• History & Evolution
• Biosynthesis & dietary uptake
• EPA and the microbiome
• EPA and inflammation
• EPA and neurology
• EPA and oncology
• EPA and 5P medicine
• References

History & Evolution

1957: first isolated (Klenk, E. and Eberhagen, D., 1957) | 1964: EPA linked to prostaglandin production (Bergström, S. et al., 1964) | 1978: discovery of cardiovascular protective effects

Eicosapentaenoic acid (EPA) is a long-chain omega-3 polyunsaturated fatty acid (PUFA) and a precursor to many lipid signaling molecules involved in inflammation and cardiometabolic regulation. The name reflects its structure: “eicosa” refers to its 20 carbon atoms, while “pentaenoic” refers to five double bonds (Castro, L. et al., 2016).

EPA was first isolated from cod liver oil in 1957 by Klenk and Eberhagen (Klenk, E. and Eberhagen, D., 1957). At the time, lipid research focused on omega-6 fatty acids, particularly arachidonic acid (AA) and linoleic acid, as their roles in cholesterol metabolism and prostaglandin synthesis made them more compelling targets (Spector, A. and Kim, H., 2019). Although Bergström later showed that EPA could also be enzymatically converted into prostaglandin E3 (Bergström, S. et al., 1964), this pathway attracted little attention and interesting early findings on omega-3 fatty acids were largely overlooked.

This changed in 1978, when Dyerberg and Bang observed notably low rates of myocardial infarction among Greenland Eskimos, who typically followed a diet of marine lipids rich in EPA and docosahexaenoic acid (DHA) (Dyerberg, J. and Bang, O., 1978).

By the 1980s, epidemiological and clinical studies linked diets rich in marine omega-3 fatty acids to lower rates of cardiovascular disease, putting EPA at the heart of vascular biology. EPA has since been shown to have protective effects against wide-ranging diseases, including obesity, diabetes, chronic obstructive pulmonary disease and certain cancers (Watabe, S. et al., 2024).

EPA is found primarily in marine microalgae, fish and other marine animals, where it accumulates through the food chain.

Biosynthesis vs. dietary uptake

In humans, EPA comes mainly from the diet, particularly oily fish, seafood and fish-based supplements. However, this is not a reliable source. Fish do not produce EPA themselves but obtain it from algae, so their EPA content varies depending on their diet and environment (Watabe, S. et al., 2024). Concerns about sustainability and contamination of fish and fish-derived supplements have increased interest in alternative sources such as algal oils (Sousa, S. et al., 2024).

EPA can also be produced endogenously, but only in small amounts. It is synthesized from the essential fatty acid alpha-linolenic acid (ALA), found in foods such as flaxseed, walnuts and vegetable oils (Watabe, S. et al., 2024). This conversion takes place mainly in the liver through a series of enzymatic reactions. The process is quite inefficient: only 1-15% of ALA is converted to EPA, and this can be further reduced by alcohol consumption, smoking, inactivity and dietary factors (Sousa, S. et al., 2024).

As a result, endogenous synthesis does not make a significant contribution to circulating EPA and, for most people, dietary intake is the main source. Recommended daily intake is around 250-500 mg per day (together with downstream metabolite DHA), rising to 700-1000 mg for pregnant and lactating women (Sousa, S. et al., 2024).

Once in circulation, EPA is incorporated into cell membranes where it can be converted into lipid mediators, including prostaglandins, leukotrienes and resolvins (Igarashi, M. et al., 2013).

EPA and the microbiome

Research shows that EPA interacts with the gut microbiome, directly affecting bacterial growth, diversity and survival. As with other PUFAs, EPA has been shown in experimental settings to have an antibacterial effect against Gram-positive species such as Bacillus cereus and Staphylococcus aureus, which are both causes of infection (Le, PNT. and Desbois, A., 2017).

Studies report a decrease in the Firmicutes/Bacteroidetes ratio and increases in genera such as Bifidobacterium, Lachnospira, Roseburia and Lactobacillus (Amedei, A. et al., 2025). EPA has also been found to modulate levels of pro-inflammatory molecules and affect short-chain fatty acid (SCFA) concentrations (Fu, Y. et al., 2021). These effects are associated with improved cardiovascular risk profiles.

In turn, beneficial bacteria including Bifidobacterium, Lactobacillus and Akkermansia have been shown to influence EPA absorption and metabolism, indicating a two-way interaction between EPA and the gut microbiome (Amedei, A. et al., 2025).

EPA and inflammation

Inflammation is one of the main pathways through which EPA affects human health.
As a precursor to lipid signaling molecules, EPA influences how inflammatory mediators are produced. In cell membranes, it partly replaces omega-6 fatty acids such as arachidonic acid, changing the pool of fatty acids available for enzymatic conversion and leading to the production of eicosanoids that are generally less pro-inflammatory (Crupi, R. and Cuzzocrea, S., 2022). EPA is also converted into specialized pro-resolving mediators, such as resolvins, which help limit and resolve inflammation. More broadly, these changes in membrane composition influence cell signaling, gene expression and the production of cytokines, adhesion molecules and other inflammatory mediators (Calder, P., 2010).

These pathways are also influenced by the exposome. Environmental factors such as diet, pollutants and oxidative stress can alter fatty acid composition and downstream inflammatory signaling (Hinman, J. et al., 2025). Studies show that higher omega-3 status, including EPA levels, is associated with increased pro-resolving and reduced pro-inflammatory lipid mediator responses following exposures such as air pollution, highlighting EPA’s role in shaping inflammatory responses to environmental stressors (Chen, H. et al., 2025).

Clinically, this translates into measurable effects on disease-related inflammation. EPA has been shown to reduce pro-inflammatory cytokines and oxidative stress, improve endothelial function and support stabilization of atherosclerotic plaques, all of which are relevant to cardiovascular risk (Crupi, R. and Cuzzocrea, S., 2022). In inflammatory conditions such as rheumatoid arthritis, higher intakes of EPA (typically with DHA) are associated with reduced inflammatory activity, although relatively high and sustained doses are often required (Calder, P., 2010). A 2024 literature review found that EPA is linked to improved metabolic and inflammatory profiles in non-communicable diet-related conditions such as obesity, diabetes and cardiovascular disease (Banaszak, M. et al., 2024).

A 2021 study using targeted lipidomics linked EPA’s anti-inflammatory effects to improvements in depression (Borsini, A. et al., 2021). Using a “depression in a dish” model of human hippocampal cells, researchers showed that EPA-derived lipid mediators protected neurons from inflammation-related damage and supported neurogenesis. In patients with major depressive disorder, higher levels of these metabolites were associated with reduced symptom severity, suggesting a mechanistic link between EPA, inflammation and depression.

EPA and neurology

EPA has been studied in neurology because of its effect on synaptic function, neuroinflammation and neuronal survival. In a preclinical study, EPA was taken up by both neurons and glial cells (Kawashima, A. et al., 2010). This increased long-term potentiation in the hippocampus and activated intracellular signaling involving phosphoinositide 3-kinase (PI3K) and protein kinase B (Akt), while reducing markers of cell death. These findings suggest a potential role in protecting against neurodegeneration and support interest in EPA in conditions such as Alzheimer’s disease, Huntington’s disease and schizophrenia.

More recent work has looked at EPA in neurodevelopmental disorders and epilepsy, where inflammation, oxidative stress and synaptic dysfunction are recurring factors (Li, M. et al., 2025). Omega-3 fatty acids, including EPA, may help in attention deficit hyperactivity disorder (ADHD), autism spectrum disorder and Tourette syndrome, but the evidence is mixed and clinical effects are often modest. For example, in ADHD, some studies report improvement in attention or behavior from higher EPA doses, while others find no clear benefit. In epilepsy, EPA and other omega-3 fatty acids have been explored mainly as adjuncts rather than stand-alone treatments, with some studies reporting reduced inflammatory markers or seizure frequency.

Omics studies suggest that lipid dysregulation, including EPA, may contribute to neurological disease progression. For example, in a 2025 longitudinal metabolomic-lipidomic study of 767 people with multiple sclerosis, decreased serum EPA was associated with slower walking speed and reduced manual dexterity (Noroozi, R. et al., 2026).

A 2026 study found that after repetitive mild traumatic brain injury, EPA accumulates in the brain under baseline conditions but is then depleted during injury-related metabolic stress (Karakaya, E. et al., 2026). This was associated with impaired endothelial repair, reduced angiogenic signaling and vascular and cognitive decline.

These findings make EPA a metabolite of interest as a marker of and potential therapeutic target in neuroinflammatory and neurovascular processes.

EPA and oncology

The relationship between EPA and cancer is unclear. The metabolite has been shown to suppress proliferation in colon, pancreatic, breast, esophageal and other cancer cell lines (Mizoguchi, K. et al., 2014), and EPA-derived resolvins seem to have an anti-tumor effect (Kiyasu, Y. et al., 2024).

However, clinical studies show mixed results. In animal models, for example, fish oil has been shown to both suppress colorectal carcinogenesis and to promote colitis-associated cancer (Kiyasu, Y. et al., 2024).

Evidence in patients is also limited. Small studies in patients undergoing chemotherapy report changes in inflammatory markers and immune function with EPA (often combined with DHA) (Munhoz, J. et al., 2025). However, findings are inconsistent, and there is currently insufficient evidence to support routine use in cancer care.

EPA and 5P medicine

EPA’s links to diet, genetics and inflammation make it an interesting metabolite for the study of chronic, non-communicable diseases, which are a top priority in population health. In epidemiological research, circulating omega-3 fatty acids, including EPA, are used as objective biomarkers of dietary intake and have been associated with cardiovascular outcomes and mortality risk across large cohorts and meta-analyses (Jiang, H. et al., 2022).

Studies also show that EPA’s effects depend on both exposure (diet) and host biology (genetics), with implications for precision medicine. For example, in the seAFOod polyp prevention trial, EPA supplementation was found to reduce adenoma risk by around 50% in individuals with a specific FADS variant, while those without the allele saw no benefit (Sun, G. et al., 2024).

EPA also seems to play a role in aging, making it increasingly relevant as populations age and the burden of chronic disease grows (Hinman, J. et al., 2025). Studies show changes in PUFA profiles and inflammatory and oxidative stress markers across the lifespan (Aiello, A. et al., 2024). In long-lived women, a lower AA to EPA ratio was associated with higher antioxidant capacity (Hinman, J. et al., 2025).

References

Aiello, A. et al. (2024). Polyunsaturated fatty acid status and markers of oxidative stress and inflammation across the lifespan: A cross-sectional study in a cohort with long-lived individuals. Experimental Gerontology, 195, 112531. DOI: https://doi.org/10.1016/j.exger.2024.112531.

Amedei, A. et al. (2025). Potential and Future Therapeutic Applications of Eicosapentaenoic/Docosahexaenoic Acid and Probiotics in Chronic Low-Grade Inflammation. Biomedicines, 13(10), 2428. DOI: https://doi.org/10.3390/biomedicines13102428.

Banaszak, M. et al. (2024). Role of Omega-3 fatty acids eicosapentaenoic (EPA) and docosahexaenoic (DHA) as modulatory and anti-inflammatory agents in noncommunicable diet-related diseases – Reports from the last 10 years. Clinical Nutrition ESPEN, 63, 240-258. DOI: https://doi.org/10.1016/j.clnesp.2024.06.053.

Bergström, S. et al. (1964). The Enzymatic Conversion of Essential Fatty Acids into Prostaglandins: PROSTAGLANDINS AND RELATED FACTORS 34. Journal of Biological Chemistry, 239, PC4006-PC4008. DOI: https://doi.org/10.1016/S0021-9258(18)91234-2.

Borsini, A. et al. (2021). Omega-3 polyunsaturated fatty acids protect against inflammation through production of LOX and CYP450 lipid mediators: relevance for major depression and for human hippocampal neurogenesis. Molecular Psychiatry, 26, 6773–6788. DOI: https://doi.org/10.1038/s41380-021-01160-8.

Calder, P. (2010). Omega-3 Fatty Acids and Inflammatory Processes. Nutrients, 2(3), 355–374. DOI: https://doi.org/10.3390/nu2030355.

Castro, L. et al. (2016). Long-chain polyunsaturated fatty acid biosynthesis in chordates: Insights into the evolution of Fads and Elovl gene repertoire. Progress in Lipid Research, 62, 25-40. DOI: https://doi.org/10.1016/j.plipres.2016.01.001.

Chen, H. et al. (2025). Omega-3 Fatty Acid Intake and Oxylipin Production in Response to Short-Term Ambient Air Pollution Exposure in Healthy Adults. Toxics, 13(12), 1063. DOI: https://doi.org/10.3390/toxics13121063.

Crupi, R. and Cuzzocrea, S. (2022). Role of EPA in Inflammation: Mechanisms, Effects, and Clinical Relevance. Biomolecules, 12(2), 242. DOI: https://doi.org/10.3390/biom12020242.

Dyerberg, J. and Bang, O. (1978). Dietary fat and thrombosis. The Lancet, 311(8056), 152. DOI: https://doi.org/10.1016/S0140-6736(78)90448-8.

Fu, Y. et al. (2021). Associations among Dietary Omega-3 Polyunsaturated Fatty Acids, the Gut Microbiota, and Intestinal Immunity. Mediators Inflamm, 8879227. DOI: https://doi.org/10.1155/2021/8879227.

Hinman, J. et al. (2025). Effects of molecular interactions between the exposome and oxylipin metabolism on healthspan. Front. Physiol., 16, DOI: https://doi.org/10.3389/fphys.2025.1584195.

Igarashi, M. et al. (2013). Kinetics of eicosapentaenoic acid in brain, heart and liver of conscious rats fed a high n-3 PUFA containing diet. Prostaglandins Leukot Essent Fatty Acids, 89(6), 403–412. DOI: https://doi.org/10.1016/j.plefa.2013.09.004.

Jiang, H. et al. (2022). Omega-3 polyunsaturated fatty acid biomarkers and risk of type 2 diabetes, cardiovascular disease, cancer, and mortality. Clin Nutr, 41(8), 798-1807. DOI: https://doi.org/10.1016/j.clnu.2022.06.034.

Karakaya, E. et al. (2026). Eicosapentaenoic acid reprograms cerebrovascular metabolism and impairs repair after brain injury, with relevance to chronic traumatic encephalopathy. Cell Reports, 117135. DOI: https://doi.org/10.1016/j.celrep.2026.117135.

Kawashima, A. et al. (2010). Effects of eicosapentaenoic acid on synaptic plasticity, fatty acid profile and phosphoinositide 3-kinase signaling in rat hippocampus and differentiated PC12 cells. J Nutr Biochem, 21(4), 268-77. DOI: https://doi.org/10.1016/j.jnutbio.2008.12.015. Epub 2009.

Kiyasu, Y. et al. (2024). EPA, DHA, and resolvin effects on cancer risk: the underexplored mechanisms. Prostaglandins Other Lipid Mediat, 174, 106854. DOI: https://doi.org/10.1016/j.prostaglandins.2024.106854.

Klenk, E. and Eberhagen, D. (1957). [Occurrence of delta5, 8, 11, 14, 17-n-eicosapentanoic acid in cod liver oil and its isolation] (Article in German). Hoppe Seylers Z Physiol Chem, 307(1), 42-8.

Le, PNT. and Desbois, A. (2017). Antibacterial Effect of Eicosapentaenoic Acid against Bacillus cereus and Staphylococcus aureus: Killing Kinetics, Selection for Resistance, and Potential Cellular Target. Mar Drugs, 15(11), 334. DOI: https://doi.org/10.3390/md15110334.

Li, M. et al. (2025). Omega-3 fatty acids: multi-target mechanisms and therapeutic applications in neurodevelopmental disorders and epilepsy. Front. Nutr. , 12, 1598588. DOI: https://doi.org/10.3389/fnut.2025.1598588.

Mizoguchi, K. et al. (2014). Induction of Apoptosis by Eicosapentaenoic Acid in Esophageal Squamous Cell Carcinoma. Anticancer Research December, 34(12), 7145-7149. PMID: 25503142.

Munhoz, J. et al. (2025). Perspective: Implications of Docosahexaenoic Acid and Eicosapentaenoic Acid Supplementation on the Immune System during Cancer Chemotherapy: Perspectives from Current Clinical Evidence. Advances in Nutrition, 16(8), 100464. DOI: https://doi.org/10.1016/j.advnut.2025.100464.

Noroozi, R. et al. (2026). Serum metabolomic-lipidomic signatures track long-term neurological performance in multiple sclerosis. Mol. Neurodegener. Adv., 2(1), DOI: https://doi.org/10.1186/s44477-025-00008-7.

Sousa, S. et al. (2024). Factors impacting the microbial production of eicosapentaenoic acid. Appl Microbiol Biotechnol, 108(1), 368. DOI: https://doi.org/10.1007/s00253-024-13209-z.

Spector, A. and Kim, H. (2019). Emergence of omega-3 fatty acids in biomedical research. Prostaglandins, Leukotrienes and Essential Fatty Acids, 140, 47-50. DOI: https://doi.org/10.1016/j.plefa.2018.11.017.

Sun, G. et al. (2024). Fatty acid desaturase insertion-deletion polymorphism rs66698963 predicts colorectal polyp prevention by the n-3 fatty acid eicosapentaenoic acid: a secondary analysis of the seAFOod polyp prevention trial. Am J Clin Nutr, 120(2), 360-368. DOI: https://doi.org/10.1016/j.ajcnut.2024.06.004.

Watabe, S. et al. (2024). Daily Consumption of α-Linolenic Acid Increases Conversion Efficiency to Eicosapentaenoic Acid in Mice. Nutrients, 16(10), 407. DOI: https://doi.org/10.3390/nu16101407.

• History & Evolution
• Biosynthesis & dietary uptake
• Homovanillic acid and the microbiome
• Homovanillic acid and neurology
• Homovanillic acid and oncology
• Homovanillic acid and 5P medicine
• References

History & Evolution

1950s: discovery | 1963: detected in brain and cerebrospinal fluid (Andén, N. et al., 1963) | 1985: metabolic pathway identified (Westerink, B., 1985)

Homovanillic acid (HVA) is a monocarboxylic acid best known for its role as a major dopamine metabolite. The name “homovanillic” refers to the compound’s relationship to vanillic acid, a similar phenolic acid, with the prefix homo- indicating the addition of a methylene group (–CH2–) between the aromatic ring and the carboxyl group.

Interest in HVA began in the 1950s and 60s, when early chromatography studies detected HVA in the urine of mammals and identified it as a marker of dopamine turnover in the brain (Armstrong, M. et al., 1956) (Williams, C. et al., 1960). In 1963, Andén et al. showed that HVA was present in the brain and cerebrospinal fluid (CSF), and introduced a fluorometric method for its detection (Andén, N. et al., 1963). Following the dopamine thread, researchers in the following decades focused on the associations between HVA, dopamine and neurological and psychiatric disorders, including Parkinson’s disease (PD), schizophrenia, depression and, more recently, neuroblastoma. Current research is directed at improving HVA’s diagnostic and prognostic utility as a biomarker in these conditions and refining techniques for its detection in different bodily fluids.

Although primarily found in humans and other mammals, as a phenolic compound HVA is also detectable in some bacteria and plants (Marhuenda-Muñoz, M. et al., 2019). It has been identified in beer, suggesting it can be produced during the fermentation process by microorganisms (Montanari, L. et al., 1999).

Biosynthesis vs. dietary uptake

HVA forms in the body as the final breakdown product of dopamine, through a two-step enzymatic process (Buleandră, M. et al., 2025). First, monoamine oxidase (MAO) breaks down dopamine into 3,4-dihydroxyphenylacetic acid (DOPAC) in dopaminergic neurons in the brain. Then, catechol-O-methyltransferase (COMT) methylates DOPAC and converts it to HVA. An alternative pathway involves COMT acting on dopamine to form 3-methoxytyramine, which MAO then breaks down to form HVA. HVA can also be biosynthesized from homovanillin through the action of the enzyme aldehyde dehydrogenase.

HVA enters the circulation from both the brain and peripheral tissues such as the lungs, liver and skeletal muscle, and is mainly excreted via the kidneys (Lambert, G. et al., 1993).

Clinically, HVA can be measured in urine, plasma and CSF. Because of the distance between site of production and site of measurement, all three matrices can be vulnerable to confounding factors (Amin, F. et al., 1992). In healthy adults, CSF concentrations typically fall in the range of hundreds of nanomoles per liter (Blennow, K. et al., 1993). Reference ranges for urinary HVA are between 0.8 to 35.0 µmol/mmol of creatinine (Newcastle Hospital). Plasma HVA has a half-life of around one hour, and may be affected by diet for several hours (Köhnke, M. et al., 2003). Fasting plasma levels vary by sex and age, and differences between females and males hold regardless of menopause and hormone treatments in transgender patients (Giltay, E. et al., 2005).

HVA may also form through dietary and microbial pathways. Dietary flavonols commonly found in tomatoes, onions, and tea, can lead to significantly elevated levels of urinary HVA (Combet, E. et al., 2011). Likewise, the microbial digestion of hydroxytyrosol (found in olive oil) can also lead to elevated levels of HVA in humans (Tuck, K. et al., 2002).

Homovanillic acid and the microbiome

Research shows links between HVA and gut microbiota. Zhao et al. found that gut bacterial species, including Bifidobacterium longum and Roseburia intestinalis, were depleted in patients with depression (Zhao, M. et al, 2024). Using metabolomics, they showed that B. longum could directly produce HVA in the gut with the substrates of mouse fodder and tyrosine. R. intestinalis promoted the growth of B. longum, in turn increasing HVA levels in the gut. R. faecis and Eubacterium rectale were also strongly correlated with HVA levels.

Another recent metabolomics study by Gątarek et al. investigated the relationship between gut-derived metabolites, including HVA, and Parkinson’s disease (Gątarek, P. et al., 2025). Here, urinary metabolite profiles indicated elevated levels of HVA and succinic acid , and reduced levels of trimethylamine N-oxide in PD patients compared to controls. These findings highlight how altered gut microbiota may influence HVA levels and in turn affect host dopamine catabolism.

Homovanillic acid and neurology

As an indicator of dopaminergic activity, HVA has long been studied as a marker in neurological and psychiatric diseases in which dopamine plays a role. In PD, HVA shows different patterns depending on the biological matrix analyzed. While urinary HVA may increase in PD patients compared to controls, CSF HVA levels are significantly lower in patients with PD, particularly those with akinesia, indicating reduced dopaminergic activity (Davidson, D. et al., 1977). HVA concentration increases with levodopa therapy, though this is not always associated with clinical improvement (Jiménez-Jiménez, F. et al., 2014).

Altered HVA has also been described in mood and cognitive disorders. CSF HVA levels are reduced in dementia with Lewy bodies and Alzheimer’s disease, compared to healthy subjects (Morimoto, S. et al., 2017). In a mouse model, administering HVA was found to improve symptoms of depression by inhibiting synaptic autophagy (Zhao, M. et al, 2024).

Autism spectrum disorder (ASD) has been linked to elevated urinary HVA using metabolomics (Gevi, F. et al., 2020). Higher HVA concentrations have also been associated with greater symptom severity, including agitation, stereotyped behaviors and reduced spontaneous activity (Kaluzna-Czaplinska, J. et al., 2010). Supplementation with vitamin B6, which has been found to be lacking in children with ASD, appears to reduce both HVA concentrations and neurological symptoms (Gątarek, P. et al., 2025). HVA has also been studied in relation to attention-deficit/hyperactivity disorder (ADHD), though findings are inconclusive (Predescu, E. et al., 2024).

In schizophrenia, plasma and CSF HVA often track with symptom severity, but the relationship is complicated. A 2021 review notes that lower CSF HVA often correlates with more severe symptoms and a poorer prognosis (Gasnier, M. et al., 2021). Other work indicates a positive correlation between HVA concentrations and symptom severity (Mazure, C. et al., 1991). Antipsychotic treatment typically causes an initial increase in HVA, followed by a gradual decrease (Gasnier, M. et al., 2021).

Homovanillic acid and oncology

HVA is a marker of catecholamine-secreting tumors, such as neuroblastoma, pheochromocytoma and other neural crest tumors. These tumors secrete excess catecholamines like dopamine, which is then broken down into HVA. Therefore, HVA levels can be a useful indicator of the tumor’s activity. Elevated urinary HVA and vanillylmandelic acid (VMA) are detected in children with neuroblastoma, making them standard tools for diagnosis, monitoring, and prognosis (Sadilkova, K. et al., 2013). A low VMA to HVA ratio is associated with more aggressive disease (Zambrano, E. and Reyes-Múgica, M., 2002).

Infant screening programs based on HVA/VMA were introduced in Japan and elsewhere in the 1980s, but later abandoned after large trials showed no survival benefit and significant overdiagnosis (Tajiri, T. et al., 2009).

While HVA remains clinically relevant in the diagnosis of neuroblastoma, a scoring system using urinary 3-methoxytyramine sulfate (3-MTS) and vanillactic acid has recently been found to be more sensitive than one based on HVA and VMA (Amano, H. et al., 2024).

Homovanillic acid and 5P medicine

HVA’s relationship to host behavior, genetics and microbial activity makes it highly relevant to 5P medicine. As a marker of dopaminergic function and treatment efficacy, HVA may help to refine and personalize different treatment strategies for patients. Advances in metabolomics now allow rapid, high-sensitivity quantification of HVA and vanillylmandelic acid in urine, streamlining diagnostic processes for a whole range of conditions (Pandya, V. and Frank, E., 2022).

At the population level, genome-wide association studies (GWAS) have identified genetic and epidemiological patterns of HVA (Luykx, J. et al., 2014). Multiomics studies are providing further insights, for example, into the causal genetic relationship between HVA and conditions such as eating disorders and schizophrenia (Wen, J. et al., 2025).

Outside the clinic, HVA also plays a role in public health. HVA and related metabolites are used in wastewater-based epidemiology, where their excretion rates correlate with catchment size and provide population-wide biomarkers for exposure and health monitoring (Pandopulos, A. et al., 2021).

References

Amano, H. et al. (2024). Scoring system for diagnosis and pretreatment risk assessment of neuroblastoma using urinary biomarker combinations. Cancer Sci, 115(5), 1634–1645 | https://doi.org/10.1111/cas.16116.

Amin, F. et al. (1992). Homovanillic Acid Measurement in Clinical Research: A Review of Methodology. Schizophrenia Bulletin, 18(1), 123-48 | https://doi.org/10.1093/schbul/18.1.123.

Andén, N. et al. (1963). On the occurrence of homovanillic acid in brain and cerebrospinal fluid and its determination by a fluorometric method. Life Sciences, 2(7), 448-458 | https://doi.org/10.1016/0024-3205(63)90132-2.

Armstrong, M. et al. (1956). The Phenolic Acids of Human Urine: Paper Chromatography of Phenolic Acids. J Biol Chem, 218(1), 293-303 | https://doi.org/10.1016/S0021-9258(18)65893-4.

Blennow, K. et al. (1993). Cerebrospinal fluid monoamine metabolites in 114 healthy individuals 18-88 years of age. Eur Neuropsychopharmacol, 3(1), 55-61 | https://doi.org/10.1016/0924-977x(93)90295-w.

Buleandră, M. et al. (2025). Electrochemical Study and Determination of Homovanillic Acid, the Final Metabolite of Dopamine, Using an Unmodified Disposable Electrode. Molecules, 30(2), 369 | https://doi.org/10.3390/molecules30020369.

Combet, E. et al. (2011). Dietary flavonols contribute to false-positive elevation of homovanillic acid, a marker of catecholamine-secreting tumors. Clinica Chimica Acta, 412(1-2), 165-169 | https://doi.org/10.1016/j.cca.2010.09.037.

Davidson, D. et al. (1977). CSF studies on the relationship between dopamine and 5-hydroxytryptamine in Parkinsonism and other movement disorders. J Neurol Neurosurg Psychiatry, 40(12), 1136–1141 | https://doi.org/10.1136/jnnp.40.12.1136.

Gątarek, P. et al. (2025). The Determination of Trimethylamine N-Oxide and Organic Acids Connected with Gut Microbiota in the Urine of Parkinson’s Disease Patients: A Pilot Study. Int J Mol Sci, 26(10), 4575 | https://doi.org/10.3390/ijms26104575.

Gasnier, M. et al. (2021). A New Look on an Old Issue: Comprehensive Review of Neurotransmitter Studies in Cerebrospinal Fluid of Patients with Schizophrenia and Antipsychotic Effect on Monoamine’s Metabolism. Clin Psychopharmacol Neurosci, 19(3), 395–410 | https://doi.org/10.9758/cpn.2021.19.3.395.

Gevi, F. et al. (2020). A metabolomics approach to investigate urine levels of neurotransmitters and related metabolites in autistic children. Biochimica et Biophysica Acta (BBA) – Molecular Basis of Disease, 1866(10), 165859 | https://doi.org/10.1016/j.bbadis.2020.165859.

Giltay, E. et al. (2005). The sex difference of plasma homovanillic acid is unaffected by cross-sex hormone administration in transsexual subjects. Journal of Endocrinology, 187(1) | https://doi.org/10.1677/joe.1.06307.

Jiménez-Jiménez, F. et al. (2014). Cerebrospinal fluid biochemical studies in patients with Parkinson’s disease: toward a potential search for biomarkers for this disease. Frontiers in Cellular Neuroscience, 8 | https://doi.org/10.3389/fncel.2014.00369.

Köhnke, M. et al. (2003). Plasma Homovanillic Acid: A Significant Association with Alcoholism is Independent of a Functional Polymorphism of the Human Catechol-O-Methyltransferase Gene. Neuropsychopharmacology, 28, 1004–1010 | https://doi.org/10.1038/sj.npp.1300107. Retrieved from Neuropsychopharmacology volume 28, pages1004–1010 (2003).

Kaluzna-Czaplinska, J. et al. (2010). Determination of homovanillic acid and vanillylmandelic acid in urine of autistic children by gas chromatography/mass spectrometry. Medical Science Monitor, 16(9), CR445-50.

Lambert, G. et al. (1993). Regional homovanillic acid production in humans. Life Sciences, 53(1), 63-75 | https://doi.org/10.1016/0024-3205(93)90612-7.

Luykx, J. et al. (2014). Genome-wide association study of monoamine metabolite levels in human cerebrospinal fluid. Mol Psychiatry, 19(2), 228-34 | https://doi.org/10.1038/mp.2012.183.

Marhuenda-Muñoz, M. et al. (2019). Microbial Phenolic Metabolites: Which Molecules Actually Have an Effect on Human Health? Nutrients, 11(11), 2725 | https://doi.org/10.3390/nu11112725.

Mazure, C. et al. (1991). Plasma Free Homovanillic Acid (HVA) as a Predictor of Clinical Response in Acute Psychosis. Biol Psychiatry, 30(5), 475-82 | https://doi.org/10.1016/0006-3223(91)90309-a.

Montanari, L. et al. (1999). Organic and Phenolic Acids in Beer. LWT – Food Science and Technology, 32(8), 535-539 | https://doi.org/10.1006/fstl.1999.0593.

Morimoto, S. et al. (2017). Homovanillic acid and 5-hydroxyindole acetic acid as biomarkers for dementia with Lewy bodies and coincident Alzheimer’s disease: An autopsy-confirmed study. PLoS ONE, 12(2), e0171524 | https://doi.org/10.1371/journal.pone.0171524.

Pandopulos, A. et al. (2021). Application of catecholamine metabolites as endogenous population biomarkers for wastewater-based epidemiology. Science of The Total Environment, 763, 142992 | https://doi.org/10.1016/j.scitotenv.2020.142992.

Pandya, V. and Frank, E. (2022). A Simple, Fast, and Reliable LC-MS/MS Method for the Measurement of Homovanillic Acid and Vanillylmandelic Acid in Urine Specimens. Methods Mol Biol, 2546, 175-183 | https://doi.org/10.1007/978-1-0716-2565-1_16.

Predescu, E. et al. (2024). Metabolomic Markers in Attention-Deficit/Hyperactivity Disorder (ADHD) among Children and Adolescents—A Systematic Review. Int J Mol Sci, 25(8), 4385 | https://doi.org/10.3390/ijms25084385.

Sadilkova, K. et al. (2013). Analysis of vanillylmandelic acid and homovanillic acid by UPLC–MS/MS in serum for diagnostic testing for neuroblastoma. Clinica Chimica Acta, 424, 253-257 | https://doi.org/10.1016/j.cca.2013.06.024.

Tajiri, T. et al. (2009). Risks and benefits of ending of mass screening for neuroblastoma at 6 months of age in Japan. Journal of Pediatric Surgery, 44(12), 2253-2257 | https://doi.org/10.1016/j.jpedsurg.2009.07.050.

Tuck, K. et al. (2002). Structural characterization of the metabolites of hydroxytyrosol, the principal phenolic component in olive oil, in rats. J Agric Food Chem, 50(8), 2404-9 | https://doi.org/10.1021/jf011264n.

Wen, J. et al. (2025). From Clinic to Mechanisms: Multi-Omics Provide New Insights into Cerebrospinal Fluid Metabolites and the Spectrum of Psychiatric Disorders. Molecular Neurobiology, 62, 9120–9132 | https://doi.org/10.1007/s12035-025-04773-0.

Williams, C. et al. (1960). In vivo alteration of the pathways of dopamine metabolism. American Journal of Physiology, 199(4), 722-726 | https://doi.org/10.1152/ajplegacy.1960.199.4.722.

Zambrano, E. and Reyes-Múgica, M. (2002). Hormonal activity may predict aggressive behavior in neuroblastoma. Pediatr Dev Pathol, 5(2), 190-9 | https://doi.org/10.1007/s10024001-0145-8.

Zhao, M. et al. (2024). Gut bacteria-driven homovanillic acid alleviates depression by modulating synaptic integrity. Cell Metabolism, 36(5), 1000-1012.e6 | https://doi.org/10.1016/j.cmet.2024.03.010.

• History & Evolution
• Biosynthesis & dietary uptake
• Bilirubin and the microbiome
• Bilirubin and neurology
• Bilirubin and oncology
• Bilirubin and 5P medicine
• References

History & Evolution

1847: discovery | 1930: first synthesis of heme | 1960s: detoxification pathway elucidated
As early as 400BCE, Hippocratic physicians described yellowing skin as a sign of liver disease – a symptom we now know to be caused by bilirubin (Berk et al. 1977). In 1847, Rudolf Virchow identified the compound responsible for jaundice, as a blood breakdown product, coining the name from bilis (Latin: bile) and rubin (Latin: red) (Lightner 2013).

In the 1930s, chemist Hans Fischer demonstrated that bilirubin originated from the porphyrin ring of hemoglobin (Hopper et al. 2021; Berk et al. 1977). Later developments in chromatographic and spectrophotometric techniques allowed researchers to separate and quantify conjugated and unconjugated bilirubin, greatly advancing clinical diagnostics (Blanckaert et al. 1976; Heirwegh et al. 1989). Understanding of heme catabolism and bilirubin production increased further in the 1960s, when specific enzymatic reactions were identified (Hopper et al. 2021).

More recently, bilirubin has emerged as more than a waste product. Research points to antioxidant and signaling properties with potential cytoprotective effects. For example, patients with Gilbert’s syndrome, a mild hereditary unconjugated hyperbilirubinemia, show lower cardiovascular and metabolic disease risk (Fevery 2008).

Biosynthesis vs. dietary uptake

Approximately 80% of bilirubin is derived from hemoglobin catabolism in bone marrow. The remaining 20% originates from the turnover of various heme-containing proteins, primarily in the liver and muscle tissues. This occurs predominantly within the mononuclear phagocyte system, especially in splenic macrophages, hepatic Kupffer cells and bone marrow monocytes (Fevery 2008; Kalakonda et al. 2022).

Heme oxygenase cleaves heme, releasing chelated iron, carbon monoxide and biliverdin (Kalakonda et al. 2022). Biliverdin reductase then rapidly reduces the biliverdin to form bilirubin, a hydrophobic, orange-yellow pigment (Kalakonda et al. 2022). Due to its poor solubility in water, unconjugated bilirubin binds tightly to serum albumin, which facilitates its transport in the bloodstream, prevents renal filtration and protects tissues from bilirubin’s potentially toxic accumulation. Under normal conditions, virtually no free bilirubin is detectable in plasma (Fevery 2008; Kalakonda et al. 2022).

In the liver, bilirubin dissociates from albumin and enters hepatocytes where it is conjugated with one or two molecules of glucuronic acid by UDP-glucuronosyltransferase 1A1 (UGT1A1). This rate-limiting reaction renders bilirubin water-soluble and marks a key detoxification step in heme metabolism (Kalakonda et al. 2022).

Only conjugated bilirubin can be actively secreted into bile, where it forms micelles with bile acids, cholesterol and phospholipids that are delivered to the small intestine (Fevery 2008). There, bacterial enzymes deconjugate and reduce bilirubin to urobilinogen and stercobilinogen. These compounds are oxidized to urobilin and stercobilin, which are excreted in feces and contribute to its characteristic brown color (Kalakonda et al. 2022). Around 10-20% of urobilinogen is reabsorbed into the portal circulation and may reenter the liver as part of the enterohepatic circulation. While the majority of reabsorbed urobilinogen is taken up again by hepatocytes and re-excreted into bile, a small proportion escapes hepatic reuptake and is ultimately excreted in urine (Gilles J. Hoilat et al. 2023). In the urinary tract, urobilinogen can spontaneously oxidize to urobilin, which gives urine its yellow color (Kalakonda et al. 2022).

Bilirubin and the microbiome

Transformation of conjugated bilirubin in the small intestine is largely dependent on the gut microbiota, particularly bacterial enzymes like β-glucuronidases (Leung et al. 2001). Recently, bilirubin reductase (BilR) was identified as the first microbial enzyme that could reduce bilirubin to urobilinogen-like compounds. BilR is expressed by several members of the Firmicutes phylum and supports microbial participation in host heme metabolism (Hall et al. 2024). However, the precise enzymatic steps responsible for converting urobilinogen to stercobilinogen remain undefined.

Microbiome immaturity or disruption can alter this process. In neonates, low BilR activity hinders bilirubin degradation, contributing to neonatal jaundice (Hall et al. 2024; You et al. 2023). In adults, inflammatory bowel disease (IBD) and other forms of dysbiosis may impair bilirubin metabolism, leading to elevated circulating levels and disrupted enterohepatic cycling (Hall et al. 2024).

Importantly, the relationship between bilirubin and the microbiome is bidirectional. By exerting oxidative stress on gram-positive bacteria, bilirubin suppresses microbial growth, while gram-negative taxa such as Bacteroidetes are often positively associated with serum bilirubin levels in human microbiome studies (Nobles et al. 2013; You et al. 2023).

Bilirubin and cardiometabolic disease

As a central product of heme catabolism, bilirubin serves not only as a biomarker of hepatic function but also as a biologically active molecule with far-reaching effects on metabolic health.

Under normal conditions, glucuronidation is the rate-limiting step in bilirubin metabolism. However, it is the canalicular secretion into bile that is most vulnerable to disruption. In conditions such as intrahepatic cholestasis or biliary obstruction, this excretory pathway is impaired, causing conjugated bilirubin to accumulate in the bloodstream (Fevery 2008). Clinically, elevated conjugated and delta bilirubin (bound to albumin) are key indicators of cholestatic liver injury (Kalakonda et al. 2022).

In contrast, unconjugated hyperbilirubinemia arises from overproduction or impaired conjugation of bilirubin, as seen in hemolytic anemia, ineffective erythropoiesis, or genetic disorders such as Crigler–Najjar and Gilbert’s syndrome (Ramírez-Mejía et al. 2024; Fevery 2008).

Mechanistic studies suggest that bilirubin acts as a metabolic buffer, through multiple pathways. It scavenges reactive oxygen species, protects vascular endothelium, and downregulates nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and cytokine production. It also reduces lipogenesis, enhances fatty acid oxidation, inhibits gluconeogenesis and improves insulin sensitivity. These actions contribute to a lower risk of cardiovascular disease, obesity and type 2 diabetes, making bilirubin a molecule of increasing interest in preventive metabolic health (Ramírez-Mejía et al. 2024).

Bilirubin and neurology

While bilirubin plays protective roles in peripheral tissues, it poses a distinct risk within the central nervous system. Unconjugated bilirubin is lipophilic and capable of crossing the blood–brain barrier, where it can accumulate in neural tissue. This is especially critical in neonates, whose immature liver conjugation systems, biliary secretory apparatus and microbiota make them highly susceptible to bilirubin neurotoxicity (Fevery 2008).

At the cellular level, unconjugated bilirubin interferes with several neural processes. It impairs mitochondrial function, disrupts energy metabolism and alters neurotransmitter synthesis, release and uptake. It also inhibits glutamate uptake by astrocytes, leading to excitotoxicity – a mechanism associated with neurodegeneration (Ramírez-Mejía et al. 2024). Moreover, bilirubin can trigger neuroinflammation, activating microglia and inducing proinflammatory signaling cascades (Zhang et al. 2023).

Despite these risks, emerging evidence suggests that moderate systemic bilirubin levels may exert neuroprotective effects via antioxidant and anti-inflammatory pathways, potentially influencing cognitive aging and neurodegenerative disease risk (Kaur et al. 2025). However, these dual properties remain context- and concentration-dependent, underlining the need for precise regulation of bilirubin metabolism to protect brain health.

Bilirubin and oncology

Recent research is repositioning bilirubin as a regulatory metabolite in cancer biology. While bilirubin’s systemic antioxidant capacity is well-known, its local effects within the tumor microenvironment and its metabolic reprogramming in cancer cells are gaining attention.

Tumors may actively reshape bilirubin metabolism to modulate intracellular redox states. In some cancers, UGT1A1 is downregulated, allowing unconjugated bilirubin to accumulate and buffer against oxidative stress from rapid proliferation or immune surveillance (Yi et al. 2025). Other tumors upregulate heme oxygenase-1 (Ren et al. 2024), shifting heme metabolism toward biliverdin and carbon monoxide, which contribute to immune modulation and angiogenesis (Loboda et al. 2015).

Bilirubin also affects immune cell function in ways that might either suppress tumor-promoting inflammation or impair antitumor immunity (Yi et al. 2025; Ren et al. 2024). These immunomodulatory effects are being explored in the context of tumor immune evasion and response to immunotherapies (Yi et al. 2025; Lee et al. 2023; Ren et al. 2024).

At the cellular level, bilirubin alters checkpoint regulators, kinase signaling, mitochondrial membrane potential and intracellular calcium homeostasis. These changes drive apoptosis, autophagy and growth arrest in cancer cells, while also disrupting invasion and vascular remodeling. Through transcriptional regulators like NF-κB, p53 and extracellular signaling-regulated kinase (ERK), bilirubin shapes the balance between tumor-promoting and tumor-suppressive signaling networks (Yi et al. 2025).

Future research will need to clarify when bilirubin acts as a protective agent and when it might be co-opted as a tumor-supportive factor.

Bilirubin and 5P medicine

Like many metabolites, bilirubin is now seen not just as a simple waste product but also as a multifunctional regulatory molecule. As we learn more about its relevance to cellular and systemic mechanisms, bilirubin will surely make its mark in 5P medicine.

Bilirubin levels are already established biomarkers in clinical diagnostics, predicting liver dysfunction, hemolytic disorders and neonatal risk for kernicterus. Moderate bilirubin elevations have also been associated with reduced cardiovascular and metabolic risk (Ramírez-Mejía et al. 2024), extending its predictive value for chronic disease susceptibility.

The molecule’s antioxidant, anti-inflammatory and immunomodulatory properties suggest new avenues for preventive strategies in oxidative stress-driven diseases such as atherosclerosis, diabetes or neurodegeneration (Ramírez-Mejía et al. 2024).

In oncology, the dual role of bilirubin as both a potential tumor-protective and tumor-supportive factor underscores the need for personalized approaches (Ren et al. 2024; Yi et al. 2025). Patient-specific bilirubin metabolism profiles could inform treatment decisions, particularly in immunotherapy and redox-targeted therapies.

References

Berk, P.D. et al.: International Symposium on Chemistry and Physiology of Bile Pigments (1977).

Blanckaert, N. et al.: Synthesis and separation by thin-layer chromatography of bilirubin-IX isomers. Their identification as tetrapyrroles and dipyrrolic ethyl anthranilate azo derivatives (1976) The Biochemical journal | https://doi.org/10.1042/bj1550405

Fevery, J.: Bilirubin in clinical practice: a review (2008) Liver International | https://doi.org/10.1111/j.1478-3231.2008.01716.x

Gilles J. Hoilat et al.: Bilirubinuria (2023). | https://www.ncbi.nlm.nih.gov/books/NBK557439/

Hall, B. et al.: BilR is a gut microbial enzyme that reduces bilirubin to urobilinogen (2024) Nature Microbiology | https://onlinelibrary.wiley.com/doi/10.1111/j.1478-3231.2008.01716.x

Heirwegh, K.P. et al.: Chromatographic analysis and structure determination of biliverdins and bilirubins (1989) Journal of chromatography | https://doi.org/10.1016/s0378-4347(00)82549-9

Hopper, C.P. et al.: A brief history of carbon monoxide and its therapeutic origins (2021) Nitric oxide : biology and chemistry | https://doi.org/10.1016/j.niox.2021.04.001

Kalakonda, A. et al.: Physiology, Bilirubin (2022). | https://www.ncbi.nlm.nih.gov/books/NBK470290/

Kaur, A. et al.: Unraveling the dual role of bilirubin in neurological Diseases: A Comprehensive exploration of its neuroprotective and neurotoxic effects (2025) Brain Research | https://doi.org/10.1016/j.brainres.2025.149472

Lee, Y. et al.: Hyaluronic acid-bilirubin nanomedicine-based combination chemoimmunotherapy (2023) Nature Communications | https://doi.org/10.1038/s41467-023-40270-5

Leung, J.W. et al.: Expression of bacterial beta-glucuronidase in human bile: an in vitro study (2001) Gastrointestinal Endoscopy | https://doi.org/10.1067/mge.2001.117546

Lightner, D.A.: Bilirubin (2013) | ISBN: 9783709116371.

Loboda, A. et al.: HO-1/CO system in tumor growth, angiogenesis and metabolism – Targeting HO-1 as an anti-tumor therapy (2015) Vascular Pharmacology | https://doi.org/10.1016/j.vph.2015.09.004

Nobles, C.L. et al.: A product of heme catabolism modulates bacterial function and survival (2013) PLoS Pathogens | https://doi.org/10.1371/journal.ppat.1003507

Ramírez-Mejía, M.M. et al.: The Multifaceted Role of Bilirubin in Liver Disease: A Literature Review (2024) Journal of Clinical and Translational Hepatology | https://doi.org/10.14218/JCTH.2024.00156

Ren, K. et al.: Prognostic and immunotherapeutic implications of bilirubin metabolism-associated genes in lung adenocarcinoma (2024) Journal of Cellular and Molecular Medicine | https://doi.org/10.1111/jcmm.18346

Ryter, S.W. et al.: Carbon monoxide and bilirubin: potential therapies for pulmonary/vascular injury and disease (2007) American Journal of Respiratory Cell and Molecular Biology | https://doi.org/10.1165/rcmb.2006-0333TR

Yi, F. et al.: Bilirubin metabolism in relation to cancer (2025) Frontiers in Oncology | https://doi.org/10.3389/fonc.2025.1570288

You, J.J. et al.: The relationship between gut microbiota and neonatal pathologic jaundice: A pilot case-control study (2023) Frontiers in Microbiology | https://doi.org/10.3389/fmicb.2023.1122172

Zhang, F. et al.: Neuroinflammation in Bilirubin Neurotoxicity (2023) Journal of integrative neuroscience | https://doi.org/10.31083/j.jin2201009

• History & Evolution
• Biosynthesis & dietary uptake
• Itaconic acid and the microbiome
• Itaconic acid, immunity and inflammation
• Itaconic acid and cardiovascular disease
• Itaconic acid and neurology
• Itaconic acid and oncology
• Itaconic acid and 5P medicine
• References

History & Evolution

1836: discovery (Baup, 1836) | 1840: first synthesis (Crasso, G., 1840) | 1960s: first industrial applications | 2012: first synthesized in mammalian cells (Sugimoto, et al., 2011)

In 1836, Swiss chemist Samuel Baup discovered an unknown compound when experimenting with the distillation of citric acid (Baup, 1836). In 1840, Gustav Crasso synthesized the same compound through the decarboxylation of aconitate, and called it itaconate – an anagram of its precursor (Crasso, 1840). This unsaturated dicarboxylic acid received little attention for almost a hundred years, until 1931, when Japanese mycologist Kinoshita discovered a strain of Aspergillus fungi in salted prune juice that produced the metabolite (Kinoshita, 1932). He named the fungus Aspergillus itaconicus.

By the 1960s, itaconic acid had caught the attention of the polymer industry, thanks to the useful double bond of its methylene group. Now, around 40,000 tons are produced each year, primarily synthesized from Aspergillus terreus (Cordes, 2015).

Scientific interest in itaconate’s biological role was renewed in the early 2010s, when Sugimoto et al. discovered that it could be synthesized in mammalian immune cells (Sugimoto et al., 2011). Since then, itaconic acid’s role as a key player in immunometabolism and inflammatory regulation has become an active area of research (Diotallevi et al., 2021).

Biosynthesis vs. dietary uptake

In humans, itaconic acid is an endogenous metabolite synthesized primarily in the mitochondria of activated macrophages (Peace et al., 2022). It is synthesized by decarboxylating cis-aconitate, an intermediate of the tricarboxylic acid (TCA) cycle. Metabolomics has shown that this occurs through the action of the enzyme cis-aconitase decarboxylase, also known as immune-responsive gene 1 (IRG1 or ACOD1) (Michelucci et al., 2013). Expression of IRG1 is upregulated during infection and inflammation, leading to increased itaconate production. Plasma concentrations are therefore highly variable, depending on immune status.

Humans do not obtain itaconic acid directly from the diet, but dietary factors may affect its production. For example, carbohydrate intake affects the TCA cycle and energy metabolism, which in turn influences the availability of itaconate precursors like cis-aconitate. Under inflammatory conditions, nutrient availability may affect itaconate synthesis.

Itaconate is the biologically active, deprotonated form of itaconic acid, which contains two carboxylic acid groups that lose protons at pH levels above 7. This gives itaconate a double negative charge and makes it the dominant form found in the bloodstream. Structurally, it resembles other dicarboxylates such as succinate, malonate, phosphoenolpyruvate and fumarate (Peace et al., 2022).

Itaconic acid and the microbiome

While itaconic acid is synthesized by environmental microbes, like some Aspergillus species, it does not appear to be produced by gut microbiota in significant amounts. That said, it has been described as a “microbiota-associated metabolite” because its production in the mitochondria of host immune cells is triggered by microbial components such as lipopolysaccharide (LPS) from Gram-negative bacteria (Fedotcheva et al., 2022).

A study using integrated multiomics approaches showed that altered itaconate metabolism leads to significant changes in gut microbiota composition. Using a mouse model, Eberhart et al. (2024) found that in response to a high-fat diet, the itaconate-synthesizing enzyme ACOD1 promotes gut dysbiosis associated with obesity and inflammation, while genetic deletion of ACOD1 protects against metabolic disease (Eberhart et al., 2024). In addition, fecal metagenomics and microbiota transplantation showed that itaconate inhibits growth of Bacteroidaceae. These findings suggest that itaconate plays a role in diet-induced obesity and positions the ACOD1–itaconate pathway as a potential therapeutic target for the inflammatory effects of obesity.

Itaconic acid also inhibits bacterial isocitrate lyase, a key enzyme in the glyoxylate shunt during bacterial infection (Peace, 2022). This limits the growth of pathogens like Mycobacterium tuberculosis, Pseudomonas indigofera and Salmonella. However, some bacteria have developed strategies to protect themselves against itaconate’s antimicrobial properties (Peace, 2022). For example, Pseudomonas aeruginosa, Yersinia pestis and M. tuberculosis can break down itaconate into pyruvate and acetyl-CoA, while P. aeruginosa and Staphylococcus aureus respond to itaconate-induced stress by shifting from LPS to extracellular polysaccharide production, leading to biofilm formation that provides intracellular protection for the bacteria.

See how the MxP® Quant 1000 assay helps scientists investigate the impact of energy metabolism on health by accurately profiling TCA metabolites with targeted metabolomics

Itaconic acid, immunity and inflammation

Itaconic acid plays an important role in immune regulation and inflammation through the antibacterial mechanisms mentioned above. It also modulates the production of reactive oxygen species (ROS), which helps defend against bacterial and viral infections. Itaconate’s antiviral properties have been found to inhibit Zika virus replication (Daniels et al., 2019).

Beyond its antimicrobial activity, itaconic acid also acts as an anti-inflammatory metabolite by targeting key metabolic pathways in activated macrophages. It inhibits succinate dehydrogenase, a key enzyme in the TCA cycle, which alters mitochondrial metabolism and promotes anti-inflammatory cytokine production (Cordes et al., 2016). It also disrupts mitochondrial energy metabolism by disrupting oxidative phosphorylation and fatty acid oxidation. Recent studies have shown that itaconic acid can promote the pentose phosphate pathway, increasing nicotinamide adenine dinucleotide phosphate (NADPH) levels and enhancing NADPH oxidase-mediated ROS production. These ROS contribute to the expression of anti-inflammatory genes, restrict pathogens like Salmonella typhimurium and reduce pro-inflammatory cytokine production (Zhu et al., 2021).

These findings have led to broader recognition of the itaconate pathway in immune regulation. A review by Li et al. (2020) describes the IRG1/itaconate axis as a central feature in immunometabolic regulation, supported by metabolomics data that link itaconate to changes in mitochondrial respiration, oxidative stress and inflammatory signaling in macrophages (Li et al., 2020).

Given these properties, itaconic acid has emerged as a metabolite of interest in immune-mediated diseases. A recent review by Luo et al. (2024) suggests itaconate and its derivatives may play a role in conditions such as rheumatoid arthritis, multiple sclerosis , type 1 diabetes mellitus and autoimmune hepatitis (Luo et al., 2025).

Itaconic acid and cardiovascular disease

Research shows that itaconate may have a protective effect in cardiovascular disease. In models of atherosclerosis, IRG1 deficiency has been shown to increase plaque burden (Harber et al., 2024), while treatment with 4-octyl itaconate (4-OI) reversed plaque inflammation in mice and reduce cardiovascular inflammation in human immune cells (Cyr et al., 2024).

Itaconate also supports cardiac repair after myocardial infarction. A study using targeted metabolomics found that after efferocytosis (the clearing of dead cells which is essential for cardiac repair), altered signaling in macrophages expressing the TREM2 receptor disrupted the TCA cycle (Gong et al., 2024). This resulting increase in itaconate production reduced cardiomyocyte apoptosis and promoted fibroblast proliferation. Increased TREM2 levels led to improved cardiac function.

Itaconic acid and neurology

Neuroinflammation is a well-established driver in the progression of numerous neurological disorders, including Alzheimer’s disease (AD), Parkinson’s disease (PD), multiple sclerosis (MS) and stroke. Itaconate and its derivatives may help counteract this neuroinflammatory activity in the central nervous system through the regulation of immune pathways including Nrf2/KEAP1, ROS production and the NLRP3 inflammasome (Kong et al., 2024).

In LPS-induced neuroinflammation, both itaconate and the related isomer mesaconate have been found to reduce inflammation in the brain and improve synaptic plasticity, linked to memory and cognitive processes (Ohm et al., 2024). A study using untargeted serum metabolomics in PD patients also identified reduced levels of circulating itaconate, suggesting a role in disease-associated immune dysregulation (Paul et al., 2023).

These findings suggest that itaconate may be a promising candidate as a biomarker and therapeutic target in neurological conditions.

Itaconic acid and oncology

Itaconic acid has also been implicated in cancer biology. In peritoneal tumors such as melanoma and ovarian carcinoma, metabolomics techniques have shown an upregulation of itaconic acid (Weiss et al., 2018). Tumor-associated macrophages were found to accumulate itaconate, resulting in increased oxidative phosphorylation and mitochondrial ROS production, and in turn promoting tumor growth. Silencing IRG1 was found to reduce the tumor burden.

However, there is also evidence that itaconic acid has anti-tumor effects (Perkovic et al., 2020) (Wang, Z. et al., 2024). A mouse model showed that dimethyl itaconate suppresses colitis-associated colorectal cancer (Wang et al., 2020). In another animal study, 4-OI was found to induce ferroptosis in the treatment of retinoblastoma (Liu et al., 2022).

Thes findings suggest that more research is needed to fully understand the role of itaconic acid in different types of cancer.

Itaconic acid and 5P medicine

In the context of 5P medicine – predictive, preventive, precision, population-based, participatory medicine – itaconic acid is again relevant due to its role in mediating immune responses and inflammation, both of which contribute to many prevalent chronic diseases. A white paper by biocrates, “Chronic diseases have a common origin”, highlights the underlying role of inflammation in complex chronic disease.

Obesity is a major public health concern and key target for preventive strategies. Dietary itaconate has been shown to reduce visceral fat accumulation in rats, highlighting a potential role in early intervention and metabolic regulation (Sakai et al., 2004).

As a biomarker of disease, itaconate levels in blood or immune cells may signal early inflammatory activity or therapeutic response, as seen in conditions like rheumatoid arthritis (Daly et al., 2020), non-alcoholic fatty liver disease (Weiss et al., 2018) and bacterial infections (Singh et al., 2021). In each of these examples, metabolomics has played a role in providing insights about the presence and behavior of this important metabolite.

Itaconic acid is a promising focus for future research, with omics technologies offering a powerful tool to support 5P medicine.

References

Baup, S.: Ueber eine neue Pyrogen-Citronensäure, und über Benennung der Pyrogen-Säuren überhaupt. (1836) Ann. Pharm | https://doi.org/10.1002/jlac.18360190107.

Cordes, T. et al.: Itaconic Acid: The Surprising Role of an Industrial Compound as a Mammalian Antimicrobial Metabolite. (2015) Annual Review of Nutrition | https://doi.org/10.1146/annurev-nutr-071714-034243.

Cordes, T. et al.: Immunoresponsive Gene 1 and Itaconate Inhibit Succinate Dehydrogenase to Modulate Intracellular Succinate Levels. (2016) J Biol Chem.| https://doi.org/10.1074/jbc.M115.685792.

Crasso, G.: Untersuchungen über das Verhalten der Citronsäure in höherer Temperatur und die daraus hervorgehenden Produkte (1840) Ann. Chem. Pharm. | https://doi.org/10.1002/jlac.18400340104.

Cyr, Y. et al.: The IRG1–itaconate axis protects from cholesterol-induced inflammation and atherosclerosis. (2024) Natl Acad Sci USA | https://doi.org/10.1073/pnas.2400675121.

Daly, R. et al.: Changes in Plasma Itaconate Elevation in Early Rheumatoid Arthritis Patients Elucidates Disease Activity Associated Macrophage Activation. (2020) Metabolites | https://doi.org/10.3390/metabo10060241.

Daniels, B. et al.: The nucleotide sensor ZBP1 and kinase RIPK3 induce the enzyme IRG1 to promote an antiviral metabolic state in neurons.(2019) Immunity | https://doi.org/10.1016/j.immuni.2018.11.017.

Diotallevi, M. et al.: Itaconate as an inflammatory mediator and therapeutic target in cardiovascular medicine. (2021) Biochem Soc Trans, | https://doi.org/10.1042/BST20210269.

Eberhart, T. et al.: ACOD1 deficiency offers protection in a mouse model of diet-induced obesity by maintaining a healthy gut microbiota. (2024) Cell Death Dis. | https://doi.org/10.1038/s41419-024-06483-2.

Fedotcheva, N. et al.: Influence of Microbial Metabolites and Itaconic Acid Involved in Bacterial Inflammation on the Activity of Mitochondrial Enzymes and the Protective Role of Alkalization.(2022) Int J Mol Sci. | https://doi.org/10.3390/ijms23169069.

Gong, S. et al.: TREM2 macrophage promotes cardiac repair in myocardial infarction by reprogramming metabolism via SLC25A53. (2024) Death Differ. | https://doi.org/10.1038/s41418-023-01252-8.

Harber, K. et al.: Targeting the ACOD1-itaconate axis stabilizes atherosclerotic plaques. (2024) Biol. | https://doi.org/10.1016/j.redox.2024.103054.

Kinoshita, K.: Über die Produktion von Itaconsäure und Mannit durch einen neuen Schimmelpilz, Aspergillus itaconicus. (1932) Acta Phytochimica | https://www.scirp.org/reference/referencespapers?referenceid=899199

Kong, X. et al.: The anti-inflammatory effects of itaconate and its derivatives in neurological disorders. (2024) Cytokine & Growth Factor Reviews, | https://doi.org/10.1016/j.cytogfr.2024.07.001.

Li, R. et al.: Itaconate: A Metabolite Regulates Inflammation Response and Oxidative Stress. (2020) Oxid Med Cell Longev.| https://doi.org/10.1155/2020/5404780.

Liu, K. et al.: Induction of autophagy-dependent ferroptosis to eliminate drug-tolerant human retinoblastoma cells. (2022) Death Dis. | https://doi.org/10.1038/s41419-022-04974-8.

Luo, Y. et al.: Metabolic Regulation of Inflammation: Exploring the Potential Benefits of Itaconate in Autoimmune Disorders. (2025) Immunology | https://doi.org/10.1111/imm.13875.

Michelucci, A. et al.: Immune-responsive gene 1 protein links metabolism to immunity by catalyzing itaconic acid production. (2013) PNAS | https://doi.org/10.1073/pnas.1218599110.

Ohm, M. et al.: The potential therapeutic role of itaconate and mesaconate on the detrimental effects of LPS-induced neuroinflammation in the brain. (2024) J Neuroinflammation | https://doi.org/10.1186/s12974-024-03188-3.

Paul, K. et al.: Untargeted serum metabolomics reveals novel metabolite associations and disruptions in amino acid and lipid metabolism in Parkinson’s disease. (2023) Mol Neurodegener. | https://doi.org/10.1186/s13024-023-00694-5.

Peace, C. et al.: The role of itaconate in host defense and inflammation. (2022) J Clin Invest. | https://doi.org/10.1172/JCI148548.

Perkovic, I. et al.: Itaconic acid hybrids as potential anticancer agents. (2020) Mol Divers. | https://doi.org/10.1007/s11030-020-10147-6.

Sakai, A. et al.: Itaconate reduces visceral fat by inhibiting fructose 2,6-bisphosphate synthesis in rat liver. (2004) Nutrition | https://doi.org/10.1016/j.nut.2004.08.007.

Singh, S. et al.: Integrative metabolomics and transcriptomics identifies itaconate as an adjunct therapy to treat ocular bacterial infection. (2021) Rep Med, | https://doi.org/10.1016/j.xcrm.2021.100277.

Sugimoto, M. et al.: Non-targeted metabolite profiling in activated macrophage secretion. (2011) Metabolomics | https://doi.org/10.1007/s11306-011-0353-9.

Wang, Q. et al.: The anti-inflammatory drug dimethyl itaconate protects against colitis-associated colorectal cancer. (2020) Journal of Molecular Medicine | https://doi.org/10.1007/s00109-020-01963-2.

Wang, Z. et al.: Cancer cell-intrinsic biosynthesis of itaconate promotes tumor immunogenicity. (2024) EMBO J. | https://doi.org/10.1038/s44318-024-00217-y.

Weiss, J. et al.: Itaconic acid mediates crosstalk between macrophage metabolism and peritoneal tumors. (2018) J Clin Invest, | https://doi.org/10.1172/JCI99169.

Zhu, X. et al.: Itaconic acid exerts anti-inflammatory and antibacterial effects via promoting pentose phosphate pathway to produce ROS. (2021) Sci Rep. | https://doi.org/10.1038/s41598-021-97352-x.

• History & Evolution
• Biosynthesis vs. dietary uptake
• NAA and the microbiome
• NAA and neurology
• NAA and oncology
• NAA and 5P medicine

History & Evolution

1956: discovery (Tallan et al. 1956) | 1959: first synthesis (Goldstein 1959)

N-acetyl-aspartic acid (NAA), or N-acetylaspartate, is one of the most abundant metabolites in the mammalian central nervous system (CNS) (Long et al. 2013). Despite extensive research since its discovery in 1956 (Tallan et al. 1956), it remains something of a biochemical enigma (Moffett et al. 2007). It is known to be involved in osmoregulation, myelin synthesis, neuronal metabolism and acetate storage for lipid metabolism, and disruption to the NAA pathway has demonstrable physiological effects (Moffett et al. 2007). However, researchers continue to debate its primary function and mechanisms of action (Bogner-Strauss 2017).

An important insight came in 1959, when Goldstein successfully synthesized NAA using acetone powders derived from rat brain tissue (Goldstein 1959). This showed that NAA is formed in neuronal mitochondria from aspartic acid and acetyl-coenzyme A (CoA).

Two further notable findings brought NAA to the attention of the neurological community (Moffett et al. 2007). First, its strong signal in magnetic resonance spectroscopy (MRS) made it a reliable marker in brain imaging. Second, researchers discovered that a deficiency of the enzyme aspartoacylase leads to NAA accumulation in the brain, causing the rare genetic disorder known as Canavan disease. These breakthroughs confirmed NAA’s role as a diagnostic tool and key player in neurological disease.

There are hints that NAA is also involved in broader cognitive processes: NAA has been nicknamed the “creativity chemical,” (Geddes 2009) and associates with memory and educational attainment (Glodzik et al. 2012).

Biosynthesis vs. dietary uptake

NAA is primarily synthesized endogenously. N-acetyl amino acids are synthesized through the breakdown of N-acetyl proteins or direct acetylation of amino acids by specific enzymes (Ramunaidu et al. 2023). In the case of NAA, N-acetylaspartate synthetase (encoded by NAT8L, a gene highly expressed in brain and connective tissue) catalyzes a reaction between acetyl-CoA and aspartic acid to produce NAA in neuronal mitochondria. NAA is transported to oligodendrocytes where it is hydrolyzed by aspartoacylase, producing L-aspartate and acetate (Moffett et al. 2007). Acetate is then incorporated into acetyl-CoA, which is essential for energy production, lipid synthesis and protein acetylation in the brain.

NAA synthesis also occurs outside the CNS (Bogner-Strauss 2017). Research shows that brown adipose tissue (BAT) expresses significant levels of NAT8L, allowing it to synthesize NAA (Huber et al. 2018). While NAA itself is not obtained directly from the diet, dietary intake can influence the availability of its precursors, such as glucose and aspartate, which are abundant in BAT. A study of mice fed high-fat and high-glucose diets found that both conditions increased NAA pathway activity in brown adipocytes compared to standard diet, in turn affecting energy and lipid metabolism (Huber et al. 2018).

NAA and the microbiome

While gut microbiota do not produce or consume NAA in significant amounts, microbial composition has been shown to influence cerebral metabolite levels, including NAA (Matsumoto et al. 2013).

Research into maternal postpartum depression (PPD) and infant neurodevelopment highlights this connection (Zhou et al. 2024). Infants born to mothers with PPD symptoms exhibited increased Veillonella and reduced Bifidobacterium, as well as lower levels of NAA and aspartate in their gut metabolome. These changes were associated with poorer neurodevelopmental scores at six months, suggesting that maternal mental health and microbiome alterations can indirectly affect brain metabolites and neurodevelopment.

Similarly, an animal model showed that fecal Ruminococcus and Butyricimonas predicted NAA levels in the brain, suggesting a potential mechanism for microbiota-brain communication in neurological conditions such as autism spectrum disorder (Mudd et al. 2017).

NAA and neurology

NAA makes up around 0.1% of the brain’s net weight (Inglese et al. 2008). Its abundance makes it a useful marker of neuronal health, measured via magnetic resonance spectroscopy (MRS). Altered levels of NAA have been observed in several brain diseases and disorders (Moffett et al. 2007).

The discovery of the connection between NAA and Canavan disease marked a major step forward in understanding the role of NAA in neurological disease (Moffett et al. 2007). In Canavan disease, a mutation in the ASPA gene impairs NAA breakdown, causing an accumulation of NAA and progressive leukodystrophy in infants. Clearly, excess NAA can be harmful. At the same time, reduced NAA levels have been linked to neuronal loss and degeneration in other conditions, for example:

Alzheimer’s disease
Patients with Alzheimer’s disease have reduced NAA levels in specific brain regions, correlating with neuronal loss and cognitive decline (Schuff et al. 2006) (Moffett et al. 2007).

Multiple sclerosis
NAA deficiency is associated with neurodegeneration and is considered a hallmark of the disease, demonstrated using MRS and HPLC analysis. A 2023 study found that in patients with MS, oxidized NAT8L mRNA limits NAT8L production and reduces NAA levels, establishing a molecular link between NAA, mRNA oxidization and MS pathogenesis (Kharel et al. 2023).

Traumatic brain injury
A review of 20 publications with metabolomics investigations into severe traumatic brain injury found that NAA levels drop after injury, reflecting neuronal damage (Fedoruk et al. 2023).

Psychiatric disorders
A 2018 review of metabolomics studies for psychosis identified NAA as a potential biomarker for psychosis (Li et al. 2018). In patients with chronic major depressive disorder, NAA concentrations were significantly reduced in several brain regions compared to healthy controls. Antidepressant treatment appeared to reduce NAA alterations in the frontal lobe.

NAA and oncology

Recent research points to the involvement of the NAA pathway in cancer biology. Elevated NAA levels have been observed in several cancer types, including non-small-cell lung cancer, breast cancer, ovarian cancer, and prostate cancer (Krause et al. 2024). A 2016 study using untargeted metabolic profiling found that NAA levels in high-grade serous ovarian cancer (HGSOC) tissue were more than 28 times higher than in normal ovarian tissue, higher than any other amino acid metabolite (Zand et al. 2016). Another multi-omics study demonstrated the accumulation of NAA in murine models of castration-resistant prostate cancer (Salji et al. 2022).

One possible mechanism is that NAA impacts key cellular processes such as histone acetylation and signaling pathways that promote tumor growth and survival. Additionally, reduced expression of NAT8L, which drives NAA synthesis, is linked to reduced cancer proliferation, further linking NAA to tumor growth  (Krause et al. 2024).

These findings suggest that NAA metabolism holds promise as a biomarker and therapeutic target oncology.

NAA and 5P medicine

Metabolomics is helping researchers understand more about the role of NAA in disease monitoring and personalized treatment planning, in both chronic and acute disease, and beyond neurology and oncology. For example, a 2022 metabolomics study identified NAA as a potential biomarker of juvenile idiopathic arthritis and cardiovascular disease risk (Lewis et al. 2022).

Another study used MRS to evaluate the impact of disease severity on brain metabolites in cases of COVID-19 (Ostojic et al. 2024). NAA levels were significantly lower in more severe cases, suggesting neuronal dysfunction.

Investigations like these reveal more about NAA pathways, offering insights for predictive diagnostics, personalized treatment and population health strategies. With continued research, NAA may soon cease to be the enigma it once was.

References

Bogner-Strauss, J.: N-Acetylaspartate Metabolism Outside the Brain: Lipogenesis, Histone Acetylation, and Cancer (2017) Frontiers in Endocrinology, 8, 240. | DOI: https://doi.org/10.3389/fendo.2017.00240.

Fedoruk, R. et al.: Metabolomics in severe traumatic brain injury: a scoping review (2023) BMC Neurosci., 24, 52. | DOI: https://doi.org/10.1186/s12868-023-00824-1.

Geddes, L. (2009, May 13). Creativity chemical favours the smart. Retrieved from New Scientist: https://www.newscientist.com/article/mg20227084-300-creativity-chemical-favours-the-smart/

Glodzik, L. et al.: The whole-brain N-acetylaspartate correlates with education in normal adults (2012) Psychiatry Research: Neuroimaging, 204(1), 49-54. | DOI: https://doi.org/10.1016/j.pscychresns.2012.04.013.

Goldstein, F.: Biosynthesis of N-Acetyl-l-aspartic Acid. Journal of Biological Chemistry (1959) 234(10), 2702-2706. | DOI: https://doi.org/10.1016/S0021-9258(18)69763-7.

Huber, K. et al.: N-acetylaspartate pathway is nutrient responsive and coordinates lipid and energy metabolism in brown adipocytes (2018) Biochim Biophys Acta Mol Cell Res., 1866(3), 337–348. | DOI: https://doi.org/10.1016/j.bbamcr.2018.08.017.

Inglese, M. et al.: Global Average Gray and White Matter N-acetylaspartate Concentration in the Human Brain (2008) Neuroimage., 41(2), 270–276. | DOI: https://doi.org/10.1016/j.neuroimage.2008.02.034.

Kharel, P. et al.: NAT8L mRNA oxidation is linked to neurodegeneration in multiple sclerosis (2023) Cell Chemical Biology, 30(3), 308 – 320.e5. | DOI: https://doi.org/10.1016/j.chembiol.2023.02.007.

Krause, N. and Wegner, A.: N-acetyl-aspartate metabolism at the interface of cancer, immunity, and neurodegeneration (2024) Current Opinion in Biotechnology, 85, 103051. | DOI: https://doi.org/10.1016/j.copbio.2023.103051.

Lewis, K. et al.: Serine, N-acetylaspartate differentiate adolescents with juvenile idiopathic arthritis compared with healthy controls: a metabolomics cross-sectional study (2022) Pediatric Rheumatology, 20(12). | DOI: https://doi.org/10.1186/s12969-022-00672-z.

Li, C. et al.: Metabolomics in patients with psychosis: A systematic review (2018) American Journal of Medical Genetics: Neuropsychiatric Genetics, 177(6), 580-588. | DOI: https://doi.org/10.1002/ajmg.b.32662.

Long. P. et al.: N-Acetylaspartate (NAA) and N-Acetylaspartylglutamate (NAAG) Promote Growth and Inhibit Differentiation of Glioma Stem-like Cells (2013) Journal of Biological Chemistry, 288(36), 26188-26200. | DOI: https://doi.org/10.1074/jbc.M113.487553.

Matsumoto, M. et al.: Cerebral low-molecular metabolites influenced by intestinal microbiota: a pilot study (2013) Front. Syst. Neurosci., 7. | DOI: https://doi.org/10.3389/fnsys.2013.00009.

Moffett, J. et al.: N-Acetylaspartate in the CNS: From Neurodiagnostics to Neurobiology (2007) Prog Neurobiol., 81(2), 89–131. | DOI: https://doi.org/10.1016/j.pneurobio.2006.12.003.

Mudd, A. et al.: Serum cortisol mediates the relationship between fecal
Ruminococcus and brain N-acetylaspartate in the young pig (2017) Gut Microbes, 8(6), 589–600. | DOI: https://doi.org/10.1080/19490976.2017.1353849.

Ostojic, J. et al.: Decreased Cerebral Creatine and N-Acetyl Aspartate Concentrations after Severe COVID-19 Infection: A Magnetic Resonance Spectroscopy Study (2024) J Clin Med., 13(14), 4128. | DOI: https://doi.org/10.3390/jcm13144128.

Ramunaidu, A. et al.: Characterization of isomeric acetyl amino acids and di-acetyl amino acids by LC/MS/MS (2023) Journal of Mass Spectrometry, 58(12), e4982. | DOI: https://doi.org/10.1002/jms.4982.

Salji, M. et al.: Multi-omics & pathway analysis identify potential roles for tumor N-acetyl aspartate accumulation in murine models of castration-resistant prostate cancer (2022) iScience, 25(4), 104056. | DOI: https://doi.org/10.1016/j.isci.2022.104056.

Schuff, N. et al.: N-Acetylaspartate As A Marker Of Neuronal Injury In Neurodegenerative Disease (2006) Adv Exp Med Biol., 576, 241–363. | DOI: https://doi.org/10.1007/0-387-30172-0_17.

Tallan, H. Moore, S. and Stein, W.: N-Acetyl-L-Aspartic Acid In Brain (1956) Journal of Biological Chemistry, 219(1), 257-264. | DOI: https://doi.org/10.1016/S0021-9258(18)65789-8.

Zand, B. et al.: Role of Increased n-acetylaspartate Levels in Cancer (2016) JNCI J Natl Cancer Inst, 108(6), djv426. | DOI: https://doi.org/10.1093/jnci/djv426.

Zhou L. et al.: Association of maternal postpartum depression symptoms with infant neurodevelopment and gut microbiota (2024) Front Psychiatry., 15, 1385229. | DOI: https://doi.org/10.3389/fpsyt.2024.1385229.

• History & Evolution
• Biosynthesis vs. dietary uptake
• Citric acid and the microbiome
• Citric acid and immune signaling
• Citric acid and bone disease
• Citric acid and urology
• Citric acid and neurology
• Citric acid and oncology
• Citric acid and 5P medicine

History & Evolution

1784: discovery of citric acid (Berovic et al. 2007) | 1893: first synthesis (Wehmer 1893) | 1937: description of role in Krebs cycle (Krebs et al. 1937)

Citrate, the conjugate base of citric acid, is a versatile metabolite found in all aerobic organisms. Citric acid was first isolated from citrus fruits in 1784 by Swedish chemist Carl Wilhelm Scheele, who identified it as the source of the sour taste of lemons (Berovic et al. 2007). More than a hundred years later, in 1893, Carl Wehmer discovered that citrate could be produced from certain fungi (Wehmer, C., 1893). When the first world war disrupted the supply of calcium citrate, scientists like James Currie built on Wehmer’s research to find a new method of citrate production based on sugar fermentation (Currie 1917). The microbial production of citric acid using the mold Aspergillus niger would be patented in the early twentieth century, paving the way for industrial applications (Di Lorenzo et al. 2022).

Citrate attracted more attention in the 1930s, when Hans Krebs and William Johnson investigated the role of citrate in carbohydrate metabolism, leading to the discovery of the citric acid cycle, also known as the tricarboxylic acid (TCA) cycle or “Krebs cycle” (Krebs et al. 1937).

Today, citrate is recognized not only for its role in cellular energy production, but also for its involvement in various physiological processes such as fatty acid metabolism, amino acid synthesis, calcium signaling, anticoagulation and the regulation of pH balance. It has been found to play role in conditions such as kidney stones, bone disease and cancer.

As natural preservatives and sweeteners, citric acid and citrates are also commonly used in foods, drinks and supplements. Citric acid is the most consumed organic acid in the world, with more than 50% used in beverages (Granchi et al. 2019). Other uses include detergents, cosmetics, pharmaceuticals and industrial chemical applications. While citric acid derived from citrus fruits is considered safe, some people report adverse reactions when consuming processed foods containing synthetic citric acid, derived from modified strains of Aspergillus niger (Sweis et al. 2018).

Biosynthesis vs. dietary uptake

Citrate is obtained through the diet and synthesized endogenously. Dietary sources include fruits and vegetables – most obviously citrus fruits – and dietary supplements. Typical daily intake is around 4g, with over 95% absorbed in the small intestine (Granchi et al. 2019). Plasma citrate levels increase within 30 minutes of ingestion, and is then filtered and reabsorbed equally quickly by the kidneys (Granchi et al. 2019).

In addition to dietary intake, plasma citrate levels are influenced by renal clearance, cellular metabolism and bone remodeling (Granchi et al. 2019). Around 10 to 35% of filtered citrate is excreted in urine, varying by age, diet and sex (Costello et al. 2016) (Shah et al. 1958). Urinary citrate decreases with acidosis and testosterone administration, and increases with alkalosis and administration of estrogens, parathyroid hormone and vitamin D (Shah et al. 1958).

While diet is a good source of plasma citrate, it is not essential: humans and animals maintain normal citrate levels even with low levels of dietary citrate, indicating other sources (Costello et al. 2016).

All cells synthesize citrate through cellular metabolism in the TCA cycle. In this cycle, the enzyme citrate synthase catalyzes the condensation of oxaloacetate with acetyl coenzyme-A (acetyl-CoA) to form citrate. Citrate then acts as a substrate for a series of chemical reactions, generating reduced nicotinamide adenine dinucleotide, flavine adenine dinucleotide, adenosine triphosphate (ATP) and eventually regenerating oxaloacetate (Icard et al. 2021). Citrate transport protein releases any excess citrate from the mitochondria (Granchi et al. 2019). Through this mechanism, citrate plays a crucial role in energy production, fatty acid synthesis and bone formation.

Around 90% of total citrate in the body is stored in bone tissue (Costello et al. 2016). During bone resorption, citrate is released into plasma, providing the main endogenous source of circulating citrate.

Citrate and the microbiome

The gut microbiome influences the availability of all metabolites involved in the TCA cycle, including citrate. Citrate participates in bacterial fermentation and contributes to regulation of acetyl-CoA, which influences lipid metabolism and fatty acid synthesis.

Citrate is a source of carbon and energy for several bacterial species (Bott 1997). In anaerobic conditions, citrate and other TCA intermediates are metabolized via citrate lyase by enterobacteria like Klebsiella pneumoniae and Escherichia coli, and lactic acid bacteria. This produces oxaloacetate, acetate and succinate, demonstrating the adaptability of these metabolic pathways under different environmental conditions.

A recent study in mice revealed several findings that suggest citric acid may be an alternative to antibiotics, which are increasingly overused (Hu 2024). Citric acid supplementation was shown to increase populations of common probiotic bacteria, such as Bifidobacteria and Lactobacillus. The study also found that citric acid promotes the expression of genes that maintain the integrity of the intestinal tight junction barrier, enhances intestinal immune function and inhibits viral infection.

Citric acid and immune signaling

In immune cells, citrate exported from mitochondria serves as a precursor for the synthesis of acetyl-CoA, which is essential for lipogenesis, histone acetylation, and antioxidant defenses (Zara et al. 2023). These processes not only sustain mitochondrial function but also link citrate to immune signaling and inflammation.

Studies have also linked citrate to the production of key inflammatory mediators, such as prostaglandins, nitric oxide and cytokines like TNF-α, IL-1β, and IL-6 (Liu et al. 2025). Furthermore, citrate regulates histone acetylation in immune cells through ATP citrate lyase, influencing the expression of genes involved in inflammatory pathways (Liu et al. 2025). These findings solidify the role of citrate in bridging metabolism, immune responses and inflammation.

Citric acid and bone disease

As noted, bone tissue serves as the main reservoir of citrate, giving it a central role in citrate homeostasis. At the same time, citrate is essential for bone health and skeletal integrity. Citrate is produced by osteoclasts and osteoblasts, but also influences their differentiation and behavior (Granchi et al. 2019). Disruptions to citrate homeostasis are associated with abnormalities in bone development and metabolism. Due to its impact on bone remodeling and resorption, citrate is a useful biomarker for orthopedic disorders (Liu et al. 2025).

Citrate has also been shown to be a promising treatment for osteoporosis. Research shows that dietary citrate increases bone mass in postmenopausal women with osteopenia (Jehle et al. 2013).

A metabolomics study investigating the relationship between citrate, bone formation and diabetes mellitus found that osteocytes increase citrate excretion in response to mechanical stress, which may promote bone formation (Villaseñor et al. 2019). This process was significantly impaired in high-glucose conditions.

Citric acid and urology

Alterations in renal clearance of citrate can lead to kidney dysfunction. In chronic kidney disease, the kidney’s ability to excrete and reabsorb citrate can be impaired, leading to lower levels of urinary citrate (Granchi et al. 2019). This increases the risk of calcium-containing kidney stones, as calcium is more likely to crystallize without citrate’s binding effect. Oral citrate therapy is commonly used to increase urinary citrate levels and help prevent and treat kidney stones, though evidence of its efficacy is unclear (Phillips et al. 2015).

A study combining metabolomics with kidney gene expression studies found that urinary excretion of citric acid cycle metabolites – particularly citrate – and renal expression of genes regulating these metabolites were reduced in non-diabetic CKD (Hallan et al. 2017).

Citric acid and neurology

Citrate is attracting interest in the treatment of Alzheimer’s disease and dementia (AD) (Chhimpa et al. 2023). Low levels of citrate synthase and citrate impair the synthesis of acetyl-CoA and acetylcholine, resulting in loss of cognitive function in patients with AD. In addition, imbalances in the TCA cycle may contribute to the accumulation of amyloid-beta, which is implicated in AD. A recent network analysis suggesting that serum citrate concentrations are associated with cognitive decline (Jaramillo-Jimenez et al. 2023). This suggests that citrate may be a suitable biomarker and potential therapeutic option for AD and dementia.

The TCA cycle is also implicated in psychiatric conditions. A recent multiomics study using untargeted metabolomics, proteomics and DNA methylation data of patients with major psychiatric disorders confirmed the role of the TCA cycle metabolites, which may help identify biomarkers of disease (Hao M. et al. 2023).

Metabolomics research has also shown higher concentrations of citrate in the cerebrospinal fluid of patients with bipolar disorder compared to controls (Smedler et al. 2022).

Citric acid and oncology

Citrate is a major player in cancer cell metabolism, offering many potential avenues for cancer diagnosis and treatment. Citrate’s role in cancer metabolism was identified as part of the Warburg effect, where cancers preferentially use glycolysis even in the presence of oxygen (Icard et al. 2021). In proliferative cancer cells, low citrate levels sustain the Warburg effect by preventing inhibition of glycolytic enzymes. At the same time, citrate supports gluconeogenesis in cells relying on oxidative metabolism. Preclinical studies have shown that high doses of citrate have various anticancer effects, such as promoting apoptosis, neutralizing extracellular acidity and inhibiting tumor growth and signaling pathways (Icard et al. 2021). It may also enhance the effectiveness of cytotoxic drugs.

Citrate has been said to play a “dual role” in cancer, both supporting cancer cell proliferation and acting as a therapeutic agent (Icard et al. The dual role of citrate in cancer 2023). This opens the door to a “Trojan horse” therapeutic strategy, exploiting cancer cells’ reliance on citrate to disrupt their metabolic processes and trigger cell death (Petillo et al. 2020).

Citrate may be a useful biomarker for certain cancers: a review of multiomics approaches to metabolic phenotyping in prostate cancer puts citrate in the spotlight (Gómez-Cebrián et al. 2022). Citrate’s unique role in prostate cancer metabolism suggests it could be used as a biomarker alongside prostate-specific antigen (Galey et al. 2024).

Citric acid and 5P medicine

As shown above, multiomics approaches are providing a deeper understanding of citrate’s role in population health, disease prevention and treatment. These strategies are proving valuable not only in chronic conditions, but also in acute and emerging diseases.

For example, in the early stages of the COVID-19 pandemic, multiomics was used to provide deeper insights into the rapidly accumulating evidence and profile disease pathophysiology. This found notable citrate clusters, which led the researchers to conclude that monitoring and supplementing citrate levels could offer beneficial effects in severe COVID-19 patients (Overmyer et al. 2020).

References

• History & Evolution
• Biosynthesis vs. dietary uptake
• Oxaloacetic acid and the microbiome
• Oxaloacetic acid and neurological diseases
• Oxaloacetic acid, metabolic disorders and cardiovascular disease
• Oxaloacetic acid and cancer
• Oxaloacetic acid and bone regeneration

History & Evolution

1890s: features in early research into acid production (Raistrick et al. 1919) | 1935: discovery of mechanism of action (Szent-Györgyi, A. et al., 1935) | 1937: description of role in Krebs cycle (Krebs et al. 1937)

Oxaloacetic acid , also known as oxalacetic acid, is a crystalline short-chain keto acid found in bacteria, plants and animals. Early research into oxaloacetic acid began in the 1890s and 1900s, during investigations into acid production in bacteria (Bentley, R., 1994). In 1919, Harold Raistrick, the “father of the study of fungal metabolites,” and Anne Clark hypothesized that oxaloacetic acid was a precursor of citrate in fungi (Raistrick et al. 1919). In 1935, Szent-Györgyi observed that C₄dicarboxylic acids, including oxaloacetic acid, acted as catalysts in oxidation reactions (Szent-Györgyi, A. et al., 1935). Building on these findings, Hans Krebs and William Johnson investigated the role of citrate in carbohydrate metabolism, leading to the discovery of the “citric acid cycle” or Krebs cycle (also referred to as the tricarboxylic acid (TCA) cycle), in which oxaloacetic acid plays a key role (Krebs et al. 1937).

Since then, oxaloacetic acid has been found to participate in many other metabolic processes, including gluconeogenesis, the urea cycle, amino acid synthesis, and fatty acid synthesis (Wishart et al. 2022). It is therefore a metabolite of interest for a wide range of diseases, including cancer, diabetes, obesity and neurodegenerative conditions. Clinical trials involving oxaloacetic acid are quite rare, partly due to challenges in producing and storing the compound.

Biosynthesis vs. dietary uptake

Oxaloacetic acid is synthesized endogenously in all living organisms. In humans and animals, this centers on the TCA cycle, which drives energy production in the mitochondria. Oxaloacetic acid is formed through the oxidation of malate, catalyzed by the enzyme malate dehydrogenase (Arnold et al. 2023). Citrate synthase then combines oxaloacetic acid with acetyl-CoA to form citrate. The next steps in the TCA cycle support the generation of reduced nicotinamide adenine dinucleotide (NADH), flavine adenine dinucleotide (FADH2) and adenosine triphosphate (ATP) (Icard et al. 2021). The production of oxaloacetic acid is the trigger for each “turn” of the TCA cycle, making it the rate-limiting substrate for the cycle (Arnold et al. 2023).

Oxaloacetic acid also plays a critical role in gluconeogenesis, where it is synthesized from pyruvate by pyruvate carboxylase in the mitochondria (Attwood et al. 1995). Interestingly, to continue along this pathway, oxaloacetic acid needs to be converted to a molecule that is more easily transported into the cytosol and then reconverted into oxaloacetic acid (Holecek 2023 ). Once this sub-cellular location issue is solved, the pathway can continue and oxaloacetic acid is converted to phosphoenolpyruvate (PEP) by the enzyme PEP carboxykinase following the reverse path of gluconeogenesis towards synthesis of glucose.

Plants and bacteria generate oxaloacetic acid via a variation of the TCA cycle, known as the glyoxylate cycle. In plants, oxaloacetic acid is synthesized in the mesophyll cells during the C₄ acid cycle which underpins photosynthesis (Ludwig et al. 2016). It is quickly converted into malate or aspartate, and then transported to bundle-sheath cells in leaves, where decarboxylation releases CO₂ for carbohydrate synthesis. In bacteria such as E. coli, oxaloacetic acid can be produced from PEP and is an essential precursor for succinate synthesis (Tan et al. 2013).

Oxaloacetic acid is found in several foods, including citrus fruits, wild rice, canola and peanuts (Wishart et al. 2022).

Oxaloacetic acid and the microbiome

Oxaloacetic acid plays a role in amino acid synthesis, particularly through transamination reactions using glutamate from the TCA and urea cycles (Shen et al. 2023). Oxaloacetic acid is directly involved in the synthesis of two non-essential amino acids, aspartate and asparagine. Aspartate is required for the synthesis of additional amino acids like methionine, lysine and threonine. The conversion of oxaloacetic acid to aspartate also generates fumarate, which is recycled into oxaloacetic acid via malate in the urea cycle.

The gut microbiome can heavily influence these processes by modulating the availability of amino acids and intermediates in the TCA and urea cycles. Oxaloacetic acid participates in bacterial fermentation in the gut, along with other TCA cycle intermediates including succinate, fumarate, citrate and malic acid (Louis et al. 2017). Gut bacteria also affect lipid metabolism by regulating the availability of acetyl-CoA, with oxaloacetic acid facilitating its transfer within cells for fatty acid synthesis.

Diet is relevant here, with research into low-carbohydrate and ketogenic diets highlighting the links between diet, microbiome and oxaloacetic acid (Longo et al. 2019). These diets reduce glucose and pyruvate availability, reducing oxaloacetic acid levels via the TCA cycle (Hartman et al. 2007). With less oxaloacetic acid, the body struggles to handle high levels of acetyl-CoA generated from fat, leading to ketone production as an alternative energy source. It has been suggested that oxaloacetic acid plays a role in the mechanism of action for the ketogenic diet’s beneficial effects on epileptic seizures (Hartman et al. 2007).

Oxaloacetic acid and neurological diseases

There is some evidence of an association between oxaloacetic acid and neurological disease, though findings are mixed and clinical trials in humans have not shown consistent benefits.

Animal studies suggest that oxaloacetic acid stimulates mitochondrial biogenesis in the brain, reduces neuroinflammation and promotes hippocampal neurogenesis (Wilkins et al. 2014). Several animal models of stroke have shown that oxaloacetic acid can improve neurologic performance and blood brain barrier integrity, and reduce lesion size (Campos et al. 2012). Oxaloacetic acid decreases blood glutamate levels, in turn reducing brain glutamate levels which are elevated in ischemic stroke. An animal model of amyotrophic lateral sclerosis (ALS) found that treatment with oxaloacetic acid preserved motor function, delayed onset of neurological symptoms and reduced neuroinflammation (Tungtur et al. 2021).

The glutamate-oxaloacetic acid link may also be relevant to potential treatments for Alzheimer’s disease (AD), Parkinson’s disease and epilepsy (Selivanov et al. 2021). The Trial of Oxaloacetic acid in Alzheimer’s Disease (TOAD) study looked at whether oxaloacetic acid could modify brain chemistry and metabolism in AD patients (Vidoni et al. 2021). However, while oxaloacetic acid was found to increase brain energy metabolism in higher doses, it did not lead to consistent changes in blood markers or improvements in cognitive scores.

A small proof-of-concept trial has shown that oxaloacetic acid treatment can significantly reduce fatigue in patients with myalgic encephalomyelitis/chronic fatigue syndrome and long COVID (Cash et al. 2022).

While these findings are encouraging, and patient trials have shown oxaloacetic acid treatment to be safe, more research is needed to validate the benefits observed in animal models.

Oxaloacetic acid, metabolic disorders and cardiovascular disease

Because of its role in cellular metabolism, gluconeogenesis and the TCA cycle, oxaloacetic acid is a metabolite of interest in the study of metabolic disorders and cardiovascular disease.

Changes in the TCA cycle have been linked to cardiovascular disease and metabolic syndrome (Calderón et al. 2013). A metabolomics study used gas chromatography-mass spectrometry to profile TCA metabolites, including oxaloacetic acid, in patients with coronary lesions, showing that risk factors like obesity, hypercholesterolemia and smoking affect TCA metabolite levels.

Oxaloacetic acid has been shown to activate the insulin signaling pathway in murine models (Wilkins et al. 2014). In humans, a 1968 study showed that oxaloacetic acid reduced hyperglycemia in type II diabetes (Yoshikawa et al. 1968), but more recent evidence is lacking. Oxaloacetic acid supplementation has been hypothesized to mimic calorie restriction, which may help to reduce fasting glucose levels and improve insulin resistance (Cash et al. 2009).

More recently, oxaloacetic acid has been shown to have a protective effect in liver disease and injury. A 2018 metabolomics study found oxaloacetic acid alleviated liver injury by scavenging reactive oxygen species, increasing ATP and inhibiting mitochondrial apoptosis (Kuang et al. 2018). Metabolomic profiling has also shown how gut microbiota may contribute to liver disease progression: a 2021 study found elevated concentrations of short chain fatty acids and their precursors, including oxaloacetic acid, in samples from non-alcoholic fatty liver disease-related hepatocellular carcinoma (Behary et al. 2021).

Oxaloacetic acid and cancer

Oxaloacetic acid is of particular interest in cancer research because of its ability to reverse the “Warburg effect,” which refers to the glycolytic processes used by cancer cells for energy generation (Samad et al. 2023). Oxaloacetic acid has been shown to inhibit lactate dehydrogenase A (LDHA) in cancer cells to reduce Warburg glycolysis, and thus impair cancer cell metabolism. Clinical studies show oxaloacetic acid can also decrease tumor development by blocking the conversion of glutamine to alpha-ketoglutarate and lowering NADPH levels.

A 2020 study used metabolomics to investigate metabolic changes in melanoma cells (Seo et al. 2020). Treatment with alpha-melanoma-stimulating hormone led to elevated levels of oxaloacetic acid and other TCA cycle metabolites, suggesting potential therapeutic targets for skin pigmentation inhibitors and anti-obesity drugs.

Beyond the lab, oxaloacetic acid supplementation has been approved by the US FDA for patients with glioblastoma multiforme and has been proposed as a safe adjunct treatment for glioma along with established chemotherapy protocols (Tucker et al. 2020).

Oxaloacetic acid and bone regeneration

A 2024 study reveals that oxaloacetic acid may be a promising option for bone regeneration therapies (Shirkoohi et al. 2024). This study found that oxaloacetic acid could be used to support the differentiation of stem cells into bone-forming cells, which may help to treat osteogenic disorders or injuries.

Given oxaloacetic acid’s central role in the TCA cycle, which is essential for numerous cellular processes – and the encouraging findings in multiple animal studies – there is a compelling case to further investigate this metabolite as a therapeutic candidate for a wide range of diseases.

References

• History & Evolution
• Biosynthesis vs. dietary uptake
• Methionine and the microbiome
• Methionine and DNA methylation
• Methionine and cardiovascular disease
• Methionine and neurogenerative disorders
• Methionine and cancer

History & Evolution

1921: discovery (Mueller, J., 1921) | 1928: structure identified (Barger, G., 1928) | 1945-7: first synthesis (Livak, J. et al., 1945).

Methionine’s story begins just over a century ago, when it was discovered in casein by JH Mueller in 1921 (Mueller, J., 1921). In 1928, Barger and Coyne determined the chemical structure of what was until then known as γ-methylthiol-α-aminobutyric acid and suggested a more straightforward name: methionine (Barger, G., 1928). By the 1930s, methionine was recognized as an essential and limiting amino acid involved in protein synthesis, with subsequent research focusing on its vital role in human and animal diets (Rose, W., 1976). In the aftermath of World War II, protein shortages and widespread malnutrition prompted the hunt for a synthetic form of methionine (Livak, J. et al., 1945).

Now, over one million tons of methionine are produced each year from petroleum, with many applications including poultry feed, flavor enhancers, cosmetics, drugs and pesticides (Neubauer, C., 2021).

Methionine is found in humans, animals, plants and bacteria. In humans, methionine has been described as a “double-edged sword” (Navik, U. et al., 2021). It plays a crucial role in cellular, metabolic, epigenetic and genomic processes, influencing a broad range of physiological functions. It has been shown to support immune function, digestion, insulin resistance, to extend lifespan, and to reduce DNA damage and carcinogenesis (Martínez, Y. et al., 2017). However, excessive amounts can be toxic, with adverse effects including increased cholesterol and hyperhomocysteinemia (Navik, U. et al., 2021); (Garlick, P., 2006).

There is a theory that not only is this ancient metabolite essential for life, but – as a self-activating and sulfur containing amino acid – it may have triggered the development of life on Earth itself (Heinen, W., 1996).

Biosynthesis vs. dietary uptake

As an essential amino acid, methionine must be obtained via the diet and gut microbiota (Parkhitko, A. et al., 2019). Primary dietary sources of methionine include eggs, fish, meat, dairy and plant protein sources (Navik, U. et al., 2021) (Rajavel, E., 2020). In bacteria, methionine is synthesized from cysteine and aspartic acid. Plasma methionine concentration in healthy subjects ranges from 13 to 45 µM (Navik, U. et al., 2021).

The Quantitative Metabolomics Database (QMDB), cataloguing the concentrations of over 600 metabolites in healthy individuals reports average levels of 28 µM for women and 30 µM for men.

Methionine is one of four sulfur-containing amino acids, and, along with cysteine, one of two incorporated into proteins (Brosnan, T., 2006). Daily dietary requirements for methionine are usually taken together with cysteine, and the total sulfur amino acid requirement (TSAA) is 13-15 mg/kg/d for adults (Rajavel, E., 2020).

Methionine metabolism involves three key stages (Parkhitko, A. et al., 2019); (Lauinger, L., 2021):
– the methionine cycle,
– the transsulfuration/transmethylation pathway
– the methionine salvage cycle

In the methionine cycle, methionine and adenosine triphosphate (ATP) are converted into the methyl donor S-adenosylmethionine (SAM) by the enzyme methionine adenosyltransferase. After transferring the methyl group, SAM is converted to S-adenosylhomocysteine (SAH) and hydrolyzed into adenosine and homocysteine. This homocysteine can be remethylated to methionine by the methionine synthase enzyme (with vitamin B12 as a coenzyme), which completes the cycle, or diverted to the transssulfuration pathway to form cysteine (Rajavel, E., 2020); (Parkhitko, A. et al., 2019). In the salvage cycle, methionine is regenerated from decarboxylated SAM, following a series of enzymatic steps involving polyamine synthesis.

Methionine is a precursor to other molecules including antioxidants such as glutathione and cystathionine, and amino acids including cysteine, carnitine, taurine and creatine (Navik, U. et al., 2021).

Methionine and the microbiome

Gut microbiota play a major role in methionine metabolism, with around 20% of dietary methionine metabolized in the gut (Wu, X. et al., 2022); (Bauchart C. et al., 2009). In turn, methionine metabolism influences gut health, as evidenced by the interactions between methionine levels, microbial composition and disease states.

A multi-omics study comparing vegan and omnivore diets found that methionine positively correlates with Bacteroides, Blautia, Dorea, Lachnoclostridium and Fusicatenibacter, with all except Bacteroides being more abundant in omnivores (Prochazkova, M. et al., 2022). Another study found that restricting sulfur amino acids like methionine increases the abundance of Firmicutes, Clostridaceae and Turicibacteraceae, while decreasing Verrucomicrobia, compared to a low-calorie diet (Nichenametla, S. et al., 2021). These findings show how methionine intake influences the diversity and abundance of bacterial species in the gut.

Animal studies have shown that restricting dietary methionine, e.g. through a plant-based diet, may improve gut health. Methionine restriction in mice was found to increase short chain fatty acid (SCFA)-producing bacteria such as Bifidobacterium, Lactobacillus, Bacteroides, Roseburia, Coprococcus, and Ruminococcus, and increase inflammation-inhibiting bacteria like Oscillospira and Corynebacterium (Yang, Y. et al., 2019). In contrast, the high fat diet reduced SCFA production and induced gut dysbiosis.

Methionine appears to play a role in several inflammatory diseases associated with disrupted gut microbiota (Wu, X. et al., 2022). For example, a study in patients with inflammatory bowel disease (IBD) showed reduced levels of methionine, serine and sarcosine, along with fewer SCFA-producing gut bacteria (Borren, N. et al., 2021). Another study found changes in methionine and homoserine levels in patients with Parkinson’s disease (PD) (Hertel, J. et al., 2019). PD patients showed an increase in Akkermansia muciniphila, which is involved in sulfur metabolism. This contributed more than 70% of potential methionine production, suggesting a clear link between gut microbiota and the methionine cycle (Wu, X. et al., 2022).

Methionine and DNA methylation

Methionine is essential for DNA methylation, a key epigenetic mechanism affecting gene expression related to growth and development (Lauinger, L., 2021). In this process, DNA methyltransferases transfer a methyl group from SAM to the C-5 position of the pyrimidine ring of cytosine (Jin, B. et al., 2011). Changes in dietary methionine change the ratio of SAM to SAH (the “methylation index”), which influences methylation capacity. This can be measured using immunoassays to detect levels of SAM and SAH in blood and tissue (Hao, X. et al., 2016).

Dysregulation of DNA methylation is associated with some cancers (Bauchart C. et al., 2009). Studies suggest that methionine supplementation can affect DNA methylation, with both positive and negative effects depending on the gene region affected (Waterland, R., 2006).

More research is needed to fully understand methionine’s role in DNA methylation. Metabolomics offers a useful tool with which to understand epigenomics and DNA methylation in more detail, especially when combined with other omics. For example, a 2020 study combined metabolomics, proteomics and genomics to show that activated T cells use methionine to synthesize SAM, identifying methionine as a key nutritional factor in shaping T helper cell activity through histone methylation (Roy, D. et al., 2020). The findings suggest restricting dietary methionine could be a viable intervention in the treatment of autoimmune conditions such as multiple sclerosis.

Layering insights from different omics analyses in this way gives a fuller picture of the interactions between metabolites, proteins, genes and environmental factors that drive disease processes.

Methionine and cardiovascular disease

Too much methionine may increase the risk of cardiovascular disease (CVD). Methionine is involved in DNA synthesis, lipid regulation, oxidative defense and cellular regulation – processes all thought to play a role in CVD and other diseases (Aissa, A. et al., 2017). Impaired DNA methylation may also be a factor, as this increases circulating LDL cholesterol and lowers HDL cholesterol, both of which are risk factors for CVD (Calderón, A. et al., 2020); (Blachier, F. et al., 2020).

Another key mechanism is linked to methionine’s downstream metabolite homocysteine, which is known to be vasotoxic (Troen, A. et al., 2003). In excess, homocysteine can cause hyperhomocysteinemia, which is a well-established risk factor for CVD. A longitudinal population-based study found that a lower concentration of methionine, higher concentration of homocysteine and lower ratio of methionine to homocysteine were associated with an increased risk of CVD (Calderón, A. et al., 2020).

Excess methionine may also damage endothelial cells and increase plasma lipid levels, which both contribute to atherosclerosis. High levels of dietary methionine have been found to increase the risk of acute coronary events in middle-aged men (Virtanen, J. et al., 2006).

By contrast, methionine restriction appears to delay development of CVD and associated risk factors such as obesity (Zhang, Y. et al., 2022).

Methionine and neurogenerative disorders

Excess methionine is linked to neurodevelopmental disorders, like autism and schizophrenia, and neurodegenerative disorders such as Alzheimer’s disease (AD) and PD (Alachkar, A. et al., 2022); (Hertel, J. et al., 2019). A 2023 study involving animal and human AD phenotypes found that methionine intake is associated with mild cognitive impairment, and methionine restriction improves cognitive function (Xi, Y. et al., 2022).

More research is needed to understand the mechanisms, but again, methionine’s role in DNA methylation and homocysteine production may be relevant. A population-based study found links between methionine, homocysteine and dementia development, suggesting that a higher methionine to homocysteine ratio may reduce brain atrophy and lower the risk of dementia (Hooshmand, B. et al., 2019).

As noted above, a longitudinal metabolomics study investigated links between PD, sulfur metabolism and the gut microbiome (Hertel, J. et al., 2019). This showed differences between methionine and cysteine production via cystathionine in PD patients and healthy subjects. Using multiomics, the researchers were able to identify patterns in microbial-host sulfur metabolism that may contribute to PD severity.

Methionine and cancer

Methionine restriction is also an “exciting potential tool in the treatment of cancer” (Wanders, D. et al., 2020). Methionine inhibits cancer cell proliferation and growth in several types of cancer, without damaging healthy cells (in the presence of homocysteine). It also appears to improve efficacy of chemotherapy and radiation therapy in animal models. A 2020 review summarized evidence showing that methionine restriction could inhibit cancer development and/or progression in prostate cancer, breast cancer and colorectal cancer (Wanders, D. et al., 2020). Possible mechanisms vary by cancer type. It may be that methionine restriction inhibits polyamine biosynthesis, induces apoptosis, alters DNA methylation and glutathione formation, and reduces activity of thymidylate synthase, the enzyme that converts dUMP to dTMP.

 
More about the Metabolite of the Month
Reference ranges of methionine in healthy humans
Quantify the impact of methionine

References

Aissa, A. et al.: Methionine-supplemented diet affects the expression of cardiovascular disease-related genes and increases inflammatory cytokines in mice heart. (2017). J Toxicol Environ Health A | DOI: 10.1080/15287394.2017.1357366

Alachkar, A. et al.: L-methionine enhances neuroinflammation and impairs neurogenesis: implication for alzheimer’s disease. (2022). Journal of Neuroimmunology | DOI: 10.1016/j.jneuroim.2022.577843

Barger, G. et al.: The amino-acid methionine; constitution and synthesis. (1928). Biochem J. | DOI: 10.1042/bj0221417

Bauchart, C. et al.: Intestinal metabolism of sulfur amino acids. (2009). Research Reviews | DOI: 10.1017/S0954422409990138

Blachier, F. et al.: Sulfur-containing amino Acids and lipid metabolism. (2020). The Journal of Nutrition | DOI: 10.1093/jn/nxaa243

Borren, N. et al.: Alterations in fecal microbiomes and serum metabolomes of fatigued patients with quiescent inflammatory bowel diseases. (2021). Clin Gastroenterol Hepatol | DOI: 10.1016/j.cgh.2020.03.013

Brosnan, T. et al.: The sulfur-containing amino acids: an overview. (2006). J Nutr | DOI: 10.1093/jn/136.6.1636S

Calderón, A. et al.: Association of homocysteine, methionine, and mthfr 677c>t polymorphism with rate of cardiovascular multimorbidity development in older adults in sweden. (2020). JAMA Netw Open | DOI: 10.1001/jamanetworkopen.2020.5316

Garlick, P. et al.: Toxicity of methionine in humans. (2006). The Journal of Nutrition | DOI: 10.1093/jn/136.6.1722S

Hao, X. et al.: Immunoassay of S-adenosylmethionine and S-adenosylhomocysteine: the methylation index as a biomarker for disease and health status. (2016). BMC Res Notes | DOI: 10.1186/s13104-016-2296-8

Heinen, W. et al.: Organic sulfur compounds resulting from the interaction of iron sulfide, hydrogen sulfide and carbon dioxide in an anaerobic aqueous environment. (1996). Orig Life Evol Biosph | DOI: 10.1007/BF01809852

Hertel, J. et al.: Integrated analyses of microbiome and longitudinal metabolome data reveal microbial-host interactions on sulfur metabolism in parkinson’s disease. (2019). Cell Rep | DOI: 10.1016/j.celrep.2019.10.035

Hooshmand, B. et al.: Association of methionine to homocysteine status with brain magnetic resonance imaging measures and risk of dementia. (2019). JAMA Psychiatry | DOI: 10.1001/jamapsychiatry.2019.1694

Jin, B. et al.: DNA methylation. (2011). Genes Cancer | DOI: 10.1177/1947601910393957

Kaiser, P. et al.: Methionine dependence of cancer. (2020). Biomolecules | DOI: 10.3390/biom10040568

Lauinger, L. et al.: Sensing and signaling of methionine metabolism. (2021). Metabolites | DOI: 10.3390/metabo11020083

Livak, J. et al.: Synthesis of dl-methionine. (1945). J Am Chem Soc | DOI: 10.1021/ja01228a050

Martínez, Y. et al.: The role of methionine on metabolism, oxidative stress, and diseases. (2017). Amino Acids | DOI: 10.1007/s00726-017-2494-2

Mueller, J. et al.: Growth-determining substances in bacteriological culture media. (1921). Proceedings of the Society for Experimental Biology and Medicine | DOI: 10.3181/00379727-18-113

Navik, U. et al.: Methionine as a double-edged sword in health and disease: Current perspective and future challenges. (2021). Ageing Research Reviews | DOI: 10.1016/j.arr.2021.101500

Neubauer, C. et al.: A planetary health perspective on synthetic methionine. (2021). The Lancet Planetary Health | DOI: 10.1016/S2542-5196(21)00138-8

Nichenametla, S. et al.: Differential Effects of Sulfur Amino Acid-Restricted and Low-Calorie Diets on Gut Microbiome Profile and Bile Acid Composition in Male C57BL6/J Mice. (2021). The Journals of Gerontology: Series A | DOI: 10.1093/gerona/glaa270

Parkhitko, A. et al.: Methionine metabolism and methyltransferases in the regulation of aging and lifespan extension across species. (2019). Aging Cell | DOI: 10.1111/acel.13034

Prochazkova, M. et al.: Vegan diet Is associated with favorable effects on the metabolic performance of intestinal microbiota: A cross-sectional multi-omics study. Front. (2022). Nutr | DOI: 10.3389/fnut.2021.783302

Rajavel, E. et al.: Methionine Nutrition and Metabolism: Insights from Animal Studies to Inform Human Nutrition. (2020). The Journal of Nutrition | DOI: 10.1093/jn/nxaa155

Rose, W. et al.: Amino Acid Requirements of Man. (1976). Nutrition Reviews | DOI: 10.1111/j.1753-4887.1976.tb05679.x

Roy, D. et al.: Methionine metabolism shapes t helper cell responses through regulation of epigenetic reprogramming. (2020). Cell Metab | DOI: 10.1016/j.cmet.2020.01.006

Troen, A. et al.: The atherogenic effect of excess methionine intake. (2003). PNAS | DOI: 10.1073/pnas.2436385100

Virtanen, J. et al.: High dietary methionine intake increases the risk of acute coronary events in middle-aged men. (2006). Nutr Metab Cardiovasc Dis | DOI: 10.1016/j.numecd.2005.05.005

Wanders, D. et al.: Methionine restriction and cancer biology. (2020). Nutrients | DOI: 10.3390/nu12030684

Waterland, R. et al.: Assessing the Effects of High Methionine Intake on DNA Methylation. (2006). The Journal of Nutrition | DOI: 10.1093/jn/136.6.1706S

Wu, X. et al.: Gut microbiota contributes to the methionine metabolism in host. (2022). Front Microbiol | DOI: 10.3389/fmicb.2022.1065668

Xi, Y. et al.: Effects of methionine intake on cognitive function in mild cognitive impairment patients and APP/PS1 Alzheimer’s Disease model mice: Role of the cystathionine-β-synthase/H2S pathway. (2022). Redox Biol | DOI: 10.1016/j.redox.2022.102595

Yang, Y. et al.: Dietary methionine restriction improves the gut microbiota and reduces intestinal permeability and inflammation in high-fat-fed mice. (2019). Food and Function | DOI: 10.1039/C9FO00766K

Zhang, Y. et al.: Methionine restriction – Association with redox homeostasis and implications on aging and diseases. (2022). Redox Biology | DOI: 10.1016/j.redox.2022.102464

• History & Evolution
• Biosynthesis vs. dietary uptake
• Indoxyl sulfate and the microbiome
• Indoxyl sulfate and nephrology
• Indoxyl sulfate and cardiovascular disease
• Indoxyl sulfate and bone disease
• Indoxyl sulfate and neurological disease

History & Evolution

1911: discovery of indoxyl sulfate (Obermayer, F. and Popper, H., 1911) | 1936: confirmation of liver’s role in indoxyl sulfate production (Houssay, B., 1936) | 1950s: discovery of role in kidney disease (Leong, S. and Sirich, T., 2016).

In 1911, Obermayer and Popper discovered high concentrations of a metabolite, then called “indican,” in patients with kidney disease (Obermayer, F. and Popper, H., 1911). Indoxyl sulfate, as it later came to be known, was initially studied as a “putrefaction” product of intestinal microbial metabolism. In the 1950s, researchers explored whether urinary excretion of indoxyl sulfate might indicate various diseases, eventually focusing on its role in kidney disease. This was a major leap forwards in understanding uremia, which had puzzled scientists for decades (Leong, S. and Sirich, T., 2016).

As a key uremic toxin, accumulation of indoxyl sulfate affects the kidneys, cardiovascular system and bones (Colombo, G. et al., 2022). Indoxyl sulfate has been linked to cognitive impairment, oxidative stress and blood-brain barrier permeability, and is also thought to play a role in remote sensing and signaling in renal and extra-renal tissues (Lowenstein, J. and Nigam, S., 2021). Recent research has focused on IS as a potential therapeutic target in chronic kidney disease and cardiovascular disease, especially when these conditions are comorbid.

Indoxyl sulfate is a precursor of indigo, which has been used as a dye for thousands of years. This connection is illustrated vividly in “purple urine bag syndrome,” where constipation or urinary tract infections cause an accumulation of bacteria in urine collection bags, leading to the conversion of indoxyl sulfate into indigo and indirubin, and resulting in purple urine (Tan, C. et al., 2008).

Biosynthesis vs. dietary uptake

Indoxyl sulfate is produced from precursors synthesized via the tryptophan-indole pathway in the gut (Banoglu, E. et al., 2001). In the first step, L-tryptophan, an amino acid obtained from dietary protein, is metabolized by tryptophanase-expressing bacteria in the intestine to produce indole. Indole is absorbed into the bloodstream and is converted to indoxyl in the liver via hydroxylation by cytochrome enzymes including CYP2E1. Sulfotransferase enzymes in the liver complete the process by converting indoxyl into indoxyl sulfate.

Research suggests that increasing dietary tryptophan (typically found in protein-rich foods) may increase indoxyl sulfate production (Lauriola, M. et al., 2023; Leong, S. and Sirich, T., 2016). I ndividuals following high-protein diets show higher levels of indoxyl sulfate in both plasma and urine compared to those on low-protein or vegetarian diets (Patel, K. et al., 2012). Conversely, very low protein diets have been shown to reduce indoxyl sulfate levels (Marzocco, S. et al., 2013). A study of 56 hemodialysis patients found that increasing dietary fiber also reduced indoxyl sulfate levels (Sirich, T. et al., 2014).

In healthy individuals, the indoxyl sulfate concentrations range is10-130 mg/day, clearing rapidly through the kidneys (Patel, K. et al., 2012). Kidney dysfunction inhibits clearing, causing indoxyl sulfate to accumulate, which contributes to renal and cardiovascular toxicity as discussed below (Hung, S. et al., 2017). The full extent of IS’ role in healthy phenotypes remains to be discovered.

Indoxyl sulfate and the microbiome

More than 85 species of bacteria produce indole through the action of tryptophanase on tryptophan (Melander, R. et al., 2014). Tryptophanase has been found in bacteria including Lactobacillus, Bifidobacterium longum, Bacteroides fragilis, Parabacteroides distasonis, Clostridium bartlettii and E. hallii (Zhang, L. and Davies, S., 2016). Even more species respond to the presence of indole, despite lacking tryptophanase and thus not producing indole themselves.



The role of the liver in indoxyl sulfate production was known as far back as 1936, when animal studies revealed that indoxyl sulfate could be produced even in the absence of a digestive tract, as long as the liver was intact (Houssay, B., 1936). More recently, metabolomics has confirmed the involvement of the gut microbiome in indoxyl sulfate production: studies comparing indoxyl sulfate levels in conventional and germ-free rats, and in hemodialysis patients with and without colons, highlight the significant role of colon bacteria (Leong, S. and Sirich, T., 2016).

Indoxyl sulfate and nephrology

As a uremic toxin, indoxyl sulfate is strongly implicated in the progression of chronic kidney disease (CKD) (Caggiano, G. et al., 2023). As renal function declines, indoxyl sulfate concentrations rise, making it a useful biomarker of CKD. Both indoxyl sulfate and p-cresol sulfate, another protein-bound uremic toxin, are linked to symptoms of renal disease such as inflammation and fibrosis (Caggiano, G. et al., 2023). Indoxyl sulfate levels increase progressively with each stage of CKD (Clark, W. et al., 2019). Patients with hospital-acquired acute kidney injury have also been found to have higher serum levels of indoxyl sulfate (Menez, S. et al., 2019). Hemodialysis is limited in its ability to remove plasma protein bond, and dialysis patients with end-stage renal disease have been found to have indoxyl sulfate levels more than 20 times higher than those of healthy subjects (Wang, Y. et al., 2020).

Emerging evidence suggests that interventions that reduce indoxyl sulfate levels may slow CKD progression, though results remain inconclusive (Wang, Y. et al., 2020). Generally, there appears to be a good indication that administering prebiotics, probiotics and synbiotics can reduce indoxyl sulfate in patients with CKD and renal failure, although further research is needed (Lim, Y. et al., 2021).

Administration of AST-120 seems to be particularly promising for CKD (Lim, Y. et al., 2021). This treatment works by adsorbing indole and p-cresol to minimize downstream indoxyl sulfate production. AST-120 has been shown to slow the decline in glomerular filtration rate of CKD patients in some clinical studies, although randomized controlled trials have been inconclusive. More research is needed to determine whether gut microbiome modulation is a viable strategy in kidney disease.

Indoxyl sulfate and cardiovascular disease

Cardiovascular disease is higher among patients with CKD than the general population. While this relationship is not entirely understood, uremic toxins are thought to play a role, acting as vascular toxins as they accumulate in the blood (Barreto, F. et al., 2009; Zhao, Y. and Wang, Z., 2020). IS disrupts endothelial integrity and is linked to pro-oxidant, pro-inflammatory and prothrombotic processes, contributing to atherosclerosis, aortic calcification and vascular disease (Lano, G. et al., 2020). Indoxyl sulfateis also an agonist for the aryl hydrocarbon receptor (AhR) on vascular smooth muscle cells, further promoting the formation of atherosclerotic lesions and thrombosis (Leong, S. and Sirich, T., 2016). These mechanisms highlight indoxyl sulfate as a key player in the progression of cardiovascular complications in CKD.



Metabolomics studies have provided further insight into the role of indoxyl sulfate in cardiovascular health. For example, research shows that elevated indoxyl sulfate in plasma predicts cardiovascular events in patients with congestive heart failure and hypertrophic cardiomyopathy (Imazu, M. et al., 2020). Another study showed an association between plasma indoxyl sulfate levels and arterial stiffness in patients with type 2 diabetes (Katakami, N. et al., 2020). Diabetes can disrupt gut microbial composition and metabolism, which may contribute to increased indoxyl sulfate levels.

Indoxyl sulfate and bone disease

Patients with CKD often develop mineral bone disorders (CKD-MBD). Research suggests a link between this disorder and IS, though the precise mechanism is unclear (Liu, W. et al., 2018). One possibility is that the accumulation of uremic toxins weakens bone quality and quantity, potentially leading to uremic osteoporosis. Bone responses to parathyroid hormone (PTH) are also progressively lower in CKD patients, with studies suggesting a connection between PTH resistance, low bone turnover and increased indoxyl sulfate levels. Furthermore, indoxyl sulfate acts on AhR and activates signaling pathways that can induce ferroptosis and disrupt osteoblast differentiation (Chen, H. et al., 2024). Uremic toxin adsorbents may be a potential therapeutic strategy to improve bone health in patients with renal disease.

Indoxyl sulfate and neurological disease

A relatively new area of interest is the link between indoxyl sulfate and neurology, particularly its involvement in Alzheimer’s disease, Parkinson’s disease, anxiety and other neurological disorders. In CKD patients, accumulation of uremic toxins can lead to cerebrovascular lesions, increasing the risk of cognitive disorders and dementia (Sankowski, B. et al., 2020).

Using metabolomics, Sankowski et al. demonstrated that patients with Parkinson’s disease had higher levels of indoxyl sulfate and p-cresol sulfate in cerebrospinal fluid compared to controls, and higher levels of another uremic toxin, trimethylamine N-oxide (TMAO), in plasma (Sankowski, B. et al., 2020).

Another metabolomics study quantified molecular markers in whole blood of dementia patients, and found elevated levels of indoxyl sulfate compared to healthy controls, again suggesting that indoxyl sulfate may be neurotoxic (Teruya, T. et al., 2021).

Indoxyl sulfate levels also positively correlate with severity of anxiety, according to an exploratory metabolomics study (Brydges, C. et al., 2021). While modulating indole levels did not seem to affect treatment outcomes, the fact that a gut microbiome-derived metabolite influenced neural processing provides further evidence of a link between the gut microbiome and neuropsychiatric disorders.

 
Quantify the impact of indoxyl sulfate
Reference ranges of indoxyl sulfate in healthy humans
Learn more about the Metabolite of the Month

References

• History & Evolution
• Biosynthesis vs. dietary uptake
• Kynurenic acid and the microbiome
• Kynurenic acid receptors
• Kynurenic acid and neurology
• Kynurenic acid and cancer
• Kynurenic acid and metabolomic diseases

History & Evolution

1853: discovery (Liebig et al. 1853) | 1904: first synthesized from tryptophan (Ellinger et al. 1904) | 1980s: mode of action discovered

Kynurenic acid (KYNA) was first discovered in 1853 by German chemist Justus von Liebig in dog urine, which inspired its name (Liebig 1853). Fifty years later, another German chemist, Ellinger, identified it as one of the first metabolites isolated from tryptophan, again from dog urine (Ellinger 1904). Initially considered little more than a by-product of the more interesting kynurenine, KYNA caught the attention of researchers investigating the brain and central nervous system (CNS) in the 1980s and 90s, when it was identified as an antagonist of ionotropic glutamate receptors (Turski et al. 2013).

KYNA is found in several mammalian organs and tissues, including the brain, retina, liver, kidneys, intestines, cardiac muscles and bodily fluids, including breast milk (Turska et al. 2022). It is also found in plants, although its role in plant physiology remains largely unexplored (Wróbel-Kwiatkowska et al. 2024).

KYNA is associated with a wealth of health benefits, including antioxidant, anti-inflammatory and neuroprotective properties. Its well-established role in neurotransmission systems links it to several neurological and cognitive conditions, such as Parkinson’s disease (PD), Alzheimer’s disease (AD), schizophrenia and mood disorders (Zhen et al. 2022). More recent evidence points to a role in energy homeostasis and in the immune and digestive systems, and it is also emerging as a potential biomarker of metabolic disease and endocrine disease (Zhen et al. 2022).

Biosynthesis vs. dietary uptake

KYNA is primarily synthesized intracellularly via the kynurenine pathway. While this process occurs endogenously, it relies on tryptophan, an exogenous essential amino acid that must be obtained through diet (Friedman et al. 2018). Dietary sources of tryptophan include seeds, soybeans, dairy, meat, fish, eggs and legumes. KYNA itself is usually found in quite low concentrations in food, with the exception of honey, broccoli and potatoes (Turski et al. 2009).

The kynurenine pathway metabolizes more than 90% of dietary tryptophan in the liver to form kynurenine, KYNA, quinolinic acid (QUIN) and other metabolites (Hou et al. 2023, Stone et al. 2013). In humans, kynurenine is primarily synthesized endogenously from tryptophan. This reaction is catalyzed by the enzyme tryptophan 2,3-dioxygenase (TDO) in the liver and, to a lesser extent, by indoleamine 2,3-dioxygenase (IDO) in other organs including immune cells (Savitz et al. 2020). Alternative mechanisms that may influence KYNA production in the brain involve D-amino acids, D-amino acid oxidase, and the effects of free radicals (Ramos-Chávez et al. 2018).

KYNA is synthesized in the gut microbiome, absorbed through the lumen of the gastrointestinal (GI) tract and transported to the liver, kidneys and other organs via the bloodstream (Turska et al. 2022). Concentration levels in digestive fluids increase steadily following ingestion, with blood levels peaking at around 15-30 minutes following ingestion (Sadok et al. 2023). Any that is not absorbed is excreted in urine.

Kynurenic acid and the microbiome

In the GI tract, tryptophan metabolism is modulated by gut microbiota including the bacterial phyla Actinobacteria, Bacteroides, Firmicutes, Fusobacteria, and Proteobacteria (Dehhaghi et al. 2020). Gut microbiota can influence the kynurenine pathway by regulating IDO-1 activity or tryptophan availability (Dehhaghi et al. 2019). For example, administering Bifidobacterium infantis to germ-free mice has been shown to increase KYNA levels (Desbonnet et al. 2008). Similarly, IDO and TDO have been found to reduce the kynurenine-to-tryptophan ratio in germ-free animals, but when those animals were exposed to microbiota, the enzyme activity normalized (Cryan et al. 2012).

The production of kynurenine metabolites is known to affect host physiology, with multiple studies demonstrating how manipulating microbial populations influences the kynurenine pathway and neural, endocrine and immune pathways (Cryan et al. 2012) (Dehhaghi et al. 2019). Differences in gut microbiota of healthy individuals and those with clinical conditions have been observed in diseases associated with the kynurenine pathway, such as anxiety, depression, schizophrenia, AD, PD, multiple sclerosis, ASD and irritable bowel syndrome (Purton et al. 2021).

There is still much to learn about the role of specific bacteria in producing KYNA and related therapeutic possibilities (Dehhaghi et al. 2019). A 2021 systematic review by Purton et al. showed that probiotics could modulate metabolite activity in the kynurenine pathway. However, evidence regarding the effect of prebiotics on this pathway remains limited (Purton et al. 2021).

Find out more about neurometabolomics, an evolving approach to better understand the role of KYNA and other metabolites in brain processes and function.

Kynurenic acid receptors

KYNA is a ligand to several receptors, acting as a signaling molecule for multiple physiological processes:

• It is an antagonist of the three ionotropic glutamate receptors N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainite, which mediate its effects in the brain (Schwarcz, R. et al. 2012). This makes it a useful test for glutamate involvement in synaptic transmission, for example in cases of stroke (Stone et al. 2000).
• KYNA is also thought to be an antagonist of the alpha7 nicotinic acetylcholine receptor (α7nAChR) (Albuquerque et al. 2013). However, this hypothesis is controversial: the results of studies showing that KYNA blocks NMDA receptors but not nicotinic receptors have not been consistently reproducible, while investigations using KYNA analogs may be vulnerable to misinterpretation (Stone et al. 2020). Given the role of α7nAChR in CNS disorders, there’s a strong case for more research to better understand if and how KYNA may block its action.
• KYNA has been more assuredly identified as an agonist of the G-protein coupled receptor (GPCR) GPR35, which is primarily present in the GI tract (Turski et al. 2013). Research in this area has led to the suggestion that KYNA could have a mediating effect on GI disorders, including ulcers, colitis and colon obstruction. KYNA’s role in modulating GPR35 also links it to immune system regulation, as does its role as an agonist of the aryl hydrocarbon receptor (AhR) (Tsuji et al. 2023). In 2020, researchers using DCyFIR, a CRISPR-based GPCR screening platform for ligand and drug discovery, found that KYNA activates hydroxycarboxylic acid receptor (HCAR3) in addition to GPR35 (Kapolka et al. 2020).

Because of its varied signaling effects on the immune system, inflammation, neuropathology, cancer and gut homeostasis, KYNA has been described as a “double-edged sword” (Wirthgen et al. 2017). Its anti-inflammatory and immunosuppressive functions are beneficial in some respects, but elevated levels of circulating KYNA are also associated with several diseases, as discussed below.

Interestingly, there appears to be a sex-specific association between circulating KYNA and immune response: a study using untargeted metabolomics found that male patients with COVID-19 had higher levels of KYNA than females, correlating with age, inflammation and disease severity (Cai et al. 2021). This may explain a potential link between KYNA levels and the differing outcomes in men and women with COVID-19.

Kynurenic acid and neurology

KYNA levels are altered in various mental disorders, with decreased levels observed in patients with affective psychosis, chronic schizophrenia, AD, cluster headaches, and chronic migraines (Wirthgen et al. 2017). Patients with PD show increased ratios of KYNA and KYNA/kynurenine, along with lower ratios of QUIN and QUIN/KYNA (Chang et al. 2018). Elevated KYNA in the cerebrospinal fluid of schizophrenia patients and its correlation with inflammatory biomarkers in AD suggest a link between KYNA and neuroinflammation (Wennström et al. 2014).
These findings highlight KYNA’s role in the pathophysiology of neurodegeneration and mental disorders, potentially through mechanisms involving oxidative stress, glutamatergic excitotoxicity, and inflammation. The participation of gut microbiota associated with the alternative routes to KYNA production involving D-amino acids and free radicals may also be relevant here (Ramos-Chávez et al. 2018).

The white-paper, Complex chronic diseases have a common origin, published by biocrates in 2023, includes a discussion of the relationship between KYNA, NDMA receptors and major depressive disorder (MDD). While KYNA inhibits the NMDA receptor and is considered neuroprotective, its fellow kynurenine metabolite quinolinic acid (QUIN) has the opposite effect and is considered neurotoxic. The QUIN/KYNA ratio is increased in the serum of patients with MDD) compared to healthy subjects, and correlates with the duration of remission, suggesting a role for these metabolites in disease progression (Savitz et al. 2015).

A metabolomics study by Erabi et al. found that KYNA was decreased in MDD, and lower levels showed a better therapeutic response to escitalopram (Erabi et al. 2020). This suggests KYNA is a potential overlapping biomarker in diagnosis and predicting therapeutic response.

KYNA does not effectively cross the blood-brain barrier, therefore limiting the potential efficacy of oral administration of KYNA in mitigating neurological and psychological diseases (Turska et al. 2022).

Kynurenic acid and cancer

Kynurenic acid’s effects in the periphery are not as well understood as its role in the brain. Still, there is a growing – if ambiguous – body of evidence confirming the presence of KYNA in several types of cancer in tumor tissue from patients with colon adenocarcinoma, glioblastoma, renal cell carcinoma (RCC) and oral squamous cell carcinoma (SCC) (Walczak et al. 2020). Elevated levels of KYNA have been found in the serum of patients with colon adenocarcinoma and lung cancer, and have been associated with invasiveness of lung cancer. In other cases, such as glioblastoma, cervical and prostate cancer, KYNA levels have been found to be lower than in healthy controls.

The precise mechanisms remain unclear. One possibility is that KYNA’s action on GPR35 initiates carcinogenesis and influences cell proliferation, survival and metastasis (Basson et al. 2023). KYNA may also interfere with nicotinamide adenine dinucleotide (NAD+) pathways, which is central to cancer biology. And as noted, KYNA activates the AhR pathway, which is also commonly activated in cancer patients, though data suggests this can be both pro- and anti-metastatic (Basson et al. 2023).

Multiple clinical trials are investigating the kynurenine pathway as a potential target for cancer therapies. More research is needed to understand where KYNA fits in the puzzle.

Kynurenic acid and metabolic diseases

Metabolomic and epidemiological research has shown that KYNA may play a role in diabetes and other metabolic diseases, and may protect against obesity and nonalcoholic fatty liver disease (Zhen et al. 2022). Deficiency in dietary KYNA is associated with adipose tissue dysfunction, with links to insulin sensitivity, energy homeostasis and lipid and carbohydrate metabolism (Tomaszewska et al. 2019). In contrast, KYNA synthesis may be influenced by physical exercise, potentially resulting in increased thermogenesis and helping to limit weight gain, insulin resistance and inflammation (Zhen et al. 2022).
Metabolomics research is increasing our understanding of the role of KYNA in metabolic disease and in turn revealing potential therapeutic targets. For example, a recent metabolomics study using an animal model found that KYNA production increases following hepatic ischemia-reperfusion injury, which diverts resources from NAD synthesis (Xu. et al. 2022). Augmenting NAD levels were found to mitigate oxidative stress, inflammation and cell death in the liver.

References