Diet- and microbiota-related metabolite, 5-aminovaleric acid betaine (5-AVAB), in health and disease
Authors: Retu Haikonen, Olli Kärkkäinen, Ville Koistinen, Kati Hanhineva
Journal: Trends in Endocrinology and Metabolism (2022)

5-Aminovaleric acid betaine (5-AVAB) is a trimethylated compound that has recently emerged as a metabolically important compound in numerous studies.Lack of microbiota reduces 5-AVAB and certain microbiota taxa are essential for 5-AVAB production.Diets influence levels of 5-AVAB, with certain diets promoting bacterial biosynthesis of 5-AVAB and some readily containing 5-AVAB.Current evidence for the metabolic role of 5-AVAB is rather contradictory, including both beneficial and harmful observations.A strong relationship of 5-AVAB is shown (e.g., with gut–brain axis, liver and heart function, and adiposity). 5-Aminovaleric acid betaine (5-AVAB) is a trimethylated compound associated with the gut microbiota, potentially produced endogenously, and related to the dietary intake of certain foods such as whole grains. 5-AVAB accumulates within the metabolically active tissues and has been typically found in higher concentrations in the heart, muscle, and brown adipose tissue. Furthermore, 5-AVAB has been associated with positive health effects such as fetal brain development, insulin secretion, and reduced cancer risk. However, it also has been linked with some negative health outcomes such as cardiovascular disease and fatty liver disease. At the cellular level, 5-AVAB can influence cellular energy metabolism by reducing β-oxidation of fatty acids. This review will focus on the metabolic role of 5-AVAB with respect to both physiology and pathology. Moreover, the analytics and origin of 5-AVAB and related compounds will be reviewed. 5-Aminovaleric acid betaine (5-AVAB) is a trimethylated compound associated with the gut microbiota, potentially produced endogenously, and related to the dietary intake of certain foods such as whole grains. 5-AVAB accumulates within the metabolically active tissues and has been typically found in higher concentrations in the heart, muscle, and brown adipose tissue. Furthermore, 5-AVAB has been associated with positive health effects such as fetal brain development, insulin secretion, and reduced cancer risk. However, it also has been linked with some negative health outcomes such as cardiovascular disease and fatty liver disease. At the cellular level, 5-AVAB can influence cellular energy metabolism by reducing β-oxidation of fatty acids. This review will focus on the metabolic role of 5-AVAB with respect to both physiology and pathology. Moreover, the analytics and origin of 5-AVAB and related compounds will be reviewed. Recent advances in mass spectrometry (MS)-based analytical methods and the increased use of metabolomics approaches have opened novel avenues to permit a much more detailed molecular-level assessment of human metabolism and its linkage with gut microbiota, as well as cellular metabolic routes. Novel compounds, and even compound groups, with putative roles in metabolic processes, are continually being described. One such compound is 5-AVAB, which belongs to the trimethylated compounds commonly called betaines (see Glossary). Glycine betaine [1.Craig S.A. Betaine in human nutrition.Am. J. Clin. Nutr. 2004; 80: 539-549Crossref PubMed Google Scholar] is the best-known betaine compound, but a wealth of information is currently accumulating on the metabolic roles of several other trimethylated compounds, including 5-AVAB, pipecolic acid betaine [2.Vuong H.E. et al.The maternal microbiome modulates fetal neurodevelopment in mice.Nature. 2020; 586: 281-286Crossref PubMed Scopus (88) Google Scholar], proline betaine [3.Cheng F. et al.A review of pharmacological and pharmacokinetic properties of stachydrine.Pharmacol. Res. 2020; 155104755Crossref PubMed Scopus (16) Google Scholar], ergothioneine [4.Borodina I. et al.The biology of ergothioneine, an antioxidant nutraceutical.Nutr. Res. Rev. 2020; 33: 190-217Crossref PubMed Scopus (53) Google Scholar], and trimethylamine N-oxide (TMAO) [5.Janeiro M.H. et al.Implication of trimethylamine N-oxide (TMAO) in disease: potential biomarker or new therapeutic target.Nutrients. 2018; 10: 1398Crossref Scopus (202) Google Scholar]. Several studies have reported positive associations between the gut microbiota and the abundance of 5-AVAB [2.Vuong H.E. et al.The maternal microbiome modulates fetal neurodevelopment in mice.Nature. 2020; 586: 281-286Crossref PubMed Scopus (88) Google Scholar,6.Koistinen V.M. et al.Contribution of gut microbiota to metabolism of dietary glycine betaine in mice and in vitro colonic fermentation.Microbiome. 2019; 7 (103–102)Crossref PubMed Scopus (27) Google Scholar,7.Zhao M. et al.TMAVA, a metabolite of intestinal microbes, is increased in plasma from patients with liver steatosis, inhibits γ-butyrobetaine hydroxylase, and exacerbates fatty liver in mice.Gastroenterology. 2020; 158: 2266-2281Abstract Full Text Full Text PDF PubMed Scopus (32) Google Scholar], and there is convincing evidence that this metabolite is at least partially synthesized by gut microbes. However, 5-AVAB has also been reported to be present in certain food sources, including milk and meat [8.Salzano A. et al.Effect of breeding techniques and prolonged post dry aging maturation process on biomolecule levels in raw buffalo meat.Vet. Sci. 2021; 8: 66Crossref PubMed Scopus (1) Google Scholar,9.Servillo L. et al.Ruminant meat and milk contain δ-valerobetaine, another precursor of trimethylamine N-oxide (TMAO) like γ-butyrobetaine.Food Chem. 2018; 260: 193-199Crossref PubMed Scopus (32) Google Scholar] and the kinds of marine algae used as foodstuffs [10.Blunden G. et al.Betaines and tertiary sulphonium compounds from 62 species of marine algae.Biochem. Syst. Ecol. 1992; 20: 373-388Crossref Google Scholar]. During recent years, 5-AVAB has been linked with whole-grain consumption [11.Kärkkäinen O. et al.Diets rich in whole grains increase betainized compounds associated with glucose metabolism.Am. J. Clin. Nutr. 2018; 108: 971-979Crossref PubMed Scopus (28) Google Scholar], demonstrated to have a role in fetal tissues [12.Pessa-Morikawa T. et al.Maternal microbiota-derived metabolic profile in fetal murine intestine, brain and placenta.BMC Microbiol. 2022; 22: 46Crossref PubMed Scopus (1) Google Scholar], and claimed to be a vital part of brain development [2.Vuong H.E. et al.The maternal microbiome modulates fetal neurodevelopment in mice.Nature. 2020; 586: 281-286Crossref PubMed Scopus (88) Google Scholar]. Additionally, liver health status has been connected to 5-AVAB [7.Zhao M. et al.TMAVA, a metabolite of intestinal microbes, is increased in plasma from patients with liver steatosis, inhibits γ-butyrobetaine hydroxylase, and exacerbates fatty liver in mice.Gastroenterology. 2020; 158: 2266-2281Abstract Full Text Full Text PDF PubMed Scopus (32) Google Scholar]. Here, we review the origin of 5-AVAB from different sources, before discussing the multiple roles that 5-AVAB has been suggested to have in the body and tissues. The importance of gut flora in synthesizing 5-AVAB has been reported recently in several publications, and the key role of the gut microbiota has been demonstrated in studies with germ-free (GF) mice, where 5-AVAB levels were significantly reduced in plasma and tissue samples as compared with conventionally housed mice [2.Vuong H.E. et al.The maternal microbiome modulates fetal neurodevelopment in mice.Nature. 2020; 586: 281-286Crossref PubMed Scopus (88) Google Scholar,6.Koistinen V.M. et al.Contribution of gut microbiota to metabolism of dietary glycine betaine in mice and in vitro colonic fermentation.Microbiome. 2019; 7 (103–102)Crossref PubMed Scopus (27) Google Scholar,7.Zhao M. et al.TMAVA, a metabolite of intestinal microbes, is increased in plasma from patients with liver steatosis, inhibits γ-butyrobetaine hydroxylase, and exacerbates fatty liver in mice.Gastroenterology. 2020; 158: 2266-2281Abstract Full Text Full Text PDF PubMed Scopus (32) Google Scholar,12.Pessa-Morikawa T. et al.Maternal microbiota-derived metabolic profile in fetal murine intestine, brain and placenta.BMC Microbiol. 2022; 22: 46Crossref PubMed Scopus (1) Google Scholar,13.Cheema M.U. Pluznick J.L. Gut microbiota plays a central role to modulate the plasma and fecal metabolomes in response to angiotensin II.Hypertension. 2019; 74: 184-193Crossref PubMed Scopus (31) Google Scholar]. Furthermore, the absence of maternal microbiota decreased 5-AVAB levels in fetal tissues, indicating that decreased synthesis of 5-AVAB is one outcome evoked by the absence of a normal microbiome in the gut [2.Vuong H.E. et al.The maternal microbiome modulates fetal neurodevelopment in mice.Nature. 2020; 586: 281-286Crossref PubMed Scopus (88) Google Scholar,12.Pessa-Morikawa T. et al.Maternal microbiota-derived metabolic profile in fetal murine intestine, brain and placenta.BMC Microbiol. 2022; 22: 46Crossref PubMed Scopus (1) Google Scholar]. Notably, most of the listed publications tend to utilize several nomenclatures for 5-AVAB (Box 1), thus hindering the efficient utilization of the existing information.Box 1Uniform naming is essential for 5-AVAB5-AVAB has been analyzed and reported during recent years concomitantly in several laboratories, resulting in extensive heterogeneity in its nomenclature. So far, it has been annotated at least as 5-AVAB [45.Kärkkäinen O. et al.Whole grain intake associated molecule 5-aminovaleric acid betaine decreases β-oxidation of fatty acids in mouse cardiomyocytes.Sci. Rep. 2018; 8: 13036-13037Crossref PubMed Scopus (16) Google Scholar], aminovaleric acid betaine [92.Wishart D.S. et al.HMDB 4.0: the human metabolome database for 2018.Nucleic Acids Res. 2018; 46: D608-D617Crossref PubMed Scopus (1758) Google Scholar], δ-valerobetaine (δVB) [44.Servillo L. et al.Carnitine precursors and short-chain acylcarnitines in water buffalo milk.J. Agric. Food Chem. 2018; 66: 8142-8149Crossref PubMed Scopus (14) Google Scholar], δ-aminovaleric acid betaine (δ-AVAB) [49.Blunden G. et al.The characterisation and quantitative estimation of betaines in commercial seaweed extracts.Bot. Mar. 1986; 29: 155Crossref Google Scholar], 5-trimethylammoniopentanoate [35.Hoppel C.L. et al.N6-Trimethyl-lysine metabolism. 3-Hydroxy-N6-trimethyl-lysine and carnitine biosynthesis.Biochem. J. 1980; 188: 509-519Crossref PubMed Scopus (21) Google Scholar], N-trimethyl 5-aminovalerate [80.Haukka J.K. et al.Metabolomic profile predicts development of microalbuminuria in individuals with type 1 diabetes.Sci. Rep. 2018; 8 (13853–10)Crossref Scopus (25) Google Scholar], trimethyl-N-aminovalerate [17.Pallister T. et al.Metabolites of milk intake: a metabolomic approach in UK twins with findings replicated in two European cohorts.Eur. J. Nutr. 2016; 56: 2379-2391Crossref PubMed Scopus (14) Google Scholar], 5-trimethylaminovalerate (GBB-5) [93.Rydzik A.M. et al.Comparison of the substrate selectivity and biochemical properties of human and bacterial γ-butyrobetaine hydroxylase.Org. Biomol. Chem. 2014; 12: 6354-6358Crossref PubMed Google Scholar], N,N,N-trimethyl-5-aminovaleric acid (TMAVA) [7.Zhao M. et al.TMAVA, a metabolite of intestinal microbes, is increased in plasma from patients with liver steatosis, inhibits γ-butyrobetaine hydroxylase, and exacerbates fatty liver in mice.Gastroenterology. 2020; 158: 2266-2281Abstract Full Text Full Text PDF PubMed Scopus (32) Google Scholar], N,N,N-trimethyl-5-aminovalerate (TMAV) [2.Vuong H.E. et al.The maternal microbiome modulates fetal neurodevelopment in mice.Nature. 2020; 586: 281-286Crossref PubMed Scopus (88) Google Scholar], δ-N-trimethylaminovaleric acid [30.Rebouche C.J. et al.є-N-Trimethyllysine availability regulates the rate of carnitine biosynthesis in the growing rat.J. Nutr. 1986; 116: 751-759Crossref PubMed Scopus (48) Google Scholar], 5-N-trimethylaminopentanoate [34.Cox R.A. Hoppel C.L. Biosynthesis of carnitine and 4-N-trimethylaminobutyrate from 6-N-trimethyl-lysine.136. 1973; : 1083-1090Google Scholar], and X-21365 (on the metabolon platform) [64.Adam J. et al.Metformin effect on nontargeted metabolite profiles in patients with type 2 diabetes and in multiple murine tissues.Diabetes (New York, N.Y.). 2016; 65: 3776-3785Google Scholar] (see Table 1 in main text).We propose the uniform name of 5-AVAB and the systematic use of Chemical Abstract Service (CAS) (6778-33-2) and PubChem ID (14274897) in the future to efficiently find all the related articles and authors. This name describes the molecular features comprehensively and the abbreviation gives a short but unique description of the molecule, as the term betaine in the nomenclature is commonly used for compounds with a quaternary ammonium where hydrogens have been substituted with methyl groups such as glycine betaine, proline betaine, and pipecolic acid betaine. Ideally, the nomenclature should not be too complex, while still describing the compound accurately. Furthermore, the IUPAC International Chemical Identifier (InChI) InChI=1S/C8H17NO2/c1-9(2,3)7-5-4-6-8(10)11/h4-7H2,1-3H3 and InChI Key CDLVFVFTRQPQFU-UHFFFAOYSA-N should be used to increase knowledge. The scientific community should avoid using multiple names for a single molecule because it complicates database and publication searches. Additionally, the Greek alphabet should also be avoided in names and abbreviations for two reasons. First, Greek letters can be written with one character or with Latin letters, for example, δVB or delta-valerobetaine, respectively. Second, the encoding used in a lot of software cannot accommodate Greek characters, meaning that this information could be lost during data processing. For the aforementioned reasons, 5-AVAB is the preferred nomenclature for the compound in question. 5-AVAB has been analyzed and reported during recent years concomitantly in several laboratories, resulting in extensive heterogeneity in its nomenclature. So far, it has been annotated at least as 5-AVAB [45.Kärkkäinen O. et al.Whole grain intake associated molecule 5-aminovaleric acid betaine decreases β-oxidation of fatty acids in mouse cardiomyocytes.Sci. Rep. 2018; 8: 13036-13037Crossref PubMed Scopus (16) Google Scholar], aminovaleric acid betaine [92.Wishart D.S. et al.HMDB 4.0: the human metabolome database for 2018.Nucleic Acids Res. 2018; 46: D608-D617Crossref PubMed Scopus (1758) Google Scholar], δ-valerobetaine (δVB) [44.Servillo L. et al.Carnitine precursors and short-chain acylcarnitines in water buffalo milk.J. Agric. Food Chem. 2018; 66: 8142-8149Crossref PubMed Scopus (14) Google Scholar], δ-aminovaleric acid betaine (δ-AVAB) [49.Blunden G. et al.The characterisation and quantitative estimation of betaines in commercial seaweed extracts.Bot. Mar. 1986; 29: 155Crossref Google Scholar], 5-trimethylammoniopentanoate [35.Hoppel C.L. et al.N6-Trimethyl-lysine metabolism. 3-Hydroxy-N6-trimethyl-lysine and carnitine biosynthesis.Biochem. J. 1980; 188: 509-519Crossref PubMed Scopus (21) Google Scholar], N-trimethyl 5-aminovalerate [80.Haukka J.K. et al.Metabolomic profile predicts development of microalbuminuria in individuals with type 1 diabetes.Sci. Rep. 2018; 8 (13853–10)Crossref Scopus (25) Google Scholar], trimethyl-N-aminovalerate [17.Pallister T. et al.Metabolites of milk intake: a metabolomic approach in UK twins with findings replicated in two European cohorts.Eur. J. Nutr. 2016; 56: 2379-2391Crossref PubMed Scopus (14) Google Scholar], 5-trimethylaminovalerate (GBB-5) [93.Rydzik A.M. et al.Comparison of the substrate selectivity and biochemical properties of human and bacterial γ-butyrobetaine hydroxylase.Org. Biomol. Chem. 2014; 12: 6354-6358Crossref PubMed Google Scholar], N,N,N-trimethyl-5-aminovaleric acid (TMAVA) [7.Zhao M. et al.TMAVA, a metabolite of intestinal microbes, is increased in plasma from patients with liver steatosis, inhibits γ-butyrobetaine hydroxylase, and exacerbates fatty liver in mice.Gastroenterology. 2020; 158: 2266-2281Abstract Full Text Full Text PDF PubMed Scopus (32) Google Scholar], N,N,N-trimethyl-5-aminovalerate (TMAV) [2.Vuong H.E. et al.The maternal microbiome modulates fetal neurodevelopment in mice.Nature. 2020; 586: 281-286Crossref PubMed Scopus (88) Google Scholar], δ-N-trimethylaminovaleric acid [30.Rebouche C.J. et al.є-N-Trimethyllysine availability regulates the rate of carnitine biosynthesis in the growing rat.J. Nutr. 1986; 116: 751-759Crossref PubMed Scopus (48) Google Scholar], 5-N-trimethylaminopentanoate [34.Cox R.A. Hoppel C.L. Biosynthesis of carnitine and 4-N-trimethylaminobutyrate from 6-N-trimethyl-lysine.136. 1973; : 1083-1090Google Scholar], and X-21365 (on the metabolon platform) [64.Adam J. et al.Metformin effect on nontargeted metabolite profiles in patients with type 2 diabetes and in multiple murine tissues.Diabetes (New York, N.Y.). 2016; 65: 3776-3785Google Scholar] (see Table 1 in main text). We propose the uniform name of 5-AVAB and the systematic use of Chemical Abstract Service (CAS) (6778-33-2) and PubChem ID (14274897) in the future to efficiently find all the related articles and authors. This name describes the molecular features comprehensively and the abbreviation gives a short but unique description of the molecule, as the term betaine in the nomenclature is commonly used for compounds with a quaternary ammonium where hydrogens have been substituted with methyl groups such as glycine betaine, proline betaine, and pipecolic acid betaine. Ideally, the nomenclature should not be too complex, while still describing the compound accurately. Furthermore, the IUPAC International Chemical Identifier (InChI) InChI=1S/C8H17NO2/c1-9(2,3)7-5-4-6-8(10)11/h4-7H2,1-3H3 and InChI Key CDLVFVFTRQPQFU-UHFFFAOYSA-N should be used to increase knowledge. The scientific community should avoid using multiple names for a single molecule because it complicates database and publication searches. Additionally, the Greek alphabet should also be avoided in names and abbreviations for two reasons. First, Greek letters can be written with one character or with Latin letters, for example, δVB or delta-valerobetaine, respectively. Second, the encoding used in a lot of software cannot accommodate Greek characters, meaning that this information could be lost during data processing. For the aforementioned reasons, 5-AVAB is the preferred nomenclature for the compound in question. Bacterial species in the gut probably have similar properties to halophilic microorganisms, which can synthesize a significant amount of glycine betaine from glycine by adding methyl groups, for example, from a methionine source. These halophilic microorganisms, namely, Actinopolyspora halophila and Ectothiorhodospira halochloris, utilize a three-step catalytic procedure with two methyltransferase enzymes, glycine sarcosine methyltransferase and sarcosine dimethylglycine methyltransferase [14.Nyyssölä A. et al.Extreme halophiles synthesize betaine from glycine by methylation.J. Biol. Chem. 2000; 275: 22196-22201Abstract Full Text Full Text PDF PubMed Scopus (138) Google Scholar,15.Nyyssölä A. et al.Characterization of glycine sarcosine N-methyltransferase and sarcosine dimethylglycine N-methyltransferase.Appl. Environ. Microbiol. 2001; 67: 2044-2050Crossref PubMed Scopus (36) Google Scholar]. However, glycine betaine can also act as a methyl donor, similar to the transformation of homocysteine into methionine [1.Craig S.A. Betaine in human nutrition.Am. J. Clin. Nutr. 2004; 80: 539-549Crossref PubMed Google Scholar,16.Ueland P.M. et al.Betaine: a key modulator of one-carbon metabolism and homocysteine status.Clin. Chem. Lab. Med. 2005; 43: 1069-1075Crossref PubMed Scopus (166) Google Scholar]. Pallister et al. [17.Pallister T. et al.Metabolites of milk intake: a metabolomic approach in UK twins with findings replicated in two European cohorts.Eur. J. Nutr. 2016; 56: 2379-2391Crossref PubMed Scopus (14) Google Scholar] suggested that 5-AVAB was likely to be a methylated product of 5-aminovaleric acid (5-AVA). In addition, we propose that glycine betaine can act as a methyl substrate donor in biosynthesis pathways either in microorganisms or endogenously, where 5-AVAB is formed from 5-AVA (Figure 1A ) [6.Koistinen V.M. et al.Contribution of gut microbiota to metabolism of dietary glycine betaine in mice and in vitro colonic fermentation.Microbiome. 2019; 7 (103–102)Crossref PubMed Scopus (27) Google Scholar]. However, this pathway still needs to be fully characterized, for instance, with an in vitro fermentation model based on human gut microbiota and isotope-labeled glycine betaine or other methyl donors. Although there have been rather few mechanistic studies demonstrating in vitro production of 5-AVAB by certain microbial species, the normal microbiome contains many important bacteria, abundances of which have been correlated with 5-AVAB syntheses, such as bifidobacteria and Coriobacteriaceae [6.Koistinen V.M. et al.Contribution of gut microbiota to metabolism of dietary glycine betaine in mice and in vitro colonic fermentation.Microbiome. 2019; 7 (103–102)Crossref PubMed Scopus (27) Google Scholar]. Bifidobacteria are known to be beneficial bacteria for human health [18.Mitsuoka T. Bifidobacteria and their role in human health.J. Ind. Microbiol. 1990; 6: 263-267Crossref Scopus (301) Google Scholar], and Coriobacteriaceae can confer protection against liver pathologies [19.Clavel T. et al.Intestinal microbiota in metabolic diseases.Gut Microbes. 2014; 5: 544-551Crossref PubMed Scopus (109) Google Scholar]. Further evidence supporting the role of microbiota in the production of 5-AVAB from dietary precursors has been obtained from studies demonstrating that even though 5-AVAB is absent in cereal-based foods [20.Koistinen V.M. et al.Metabolic profiling of sourdough fermented wheat and rye bread.Sci. Rep. 2018; 8 (5684–5611)Crossref Scopus (43) Google Scholar], the addition of cereal bran or whole grains to the diet has been associated with increased circulating and tissue levels of 5-AVAB [6.Koistinen V.M. et al.Contribution of gut microbiota to metabolism of dietary glycine betaine in mice and in vitro colonic fermentation.Microbiome. 2019; 7 (103–102)Crossref PubMed Scopus (27) Google Scholar,11.Kärkkäinen O. et al.Diets rich in whole grains increase betainized compounds associated with glucose metabolism.Am. J. Clin. Nutr. 2018; 108: 971-979Crossref PubMed Scopus (28) Google Scholar]. In the absence of the appropriate gut flora, then there are no microbes able to produce 5-AVAB, even though there could be starting materials present in the gut. Thus, the observed 5-AVAB traces detected in GF mice [6.Koistinen V.M. et al.Contribution of gut microbiota to metabolism of dietary glycine betaine in mice and in vitro colonic fermentation.Microbiome. 2019; 7 (103–102)Crossref PubMed Scopus (27) Google Scholar] have been directly obtained from nutrition or potentially produced via endogenous metabolism. One of the most likely precursors en route to 5-AVAB biosynthesis is 5-AVA, which can also be synthesized by the gut microbiome. There is good evidence that bacteria belonging to the genus Pseudomonas can produce 5-AVA using l-lysine as the starting material [21.Fothergill J.C. Guest J.R. Catabolism of L-lysine by Pseudomonas aeruginosa.J. Gen. 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Proline is reduced directly to 5-AVA by proline reductase (EC 1.21.4.1) as part of the Stickland reaction [25.Bouillaut L. et al.Proline-dependent regulation of Clostridium difficile Stickland metabolism.J. Bacteriol. 2013; 195: 844-854Crossref PubMed Scopus (83) Google Scholar]. Additionally, arginine and ornithine treatment is demonstrated to elevate 5-AVA concentration in Clostridia bacteria [26.Wildenauer F.X. Winter J. Fermentation of isoleucine and arginine by pure and syntrophic cultures of Clostridium sporogenes.FEMS Microbiol. Lett. 1986; 38: 373-379Crossref Google Scholar,27.Costilow R.N. Laycock L. Proline as an intermediate in the reductive deamination of ornithine to δ-aminovaleric acid.J. Bacteriol. 1968; 96: 1011-1020Crossref PubMed Google Scholar]. 5-AVA biosynthesis from arginine utilizes a Stickland reaction, ornithine cyclodeaminase (EC 4.3.1.12), and proline reductase in Clostridia bacteria with proline and ornithine as intermediates (Figure 1C,D) [27.Costilow R.N. Laycock L. Proline as an intermediate in the reductive deamination of ornithine to δ-aminovaleric acid.J. Bacteriol. 1968; 96: 1011-1020Crossref PubMed Google Scholar, 28.Costilow R.N. Laycock L. Ornithine cyclase (deaminating).J. Biol. Chem. 1971; 246: 6655-6660Abstract Full Text PDF PubMed Google Scholar, 29.Muth W.L. Costilow R.N. Ornithine cyclase (deaminating).J. Biol. Chem. 1974; 249: 7463-7467Abstract Full Text PDF PubMed Google Scholar]. 5-AVA can be further converted to 5-AVAB [2.Vuong H.E. et al.The maternal microbiome modulates fetal neurodevelopment in mice.Nature. 2020; 586: 281-286Crossref PubMed Scopus (88) Google Scholar], potentially by glycine sarcosine methyltransferase and sarcosine dimethylglycine methyltransferase (Figure 1A). Furthermore, 5-AVAB can be biosynthesized directly without the need for a 5-AVA intermediate from trimethyllysine by Enterococcus faecalis utilizing the DavB and DavA enzyme pathway (Figure 1E) [7.Zhao M. et al.TMAVA, a metabolite of intestinal microbes, is increased in plasma from patients with liver steatosis, inhibits γ-butyrobetaine hydroxylase, and exacerbates fatty liver in mice.Gastroenterology. 2020; 158: 2266-2281Abstract Full Text Full Text PDF PubMed Scopus (32) Google Scholar], similarly as demonstrated for lysine (Figure 1B). In addition, trimethyllysine can be directly converted to 5-AVAB by a Stickland reaction (Figure 1F), as demonstrated by Servillo et al. [9.Servillo L. et al.Ruminant meat and milk contain δ-valerobetaine, another precursor of trimethylamine N-oxide (TMAO) like γ-butyrobetaine.Food Chem. 2018; 260: 193-199Crossref PubMed Scopus (32) Google Scholar]. They showed that the bovine ruminal microbiome forms 5-AVAB after just 2 h of incubation and, according to their hypothesis, there is an oxidation–reduction reaction, which can catalyze 5-AVAB production. Moreover, Rebouche et al. [30.Rebouche C.J. et al.є-N-Trimethyllysine availability regulates the rate of carnitine biosynthesis in the growing rat.J. Nutr. 1986; 116: 751-759Crossref PubMed Scopus (48) Google Scholar] reported an increased amount of methyl-3H-labeled 5-AVAB in rat feces after the animals were fed with methyl-3H-labeled trimethyllysine, thus demonstrating 5-AVAB biosynthesis in microbiota per se. However, the chemical transformation and microbiome species behind 5-AVAB synthesis need to be characterized more thoroughly to identify possible similarities in the 5-AVAB biosynthetic capacity between other species and the human gut microbiome. Although the origin of 5-AVAB is not fully clear, the biosynthesis of other betaines, for example, glycine betaine, has been better characterized. For instance, there are at least two well-characterized glycine betaine synthetic pathways in the body that could be extended to 5-AVAB production. The first pathway produces glycine betaine by utilizing two methyltransferase enzymes, and this may also be possible in the biosynthesis of 5-AVAB (Figure 1A). The second glycine betaine biosynthesis pathway that is typically encountered in bacteria also takes place endogenously in the inner membrane of mitochondria. This pathway utilizes choline as its starting material and produces glycine betaine by oxidation [31.Boch J. et al.Synthesis of the osmoprotectant glycine betaine in Bacillus subtilis: characterization of the gbsAB genes.J. Bacteriol. 1996; 178: 5121-5129Crossref PubMed Scopus (111) Google Scholar, 32.Pocard J.-A. et al.Molecular characterization of the bet genes encoding glycine betaine synthesis in Sinorhizobium meliloti 102F34.Microbiology (Soc. Gen. Microbiol.). 1997; 143: 1369-1379Google Scholar, 33.Salvi F. Gadda G. Human choline dehydrogenase: medical promises and biochemical challenges.Arch. Biochem. Biophys. 2013; 537: 243-252Crossref PubMed Scopus (27) Google Scholar]. Similarly, 5-hydroxypentyltrimethylammonium could be oxidized to produce 5-AVAB, although this pathway has not been examined in