Short-chain fatty acids: definition, synthesis and function

Short-chain fatty acids or SCFAs are saturated fatty acids with a straight or branched carbon-chain made of 2-5 carbon atoms, and are acetic acid, propionic acid, butyric acid, isobutyric acid, valeric acid, isovaleric acid, and 2-methylbutyric acid.^[1]
In humans, they are, along with secondary bile salts, the main metabolites produced by bacteria of the gut microbiota in the cecum and colon, and derive almost entirely from the anaerobic fermentation of non-digestible carbohydrates.^[11] The most abundant are acetic acid, propionic acid and butyric acid, which represent 90-95 percent of the produced SCFAs.^[6] The remaining percentage is made of the branched SCFAs.
They are the major anions present in the colon. Their concentration is higher in the cecum and in the proximal colon than in the distal part, where the substrates for their synthesis are depleted.^[1][2][4] They are able to reduce colonic pH value and thus acidify the stool.
About 90-95 percent of the SCFAs are absorbed in the cecum and colon, whereas 5-10 percent are excreted with the feces.^[13]
They are thought to provide about 70 percent of the energy needs of colonocytes.^[4]
Short-chain fatty acids are able to modulate the physiology and composition of the gut microbiota.^[7] Furthermore, a growing body of research suggests that they play a important role in maintaining human health.^[4]

Contents

• Sources
• Properties
• Health effects
• Synthesis
  • Synthesis of acetic acid
  • Synthesis of propionic acid
  • Synthesis of butyric acid
  • Synthesis from amino acids
  • Endogenous synthesis
• Membrane receptors
• Absorption
• Role in colonocytes
• Transport
• Hepatic and extrahepatic metabolism
• References

Sources

Like medium-chain fatty acids and long-chain fatty acids, short-chain fatty acids are present in animal and plant tissues mostly in the form of triglycerides, although in much lower amounts than long-chain ones.
In adults, the main food source is milk and dairy products, where butyric acid is the SCFA present with the highest concentration. Other sources are some vegetable oils, such as palm kernel oil and coconut oil.
In breastfed infants, the main source is breast milk.^[10]
However, for humans, and most mammals, the most important source is the anaerobic fermentation of fibers and resistant starch, namely, indigestible carbohydrates, by the bacteria of the gut microbiota.^[6] Approximately 500-600 mM of SCFAs are produced through this pathway per day. Acetic, propionic and butyric acids are present in a molar ratio of about 60:20:20, respectively, although the relative proportion of each depends on the microbiota composition, the substrate, and the intestinal transit time.^[7][11]

Properties

Short-chain fatty acids have carbon chains made of 2-5 carbon atoms, a characteristic that strongly affects the physical properties.^[9]
Acetic acid, propionic acid, butyric acid and valeric acid are straight-chain fatty acids, whereas isobutyric acid, isovaleric acid and 2-methylbutyric acid are branched-chain fatty acids.

They are small molecules, and are the smallest among all lipids.
They are liquid at room temperature, and are soluble in polar solvents such as water, unlike saturated fatty acids with longer carbon chains, whose solubility in polar solvents, considering those with straight chain, decreases as the length of the chain increases, as the hydrophobic part of the molecule is the carbon chain, whereas the carboxyl group is polar.^[10]
Finally, it should also be noted that butyric acid and isobutyric acid, which have the molecular formula C₄H₈O₂, are an example of chain isomerism, as well as valeric acid, isovaleric acid and 2-methylbutyric acid, which have the formula molecular C₅H₁₀O₂.

Health effects

Short-chain fatty acids appear to play a crucial role in maintaining human health.^[4] Their activity seems to occur through direct and/or indirect effects on cellular processes such as proliferation, differentiation and gene expression, thus contributing to the regulation of processes such as glucose homeostasis, intestinal and immune function, and the regulation of the gut-brain axis.^[6] Their health effects seem to be confirmed also by studies showing that intestinal dysbiosis appears to be implicated in metabolic pathologies, such as disorders involving glucose homeostasis, and behavioral and neurological pathologies, such as depression, and Alzheimer’s and Parkinson’s.^[11]

Synthesis

In humans, the enzyme equipment carrying out carbohydrate digestion lacks the enzymes capable of digesting fiber and resistant starch, the latter so called precisely because it resists the action of alpha-amylase. On the contrary, the bacteria of the gut microbiota code for a large number of different glycoside hydrolases, more than 260, which also hydrolyze fibers and resistant starch, releasing the constituent monosaccharides.^[7] Hexoses and deoxyhexoses enter glycolysis, and pentoses enter the pentose phosphate pathway, to give pyruvate, the conjugate base of pyruvic acid, which is the main precursor for the synthesis of short-chain fatty acids.^[2][4][5]
The synthesis of SCFAs is affected by several factors; below are some examples.^[4][11][13]

• The fiber content of the diet. For example, a diet rich in fibers, such as the Mediterranean diet may influence their synthesis.
• The composition of the gut microbiota.
• The pH of the intestinal lumen, as the bacteria that produce butyric acid dominate at pH value around 5.5, while the bacteria that produce acetic and propionic acids dominate at pH value around 6.5.
• The gut transit time.
• The amount of oxygen in the intestinal lumen.

Acetic acid and propionic acid are mainly produced by species of the phylum Bacteroides, while butyric acid, for whose synthesis resistant starch is particularly important, by species of the phylum Firmicutes.^[6]

Synthesis of acetic acid

Acetic acid, the most abundant SCFA in the colon, can be synthesized via the Wood-Ljungdahl pathway in the reductive direction, through the reduction of CO₂ to acetate, or from acetyl-CoA, the most important metabolic pathway, responsible of the production of about two thirds of butyric acid present in the intestinal lumen.^[4]

Synthesis of propionic acid

Propionic acid can be synthesized through three different metabolic pathways: the acrylate and succinate pathways, which use lactic acid produced by other bacteria, and the propanediol pathway, in which the precursors are deoxyhexoses.^[2][3][10]
The acrylate pathway converts lactic acid to propionyl-CoA, via lactoyl-CoA. In the final step, propionyl-CoA is hydrolyzed to propionic acid.
In the succinate pathway, lactate is reduced to pyruvate, which is carboxylated to oxaloacetate, which, through a pathway that has malate, fumarate, succinate, and methylmalonyl-CoA as intermediates, is converted to propionyl-CoA, which in turn is hydrolyzed to propionic acid. The succinate pathway is thought to be the dominant pathway for propionic acid synthesis in the gut.
In the propanediol pathway, some deoxyhexoses, such as fucose and rhamnose, are converted via 1,2-propanediol to propionyl-CoA and then propionic acid.

Synthesis of butyric acid

The synthesis of butyric acid can follow two routes.^[7][10]
In most butyric acid-producing bacteria, the short-chain fatty acid is synthesized through a pathway that begins with the condensation of two acetyl-CoA to acetoacetyl-CoA, which, through a pathway that has beta-hydroxybutyryl-CoA and crotonyl-CoA as intermediates, is converted to butyryl-CoA. The final step is the release of butyric acid from butyryl-CoA.^[8]
In a small number of bacterial species, butyryl-CoA is converted to butyryl phosphate, from which butyric acid is released.^[2]

Synthesis from amino acids

Acetic acid, propionic acid and butyric acid can also be produced from amino acids obtained from peptide and protein degradation, although the amount produced by these pathways is small.^[7]
These synthesis occur in the distal part of the colon, often by non-commensal bacteria, as in the case of glutamate and lysine fermentation.^[3] Different short-chain fatty acids are produced by the metabolism of different amino acids; below are some examples.^[4]

• Glutamic acid mainly produces acetic acid and butyric acid.
• Aspartic acid mainly produces acetic acid and propionic acid.
• The basic amino acids lysine, arginine and histidine produce acetic acid and butyric acid.
• Cysteine produces acetic, propionic and butyric acids.
• Methionine mainly produce propionic acid and butyric acid.
• Branched-chain fatty acids derive from the branched-chain amino acids leucine, isoleucine and valine.

The pH of the intestinal lumen influences the metabolism of proteins by the gut microbiota; for example, their breakdown into amino acids is more likely at neutral or weakly alkaline pH.
It should be noted that potentially toxic compounds, such as ammonia, sulphites and phenols, are also produced from the intestinal metabolism of amino acids.

Endogenous synthesis

Mammals, and therefore humans, have the enzymatic equipment for the endogenous synthesis of short-chain fatty acids. The synthesis occurs mainly in the liver, by beta-oxidation cycles which lead to the formation of acyl-CoA with a shorter carbon chain than the starting fatty acid. The acil-CoA is then hydrolyzed to fatty acid and CoA by an acyl-CoA thioesterases (EC 3.1.2.20).^[10][12]

Membrane receptors

Short-chain fatty acids can bind to specific receptors on the plasma membrane, the G protein-coupled receptors, including GPR41, GPR43 and GPR109A.^[6]
The effects produced by the binding of the SCFAs on the receptors depend on the cell type. For example, binding to receptors on intestinal L cells is associated with the release of glucagon-like peptide-1, or GLP-1, and peptide YY, hormones that affect appetite and food intake. The binding to enterochromaffin cells induces the release of serotonin, which may affect intestinal motility. Finally, binding to receptors on pancreatic beta-cells increases insulin release.^[11]
The different short-chain fatty acids have different ability to activate the receptors: GPR43 is more likely to be activated by acetic and propionic acids, GPR41 by propionic and butyric acids, while GPR109A by butyric acid.^[4]

Absorption

About 90 percent of the short-chain fatty acids present in the intestinal lumen are absorbed by colonocytes. The passage across the plasma membrane can occur by passive diffusion or by active transport mediated by two types of membrane transporters: the H⁺-dependent and Na⁺-dependent monocarboxylate transporters, or MCTs and SMCTs, respectively.^[6][7]
Passive transport affects protonated forms of SCFAs, so it is influenced by colonic pH value. A weak acidification of the intestinal lumen, which can be due to the metabolic activity of the microorganisms, increases the prevalence of the protonated form and therefore of passive transport.^[10]

Role in colonocytes

In colonocytes, short-chain fatty acids have energy and regulatory function.
When used for energy purposes, acetic acid and butyric acid are converted to acetyl-CoA, and propionic acid to propionyl-CoA. Through the production of ATP, SCFAs contribute to the maintenance of cellular homeostasis, but also, for example, to the maintenance of the integrity of the tight junctions at the cell apex, and therefore of the integrity of the intestinal barrier.^[11] Of the three major short-chain fatty acids produced by gut microbiota, butyric acid is the major source of energy for colonocytes, while acetic and propionic acids are poorly metabolized and mostly drained by the portal vein.^[2][13]
Considering the regulatory role, SCFAs are, for example, capable of inhibiting histone deacetylases (EC 3.5.1.98), enzymes that catalyze the removal of acetyl groups from lysine residues of histone proteins, acetyl groups previously inserted by histone acetyltransferase (EC 2.3.1.48).^[7] The R groups of deacetylated lysines have positive charges, which allows histone proteins to wrap the DNA more tightly. This makes the nucleosome more compact, and consequently more difficult to carry out transcription and gene expression. The different short-chain fatty acids have different abilities to inhibit histone deacetylases:

• • up to 80 percent for butyric acid;
  • up to 60 percent for propionic acid;
  • acetic acid has the lowest inhibitory rate.^[4]

This mode of action on histone deacetylases has been observed not only in the gut and associated immune tissue, but also in the central and peripheral nervous systems.^[11]

Transport

Short-chain fatty acids that have not been utilized by colonocytes leave the cell by passive diffusion and active transport across the basolateral membrane, and enter the portal circulation where acetic acid reaches the highest concentration, about 260 mM/L, while propionic and butyric acids reach concentrations of about 30 mM/L.^[2]
In the rectum, a small amount of these lipids can pass directly into the systemic circulation, thus bypassing the liver, via the internal iliac vein.^[13]
Unlike long-chain fatty acids, short- and medium-chains fatty acids are present in the circulation in free form, namely, as non-esterified fatty acids, and, bound to albumin, reach the liver. Subsequent cell uptake and intracellular transport do not require fatty acid transport proteins, plasma membrane fatty acid translocases, or cytosolic fatty acid binding proteins. Therefore, their oxidation may be much faster than that of long-chain fatty acids and the longest members of medium-chain fatty acids, namely, fatty acid with carbon chains longer than 8 carbon atoms.^[10]

Hepatic and extrahepatic metabolism

The liver is an important site for short-chain fatty acids metabolism.^[13]
It can to absorb about 40 percent of the acetic acid and 80 percent of the propionic acid from the portal vein. Propionic acid is mostly metabolized in the liver, where it can also be used as a substrate for gluconeogenesis.^[7]
A small amount of the gut derived SCFAs, about 36 percent for acetic acid, 9 percent for propionic acid and only 2 percent for butyric acid, reach, through the systemic circulation, the peripheral tissues. In muscle, acetic acid can be used for lipid synthesis or be oxidized for energy production. Furthermore, it is thought that SCFA concentrations in the systemic circulation, even if small, are capable of influencing the metabolism and physiology of peripheral cells and tissues.

References

1. ^ ^{a b} Abdul Rahim M.B.H., Chilloux J., Martinez-Gili L., Neves A.L., Myridakis A., Gooderham N., Dumas M.-E. Diet-induced metabolic changes of the human gut microbiome: importance of short-chain fatty acids, methylamines and indoles. Acta Diabetol 2019;56:493-500. doi:10.1007/s00592-019-01312-x
2. ^ ^{a b c d e} Deleu S., Machiels K., Raes J., Verbeke K., Vermeire S. Short chain fatty acids and its producing organisms: an overlooked therapy for IBD? EBioMedicine 2021;66:103293. doi:10.1016/j.ebiom.2021.103293
3. ^ ^{a b} Koh A., De Vadder F., Kovatcheva-Datchary P., Bäckhed F. From dietary fiber to host physiology: short-chain fatty acids as key bacterial metabolites. Cell 2016;165(6):1332-1345. doi:10.1016/j.cell.2016.05.041
4. ^ ^{a b c d e f g h i j} Liu X.F., Shao J.H., Liao Y.T., Wang L.N., Jia Y., Dong P.J., Liu Z.Z., He D.D., Li C., Zhang X. Regulation of short-chain fatty acids in the immune system. Front Immunol 2023;14:1186892. doi:10.3389/fimmu.2023.1186892
5. ^{^} Miller T.L., Wolin M.J. Pathways of acetate, propionate, and butyrate formation by the human fecal microbial flora. Appl Environ Microbiol 1996;62(5):1589-92. doi:10.1128/aem.62.5.1589-1592
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7. ^ ^{a b c d e f g h} Portincasa P., Bonfrate L., Vacca M., De Angelis M., Farella I., Lanza E., Khalil M.,Wang D.Q.-H., Sperandio M., Di Ciaula A. Gut microbiota and short chain fatty acids: implications in glucose homeostasis. Int J Mol Sci 2022;23:1105. doi:10.3390/ijms23031105
8. ^{^} Pryde S.E., Duncan S.H., Hold G.L., Stewart C.S., Flint H.J. The microbiology of butyrate formation in the human colon. FEMS Microbiol Lett 2002;217(2):133-9. doi:10.1111/j.1574-6968.2002.tb11467.x
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11. ^ ^{a b c d e f g} Silva Y.P., Bernardi A., Frozza R.L. The role of short-chain fatty acids from gut microbiota in gut-brain communication. Front Endocrinol (Lausanne) 2020;11:25. doi:10.3389/fendo.2020.00025
12. ^{^} Trefely S., Lovell C.D., Snyder N.W., Wellen K.E. Compartmentalised acyl-CoA metabolism and roles in chromatin regulation. Mol Metab 2020;38:100941. doi:10.1016/j.molmet.2020.01.005
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