Fatty Acids and Fatty Acid Metabolism

What are Fatty Acids?

Fatty acids are a class of carboxylic acid compounds, consisting of hydrocarbon groups consisting of carbon and hydrogen connected to carboxyl groups. Under sufficient oxygen supply, they can be oxidized and decomposed into CO₂ and H₂O, releasing large amounts of energy. Therefore, fatty acids are one of the main energy sources of the body.

Classification of Fatty Acids

1. Divided according to the number of carbon atoms on the carbon chain.

① Short chain fatty acids (SCFA, C≤6): The number of carbon atoms on the carbon chain is between 1~6. It is also called Volatile fatty acids (VFA) because of its strong volatile energy. The common short-chain fatty acids are: acetic acid, propionic acid, butyric acid, isobutyric acid, valeric acid, isovaleric acid, and capric acid. Short-chain fatty acids are the main metabolites of microorganisms and can inhibit the proliferation of certain bacteria to maintain the stability of microflora in low pH environment. Research on these fatty acids is mainly related to the metabolism of intestinal flora, host microbial co-metabolism and the development of microbial fermentation, food nutrition, drugs or health products.

② Mid chain fatty acids (MCFA, 6 < C ≤ 12): the number of carbon atoms in the carbon chain is between 6 and 12. The common medium chain fatty acids are: octanoic acid, capric acid, undecanoic acid, dodecanoic acid, etc.

③ Long chain fatty acids (LCFA, C>12): the number of carbon atoms on the carbon chain is 12 or more. Common long-chain fatty acids include: myristic acid, myristic acid, palmitoleic acid, stearic acid, oleic acid, etc.

2. Divided according to the number of double bonds between carbon atoms in the carbon chain.

① Saturated fatty acids: no double bond between carbon and carbon in the carbon chain

② Monounsaturated fatty acids: on the carbon chain, there is a double bond between carbon and carbon

③ Polyunsaturated fatty acid: there are two or more double bonds between carbon and carbon in the carbon chain

Fatty acids classification (Johnston et al., 2017)

Biochemistry of Fatty Acids

Fatty acids, essential components of lipids, play pivotal roles in cellular structure, energy metabolism, and signaling processes within biological systems. Understanding their molecular structure, functionality in cell membranes, and contributions to energy storage and insulation provides insights into their fundamental biochemical significance.

Molecular Structure of Fatty Acids

Fatty acids are carboxylic acids characterized by a long hydrocarbon chain ranging from 4 to 24 carbons in length. At one end of the chain is a carboxyl group (–COOH), which is hydrophilic and ionized in biological fluids. The hydrocarbon chain, typically hydrophobic and non-polar, varies in saturation—whether it contains single bonds (saturated fatty acids), one double bond (monounsaturated fatty acids), or multiple double bonds (polyunsaturated fatty acids).

Saturated Fatty Acids

Saturated fatty acids have no double bonds between carbon atoms along their hydrocarbon chains. This structure allows them to pack closely together, making them solid at room temperature. Examples include palmitic acid (16:0) and stearic acid (18:0), commonly found in animal fats and some vegetable oils.

Unsaturated Fatty Acids

Unsaturated fatty acids contain one or more double bonds between carbon atoms in their hydrocarbon chains, introducing kinks that disrupt tight packing. Monounsaturated fatty acids (e.g., oleic acid, 18:1) have one double bond, while polyunsaturated fatty acids (e.g., linoleic acid, 18:2; alpha-linolenic acid, 18:3) have two or more double bonds. These characteristics impart fluidity to lipid membranes and influence membrane permeability and function.

Functionality in Cell Membranes and Lipid Bilayers

Fatty acids serve as integral components of phospholipids and glycolipids, the primary constituents of cellular membranes. Phospholipids consist of a glycerol backbone linked to two fatty acids and a phosphate group, creating a polar "head" and non-polar "tails." This amphipathic structure allows phospholipids to form lipid bilayers, the basic framework of cell membranes.

Membrane Fluidity and Stability

The composition of fatty acids within phospholipids profoundly influences membrane fluidity and stability. Saturated fatty acids contribute rigidity and lower permeability, making membranes less fluid. In contrast, unsaturated fatty acids increase membrane fluidity by introducing kinks that prevent close packing of lipid molecules. This fluidity is crucial for membrane flexibility and the function of membrane proteins and receptors involved in cell signaling and transport.

Role in Energy Storage and Insulation

Beyond their structural roles, fatty acids are vital for energy storage and thermal insulation in organisms. Excess dietary fats and carbohydrates are converted into fatty acids and stored as triglycerides in adipose tissue. Triglycerides consist of three fatty acid molecules esterified to a glycerol backbone, forming a compact and efficient energy storage molecule. During energy-demanding activities, such as fasting or exercise, triglycerides are hydrolyzed into fatty acids and glycerol, which are released into the bloodstream to provide energy via beta-oxidation in mitochondria.

Adipose Tissue as an Energy Reservoir

Adipose tissue serves as a dynamic reservoir for fatty acids, responding to hormonal signals (e.g., insulin, glucagon, adrenaline) that regulate lipolysis (breakdown of triglycerides) and lipogenesis (synthesis of triglycerides). This regulatory balance ensures a constant supply of energy substrates to meet metabolic demands throughout the body.

Sources of Fatty Acids

Fatty acids are essential nutrients obtained from dietary sources and synthesized endogenously within the body. Understanding the origins and types of fatty acids is crucial for appreciating their dietary importance and physiological roles.

Dietary Sources

Animal Fats

Animal fats, such as those found in meat, butter, and dairy products, are rich sources of saturated fatty acids (SFAs) and monounsaturated fatty acids (MUFAs). For instance, palmitic acid (16:0) and stearic acid (18:0) are prevalent SFAs, while oleic acid (18:1) is a common MUFA found in beef, pork, and dairy fats. These fats contribute to the flavor, texture, and nutritional profile of animal-derived foods.

Plant Oils

Plant oils, derived from seeds, nuts, and fruits, provide a variety of fatty acids, including both MUFAs and polyunsaturated fatty acids (PUFAs). Olive oil, rich in oleic acid (18:1), is a prominent example of a MUFA source. Meanwhile, oils like sunflower oil and soybean oil contain linoleic acid (18:2) and alpha-linolenic acid (18:3), essential PUFAs crucial for human health.

Seafood

Fish and seafood are renowned for their high content of omega-3 fatty acids, particularly eicosapentaenoic acid (EPA, 20:5) and docosahexaenoic acid (DHA, 22:6). These long-chain PUFAs play critical roles in brain development, cardiovascular health, and inflammation regulation. Fatty fish such as salmon, mackerel, and sardines are primary dietary sources of EPA and DHA.

Endogenous Synthesis in the Body

Fatty Acid Synthesis

In addition to dietary intake, fatty acids can be synthesized endogenously through fatty acid synthesis pathways primarily occurring in the liver and adipose tissue. This process begins with acetyl-CoA, derived from carbohydrate and protein metabolism, which undergoes a series of enzymatic reactions to form long-chain fatty acids, predominantly palmitic acid (16:0).

Regulation and Significance

Endogenous fatty acid synthesis is tightly regulated by hormonal signals, such as insulin, which promotes lipogenesis (synthesis of fatty acids) during times of nutrient abundance. Conversely, fasting or low-energy states stimulate lipolysis (breakdown of triglycerides) to release fatty acids for energy production, highlighting the dynamic regulation of lipid metabolism in response to metabolic demands.

Essential Fatty Acids and Their Importance

Among the dietary fatty acids, omega-3 and omega-6 fatty acids are classified as essential fatty acids (EFAs) because the human body cannot synthesize them de novo. Linoleic acid (18:2, omega-6) and alpha-linolenic acid (18:3, omega-3) must be obtained from dietary sources to maintain optimal health and well-being.

• Omega-6 Fatty Acids: Found abundantly in vegetable oils (e.g., soybean oil, sunflower oil), omega-6 fatty acids play roles in immune response, inflammation regulation, and skin barrier function.
• Omega-3 Fatty Acids: Predominantly sourced from fatty fish (e.g., salmon, trout, mackerel), omega-3 fatty acids are crucial for cardiovascular health, brain function, and reducing inflammation. EPA and DHA derived from fish oils are particularly bioavailable and beneficial.

Transport and Storage of Fatty Acids

Binding to Serum Albumin and Transport in Blood

Upon absorption from the digestive tract or release from adipose tissue, fatty acids are transported in the bloodstream bound to serum albumin, a carrier protein abundant in plasma. This binding is essential for solubilizing fatty acids in the aqueous environment of blood plasma, where they exist predominantly in their ionized form as fatty acid-albumin complexes.

• Mechanism: Serum albumin features multiple binding sites for fatty acids, accommodating various chain lengths and degrees of saturation. This reversible binding enables fatty acids to be transported throughout the body while protecting them from enzymatic degradation and facilitating their delivery to tissues.
• Significance: The binding of fatty acids to serum albumin ensures their availability for energy production in organs such as the heart, liver, and skeletal muscles. This process is tightly regulated by hormonal signals (e.g., insulin, glucagon) and nutritional status to meet metabolic demands.

Storage as Triglycerides in Adipose Tissue

Excess dietary fats and carbohydrates, once absorbed and metabolized, are converted into triglycerides (TGs) within hepatocytes (liver cells) or adipocytes (fat cells) for storage. Triglycerides consist of three fatty acid molecules esterified to a glycerol backbone, forming a compact and efficient energy reservoir in adipose tissue.

• Lipogenesis: Under conditions of nutrient abundance (e.g., after a meal), insulin stimulates lipogenesis—the synthesis of triglycerides from fatty acids and glycerol. This process involves the enzymatic esterification of fatty acids with glycerol-3-phosphate to form TGs, which are then packaged into lipid droplets within adipocytes.
• Lipolysis: During periods of energy demand (e.g., fasting, exercise), lipolysis is initiated to mobilize stored triglycerides. Hormones such as glucagon and adrenaline stimulate lipolysis, which involves the hydrolysis of TGs into glycerol and free fatty acids. These fatty acids are released into the bloodstream for transport to tissues requiring energy.

Mobilization and Utilization During Energy Demand

Beta-Oxidation in Mitochondria

Once released into the bloodstream, free fatty acids are taken up by cells, particularly muscle and liver cells, where they undergo beta-oxidation—a series of enzymatic reactions occurring in mitochondria.

• Process: Beta-oxidation involves the sequential removal of two-carbon acetyl-CoA units from the fatty acid chain, producing acetyl-CoA molecules that enter the citric acid cycle (Krebs cycle) for ATP production through oxidative phosphorylation.
• Energy Yield: Fatty acid oxidation yields a higher energy yield per carbon compared to glucose metabolism, making it a preferred energy source during prolonged exercise or fasting periods. This metabolic flexibility ensures efficient energy utilization while sparing glucose for essential functions in the brain and red blood cells.

Regulatory Mechanisms and Clinical Implications

Hormonal Regulation

• Insulin: Promotes storage of fatty acids as triglycerides in adipose tissue during times of nutrient excess.
• Glucagon and Adrenaline: Stimulate lipolysis and fatty acid release from adipose tissue during energy-demanding activities.

Clinical Relevance

Understanding the transport and storage of fatty acids is crucial for managing metabolic disorders such as obesity, diabetes, and cardiovascular disease. Dysregulation in fatty acid metabolism can lead to lipid accumulation in tissues (lipotoxicity) and contribute to insulin resistance and inflammation.

What is Fatty Acid Metabolism?

The energy stored in fats is mainly in fatty acids. The fatty acids produced by lipolysis bind to clear proteins (albumin) to produce very high density lipoproteins, which are transported via the circulation to the tissues that require energy. Tissues take up fatty acids via various transporter proteins (CD36, FATP, FABP, etc.).

Fatty acid metabolism consists of various metabolic processes involving or closely related to fatty acids. These processes can be mainly divided into (1) catabolic processes that produce energy and (2) anabolic processes.

Fatty Acid Catabolism: Beta-Oxidation

Fatty acid catabolism, primarily through beta-oxidation, is a vital metabolic process that converts fatty acids into acetyl-CoA, generating energy in the form of ATP. This process is essential for maintaining energy homeostasis and supporting various physiological functions.

Beta-Oxidation

Beta-oxidation is a series of enzymatic reactions that occur in the mitochondrial matrix, where fatty acids are sequentially broken down into acetyl-CoA units. This process involves four main steps:

Activation: Fatty acids are first activated in the cytoplasm through esterification with coenzyme A (CoA) to form fatty acyl-CoA molecules. This step requires ATP hydrolysis and is catalyzed by acyl-CoA synthetase enzymes.

Transport into Mitochondria: Fatty acyl-CoA molecules are then transported into the mitochondrial matrix via the carnitine shuttle system. This process involves the conversion of fatty acyl-CoA to fatty acyl-carnitine by carnitine palmitoyltransferase I (CPT-I) in the outer mitochondrial membrane, followed by transport via carnitine translocase and reconversion back to fatty acyl-CoA by CPT-II in the inner mitochondrial membrane.

Beta-Oxidation Steps: Once inside the mitochondrial matrix, beta-oxidation proceeds through a repetitive cycle of four enzymatic reactions:

• Oxidation: The first step involves the oxidation of the fatty acyl-CoA molecule by acyl-CoA dehydrogenase, resulting in the formation of trans-2-enoyl-CoA and reducing equivalents in the form of FADH2.
• Hydration: The trans-2-enoyl-CoA is then hydrated by enoyl-CoA hydratase, forming L-3-hydroxyacyl-CoA.
• Oxidation (again): The L-3-hydroxyacyl-CoA undergoes further oxidation by hydroxyacyl-CoA dehydrogenase, yielding 3-ketoacyl-CoA and reducing equivalents in the form of NADH.
• Thiolysis: Finally, thiolase cleaves the 3-ketoacyl-CoA into acetyl-CoA and a shortened acyl-CoA molecule, ready to undergo another round of beta-oxidation.

Acetyl-CoA Production: Each cycle of beta-oxidation shortens the fatty acyl-CoA chain by two carbons, generating one molecule of acetyl-CoA per cycle. The acetyl-CoA molecules produced then enter the citric acid cycle (Krebs cycle) to generate ATP through oxidative phosphorylation.

Regulation of Beta-Oxidation

Beta-oxidation is tightly regulated to meet cellular energy demands and maintain metabolic balance. Key regulatory factors include:

• Substrate Availability: Levels of fatty acyl-CoA substrates influenced by dietary intake and lipolysis.
• Hormonal Regulation: Insulin promotes fatty acid synthesis and storage during periods of nutrient abundance, whereas glucagon and adrenaline stimulate lipolysis and beta-oxidation during fasting or energy-demanding conditions.
• Enzyme Activity: The activities of enzymes involved in beta-oxidation (e.g., acyl-CoA dehydrogenase, carnitine palmitoyltransferase) are regulated by allosteric mechanisms and post-translational modifications.

Diagram of fatty acid catabolism (Boer et al., 2006).

Fatty Acid Anabolism: Synthesis and Regulation

Fatty acid anabolism, also known as lipogenesis, is the biochemical process by which fatty acids are synthesized from acetyl-CoA and ultimately incorporated into triglycerides and phospholipids. This process is vital for energy storage, membrane structure, and the production of lipid-based signaling molecules within cells.

Fatty Acid Synthesis

Fatty acid synthesis primarily occurs in the liver, adipose tissue, and lactating mammary glands, where it serves to convert excess carbohydrates and proteins into fatty acids for storage as triglycerides.

Key Steps in Fatty Acid Synthesis:

1. Acetyl-CoA Formation: The precursor for fatty acid synthesis is acetyl-CoA, which is generated from pyruvate through glycolysis or from citrate in the mitochondria during the citric acid cycle (Krebs cycle).

2. Formation of Malonyl-CoA: Acetyl-CoA is carboxylated to form malonyl-CoA by acetyl-CoA carboxylase (ACC), a rate-limiting enzyme in fatty acid synthesis. This step requires ATP and biotin as cofactors.

3. Fatty Acid Chain Elongation: Fatty acid synthesis occurs in the cytoplasm via a series of enzymatic reactions catalyzed by fatty acid synthase (FAS). FAS is a multifunctional enzyme complex that sequentially adds two-carbon units from malonyl-CoA to a growing acyl chain bound to acyl carrier protein (ACP).

4. Formation of Palmitate (16:0): After several rounds of condensation, reduction, dehydration, and reduction reactions, a 16-carbon saturated fatty acid, palmitic acid (16:0), is synthesized.

5. Formation of Longer and Unsaturated Fatty Acids: Further modification of palmitic acid can occur to produce longer-chain fatty acids (e.g., stearic acid, 18:0) and unsaturated fatty acids (e.g., oleic acid, 18:1) through desaturation reactions catalyzed by desaturase enzymes.

Regulation of Fatty Acid Synthesis

Fatty acid synthesis is tightly regulated to maintain lipid homeostasis and respond to metabolic demands:

• Regulation of Acetyl-CoA Carboxylase (ACC): ACC activity is regulated by hormonal signals, such as insulin (which activates ACC) and glucagon (which inhibits ACC), as well as by nutritional factors reflecting cellular energy status.
• Regulation of Fatty Acid Synthase (FAS): FAS activity is regulated at multiple levels, including transcriptional control, post-translational modifications, and allosteric regulation by metabolites like citrate (which activates FAS) and palmitoyl-CoA (which inhibits FAS).
• Role of Sterol Regulatory Element-Binding Proteins (SREBPs): SREBPs are transcription factors that regulate the expression of enzymes involved in fatty acid and cholesterol synthesis in response to cellular lipid levels and nutritional signals.

Physiological Functions of Lipogenesis

Energy Storage:

Fatty acid synthesis facilitates the conversion of excess carbohydrates and proteins into triglycerides for storage in adipose tissue, providing a highly efficient energy reserve.

Membrane Structure:

Synthesized fatty acids are essential components of phospholipids and glycolipids, maintaining the structural integrity and fluidity of cellular membranes.

Signaling Molecules:

Lipogenesis generates lipid-derived signaling molecules, such as prostaglandins and leukotrienes, which play roles in inflammation, immune response, and cell signaling pathways.

Biosynthesis and degradation of fatty acids and membrane lipids (Janßen et al., 2014)

Fatty Acid Metabolites Analysis

Lipid metabolism is involved in a variety of cellular processes critical to cellular transformation, tumor development and disease progression. In cancer research, fatty acid metabolism affects cancer cell biology in many ways, particularly the synthesis of membrane lipids, including glycerophospholipids and signal transduction intermediates such as phosphatidylinositol (4,5)-biphosphate, diacylglycerol (DAG) and phospholipids that promote mitogenic and/or oncogenic signaling. Fatty acids are also substrates for mitochondrial ATP and NADH synthesis, arachidonate production and post-translational protein-lipid modifications of signaling proteins. Cancer cells can acquire fatty acids from a range of intracellular and extracellular sources. These alterations in fatty acid metabolism are a feature of tumorigenesis and metastasis.

Creative Proteomics offers fatty acid and fatty acid metabolite analysis services based on our GC-FID and LC-MS platforms. Our services include but are not limited to.

• Absolute qualitative quantification of 7 short-chain fatty acids using GC-MS technology and standards
• Relative quantification (or absolute quantification) of 50 medium and long chain fatty acids using GC-MS technology
• Absolute quantification of 37 free fatty acids using LC-MS/MS technology platform

References

1. Johnston, M. R., & Sobhi, H. F. (2017). Advances in fatty acid analysis for clinical investigation and diagnosis using GC/MS methodology. J. Biochem. Analyt. Stud, 3.
2. De Boer, V. C. J., Van Schothorst,. (2006). Chronic quercetin exposure affects fatty acid catabolism in rat lung. Cellular and Molecular Life Sciences CMLS, 63(23), 2847-2858.
3. Janßen, H. J., & Steinbüchel, A. (2014). Fatty acid synthesis in Escherichia coli and its applications towards the production of fatty acid based biofuels. Biotechnology for biofuels, 7(1), 1-26.