Inbiochemistry andmetabolism,beta oxidation (also β-oxidation) is thecatabolic process by whichfatty acid molecules are broken down in thecytosol in prokaryotes and in themitochondria in eukaryotes to generateacetyl-CoA. Acetyl-CoA enters thecitric acid cycle, generatingNADH andFADH₂, which are electron carriers used in theelectron transport chain. It is named as such because thebeta carbon of the fatty acid chain undergoes oxidation and is converted to acarbonyl group to start the cycle all over again. Beta-oxidation is primarily facilitated by themitochondrial trifunctional protein, an enzyme complex associated with theinner mitochondrial membrane, althoughvery long chain fatty acids are oxidized inperoxisomes.

The overall reaction for one cycle of beta oxidation is:

C_n-acyl-CoA + FAD + NAD⁺ + H₂O + CoA → C_n-2-acyl-CoA + FADH₂ + NADH + H⁺ + acetyl-CoA

Free fatty acids cannot penetrate any biological membrane due to their negative charge. Free fatty acids must cross the cell membrane through specifictransport proteins, such as theSLC27 family fatty acid transport protein.^[1] Once in thecytosol, the following processes bring fatty acids into the mitochondrial matrix so that beta-oxidation can take place.

1. Long-chain-fatty-acid—CoA ligase catalyzes the reaction between a fatty acid withATP to give a fatty acyl adenylate, plus inorganic pyrophosphate, which then reacts with freecoenzyme A to give afatty acyl-CoA ester andAMP.
2. If the fatty acyl-CoA has a long chain, then thecarnitine shuttle must be utilized (shown in the table below):
   • Acyl-CoA is transferred to the hydroxyl group of carnitine bycarnitine palmitoyltransferase I, located on the cytosolic faces of theouter andinner mitochondrial membranes.
   • Acyl-carnitine is shuttled inside by acarnitine-acylcarnitine translocase, as a carnitine is shuttled outside.
   • Acyl-carnitine is converted back to acyl-CoA bycarnitine palmitoyltransferase II, located on the interior face of theinner mitochondrial membrane. The liberated carnitine is shuttled back to the cytosol, as an acyl-carnitine is shuttled into the matrix.
3. If the fatty acyl-CoA contains a short chain, theseshort-chain fatty acids can simply diffuse through the inner mitochondrial membrane.

Once the fatty acid is inside themitochondrial matrix, beta-oxidation occurs by cleaving two carbons every cycle to form acetyl-CoA. The process consists of 4 steps.^[2]

1. A long-chain fatty acid isdehydrogenated to create a transdouble bond between C2 and C3. This is catalyzed byacyl CoA dehydrogenase to produce trans-delta 2-enoyl CoA. It uses FAD as an electron acceptor and it is reduced to FADH₂.
2. Trans-delta 2-enoyl CoA is hydrated at the double bond to produce L-3-hydroxyacyl CoA byenoyl-CoA hydratase.
3. L-3-hydroxyacyl CoA is dehydrogenated again to create 3-ketoacyl CoA by 3-hydroxyacyl CoA dehydrogenase. This enzyme uses NAD as an electron acceptor.
4. Thiolysis occurs between C2 and C3 (alpha and beta carbons) of 3-ketoacyl CoA. Thiolase enzyme catalyzes the reaction when a new molecule of coenzyme A breaks the bond by nucleophilic attack on C3. This releases the first two carbon units, as acetyl CoA, and a fatty acyl CoA minus two carbons. The process continues until all of the carbons in the fatty acid are turned into acetyl CoA.

This acetyl-CoA then enters the mitochondrial tricarboxylic acid cycle (TCA cycle). Both the fatty acid beta-oxidation and the TCA cycle produce NADH and FADH₂, which are used by the electron transport chain to generate ATP.

Fatty acids are oxidized by most of the tissues in the body. However, some tissues such as thered blood cells of mammals (which do not contain mitochondria) and cells of thecentral nervous system do not use fatty acids for their energy requirements, but instead use carbohydrates (red blood cells and neurons) orketone bodies (neurons only).

Because many fatty acids are not fully saturated or do not have an even number of carbons, several different mechanisms have evolved, described below.

Once inside the mitochondria, each cycle of β-oxidation, liberating a two carbon unit (acetyl-CoA), occurs in a sequence of four reactions:^[3]

This process continues until the entire chain is cleaved into acetyl CoA units. The final cycle produces two separate acetyl CoAs, instead of one acyl CoA and one acetyl CoA. For every cycle, the Acyl CoA unit is shortened by two carbon atoms. Concomitantly, one molecule of FADH₂, NADH and acetyl CoA are formed.

Fatty acids with an odd number of carbons are found in the lipids of plants and some marine organisms. Many ruminant animals form a large amount of 3-carbon propionate during the fermentation of carbohydrates in the rumen.^[4] Long-chain fatty acids with an odd number of carbon atoms are found particularly in ruminant fat and milk.^[5]

Chains with an odd-number ofcarbons are oxidized in the same manner as even-numbered chains, but the final products arepropionyl-CoA and acetyl-CoA.

Propionyl-CoA is first carboxylated using abicarbonateion into a D-stereoisomer ofmethylmalonyl-CoA. This reaction involves abiotinco-factor, ATP and the enzymepropionyl-CoA carboxylase.^[6] The bicarbonate ion's carbon is added to the middle carbon of propionyl-CoA, forming a D-methylmalonyl-CoA. However, the D-conformation is enzymatically converted into the L-conformation bymethylmalonyl-CoA epimerase. It then undergoes intramolecular rearrangement, which is catalyzed bymethylmalonyl-CoA mutase (requiring B₁₂ as a coenzyme) to form succinyl-CoA. Thesuccinyl-CoA formed then enters thecitric acid cycle.

However, whereas acetyl-CoA enters the citric acid cycle by condensing with an existing molecule ofoxaloacetate, succinyl-CoA enters the cycle as a principal in its own right. Thus, the succinate just adds to the population of circulating molecules in the cycle and undergoes no net metabolization while in it. When this infusion of citric acid cycle intermediates exceedscataplerotic demand (such as foraspartate orglutamate synthesis), some of them can be extracted to thegluconeogenesis pathway, in the liver and kidneys, throughphosphoenolpyruvate carboxykinase, and converted to free glucose.^[7]

β-Oxidation of unsaturated fatty acids poses a problem since the location of a cis-bond can prevent the formation of a trans-Δ² bond which is essential for continuation of β-Oxidation as this conformation is ideal for enzyme catalysis. This is handled by additional two enzymes,Enoyl CoA isomerase and2,4 Dienoyl CoA reductase.^[8]

β-oxidation occurs normally until the acyl CoA (because of the presence of a double bond) is not an appropriate substrate foracyl CoA dehydrogenase, orenoyl CoA hydratase:

• If the acyl CoA contains acis-Δ³ bond, thencis-Δ³-Enoyl CoA isomerase will convert the bond to atrans-Δ² bond, which is a regular substrate.
• If the acyl CoA contains acis-Δ⁴ double bond, then its dehydrogenation yields a 2,4-dienoyl intermediate, which is not a substrate for enoyl CoA hydratase. However, the enzyme2,4 Dienoyl CoA reductase reduces the intermediate, using NADPH, intotrans-Δ³-enoyl CoA. This compound is converted into a suitable intermediate by 3,2-Enoyl CoA isomerase and β-Oxidation continues.

Fatty acid oxidation also occurs inperoxisomes when the fatty acid chains are too long to be processed by the mitochondria. The same enzymes are used in peroxisomes as in the mitochondrial matrix and acetyl-CoA is generated. Very long chain (greater than C-22) fatty acids, branched fatty acids,^[9] someprostaglandins andleukotrienes^[10] undergo initial oxidation in peroxisomes untiloctanoyl-CoA is formed, at which point it undergoes mitochondrial oxidation.^[11]

One significant difference is that oxidation in peroxisomes is not coupled toATP synthesis. Instead, the high-potential electrons are transferred to O₂, which yieldshydrogen peroxide. The enzymecatalase, found primarily in peroxisomes and thecytosol oferythrocytes (and sometimes inmitochondria^[12]), converts thehydrogen peroxide intowater andoxygen.

Peroxisomal β-oxidation also requires enzymes specific to the peroxisome and to very long fatty acids. There are four key differences between the enzymes used for mitochondrial and peroxisomal β-oxidation:

1. The NADH formed in the third oxidative step cannot be reoxidized in the peroxisome, so reducing equivalents are exported to the cytosol.
2. β-oxidation in the peroxisome requires the use of a peroxisomalcarnitine acyltransferase (instead of carnitine acyltransferase I and II used by the mitochondria) for transport of the activated acyl group into the mitochondria for further breakdown.
3. The first oxidation step in the peroxisome is catalyzed by the enzymeacyl-CoA oxidase.
4. Theβ-ketothiolase used in peroxisomal β-oxidation has an altered substrate specificity, different from the mitochondrialβ-ketothiolase.

Peroxisomal oxidation is induced by a high-fat diet and administration of hypolipidemic drugs likeclofibrate.

Theoretically, the ATP yield for each oxidation cycle where two carbons are broken down at a time is 17, as each NADH produces 3 ATP, FADH₂ produces 2 ATP and a full rotation of Acetyl-CoA in citric acid cycle produces 12 ATP.^[13] In practice, it is closer to 14 ATP for a full oxidation cycle as 2.5 ATP per NADH molecule is produced, 1.5 ATP per each FADH₂ molecule is produced and Acetyl-CoA produces 10 ATP per rotation of the citric acid cycle^[13](according to theP/O ratio). This breakdown is as follows:

For an even-numbered saturated fat (C_n), 0.5 * n - 1 oxidations are necessary, and the final process yields an additional acetyl CoA. In addition, two equivalents ofATP are lost during the activation of the fatty acid. Therefore, the total ATP yield can be stated as:

${\displaystyle (0.5\times n-1)\times 14+10-2=ATP}$ ^[14]

or

${\displaystyle 7n-6}$ 

For instance, the ATP yield ofpalmitate (C₁₆,n = 16) is:

${\displaystyle 7\times 16-6=106ATP}$ 

Represented in table form:

For an odd-numbered saturated fat (C_n), 0.5 * n - 1.5 oxidations are necessary, and the final process yields 8 acetyl CoA and 1 propionyl CoA. It is then converted to a succinyl CoA by a carboxylation reaction and generates additional 5 ATP (1 ATP is consumed in carboxylation process generating a net of 4 ATP). In addition, two equivalents ofATP are lost during the activation of the fatty acid. Therefore, the total ATP yield can be stated as:

${\displaystyle (0.5\times n-1.5)\times 14+4-2=ATP}$ 

or

${\displaystyle 7n-19}$ 

For instance, the ATP yield ofNonadecylic acid (C₁₉,n = 19) is:

${\displaystyle 7\times 19-19=114ATP}$ 

Represented in table form:

There are at least 25 enzymes and specific transport proteins in the β-oxidation pathway.^[16] Of these, 18 have been associated with human disease asinborn errors of metabolism.

Furthermore, studies indicate that lipid disorders are involved in diverse aspects of tumorigenesis, and fatty acid metabolism makes malignant cells more resistant to a hypoxic environment. Accordingly, cancer cells can display irregular lipid metabolism with regard to both fatty acid synthesis and mitochondrialfatty acid oxidation (FAO) that are involved in diverse aspects of tumorigenesis and cell growth.^[17] Several specific β-oxidation disorders have been identified.

Medium-chain acyl-coenzyme A dehydrogenase (MCAD) deficiency^[18] is the most common fatty acid β-oxidation disorder and a prevalent metabolic congenital error It is often identified through newborn screening. Although children are normal at birth, symptoms usually emerge between three months and two years of age, with some cases appearing in adulthood.

Medium-chain acyl-CoA dehydrogenase (MCAD) plays a crucial role in mitochondrial fatty acid β-oxidation, a process vital for generating energy during extended fasting or high-energy demand periods. This process, especially important when liver glycogen is depleted, supports hepatic ketogenesis. The specific step catalyzed by MCAD involves the dehydrogenation of acyl-CoA. This step converts medium-chain acyl-CoA to trans-2-enoyl-CoA, which is then further metabolized to produce energy in the form of ATP.

Symptoms

• Affected children, who seem healthy initially, may experience symptoms like low blood sugar without ketones (hypoketotic hypoglycemia) andvomiting
• Can escalate tolethargy,seizures andcoma, typically triggered by illness
• Acute episodes may also involve enlarged liver (hepatomegaly) and liver issues
• Sudden death

Treatments

• Administering simple carbohydrates
• Avoiding fasting
• Frequent feedings for infants
• For toddlers, a diet with less than 30% of total energy from fat
• Administering 2 g/kg of uncooked cornstarch at bedtime for sufficient overnight glucose
• Preventing hypoglycemia, especially due to excessive fasting.
• Avoiding infant formulas with medium-chain triglycerides as the main fat source

Long-chain hydroxyacyl-CoA dehydrogenase (LCHAD) deficiency^[19] is a mitochondrial effect of impaired enzyme function.

LCHAD performs the dehydrogenation of hydroxyacyl-CoA derivatives, facilitating the removal of hydrogen and the formation of aketo group. This reaction is essential for the subsequent steps in beta oxidation that lead to the production of acetyl-CoA, NADH, and FADH2, which are important for generating ATP, the energy currency of the cell.

Long-chain hydroxyacyl-CoA dehydrogenase (LCHAD) deficiency is a condition that affects mitochondrial function due to enzyme impairments. LCHAD deficiency is specifically caused by a shortfall in the enzymelong-chain 3-hydroxyacyl-CoA dehydrogenase. This leads to the body's inability to transform specific fats into energy, especially during fasting periods.

Symptoms

• Severe Phenotype: symptoms appear soon after birth and includehypoglycemia,hepatomegaly, brain dysfunction (encephalopathy) and oftencardiomyopathy
• Intermediate Phenotype: characterized byhypoketotic hypoglycemia and is triggered by infection or fasting during infancy
• Mild (Late-Onset) Phenotype: presents as muscle weakness (myopathy) and nerve disease (neuropathy)
• Long-Term Complications: can includeperipheral neuropathy and eye damage (retinopathy)

Treatments

• Regular feeding to avoid fasting
• Use ofmedium-chain triglyceride (MCT) ortriheptanoin supplements andcarnitine supplements
• Low-fat diet
• Hospitalization with intravenous fluids containing at least 10% dextrose
• Bicarbonate therapy for severemetabolic acidosis
• Management of high ammonia levels and muscle breakdown
• Cardiomyopathy management
• Regular monitoring of nutrition, blood and liver tests with annual fatty acid profile
• Growth, development, heart and neurological assessments and eye evaluations

Very long-chain acyl-coenzyme A dehydrogenase deficiency (VLCAD deficiency) is a genetic disorder that affects the body's ability to break down certain fats. In the β-oxidation cycle, VLCAD's role involves the removal of two hydrogen atoms from the acyl-CoA molecule, forming a double bond and converting it into trans-2-enoyl-CoA. This crucial first step in the cycle is essential for the fatty acid to undergo further processing and energy production. When there is a deficiency in VLCAD, the body struggles to effectively break down long-chain fatty acids. This can lead to a buildup of these fats and a shortage of energy, particularly during periods of fasting or increased physical activity.^[20]

Symptoms

• Severe Early-Onset Cardiac and Multiorgan Failure Form: symptoms appear within days of birth and includehypertrophic/dilated cardiomyopathy, fluid around heart (pericardial effusion), heart rhythm problems (arrhythmias),hepatomegaly and occasionalintermittent hypoglycemia

• Hepatic or Hypoketotic Hypoglycemic Form: typically appears in early childhood withhypoketotic hypoglycemia

• Later-Onset Episodic Myopathic Form: presents with muscle breakdown after exercise (intermittent rhabdomyolysis), muscle cramps and pain, exercise intolerance andlow blood sugar

Treatments

• Low-fat diet

• Use ofmedium-chain triglyceride (MCT) supplements

• Regular, frequent feeding, especially for infants and children
• Snacks high in complex carbohydrates before bedtime
• Guided and limited exercise for older individuals
• Administration of high-energy fluids intravenously

• AvoidingL-carnitine and IV fats
• Plenty of fluids and urine alkalization for muscle breakdown

• Fatty acid metabolism
• Fatty-acid metabolism disorder
• Lipolysis
• Omega oxidation
• Alpha oxidation

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• Jain, P.; Singh, S.; Arya, A. (2021)."Integrated formulae for calculating fatty acid ATP yield".Biochemistry and Molecular Biology Education.49 (3):492–499.doi:10.1002/bmb.21486.PMID 33427394.S2CID 231577993.