutr. 133(6), 1806–1814 (2003).
- 16 . Gas chromatographic and gas chromatographic–mass spectrometric analysis of organic acids in plasma of patients with chronic renal failure. J. Chromatogr. 289, 259–266 (1984).
- 17 . Abnormal blood constituents in acute renal failure. Clin. Chim. Acta 7, 623–633 (1962).
- 18 Identification of plasma and urinary metabolites and catabolites derived from orange juice (poly)phenols: analysis by high-performance liquid chromatography–high-resolution mass spectrometry. J. Agric. Food Chem. 64(28), 5724–5735 (2016).
- 19 Virtual computational chemistry laboratory – design and description. J. Comput. Aided Mol. Des. 19(6), 453–463 (2005).
- 20 Acute decreases in cerebrospinal fluid glutathione levels after intracerebroventricular morphine for cancer pain. Anesth. Analg. 89(5), 1209–1215 (1999).
- 21 Role of intestinal microbiota in the generation of polyphenol-derived phenolic acid mediated attenuation of Alzheimer's disease β-amyloid oligomerization. Mol. Nutr. Food Res. 59(6), 1025–1040 (2015).
- 22 . Quantitative determination of endogenous compounds in biological samples using chromatographic techniques. Trends Anal. Chem. 27(10), 924–933 (2008).
- 23 . Standard operating procedures for pre-analytical handling of blood and urine for metabolomic studies and biobanks. J. Biomol. NMR 49(3–4), 231–243 (2011).
- 24 . Targeted metabolomics analysis identifies intestinal microbiota-derived urinary biomarkers of colonization resistance in antibiotic-treated mice. Antimicrob. Agents Chemother. 61(8), e0047–17 (2017).
- 25 . Measurement of trimethylamine-N-oxide by stable isotope dilution liquid chromatography tandem mass spectrometry. Anal. Biochem. 455, 35–40 (2014).
- 26 Furoxans (oxadiazole-4 N-oxides) with attenuated reactivity are neuroprotective, cross the blood–brain barrier, and improve passive avoidance memory. J. Med. Chem. 61(10), 4593–4607 (2018).
- 27 . GC–MS determination of organic acids with solvent extraction after cation-exchange chromatography. Clin. Chem. 43(12), 2256–2261 (1997).
- 28 . Secretory activity and aryl acid content of serum, urine, and cerebrospinal fluid in normal and uremic man. J. Lab. Clin. Med. 85(5), 723–731 (1975).
- 29 . Precursors and metabolites of phenylethylamine, m and p-tyramine and tryptamine in human lumbar and cisternal cerebrospinal fluid. J. Neurol. Neurosurg. Psychiatry 45(7), 633–639 (1982).
- 30 Cerebrospinal fluid metabolome in mood disorders-remission state has a unique metabolic profile. Sci. Rep. 2, 667 (2012).
- 31 . Mass-fragmentography of nanogram quantities of biogenic amine metabolites in human cerebrospinal fluid and whole rat brain. Biomed. Mass Spectrom. 2, 183–189 (1975).
- 32 . The effect of age on concentrations of monoamines, amino acids, and their related substances in the cerebrospinal fluid. J. Neural. Transm. Park. Dis. Dement. Sect. 5(3), 215–226 (1993).
- 33 HMDB 4.0: the human metabolome database for 2018. Nucleic Acids Res. 46(D1), D608–D617 (2018).
- 34 . Determination of kynurenine by a simple gas–liquid chromatographic method applicable to urine, plasma, brain and cerebrospinal fluid. J. Chromatogr. 146(1), 33–41 (1978).
- 35 Activation of the kynurenine pathway and increased production of the excitotoxin quinolinic acid following traumatic brain injury in humans. J. Neuroinflammation 12, 110 (2015).
- 36 . The origin of indoleacetic acid and indolepropionic acid in rat and human cerebrospinal fluid. J. Neurochem. 34(5), 1087–1092 (1980).
- 37 . Large neutral amino acids levels in primate cerebrospinal fluid do not confirm competitive transport under baseline conditions. Brain Res. 1648(Pt A), 372–379 (2016).
- 38 . A HILIC-MS/MS method for the simultaneous determination of seven organic acids in rat urine as biomarkers of exposure to realgar. J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 905, 37–42 (2012).
- 39 . Urinary metabolites of coumarin and o-coumaric acid. J. Biol. Chem. 234(4), 946–948 (1959).
- 40 . β-p-hydroxyphenylhydracrylic acid as a urinary constituent in a patient with gastrointestinal disease. Clin. Chim. Acta 47(2), 307–314 (1973).
- 41 . Aromatic acids in urine of healthy infants, persistent hyperphenylalaninemia, and phenylketonuria, before and after phenylalanine load. Pediatr. Res. 8(7), 704–709 (1974).
- 42 . Aberrant amino acid transport in fibroblasts from children with autism. Neurosci. Lett. 418(1), 82–86 (2007).
- 43 . A simplified method to quantify dysregulated tyrosine transport in schizophrenia. Schizophr. Res. 150(2–3), 386–391 (2013).
- 44 Antioxidants in health, disease and aging. CNS Neurol. Disord. Drug Targets 10(2), 192–207 (2011).
- 45 Epigenetic modulation of inflammation and synaptic plasticity promotes resilience against stress in mice. Nat. Commun. 9(1), 477 (2018).
- 46 . Recent developments in understanding the role of the gut microbiota in brain health and disease. Ann. N. Y. Acad. Sci. 1420(1), 5–20 (2017).
- 47 . The neuro-endocrinological role of microbial glutamate and GABA signaling. Front. Microbiol. 7, 1934 (2016).
- 48 Recent findings within the microbiota–gut–brain–endocrine metabolic interactome. Pathol. Lab. Med. Int. 9, 21–30 (2017).