Chapter 10

Lipids

Textbook pages 1284–1366 (Lehninger, 8e) · 25 MCQs below · Source: printed chapter text extracted from the PDF

CHAPTER 10 LIPIDS organized into eight general categories of chemical structure (listed in Table 10-2, at the end of this chapter). We discuss the energy-yielding oxidation of lipids in Chapter 17 and their synthesis in Chapter 21. Here, we will focus on four principles of cellular lipid function: Fatty acids are water-insoluble hydrocarbons used for cellular energy storage. Fatty acids are highly reduced and thus provide a rich source of stored chemical energy for cells. Storage of hydrophobic fats as triacylglycerols is also highly efﬁcient because water is not needed to hydrate the stored fats. Membrane lipids are composed of hydrophobic tails attached to polar head groups. Cellular membranes are composed of a variety of lipids, including glycerophospholipids and sterols. These lipids are used for structuring membranes as well as for displaying molecules on the membrane surfaces for signaling and molecular recognition. Lipids have uses in the cell beyond energy storage and construction of membranes. Many lipids are present in the cell at smaller amounts than those making up membranes or being stored as fat. These lipids can function as cellular messengers, hormones, electron carriers, or pigments. The chemical properties of lipids are related to their structure and composition. As in studies of other biomolecules, methods such as enzymatic, chromatographic, and mass spectrometry can all be used to identify lipids and determine their atomic structure. 10.1 Storage Lipids The fats and oils almost universally used as stored forms of energy in living organisms are derivatives of fatty acids. The fatty acids are hydrocarbon derivatives, at about the same low oxidation state (that is, as highly reduced) as the hydrocarbons in fossil fuels. The cellular oxidation of fatty acids (to CO2 and H2O), like the controlled, rapid burning of fossil fuels in internal combustion engines, is highly exergonic. We introduce here the structures and nomenclature of the fatty acids most commonly found in living organisms. We describe two types of fatty acid–containing compounds, triacylglycerols and waxes, to illustrate the diversity of structures and physical properties in this family of compounds. Fatty Acids Are Hydrocarbon Derivatives Fatty acids are carboxylic acids with hydrocarbon chains ranging from 4 to 36 carbons long (C4 to C36). In some fatty acids, this chain is unbranched and fully saturated (contains no double bonds); in others, the chain contains one or more double bonds (Table 10-1). A few contain three-carbon rings, hydroxyl groups, or methyl-group branches. TABLE 10-1 Some Naturally Occurring Fatty Acids: Structure, Properties, and Nomenclature Solubility at 30(mg/g solvent Carbonskeleton Structure Systematic name Commonname(derivation) Meltingpoint(°C) Water Benz 12:0 CH3(CH2)10COOH n-Dodecanoic acid Lauric acid (Latin laurus, “laurel plant”) 44.2 0.063 2,6 14:0 CH3(CH2)12COOH n-Tetradecanoic acid Myristic acid (Latin Myristica, nutmeg genus) 53.9 0.024 87 16:0 CH3(CH2)14COOH n-Hexadecanoic acid Palmitic acid (Latin palma “palm tree”) 63.1 0.0083 34 18:0 CH3(CH2)16COOH n-Octadecanoic acid Stearic acid (Greek stear, “hard fat”) 69.6 0.0034 12 20:0 CH3(CH2)18COOH n-Eicosanoic acid Arachidic acid (Latin 76.5 a b Arachis, legume genus) 24:0 CH3(CH2)22COOH n-Tetracosanoic acid Lignoceric acid (Latin lignum, “wood” + cera, “wax”) 86.0 16:1(Δ9) CH3(CH2)5CH═CH(CH2)7COOH cis-9-Hexadecenoic acid Palmitoleic acid 1 to – 0.5 18:1(Δ9) CH3(CH2)7CH═CH(CH2)7COOH cis-9-Octadecenoic acid Oleic acid (Latin oleum, “oil”) 13.4 18:2(Δ9,12) CH3(CH2)4CH═CHCH2CH═CH(CH2)7COOH cis-,cis-9,12- Octadecadienoic acid Linoleic acid (Greek linon, “flax”) 1–5 18:3(Δ9,12,15) CH3CH2CH═CHCH2CH═CHCH2CH═CH(CH2)7COOH cis-,cis-,cis-9,12, 15- Octadecatrienoic acid α -Linolenic acid –11 20:4(Δ5,8,11,14) CH3(CH2)4CH═CHCH2CH═CHCH2 CH═CHCH2CH═CH(CH2)3COOH cis-,cis-,cis-,cis-5,8,11, 14-Icosatetraenoic acid Arachidonic acid –49.5 All acids are shown in their nonionized form. At pH 7, all free fatty acids have an ionized carboxylate. Note that numbering of carbon atoms begins at the carboxyl carbon. The prefix n- indicates the “normal” unbranched structure. For instance, “dodecanoic” simply indicates 12 carbon atoms, which could be arranged in a variety of brancheforms; “n-dodecanic” specifies the linear unbranched form. For unsaturated fatty acids, the configuration of each double bond is indicated; in biological fatty acids theconfiguration is almost always cis. KEY CONVENTION A simpliﬁed nomenclature for unbranched fatty acids speciﬁes the chain length and number of double bonds, separated by a colon. For example, the 16-carbon saturated palmitic acid is abbreviated 16:0, and the 18-carbon oleic (octadecenoic) acid, with one double bond (shown below), is 18:1. Each line segment of the zigzag in the structure represents a single bond between adjacent carbons. The carboxyl carbon is assigned the number 1 (C-1), and the α carbon next to it is C-2. The positions of any double bonds, designated Δ (delta), are speciﬁed relative to C-1 by a superscript number indicating the lower-numbered carbon in the double bond. By this convention, oleic acid, with a double bond between C-9 and C-10, is designated 18:1(Δ9); a 20-carbon fatty acid with one double bond between C-9 and C-10 and another between C-12 and C-13 is designated 20:2(Δ9,12). a b The most commonly occurring fatty acids have even numbers of carbon atoms in an unbranched chain of 12 to 24 carbons (Table 10-1). As we shall see in Chapter 21, the even number of carbons results from the mode of synthesis of these compounds, which involves successive condensations of two-carbon acetyl units. There is also a common pattern in the location of double bonds; in most monounsaturated fatty acids the double bond is between C-9 and C-10 (Δ9), and the other double bonds of polyunsaturated fatty acids are generally Δ12 and Δ15. (Arachidonic acid is an exception to this generalization; see Table 10-1.) The double bonds of polyunsaturated fatty acids are almost never conjugated (alternating single and double bonds, as in —CH═CH—CH═CH— ), but are separated by a methylene group: —CH═CH—CH2—CH═CH—. In nearly all naturally occurring unsaturated fatty acids, the double bonds are in the cis conﬁguration. Notable exceptions include the trans fatty acids produced by fermentation in the rumen of dairy animals. Humans can ingest these trans fats from dairy products and meat. KEY CONVENTION The family of polyunsaturated fatty acids (PUFAs) with a double bond between the third carbon and the fourth carbon from the methyl end of the chain are of special importance in human nutrition. Because the physiological role of PUFAs is related more to the position of the ﬁrst double bond near the methyl end of the chain than to that near the carboxyl end, an alternative nomenclature is sometimes used for these fatty acids. The carbon of the methyl group — that is, the carbon most distant from the carboxyl group — is called the ω (omega; the last letter in the Greek alphabet) carbon and is given the number 1 (C-1); the carboxyl carbon in this convention has the highest number. The positions of the double bonds are indicated relative to the ω carbon. In this convention, PUFAs with a double bond between C-3 and C-4 are called omega-3 (ω -3) fatty acids, and those with a double bond between C-6 and C-7 are omega-6 (ω -6) fatty acids. Shown below is eicosapentaenoic acid, which can be designated as 20:5(Δ5,8,11,14,17) by the standard nomenclature but is also referred to as an omega-3 fatty acid, emphasizing the biologically important double bond in the omega-3 position.

Humans require the omega-3 PUFA α -linolenic acid (ALA; 18:3(Δ9,12,15), in the standard convention), but do not have the enzymatic capacity to synthesize it and must therefore obtain ALA from the diet. Humans also use ALA to synthesize two other omega-3 PUFAs important in cellular function: eicosapentaenoic acid (EPA; 20:5(Δ5,8,11,14,17), shown in the Key Convention above) and docosahexaenoic acid (DHA; 22:6(Δ4,7,10,13,16,19)). An imbalance of omega-6 and omega-3 PUFAs in the diet is associated with an increased risk of cardiovascular disease. The optimal dietary ratio of omega-6 to omega-3 PUFAs is between 1:1 and 4:1, but the ratio in the diets of most North Americans is closer to 10:1 to 30:1. The “Mediterranean diet,” which has been associated with lowered cardiovascular risk, is richer in omega-3 PUFAs, obtained in leafy vegetables (salads) and ﬁsh oils. Fish oils especially rich in EPA and DHA are a common dietary supplement even though their exact role in preventing cardiovascular disease is controversial. The physical properties of the fatty acids, and of compounds that contain them, are largely determined by the length and degree of unsaturation of the hydrocarbon chain. The nonpolar hydrocarbon chain accounts for the poor solubility of fatty acids in water. Lauric acid (12:0, Mr 200), for example, has a solubility in water of 0.063 mg/g — much less than that of glucose (Mr 180), which is 1,100 mg/g. The longer the fatty acyl chain and the fewer the double bonds, the lower the solubility in water. The carboxylic acid group is polar (and ionized at neutral pH) and accounts for the slight solubility of short-chain fatty acids in water. Melting points are also strongly inﬂuenced by the length and degree of unsaturation of the hydrocarbon chain. At room temperature (25 °C), the saturated fatty acids from 12:0 to 24:0 have a waxy consistency, whereas unsaturated fatty acids of these lengths are oily liquids. This difference in melting points is due to different degrees of packing of the fatty acid molecules (Fig. 10-1). In the fully saturated compounds, free rotation around each carbon–carbon bond gives the hydrocarbon chain great ﬂexibility; the most stable conformation is the fully extended form, in which the steric hindrance of neighboring atoms is minimized. These molecules can pack together tightly in nearly crystalline arrays, with atoms along their lengths in van der Waals contact with the atoms of neighboring molecules. In unsaturated fatty acids, a cis double bond forces a kink in the hydrocarbon chain. Fatty acids with one or several such kinks cannot pack together as tightly as fully saturated fatty acids, and their interactions with each other are therefore weaker. Because less thermal energy is needed to disorder these poorly ordered arrays of unsaturated fatty acids, they have markedly lower melting points than saturated fatty acids of the same chain length (Table 10-1). FIGURE 10-1 The packing of fatty acids into stable aggregates. The extent of packing depends on the degree of saturation. (a) Two representations of the fully saturated acid stearic acid, 18:0 (stearate at pH 7), in its usual extended conformation. (b) The cis double bond (red) in oleic acid, 18:1(Δ9) (oleate), restricts rotation and introduces a rigid bend in the hydrocarbon tail. All other bonds in the chain are free to rotate. (c) Fully saturated fatty acids in the extended form pack into nearly crystalline arrays, stabilized by extensive hydrophobic interaction. (d) The presence of one or more fatty acids with cis double bonds (red) interferes with this tight packing and results in less stable aggregates. In vertebrates, free fatty acids (unesteriﬁed fatty acids, with a free carboxylate group) can circulate in the blood bound noncovalently to carrier proteins such as serum albumin. However, fatty acids are present in blood plasma mostly as carboxylic acid derivatives such as esters or amides. Lacking the charged carboxylate group, these fatty acid derivatives are generally even less soluble in water than are the free fatty acids and are transported through the blood primarily as lipoprotein particles (see Chapter 21). Triacylglycerols Are Fatty Acid Esters of Glycerol The simplest lipids constructed from fatty acids are the triacylglycerols, also referred to as triglycerides, fats, or neutral fats. Triacylglycerols are composed of three fatty acids, each in ester linkage with a single glycerol (Fig. 10-2). Those containing the same kind of fatty acid in all three positions are called simple triacylglycerols and are named aer the fatty acid they contain. Simple triacylglycerols of 16:0, 18:0, and 18:1, for example, are tripalmitin, tristearin, and triolein, respectively. Most naturally occurring triacylglycerols are mixed; they contain two or three different fatty acids. To name these compounds unambiguously, the name and position of each fatty acid must be speciﬁed. FIGURE 10-2 Glycerol and a triacylglycerol. The mixed triacylglycerol shown here has three diﬀerent fatty acids attached to the glycerol backbone. When glycerol has diﬀerent fatty acids at C-1 and C-3, C-2 is a chiral center. Because the polar hydroxyls of glycerol and the polar carboxylates of the fatty acids are bound in ester linkages, triacylglycerols are nonpolar, hydrophobic molecules, essentially insoluble in water. Lipids have lower speciﬁc gravities than water, which explains why mixtures of oil and water (oil- and-vinegar salad dressing, for example) have two phases: oil, with the lower speciﬁc gravity, ﬂoats on the aqueous phase. Triacylglycerols Provide Stored Energy and Insulation In most eukaryotic cells, triacylglycerols form a separate phase of microscopic, oily droplets in the aqueous cytosol, serving as depots of metabolic fuel. In vertebrates, specialized cells called adipocytes, or fat cells, store large amounts of triacylglycerols as fat droplets that nearly ﬁll the cell (Fig. 10-3a). Triacylglycerols are also stored as oils in the seeds of many types of plants, providing energy and biosynthetic precursors during seed germination (Fig. 10-3b). Adipocytes and germinating seeds contain lipases, enzymes that catalyze the hydrolysis of stored triacylglycerols, releasing fatty acids for export to sites where they are required as fuel. FIGURE 10-3 Fat stores in cells. (a) Cross section of human white adipose tissue. Each cell contains a fat droplet (white) so large that it squeezes the nucleus (stained red) against the plasma membrane. (b) Cross section of a cotyledon cell from a seed of the plant Arabidopsis. The large dark structures are protein bodies, which are surrounded by stored oils in the light-colored oil bodies. There are two signiﬁcant advantages to using triacylglycerols as stored fuels, rather than polysaccharides such as glycogen and starch. First, the carbon atoms of fatty acids are more reduced than those of sugars, so oxidation of triacylglycerols yields more than twice as much energy, gram for gram, as the oxidation of carbohydrates. Second, because triacylglycerols are hydrophobic and therefore unhydrated, the organism that carries stored fuel in the form of fat does not have to carry the extra weight of water of hydration that is associated with stored polysaccharides (2 g per gram of polysaccharide). Humans have fat tissue (composed primarily of adipocytes) under the skin, in the abdominal cavity, and in the mammary glands. Moderately obese people with 15 to 20 kg of triacylglycerols deposited in their adipocytes could meet their energy needs for months by drawing on their fat stores. In contrast, the human body can store less than a day’s energy supply in the form of glycogen. Carbohydrates such as glucose do offer certain advantages as quick sources of metabolic energy; one of those advantages is their ready solubility in water. In some animals, triacylglycerols stored under the skin serve not only as energy stores but also as insulation against low temperatures. Seals, walruses, penguins, and other warm-blooded polar animals are amply padded with triacylglycerols. In hibernating animals (bears, for example), the huge fat reserves accumulated before hibernation serve the dual purposes of insulation and energy storage (see Box 17-1). Partial Hydrogenation of Cooking Oils Improves TheirStability but Creates Fatty Acids with Harmful HealthEﬀects Most natural fats, such as those in vegetable oils, dairy products, and animal fat, are complex mixtures of simple and mixed triacylglycerols. These contain a variety of fatty acids differing in chain length and degree of saturation (Fig. 10-4). Vegetable oils such as corn (maize) oil and olive oil are composed largely of triacylglycerols with unsaturated fatty acids and thus are liquids at room temperature. Triacylglycerols containing only saturated fatty acids, such as tristearin, the major component of beef fat, are white, greasy solids at room temperature. FIGURE 10-4 Fatty acid composition of three food fats. Olive oil, butter, and beef fat consist of mixtures of triacylglycerols diﬀering in their fatty acid composition. The melting points of these fats — and hence their physical state at room temperature (25 °C) — are a direct function of their fatty acid composition. Olive oil has a high proportion of long-chain (C16 and C18) unsaturated fatty acids, which accounts for its liquid state at 25 °C. The higher proportion of long-chain (C16 and C18) saturated fatty acids in butter increases its melting point, so butter is a so solid at room temperature. Beef fat, with an even higher proportion of long-chain saturated fatty acids, is a hard solid. When lipid-rich foods are exposed too long to the oxygen in air, they may spoil and become rancid. The unpleasant taste and smell associated with rancidity result from the oxidative cleavage of double bonds in unsaturated fatty acids, which produces aldehydes and carboxylic acids of shorter chain length and therefore higher volatility; these compounds pass readily through the air to your nose. Throughout the twentieth century, partial hydrogenation of commercial vegetable oils was used to improve shelf life and to increase the stability of cooking oils at the high temperatures used in deep-frying. This process converts many of the cis double bonds in the fatty acids to single bonds and increases the melting temperature of the oils so that they are more nearly solid at room temperature (margarine is produced from vegetable oil in this way). Partial hydrogenation, however, has another, undesirable, effect: some cis double bonds are converted to trans double bonds. There is now strong evidence that dietary intake of trans fatty acids (oen referred to simply as “trans fats”) leads to a higher incidence of cardiovascular disease, and that avoiding these fats in the diet substantially reduces the risk of coronary heart disease. Dietary trans fatty acids raise the level of triacylglycerols and of LDL (“bad”) cholesterol in the blood, and lower the level of HDL (“good”) cholesterol, and these changes alone are enough to increase the risk of coronary heart disease. But trans fatty acids may have further adverse effects. They seem, for example, to increase the body’s inﬂammatory response, which is another risk factor for heart disease. (See Chapter 21 for descriptions of LDL and HDL — low-density and high-density lipoprotein — cholesterol and their health effects.) Regulatory agencies around the world now limit or ban the use of trans fatty acids in prepared and packaged foods.

Waxes Serve as Energy Stores and Water Repellents Biological waxes are esters of long-chain (C14 to C36) saturated and unsaturated fatty acids with long-chain (C16 to C30) alcohols (Fig. 10-5). Their melting points (60 to 100 °C) are generally higher than those of triacylglycerols. In plankton, the free-ﬂoating microorganisms at the bottom of the food chain for marine animals, waxes are the chief storage form of metabolic fuel. FIGURE 10-5 Biological wax. (a) Triacontanoylpalmitate, the major component of beeswax, is an ester of palmitic acid with the alcohol triacontanol. (b) The beeswax of a honeycomb is firm at 25 °C and completely impervious to water. The term “wax” originates in the Old English weax, meaning “the material of the honeycomb.” Waxes also serve a diversity of other functions related to their water-repellent properties and their ﬁrm consistency. Certain skin glands of vertebrates secrete waxes to protect hair and skin and keep it pliable, lubricated, and waterproof. Birds, particularly waterfowl, secrete waxes from preen glands to keep their feathers water-repellent. The shiny leaves of holly, rhododendrons, poison ivy, and many tropical plants are coated with a thick layer of waxes, which prevents excessive evaporation of water and protects against parasites. Biological waxes ﬁnd a variety of applications in the pharmaceutical, cosmetic, and other industries. Lanolin (from lamb’s wool), beeswax (Fig. 10-5), carnauba wax (from a Brazilian palm tree), and wax extracted from the seeds of the jojoba bush are widely used in the manufacture of lotions, ointments, and polishes. SUMMARY 10.1 Storage Lipids Lipids are water-insoluble cellular components of diverse structure that can be extracted fromtissues by nonpolar solvents. Almost all fatty acids, the hydrocarbon components of many lipids,have an even number of carbon atoms (usually 12 to 24); they are either saturated or unsaturated,with double bonds almost always in the cis conﬁguration. Triacylglycerols contain three fatty acid molecules esteriﬁed to the three hydroxyl groups ofglycerol. Simple triacylglycerols contain only one type of fatty acid; mixed triacylglycerols containtwo or three types. Triacylglycerols are present in many foods and are primarily used for storing energy andproviding insulation in animals. Lipases release fatty acids from storage so they can be used as fuel. Since many natural fats can easily turn rancid, partial hydrogenation has been used to extendtheir shelf life. This process can produce trans fatty acids, which increase risk for coronary heartdisease. As a result, their use in prepared and processed foods has become highly regulated. Waxes are esters of long-chain fatty acids and long-chain alcohols with diverse uses in biologyand industry. 10.2 Structural Lipids in Membranes The central architectural feature of biological membranes is a double layer of lipids, which acts as a barrier to the passage of polar molecules and ions. Membrane lipids are amphipathic: one end of the molecule is hydrophobic, the other hydrophilic. The association of their hydrophobic regions with each other and their hydrophilic interactions with water direct their packing into sheets called membrane bilayers. In this section, we describe four general types of membrane lipids: phospholipids, which have hydrophobic regions composed of two fatty acids joined to glycerol or sphingosine; glycolipids, which contain a simple sugar or a complex oligosaccharide at the polar ends; archaeal tetraether lipids, which have two very long alkyl chains ether-linked to glycerol at both ends; and sterols, compounds characterized by a rigid system of four fused hydrocarbon rings (Fig. 10-6). Within these groups of membrane lipids, enormous diversity results from various combinations of fatty acid “tails” and polar “heads.” The arrangement of these lipids in membranes, and their structural and functional roles therein, are considered in the next chapter. FIGURE 10-6 Some common types of storage and membrane lipids. The triacylglycerol, phospholipid, glycolipid, and archaeal ether lipid types shown here have either glycerol or sphingosine as the backbone (light red screen), to which are attached one or more long-chain alkyl groups (yellow) and a polar head group (blue). Sterols have a core composed of four fused carbocylic rings called the steroid nucleus. In triacylglycerols, glycerophospholipids, galactolipids, and sulfolipids, the alkyl groups are fatty acids in ester linkage. Sphingolipids contain a single fatty acid, in amide linkage to the sphingosine backbone. The membrane lipids of archaea are variable; that shown here has two very long, branched alkyl chains, each end in ether linkage with a glycerol moiety. Sterols have many diﬀerent types of alkyl chains that can be attached to the steroid D ring at C-17. In phospholipids, the polar head group is joined through a phosphodiester, whereas glycolipids have a direct glycosidic linkage between the head- group sugar and the backbone glycerol. Many sterols contain a polar head group such as a carbonyl or hydroxyl group attached at C-3 of the steroid A ring. Glycerophospholipids Are Derivatives of Phosphatidic Acid Glycerophospholipids, also called phosphoglycerides, are membrane lipids in which two fatty acids are attached in ester linkage to the ﬁrst and second carbons of glycerol, and a highly polar or charged group is attached through a phosphodiester linkage to the third carbon. Glycerol is prochiral; it has no asymmetric carbons, but attachment of phosphate at one end converts it into a chiral compound, which can be correctly named either L-glycerol 3-phosphate, D-glycerol 1-phosphate, or sn- glycerol 3-phosphate (Fig. 10-7). Glycerophospholipids are named as derivatives of the parent compound, phosphatidic acid (Fig. 10- 8), according to the polar alcohol in the head group. Phosphatidylcholine and phosphatidylethanolamine have choline and ethanolamine as their polar head groups, for example. Cardiolipin is a two-tailed glycerophospholipid in which two phosphatidic acid moieties share the same glycerol as their head group (Fig. 10-8). Cardiolipin is found in most bacterial membranes; in eukaryotic cells, cardiolipin is located almost exclusively in the inner mitochondrial membrane (where it is synthesized), a location consistent with the endosymbiosis hypothesis for the origin of these organelles (see Fig. 1-37). FIGURE 10-7 -Glycerol 3-phosphate, the backbone of phospholipids. Glycerol itself is not chiral, as it has a plane of symmetry through C-2. However, glycerol is prochiral — it can be converted to a chiral compound by adding a substituent such as phosphate to either of the — CH2OH groups. One unambiguous nomenclature for glycerol phosphate is the , system (described on p. 73), in which the isomers are named according to their stereochemical relationships to glyceraldehyde isomers. By this system, the stereoisomer of glycerol phosphate found in most lipids is correctly named either -glycerol 3-phosphate or -glycerol 1-phosphate. Another way to specify stereoisomers is the sn (stereospecific numbering) system, in which C-1 is, by definition, the group of the prochiral compound that occupies the pro-S position. The common form of glycerol phosphate in phospholipids is, by this system, sn-glycerol 3-phosphate (in which C-2 has the R configuration). In archaea, the glycerol in lipids has the other configuration; it is -glycerol 3-phosphate. FIGURE 10-8 Glycerophospholipids. The common glycerophospholipids are diacylglycerols linked to head-group alcohols through a phosphodiester bond. Phosphatidic acid, a phosphomonoester, is the parent compound. Each derivative is named for the head-group alcohol, with the prefix “phosphatidyl-.” In cardiolipin, two phosphatidic acids share a single glycerol (R1 and R2 are fatty acyl groups), resulting in a symmetric molecule. *Note that phosphate esters each have a charge of about –1.5; one of their — OH groups is only partially ionized at pH 7. In all glycerophospholipids, the head group is joined to glycerol through a phosphodiester bond, in which the phosphate group bears a negative charge at neutral pH. The polar alcohol may be negatively charged (as in phosphatidylinositol 4,5-bisphosphate), neutral (phosphatidylserine), or positively charged (phosphatidylcholine, phosphatidylethanolamine). As we shall see in Chapter 11, these charges contribute greatly to the surface properties of membranes. The fatty acids in glycerophospholipids can be any of a wide variety, so a given phospholipid (phosphatidylcholine, for example) may consist of several molecular species, each with its unique complement of fatty acids. The distribution of molecular species is speciﬁc to the organism, to the particular tissue within the organism, and to the particular glycerophospholipids in the same cell or tissue. In general, glycerophospholipids contain a C16 or C18 saturated fatty acid at C-1 and a C18 or C20 unsaturated fatty acid at C-2. With few exceptions, the biological signiﬁcance of the variation in fatty acids and head groups is not yet understood. Some Glycerophospholipids Have Ether-Linked Fatty Acids Some animal tissues and some unicellular organisms are rich in ether lipids, in which one of the two acyl chains is attached to glycerol in ether, rather than ester, linkage. The ether-linked chain may be saturated, as in the alkyl ether lipids, or it may contain a double bond between C-1 and C-2, as in plasmalogens (Fig. 10-9). Vertebrate heart tissue is uniquely enriched in ether lipids; about half of the heart phospholipids are plasmalogens. The membranes of halophilic bacteria, ciliated protists, and certain invertebrates also contain high proportions of ether lipids. The functional signiﬁcance of ether lipids in these membranes is unknown; perhaps their resistance to the phospholipases that cleave ester-linked fatty acids from membrane lipids is important in some roles. FIGURE 10-9 Ether lipids. Plasmalogens have an ether-linked alkenyl chain where most glycerophospholipids have an ester-linked fatty acid (compare to Fig. 10-8). Platelet-activating factor has a long ether-linked alkyl chain at C-1 of glycerol, but C-2 is ester-linked to acetic acid, which makes the compound much more water-soluble than most glycerophospholipids and plasmalogens. The head-group alcohol is ethanolamine in plasmalogens and choline in platelet-activating factor. At least one ether lipid, platelet-activating factor, is a potent molecular signal. It is released from leukocytes called basophils and stimulates platelet aggregation and the release of serotonin (a vasoconstrictor) from platelets. It also exerts a variety of effects on liver, smooth muscle, heart, uterine, and lung tissues and plays an important role in inﬂammation and the allergic response. Galactolipids of Plants and Ether- Linked Lipids of Archaea Are Environmental Adaptations The second group of membrane lipids in Figure 10-6, the glycolipids, includes those that predominate in plant cells: the galactolipids, in which one or two galactose residues are connected by a glycosidic linkage to C-3 of a 1,2-diacylglycerol (Fig. 10-10). Galactolipids are localized in the thylakoid membranes (internal membranes) of chloroplasts; they make up 70% to 80% of the total membrane lipids of a vascular plant, and are therefore probably the most abundant membrane lipids in the biosphere. Phosphate is oen the limiting plant nutrient in soil, and perhaps the evolutionary pressure to conserve phosphate for more critical roles favored plants that made phosphate-free lipids. Plant membranes also contain sulfolipids, in which a sulfonated glucose residue is joined to a diacylglycerol in glycosidic linkage. The sulfonate group bears a negative charge like that of the phosphate group in phospholipids. FIGURE 10-10 Two galactolipids of chloroplast thylakoid membranes. In monogalactosyldiacylglycerols (MGDGs) and digalactosyldiacylglycerols (DGDGs), both acyl groups are polyunsaturated and the head groups are uncharged. Some archaea that live in ecological niches with extreme conditions — for example, high temperatures (boiling water), low pH, or high ionic strength — have membrane lipids containing long-chain (32 carbons) branched hydrocarbons linked at each end to glycerol (Fig.10-6). These linkages are through ether bonds, which are much more stable to hydrolysis at low pH and high temperature than are the ester bonds found in the lipids of bacteria and eukaryotes. In their fully extended form, these archaeal lipids are twice the length of phospholipids and sphingolipids, and can span the full width of the plasma membrane. Sphingolipids Are Derivatives of Sphingosine Sphingolipids, a large class of membrane phospholipids and glycolipids, have a polar head group and two nonpolar tails, but unlike glycerophospholipids and galactolipids they contain no glycerol (Fig. 10-6). Sphingolipids are composed of one molecule of the long-chain amino alcohol sphingosine (also called 4- sphingenine) or one of its derivatives, one molecule of a long- chain fatty acid, and a polar head group that is joined by a glycosidic linkage in some cases and a phosphodiester in others (Fig. 10-11). FIGURE 10-11 Sphingolipids. The first three carbons at the polar end of sphingosine are analogous to the three carbons of glycerol in glycerophospholipids. The amino group at C-2 bears a fatty acid in amide linkage. The fatty acid is usually saturated or monounsaturated, with 16, 18, 22, or 24 carbon atoms. Ceramide is the parent compound for this group. Other sphingolipids diﬀer in the polar head group attached at C-1. Gangliosides have very complex oligosaccharide head groups. Standard symbols for sugars are used in this figure, as shown in Table 7-1. Carbons C-1, C-2, and C-3 of the sphingosine molecule are structurally analogous to the three carbons of glycerol in glycerophospholipids. When a fatty acid is attached in amide linkage to the — NH2 on C-2, the resulting compound is a ceramide, which is structurally similar to a diacylglycerol. Ceramides are the structural parents of all sphingolipids. There are three subclasses of sphingolipids, all derivatives of ceramide but differing in their head groups: sphingomyelins, neutral (uncharged) glycolipids, and gangliosides. Sphingomyelins contain phosphocholine or phosphoethanolamine as their polar head group and are therefore classiﬁed along with glycerophospholipids as phospholipids (Fig. 10-6). Indeed, sphingomyelins resemble phosphatidylcholines in their general properties and three- dimensional structure, and in having no net charge on their head groups (Fig. 10-12). Sphingomyelins are present in the plasma membranes of animal cells and are especially prominent in myelin, a membranous sheath that surrounds and insulates the axons of some neurons — thus the name “sphingomyelins.” FIGURE 10-12 The similar molecular structures of two classes of membrane lipid. Phosphatidylcholine (a glycerophospholipid) and sphingomyelin (a sphingolipid) have similar dimensions and physical properties, but presumably play diﬀerent roles in membranes. Glycosphingolipids, which occur largely in the outer face of plasma membranes, have head groups with one or more sugars connected directly to the — OH at C-1 of the ceramide moiety; they do not contain phosphate. Cerebrosides have a single sugar linked to ceramide; those with galactose are characteristically found in the plasma membranes of cells in neural tissue, and those with glucose are found in the plasma membranes of cells in nonneural tissues. Globosides are glycosphingolipids with two or more sugars, usually D-glucose, D-galactose, or N-acetyl-D- galactosamine. Cerebrosides and globosides are sometimes called neutral glycolipids, as they have no charge at pH 7. Gangliosides, the most complex sphingolipids, have oligosaccharides as their polar head groups and one or more residues of N-acetylneuraminic acid (Neu5Ac), a sialic acid (oen simply called “sialic acid”), at the termini. Deprotonated sialic acid gives gangliosides the negative charge at pH 7 that distinguishes them from globosides. Gangliosides with one sialic acid residue are in the GM (M for mono-) series, those with two sialic acid residues are in the GD (D for di-) series, and so on (GT, three sialic acid residues; GQ, four). Sphingolipids at Cell Surfaces Are Sites of Biological Recognition When sphingolipids were discovered more than a century ago by the physician-chemist Johann Thudichum, their biological role seemed as enigmatic as the Sphinx, for which he therefore named them. In humans, at least 60 different sphingolipids have been identiﬁed in cellular membranes. Many of these are especially prominent in the plasma membranes of neurons, and some are clearly recognition sites on the cell surface, but a speciﬁc function for only a few sphingolipids has been discovered thus far. The carbohydrate moieties of certain sphingolipids deﬁne the human blood groups and therefore determine the type of blood that individuals can safely receive in blood transfusions (Fig. 10-13). FIGURE 10-13 Glycosphingolipids as determinants of blood groups. The human blood groups (O, A, B) are determined in part by the oligosaccharide head groups of these glycosphingolipids. The same three oligosaccharides are also found attached to certain blood proteins of individuals of blood types O, A, and B, respectively. Standard symbols for sugars are used here (see Table 7-1). Gangliosides are concentrated on the outer face of cell surface plasma membranes where they present points of recognition for extracellular molecules or the surfaces of neighboring cells. The kinds and amounts of gangliosides in the plasma membrane change dramatically during embryonic development. Tumor formation induces the synthesis of a new complement of gangliosides, and very low concentrations of a speciﬁc ganglioside have been found to induce differentiation of cultured neuronal tumor cells. Guillain-Barré syndrome is a serious autoimmune disorder in which the body makes antibodies against its own gangliosides, including those in neurons. The resulting inﬂammation damages the peripheral nervous system, leading to temporary (or sometimes permanent) paralysis. In cholera, cholera toxin produced by the intestinal bacterium Vibrio cholerae enters sensitive cells aer attaching to speciﬁc gangliosides on the intestinal epithelial cell surface. Investigation of the biological roles of diverse gangliosides remains fertile ground for future research. Phospholipids and Sphingolipids Are Degraded in Lysosomes Most cells continually degrade and replace their membrane lipids. For each hydrolyzable bond in a glycerophospholipid, there is a speciﬁc hydrolytic enzyme in the lysosome (Fig. 10-14). Phospholipases of the A type remove one of the two fatty acids, producing a lysophospholipid. (These esterases do not attack the ether link of plasmalogens.) Lysophospholipases remove the remaining fatty acid. FIGURE 10-14 The specificities of phospholipases. Phospholipases A1 and A2 hydrolyze the ester bonds of intact glycerophospholipids at C-1 and C-2 of glycerol, respectively. When one of the fatty acids has been removed by a type A phospholipase, the second fatty acid is removed by a lysophospholipase (not shown). Phospholipases C and D each split one of the phosphodiester bonds in the head group. Some phospholipases act on only one type of glycerophospholipid, such as phosphatidylinositol 4,5- bisphosphate (PIP2, shown here) or phosphatidylcholine; others are less specific. Gangliosides are degraded by a set of lysosomal enzymes that catalyze the stepwise removal of sugar units, ﬁnally yielding a ceramide. A genetic defect in any of these hydrolytic enzymes leads to the accumulation of gangliosides in the cell, with severe medical consequences (Box 10-1). BOX 10-1 MEDICINE Abnormal Accumulations of Membrane Lipids: Some Inherited Human Diseases The polar lipids of membranes undergo constant metabolic turnover, the rate of their synthesis normally counterbalanced by the rate of breakdown. The breakdown of lipids is promoted by hydrolytic enzymes in lysosomes, with each enzyme capable of hydrolyzing a specific bond. When sphingolipid degradation is impaired by a defect in one of these enzymes (Fig. 1), partial breakdown products accumulate in the tissues, causing serious disease. More than 50 distinct lysosomal storage diseases have been discovered, each the result of a single mutation in one of the genes for a lysosomal protein. FIGURE 1 Pathways for the breakdown of GM1, globoside, and sphingomyelin to ceramide. A defect in the enzyme hydrolyzing a particular step is indicated by ; the disease that results from accumulation of the partial breakdown product is noted. For example, Niemann-Pick disease is caused by a rare genetic defect in the enzyme sphingomyelinase, the enzyme that cleaves phosphocholine from sphingomyelin. Sphingomyelin accumulates in the brain, spleen, and liver. The disease becomes evident in infants and causes intellectual disability and early death. More common is Tay-Sachs disease, in which ganglioside GM2 accumulates in the brain and spleen (Fig. 2) owing to lack of the enzyme hexosaminidase A. The symptoms of Tay-Sachs disease are progressive developmental delay and disability, paralysis, blindness, and death by the age of 3 or 4 years. FIGURE 2 Electron micrograph of a portion of a brain cell from an infant with Tay-Sachs disease, obtained postmortem, showing abnormal ganglioside deposits in the lysosomes. Genetic counseling can predict and avert many inheritable diseases. Tests on prospective parents can detect abnormal enzymes; then DNA testing can determine the exact nature of the defect and the risk it poses for oﬀspring. Once a pregnancy occurs, fetal cells obtained by sampling a part of the placenta (chorionic villus sampling) or the fluid surrounding the fetus (amniocentesis) can be tested in the same way. Sterols Have Four Fused Carbon Rings Sterols are structural lipids present in the membranes of most eukaryotic cells. The characteristic structure of this group of membrane lipids is the steroid nucleus, consisting of four fused rings, three with six carbons and one with ﬁve (Fig. 10-15). The steroid nucleus is almost planar and is relatively rigid; the fused rings do not allow rotation about C— C bonds. Cholesterol, the major sterol in animal tissues, is amphipathic, with a polar head group (the hydroxyl group at C-3) and a nonpolar hydrocarbon body (the steroid nucleus and the hydrocarbon side chain at C-17) about as long as a 16-carbon fatty acid in its extended form. Similar sterols are found in other eukaryotes: stigmasterol in plants and ergosterol in fungi, for example. Bacteria cannot synthesize sterols; a few bacterial species, however, can incorporate exogenous sterols into their membranes. The sterols of all eukaryotes are synthesized from simple ﬁve-carbon isoprene subunits, as are the fat-soluble vitamins, quinones, and dolichols described in Section 10.3. FIGURE 10-15 Cholesterol. In this chemical structure of cholesterol, the rings are labeled A through D to simplify reference to derivatives of the steroid nucleus. The C-3 hydroxyl group (shaded blue) is the polar head group. For storage and transport of the sterol, this hydroxyl group condenses with a fatty acid to form a sterol ester. In addition to their roles as membrane constituents, the sterols serve as precursors for a variety of products with speciﬁc biological activities. Steroid hormones, for example, are potent biological signals that regulate gene expression. Bile acids are polar derivatives of cholesterol that act as detergents in the intestine, emulsifying dietary fats to make them more readily accessible to digestive lipases. We return to cholesterol and other sterols in later chapters, to consider the structural role of cholesterol in biological membranes (Chapter 11), signaling by steroid hormones (Chapter 12), and the remarkable biosynthetic pathway to cholesterol and transport of cholesterol by lipoprotein carriers (Chapter 21). SUMMARY 10.2 Structural Lipids in Membranes Lipids with polar heads and nonpolar tails are major components of membranes. The most abundant are the glycerophospholipids, which contain fatty acids esteriﬁed to two of the hydroxyl groups of glycerol. The third hydroxyl of glycerol is esteriﬁed with the polar head group via a phosphodiester bond. Common glycerophospholipids such as phosphatidylethanolamine and phosphatidylcholine differ in the structure of their head group. Heart tissue plasmalogens and platelet-activating factor are examples of glycerophospholipids containing ether-linked acyl chains. Chloroplast membranes are rich in galactolipids, composed of a diacylglycerol with one or two linked galactose residues, and sulfolipids, diacylglycerols with a linked sulfonated sugar residue. Some archaea have unique membrane lipids, with long-chain alkyl groups ether-linked to glycerol at each end. These lipids are stable under the harsh conditions in which these archaea live. The sphingolipids contain sphingosine instead of glycerol. The three subclasses of sphingolipids are all ceramide derivatives: sphingomyelins, neutral glycolipids, and gangliosides. Sphingomyelins have choline head groups; the other classes have sugar components. Sphingolipids are abundant in the plasma membranes of neurons, and they deﬁne human blood groups. Phospholipases degrade glycerophospholipids and catalyze hydrolysis at speciﬁc positions within the structure. Sterols have four fused rings and a hydroxyl group. Cholesterol, the major sterol in animals, is both a structural component of membranes and precursor to a wide variety of steroids. 10.3 Lipids as Signals, Cofactors, and Pigments The two functional classes of lipids considered thus far (storage lipids and structural lipids) are major cellular components; membrane lipids make up 5% to 10% of the dry mass of most cells, and storage lipids make up more than 80% of the mass of an adipocyte. With some important exceptions, these lipids play a passive role in the cell; lipid fuels are stored until oxidized by enzymes, and membrane lipids form impermeable barriers around cells and cellular compartments. Another group of lipids, present in much smaller amounts, includes those with active roles in metabolic trafﬁc as metabolites and messengers. Some serve as potent signals — as hormones, carried in the blood from one tissue to another, or as intracellular messengers generated in response to an extracellular signal (hormone or growth factor). Others function as enzyme cofactors in electron- transfer reactions in chloroplasts and mitochondria, or in the transfer of sugar moieties in a variety of glycosylation reactions. A third group consists of lipids with a system of conjugated double bonds: pigment molecules that absorb visible light. Some of these act as light-capturing pigments in vision and photosynthesis; others produce natural colorations, such as the orange of pumpkins and carrots and the yellow of canary feathers. Finally, a very large group of volatile lipids produced in plants consists of signaling molecules that pass through the air, allowing plants to communicate with each other and to invite animal friends and deter foes. In this section, we describe a few representatives of these biologically active lipids. In later chapters, we consider their synthesis and biological roles in more detail. Phosphatidylinositols and Sphingosine Derivatives Act as Intracellular Signals Phosphatidylinositol (PI) and its phosphorylated derivatives (Fig. 10-16) act at several levels to regulate cell structure and metabolism. Phosphatidylinositol 4,5-bisphosphate (PIP2; Fig. 10-14) in the cytoplasmic (inner) face of plasma membranes serves as a reservoir of messenger molecules that are released inside the cell in response to extracellular signals interacting with speciﬁc surface receptors. Extracellular signals such as the hormone vasopressin activate a speciﬁc phospholipase C in the membrane, which hydrolyzes PIP2 to release two products that act as intracellular messengers: inositol 1,4,5-trisphosphate (IP3), which is water soluble, and diacylglycerol, which remains associated with the plasma membrane. IP3 triggers release of Ca2+ from the endoplasmic reticulum, and the combination of diacylglycerol and elevated cytosolic Ca2+ activates the enzyme protein kinase C. By phosphorylating speciﬁc proteins, this enzyme brings about the cell’s response to the extracellular signal. This signaling mechanism is described more fully in Chapter 12 (see Fig. 12-15). FIGURE 10-16 Phosphatidylinositol and its derivatives. (a) In phosphatidylinositol, the glycerol phospholipid is attached at C-1 of inositol. Phosphorylation of the remaining inositol hydroxyl groups forms signaling molecules such as phosphatidylinositol 4,5-bisphosphate (PIP2; see Fig. 10-14), inositol 1,4,5-bisphosphate (IP3), and phosphatidylinositol 3,4,5-trisphosphate (PIP3). (b) A useful trick for remembering the inositol carbon-numbering scheme is to compare the chair conformation of inositol to a turtle. Begin numbering with C-1 as the right front flipper and move counterclockwise around the structure, with C-2 the head and C-5 the tail. Inositol phospholipids also serve as points of nucleation for supramolecular complexes involved in signaling or in exocytosis. Certain signaling proteins bind speciﬁcally to phosphatidylinositol 3,4,5-trisphosphate (PIP3) in the plasma membrane, initiating the formation of multienzyme complexes at the membrane’s cytosolic surface. Thus, formation of PIP3 in response to extracellular signals brings the proteins together in signaling complexes at the surface of the plasma membrane (see Figs. 12-11 and 12-23). Membrane sphingolipids also can serve as sources of intracellular messengers. Both ceramide and sphingomyelin (Fig. 10-11) are potent regulators of protein kinases, and ceramide or its derivatives are involved in the regulation of cell division, differentiation, migration, and programmed cell death (also called apoptosis; see Chapter 12). Eicosanoids Carry Messages to Nearby Cells

Eicosanoids are paracrine hormones, substances that act only on cells near the point of hormone synthesis instead of being transported in the blood to act on cells in other tissues or organs. These fatty acid derivatives have a variety of dramatic effects on vertebrate tissues. They are involved in reproductive function; in the inﬂammation, fever, and pain associated with injury or disease; in the formation of blood clots and the regulation of blood pressure; in gastric acid secretion; and in various other processes important in human health or disease. Eicosanoids are derived from arachidonate (arachidonic acid; 20:4(Δ5,8,11,14)) and eicosapentaenoic acid (EPA; 20:5(Δ5,8,11,14,17)), from which they take their general name (Greek eikosi, “twenty”). There are four major classes of eicosanoids: prostaglandins, thromboxanes, leukotrienes, and lipoxins (Fig. 10-17). Eicosanoid names include letter designations for the functional groups on the ring and numbers indicating the number of double bonds in the hydrocarbon chain. FIGURE 10-17 Arachidonic acid and some eicosanoid derivatives. Arachidonic acid (arachidonate at pH 7) is the precursor of eicosanoids, including the prostaglandins, thromboxanes, leukotrienes, and lipoxins. In prostaglandin E2, C-8 and C-12 of arachidonate are joined to form the characteristic five-membered ring. In thromboxane A2, the C-8 and C-12 are joined and an oxygen atom is added to form the six-membered ring. Nonsteroidal anti-inflammatory drugs (NSAIDs) such as aspirin and ibuprofen block the formation of prostaglandins and thromboxanes from arachidonate by inhibiting the enzyme cyclooxygenase (prostaglandin H2 synthase). Leukotriene A4 has a series of three conjugated double bonds, and no cyclic moiety. Lipoxins are also noncyclic derivatives of arachidonate, with several hydroxyl groups. Prostaglandins (PG) contain a ﬁve-carbon ring, and their name derives from the prostate gland from which they were ﬁrst isolated. PGE2 and other series 2 prostaglandins are synthesized from arachidonate; series 3 prostaglandins are derived from EPA (see Fig. 21-12). Prostaglandins have an array of functions. Some stimulate contraction of the smooth muscle of the uterus during menstruation and labor. Others affect blood ﬂow to speciﬁc organs, the wake-sleep cycle, and the responsiveness of certain tissues to hormones such as epinephrine and glucagon. Prostaglandins in a third group elevate body temperature (producing fever) and cause inﬂammation and pain. The thromboxanes (TX) have a six-membered ring containing an ether. They are produced by platelets (also called thrombocytes) and act in the formation of blood clots and reduction of blood ﬂow to the site of a clot. Nonsteroidal anti-inﬂammatory drugs (NSAIDs) — aspirin, ibuprofen, and meclofenamate, for example — inhibit the enzyme cyclooxygenase or COX (also called prostaglandin H2 synthase), which catalyzes an early step in the pathway from arachidonate to series 2 prostaglandins and thromboxanes (Fig. 10-17) and from EPA to series 3 prostaglandins and thromboxanes (see Fig. 21-12). Leukotrienes (LT), ﬁrst found in leukocytes, contain three conjugated double bonds. They are powerful biological signals. For example, leukotriene D4, derived from leukotriene A4, induces contraction of the smooth muscle lining the airways to the lung. Overproduction of leukotrienes causes asthmatic attacks, and leukotriene synthesis is one target of antiasthmatic drugs such as prednisone. The strong contraction of the smooth muscle of the lungs that occurs during anaphylactic shock is part of the potentially fatal allergic reaction in individuals hypersensitive to bee stings, penicillin, or other agents. Lipoxins (LX), like leukotrienes, are linear eicosanoids. Their distinguishing feature is the presence of several hydroxyl groups along the chain (Fig. 10-17). These compounds are potent anti- inﬂammatory agents. Because their synthesis is stimulated by low doses (81 mg) of aspirin taken daily, this low dose is commonly prescribed for individuals with cardiovascular disease. Steroid Hormones Carry Messages between Tissues Steroids are oxidized derivatives of sterols; they have the sterol nucleus but lack the alkyl chain attached to ring D of cholesterol, and they are more polar than cholesterol. Steroid hormones move through the bloodstream (on protein carriers) from their site of production to target tissues, where they enter cells, bind to highly speciﬁc receptor proteins in the nucleus, and trigger changes in gene expression and thus metabolism. Because hormones have very high afﬁnity for their receptors, very low concentrations of hormones (nanomolar or less) are sufﬁcient to produce responses in target tissues. The major groups of steroid hormones are the male and female sex hormones and the hormones produced by the adrenal cortex, cortisol and aldosterone (Fig. 10-18). Prednisone is a steroid drug with strong anti-inﬂammatory activity, mediated in part by the inhibition of arachidonate release by phospholipase A2 and consequent inhibition of the synthesis of prostaglandins, thromboxanes, leukotrienes, and lipoxins. Prednisone and similar drugs have a variety of medical applications, including the treatment of asthma and rheumatoid arthritis.

FIGURE 10-18 Steroids derived from cholesterol. Vascular plants contain the steroidlike brassinolide (Fig. 10-18), a potent growth regulator that increases the rate of stem elongation and affects the orientation of cellulose microﬁbrils in the cell wall during growth. Vascular Plants Produce Thousands of Volatile Signals Plants produce thousands of different lipophilic compounds, volatile substances that are used to attract pollinators, repel herbivores, attract organisms that defend the plant against herbivores, and communicate with other plants. Jasmonate, for example, derived from the fatty acid 18:3(Δ9,12,15) in membrane lipids, triggers the plant’s defenses in response to insect-inﬂicted damage. The methyl ester of jasmonate gives the characteristic fragrance of jasmine oil, which is widely used in the perfume industry. Many plant volatiles, including geraniol (the characteristic scent of geraniums), β -pinene (pine trees), limonene (limes), and menthol, are derived from fatty acids or from compounds made by the condensation of ﬁve-carbon isoprene units. Vitamins A and D Are Hormone Precursors During early decades of the twentieth century, a major focus of research in physiological chemistry was the identiﬁcation of vitamins, compounds that are essential to the health of humans and other vertebrates but cannot be synthesized by these animals and must therefore be obtained in the diet. Early nutritional studies identiﬁed two general classes of such compounds: those soluble in nonpolar organic solvents (fat-soluble vitamins) and those that could be extracted from foods with aqueous solvents (water-soluble vitamins). Eventually, the fat-soluble group was resolved into the four vitamin groups A, D, E, and K, all of which are isoprenoid compounds synthesized by the condensation of multiple isoprene units. Two of these (D and A) serve as hormone precursors. Vitamin D3, also called cholecalciferol, is normally formed in the skin from 7-dehydrocholesterol in a photochemical reaction driven by the UV component of sunlight (Fig. 10-19a). Vitamin D3 is not itself biologically active, but it is converted by enzymes in the liver and kidney to 1α ,25-dihydroxyvitamin D3 (calcitriol), a hormone that regulates calcium uptake in the intestine and calcium levels in kidney and bone. Deﬁciency of vitamin D leads to defective bone formation and the disease rickets (Fig. 10-19b), for which administration of vitamin D produces a dramatic cure. Vitamin D2 (ergocalciferol) is a commercial product formed by UV irradiation of the ergosterol of yeast. Vitamin D2 is structurally similar to D3, with slight modiﬁcation to the side chain attached to the sterol D ring. Both have the same biological effects, and D2 is commonly added to milk and butter as a dietary supplement. The product of vitamin D metabolism, 1α ,25- dihydroxyvitamin D3, regulates gene expression by interacting with speciﬁc nuclear receptor proteins. We discuss such regulation of gene expression in more detail in Chapter 28. FIGURE 10-19 Vitamin D3 production and metabolism. (a) Cholecalciferol (vitamin D3) is produced in the skin by UV irradiation of 7-dehydrocholesterol, which breaks the bond shaded light red. In the liver, a hydroxyl group is added at C-25; in the kidney, a second hydroxylation at C-1 produces the active hormone, 1α ,25-dihydroxyvitamin D3. This hormone regulates the metabolism of Ca2+ in kidney, intestine, and bone. (b) Dietary vitamin D prevents rickets, a disease once common in cold climates where heavy clothing blocks the UV component of sunlight necessary for the production of vitamin D3 in skin. Rickets results in weak or so bones in children; it can o en be identified by bowed legs and other bone deformities. Vitamin A1 (all-trans-retinol) and its oxidized metabolites retinoic acid and retinal act in the processes of development, cell growth and differentiation, and vision (Fig. 10-20). Vitamin A1 or β -carotene in the diet can be converted enzymatically to all- trans-retinoic acid, a retinoid hormone that acts through a family of nuclear receptor proteins (RAR, RXR, PPAR) to regulate gene expression central to embryonic development, stem cell differentiation, and cell proliferation. All-trans-retinoic acid is used to treat certain types of leukemia, and it is the active ingredient in the drug tretinoin (Retin-A), used to treat severe acne and wrinkled skin. In the vertebrate eye, retinal bound to the protein opsin forms the photoreceptor pigment rhodopsin. The photochemical conversion of 11-cis-retinal to all-trans-retinal is the fundamental event in vision (see Fig. 12-19). FIGURE 10-20 Dietary β -carotene and vitamin A1 as precursors of the retinoids. (a) β -Carotene is shown with its isoprene structural units set oﬀ by dashed red lines. Symmetric cleavage of β -carotene yields two molecules of all-trans-retinal (b), which can be either further oxidized to all- trans-retinoic acid, a retinoid hormone (c), or reduced to all-trans-retinol, vitamin A1 (d). In the visual pathway, all-trans-retinol from this reaction, or obtained directly through the diet, can be converted to the aldehyde 11-cis- retinal (e). This product combines with the protein opsin to form rhodopsin (not shown), a visual pigment widespread in nature. In the dark, the retinal of rhodopsin is in the 11-cis form. When a rhodopsin molecule is excited by visible light, the 11-cis-retinal undergoes a series of photochemical reactions that convert it to all-trans-retinal (f), forcing a change in the shape of the entire rhodopsin molecule. This transformation in the rod cell of the vertebrate retina sends an electrical signal to the brain that is the basis of visual transduction. Unlike most vitamins, vitamin A can be stored for some time in the body (primarily as its ester with palmitic acid, in the liver). Vitamin A was ﬁrst isolated from ﬁsh liver oils; eggs, whole milk, and butter are also good dietary sources. Another source is β - carotene (Fig. 10-20), the pigment that gives carrots, sweet potatoes, and other yellow vegetables their characteristic color. Carotene is one of a very large number (>700) of carotenoids, natural products with a characteristic extensive system of conjugated double bonds, which makes possible their strong absorption of visible light (450–470 nm). Vitamin A deﬁciency in a pregnant woman can lead to congenital malformations and growth retardation in the infant. In adults, vitamin A is also essential to vision, immunity, and reproduction. Deﬁciency of vitamin A leads to a variety of symptoms, including dryness of the skin, eyes, and mucous membranes, and night blindness, an early symptom commonly used in diagnosing vitamin A deﬁciency. In the developing world, vitamin A deﬁciency causes an estimated 250,000 or more cases of blindness or death in children each year. One effective strategy for providing vitamin A is the metabolic engineering of rice strains to overproduce β -carotene. Rice has all the enzymatic machinery to produce β -carotene in its leaves, but these enzymes are less active in the grain. Introduction of two genes into the rice has resulted in “golden rice” with grains enriched in β -carotene (Fig. 10-21). FIGURE 10-21 Carotene-enriched rice. Worldwide, more than 250 million children and pregnant women suﬀer from vitamin A deficiency, which causes at least 250,000 cases of irreversible blindness each year in children. Half of these children end up dying within a year of losing their sight. This deficiency is particularly prevalent where rice is a staple food. An international humanitarian eﬀort — the Golden Rice Project — has made great strides in addressing this health crisis. Wild-type rice grains (le ) do not produce β -carotene, the metabolic precursor of vitamin A. Rice plants have been genetically engineered to produce β -carotene in the grain, which takes on the yellow color of the carotene (right). A diet supplemented with Golden Rice provides enough β -carotene to prevent vitamin A deficiency and its tragic health consequences. Vitamins E and K and the Lipid Quinones Are Oxidation-Reduction Cofactors Vitamin E is the collective name for a group of closely related lipids called tocopherols, all of which contain a substituted aromatic ring and a long isoprenoid side chain (Fig. 10-22a). Because they are hydrophobic, tocopherols associate with cell membranes, lipid deposits, and lipoproteins in the blood. Tocopherols are biological antioxidants. The aromatic ring reacts with and destroys the most reactive forms of oxygen radicals and other free radicals, protecting unsaturated fatty acids from oxidation and preventing oxidative damage to membrane lipids, which can cause cell fragility. Tocopherols are found in eggs and vegetable oils and are especially abundant in wheat germ. Laboratory animals fed diets depleted of vitamin E develop scaly skin, muscular weakness and wasting, and sterility. Vitamin E deﬁciency in humans is very rare; the principal symptom is fragile erythrocytes. FIGURE 10-22 Some other biologically active isoprenoid compounds or derivatives. Units derived from isoprene are set oﬀ by dashed red lines. In most mammalian tissues, ubiquinone (also called coenzyme Q) has 10 isoprene units. Dolichols of animals have 17 to 21 isoprene units (85 to 105 carbon atoms), bacterial dolichols have 11, and dolichols of plants and fungi have 14 to 24. The aromatic ring of vitamin K (Fig. 10-22b) undergoes a cycle of oxidation and reduction during the formation of active prothrombin, a blood plasma protein essential in blood clotting. Prothrombin is a proteolytic enzyme that splits peptide bonds in the blood protein ﬁbrinogen to convert it to ﬁbrin, the insoluble ﬁbrous protein that holds blood clots together (see Fig. 6-43). Vitamin K deﬁciency, which slows blood clotting, is extremely uncommon in humans, aside from a small percentage of infants who suffer from hemorrhagic disease of the newborn, a potentially fatal disorder. In the United States, newborns are routinely given a 1 mg injection of vitamin K. Vitamin K1 (phylloquinone) is found in green plant leaves; a related form, vitamin K2 (menaquinone), is formed by bacteria living in the vertebrate intestine. Warfarin (Fig. 10-22c) is a synthetic compound that inhibits the formation of active prothrombin. It is particularly poisonous to rats, causing death by internal bleeding. Ironically, this potent rodenticide is also an invaluable anticoagulant drug for treating humans at risk for excessive blood clotting, such as surgical patients and those with coronary thrombosis. Ubiquinone (also called coenzyme Q) and plastoquinone (Fig. 10- 22d, e) are isoprenoids that function as lipophilic electron carriers in the oxidation-reduction reactions that drive ATP synthesis in mitochondria and chloroplasts, respectively. Both ubiquinone and plastoquinone can accept either one or two electrons and either one or two protons (see Fig. 19-3). Dolichols Activate Sugar Precursors for Biosynthesis During assembly of the complex carbohydrates of bacterial cell walls, and during the addition of polysaccharide units to certain proteins (glycoproteins) and lipids (glycolipids) in eukaryotes, the sugar units to be added are chemically activated by attachment to isoprenoid alcohols called dolichols (Fig. 10-22f). These compounds have strong hydrophobic interactions with membrane lipids, anchoring the attached sugars to the membrane, where they participate in sugar-transfer reactions. Many Natural Pigments Are Lipidic Conjugated Dienes Conjugated dienes have carbon chains with alternating single and double bonds. Because this structural arrangement allows the delocalization of electrons, the compounds can be excited by low- energy electromagnetic radiation (visible light), giving them colors visible to humans and other animals. Carotene (Fig. 10-20) is yellow-orange; similar compounds give bird feathers their striking reds, oranges, and yellows (Fig. 10-23). Like sterols, steroids, dolichols, vitamins A, E, D, and K, ubiquinone, and plastoquinone, these pigments are synthesized from ﬁve-carbon isoprene derivatives; the biosynthetic pathway is described in detail in Chapter 21. FIGURE 10-23 Lipids as pigments in plants and bird feathers. Compounds with long conjugated systems absorb light in the visible region of the spectrum. Subtle diﬀerences in the chemistry of these compounds produce pigments of strikingly diﬀerent colors. Birds acquire the pigments that color their feathers red or yellow by eating plant materials that contain carotenoid pigments, such as canthaxanthin and zeaxanthin. The diﬀerences in pigmentation between male and female birds are the result of diﬀerences in intestinal uptake and processing of carotenoids. Polyketides Are Natural Products with Potent Biological Activities Polyketides are a diverse group of lipids with biosynthetic pathways (Claisen condensations) similar to those for fatty acids. They are secondary metabolites, compounds that are not central to an organism’s metabolism but serve some subsidiary function that gives the organism an advantage in some ecological niche. Many polyketides ﬁnd use in medicine as antibiotics (erythromycin), antifungals (amphotericin B), or inhibitors of cholesterol synthesis (lovastatin) (Fig. 10-24).

FIGURE 10-24 Three polyketide natural products used in human medicine. SUMMARY 10.3 Lipids as Signals, Cofactors, and Pigments Some types of lipids, although present in relatively small quantities, play critical roles as cofactors or signals. Membrane sphingolipids and derivatives of phosphatidylinositol such as PIP2 and PIP3 are used as signaling molecules or for nucleating formation of multiprotein complexes. Prostaglandins, thromboxanes, leukotrienes, and lipoxins, all of which are eicosanoids derived from arachidonate, are extremely potent hormones involved in reproduction, inﬂammation, regulating blood pressure, and other bodily processes. NSAIDs inhibit formation of some prostaglandins and thromboxanes. Steroid hormones, such as the sex hormones, are derived from sterols. They serve as powerful biological signals and move through the bloodstream to alter gene expression in target cells. Plants produce volatile lipids to attract or repel other organisms and for communication. Many of these lipids are used as fragrances in perfumes. Vitamins D, A, E, and K are fat-soluble compounds made up of isoprene units. All play essential roles in the metabolism or physiology of animals. Vitamin D is precursor to a hormone that regulates calcium metabolism. Vitamin A furnishes the visual pigment of the vertebrate eye and is a regulator of gene expression during epithelial cell growth. Vitamins E and K and quinones can be oxidized or reduced by cells. Vitamin E functions in the protection of membrane lipids from oxidative damage, and vitamin K is essential for blood clotting. Quinones are essential for carrying electrons during the reactions that drive ATP synthesis in mitochondria and chloroplasts. Dolichols activate and anchor sugars to cellular membranes; the sugar groups are then used in the synthesis of complex carbohydrates, glycolipids, and glycoproteins. Lipidic conjugated dienes serve as pigments in ﬂowers and fruits and give bird feathers their striking colors. Polyketides are natural products widely used in medicine. 10.4 Working with Lipids Because lipids are insoluble in water, their extraction and subsequent fractionation require the use of organic solvents and some techniques not commonly used in the puriﬁcation of water- soluble molecules such as proteins and carbohydrates. In general, complex mixtures of lipids are separated by differences in polarity or solubility in nonpolar solvents. Lipids that contain ester- or amide-linked fatty acids can be hydrolyzed by treatment with acid or alkali or with speciﬁc hydrolytic enzymes (phospholipases, glycosidases) to yield their components for analysis. Some methods commonly used in lipid analysis are shown in Figure 10-25 and discussed below. FIGURE 10-25 Common procedures in the extraction, separation, and identification of cellular lipids. (a) Tissue is homogenized in a chloroform/methanol/water mixture, which upon addition of water and removal of unextractable sediment by centrifugation yields two phases. (b) Major classes of extracted lipids in the chloroform phase may first be separated by thin-layer chromatography (TLC), in which lipids are carried up a silica gel–coated plate by a rising solvent front, with less-polar lipids traveling farther than more-polar or charged lipids, or by adsorption chromatography on a column of silica gel, through which solvents of increasing polarity are passed. For example, column chromatography with appropriate solvents can be used to separate closely related lipid species such as phosphatidylserine, phosphatidylglycerol, and phosphatidylinositol. Once separated, each lipid’s complement of fatty acids can be determined by mass spectrometry. (c) Alternatively, in the “shotgun” approach, an unfractionated extract of lipids can be directly subjected to high-resolution mass spectrometry of diﬀerent types and under diﬀerent conditions to determine the total composition of all the lipids — that is, the lipidome. Lipid Extraction Requires Organic Solvents Neutral lipids (triacylglycerols, waxes, pigments, and so forth) are readily extracted from tissues with ethyl ether, chloroform, or benzene, solvents that do not permit lipid clustering driven by the hydrophobic effect. Membrane lipids are more effectively extracted by more polar organic solvents, such as ethanol or methanol, which reduce the hydrophobic interactions among lipid molecules while also weakening the hydrogen bonds and electrostatic interactions that bind membrane lipids to membrane proteins. A commonly used extractant is a mixture of chloroform, methanol, and water, initially in volume proportions (1:2:0.8) that are miscible, producing a single phase. Aer tissue is homogenized in this solvent to extract all lipids, more water is added to the resulting extract, and the mixture separates into two phases: methanol/water (top phase) and chloroform (bottom phase). The lipids remain in the chloroform layer, and the more polar molecules such as proteins and sugars partition into the methanol/water layer (Fig. 10-25a). Adsorption Chromatography Separates Lipids of Diﬀerent Polarity Complex mixtures of tissue lipids can be fractionated by chromatographic procedures based on the different polarities of each class of lipid (Fig. 10-25b). In adsorption chromatography, an insoluble, polar material such as silica gel (a form of silicic acid, Si(OH)4) is packed into a glass column, and the lipid mixture (in chloroform solution) is applied to the top of the column. (In high- performance liquid chromatography, or HPLC, the column is of smaller diameter and solvents are forced through the column under high pressure.) The polar lipids bind tightly to the polar silicic acid, but the neutral lipids pass directly through the column and emerge in the ﬁrst chloroform wash. The polar lipids are then eluted, in order of increasing polarity, by washing the column with solvents of progressively higher polarity. Uncharged but polar lipids (cerebrosides, for example) are eluted with acetone, and very polar or charged lipids (such as glycerophospholipids) are eluted with methanol. Thin-layer chromatography (TLC) on silicic acid employs the same principle (Fig. 10-25b). A thin layer of silica gel is spread onto a glass plate, to which it adheres. A small sample of lipids dissolved in chloroform is applied near one edge of the plate, which is dipped in a shallow container of an organic solvent or solvent mixture; the entire setup is enclosed in a chamber saturated with the solvent vapor. As the solvent rises on the plate by capillary action, it carries lipids with it. The less polar lipids move farthest, as they have less tendency to bind to the silicic acid. The separated lipids can be detected by spraying the plate with a dye (rhodamine) that ﬂuoresces when associated with lipids, or by exposing the plate to iodine fumes. Iodine reacts reversibly with the double bonds in fatty acids, such that lipids containing unsaturated fatty acids develop a yellow or brown color. Several other spray reagents are also useful in detecting speciﬁc lipids. For subsequent analysis, regions containing separated lipids can be scraped from the plate and the lipids recovered by extraction with an organic solvent. Gas Chromatography Resolves Mixtures of Volatile Lipid Derivatives Gas chromatography (GC) separates volatile components of a mixture according to their relative tendencies to dissolve in the inert material packed in the chromatography column or to volatilize and move through the column, carried by a current of an inert gas such as helium. Some lipids are naturally volatile, but most must ﬁrst be derivatized to increase their volatility (that is, lower their boiling point). For an analysis of the fatty acids in a sample of phospholipids, the lipids are ﬁrst transesteriﬁed: heated in a methanol/HCl or methanol/NaOH mixture to convert fatty acids esteriﬁed to glycerol into their methyl esters. These fatty acyl methyl esters are then loaded onto the GC column, and the column is heated to volatilize the compounds. Those fatty acyl esters that are most soluble in the column material will partition into (dissolve in) that material; the less-soluble lipids are carried by the stream of inert gas and emerge ﬁrst from the column. The order of elution depends on the nature of the solid adsorbent in the column and on the boiling point of the components of the lipid mixture. With these techniques, mixtures of fatty acids of various chain lengths and various degrees of unsaturation can be completely resolved. Specific Hydrolysis Aids in Determination of Lipid Structure Certain classes of lipids are susceptible to degradation under speciﬁc conditions. For example, all ester-linked fatty acids in triacylglycerols, phospholipids, and sterol esters are released by mild acid or alkaline treatment, and somewhat harsher hydrolysis conditions release amide-bound fatty acids from sphingolipids. Enzymes that speciﬁcally hydrolyze certain lipids are also useful in the determination of lipid structure. Phospholipases A, C, and D (Fig. 10-14) each split particular bonds in phospholipids and yield products with characteristic solubilities and chromatographic behaviors. Phospholipase C, for example, releases a water-soluble phosphoryl alcohol (such as phosphocholine from phosphatidylcholine) and a chloroform- soluble diacylglycerol, each of which can be characterized separately to determine the structure of the intact phospholipid. The combination of speciﬁc hydrolysis with characterization of the products by TLC, GC, or HPLC oen allows determination of a lipid structure. Mass Spectrometry Reveals Complete Lipid Structure To establish unambiguously the length of a hydrocarbon chain or the position of double bonds, mass spectrometric analysis of lipids or their volatile derivatives is invaluable. The chemical properties of similar lipids (for example, two fatty acids of similar length unsaturated at different positions, or two isoprenoids with different numbers of isoprene units) are very much alike, and their order of elution from the various chromatographic procedures oen does not distinguish between them. When the eluate from a chromatography column is sampled by mass spectrometry, however, the components of a lipid mixture can be simultaneously separated and identiﬁed by their unique pattern of fragmentation (Fig. 10-26). With the increased resolution of mass spectrometry, it is possible to identify individual lipids in very complex mixtures without ﬁrst fractionating the lipids in a crude extract. This “shotgun” method (Fig. 10-25c) avoids losses during the preliminary separation of lipid subclasses, and it is faster. FIGURE 10-26 Determination of fatty acid structure by mass spectrometry. The fatty acid is first converted to a derivative that minimizes migration of the double bonds when the molecule is fragmented by electron bombardment. The derivative shown here is a picolinyl ester of linoleic acid — 18:2(Δ9,12) (Mr 371) — in which the alcohol is picolinol (red). When bombarded with a stream of electrons, this molecule is volatilized and converted to a parent ion (M +; Mr 371), in which the N atom bears the positive charge, and a series of smaller fragments produced by breakage of C— C bonds in the fatty acid. The mass spectrometer separates these charged fragments according to their mass/charge ratio (m/z). The prominent ions at m/z= 92,108,151, and 164 contain the pyridine ring of the picolinol and various fragments of the carboxyl group, showing that the compound is indeed a picolinyl ester. The molecular ion, M + (m/z= 371), confirms the presence of a C18 fatty acid with two double bonds. The uniform series of ions 14 atomic mass units (u) apart represents loss of each successive methyl and methylene group from the methyl end of the acyl chain (beginning at C-18; the right end of the molecule as shown here), until the ion at m/z= 300 is reached. This is followed by a gap of 26 u for the carbons of the terminal double bond, at m/z= 274; a further gap of 14 u for the C-11 methylene group, at m/z= 260; and so forth. By this means, the entire structure is determined, although these data alone do not reveal the configuration (cis or trans) of the double bonds. [Information from W. W. Christie, Lipid Technol. 8:64, 1996.] Lipidomics Seeks to Catalog All Lipids and Their Functions As lipid biochemists have become aware of the thousands of different naturally occurring lipids, they have created a database analogous to the Protein Data Bank. The LIPID MAPS Lipidomics Gateway (www.lipidmaps.org) has its own classiﬁcation system that places each lipid species in one of eight chemical categories, each designated by two letters (Table 10-2). Within each category, ﬁner distinctions are indicated by numbered classes and subclasses. For example, all glycerophosphocholines are GP01. The subgroup of glycerophosphocholines with two fatty acids in ester linkage is designated GP0101; the subgroup with one fatty acid ether-linked at position 1 and one ester-linked at position 2 is GP0102. The speciﬁc fatty acids are designated by numbers that give every lipid its own unique identiﬁer, so that each individual lipid, including lipid types not yet discovered, can be unambiguously described in terms of a 12-character identiﬁer, the LM_ID. One factor used in this classiﬁcation system is the nature of the biosynthetic precursor. For example, prenol lipids (such as dolichols and vitamins E and K) are formed from isoprenyl precursors. TABLE 10-2 Eight Major Categories of Biological Lipids Category Category code Examples Fatty acids FA Oleate, stearoyl-CoA, palmitoylcarnitine Glycerolipids GL Di- and triacylglycerols Glycerophospholipids GP Phosphatidylcholine, phosphatidylserine, phosphatidyethanoloamine Sphingolipids SP Sphingomyelin, ganglioside GM2 Sterol lipids ST Cholesterol, progesterone, bile acids Prenol lipids PR Farnesol, geraniol, retinol, ubiquinone Saccharolipids SL Lipopolysaccharide Polyketides PK Tetracycline, erythromycin, aflatoxin B1 The eight chemical categories in Table 10-2 do not coincide perfectly with the less formal categorization according to biological function that we have used in this chapter. For example, the structural lipids of membranes include both glycerophospholipids and sphingolipids, which are separate categories in Table 10-2. Each method of classiﬁcation has its advantages. The application of mass spectrometric techniques with high throughput and high resolution can provide quantitative catalogs of all the lipids present in a speciﬁc cell type under particular conditions — the lipidome — and of the ways in which the lipidome changes with differentiation, disease such as cancer, or drug treatment. An animal cell contains more than a thousand different lipid species, each presumably having a speciﬁc function. These functions are known for a growing number of lipids, but the still largely unexplored lipidome offers a rich source of new problems for the next generation of biochemists and cell biologists to solve. SUMMARY 10.4 Working with Lipids In the determination of lipid composition, lipids can ﬁrst be extracted from tissues with organic solvents. Lipids in mixtures can be separated on the basis of their polarity and interactions with polar materials such as silica, using adsorption chromatography methods such as HPLC or TLC. GC volatilizes lipids so that they can be carried by a stream of inert gas and resolved based on their ability to partition into a soluble column material. Phospholipases speciﬁc for one of the bonds in a phospholipid can be used to generate simpler compounds for subsequent analysis. High-resolution mass spectrometry allows the analysis of crude mixtures of lipids without prefractionation — the “shotgun” approach. Lipidomics combines powerful analytical techniques to determine the full complement of lipids in a cell or a tissue (the lipidome) and to assemble annotated databases that allow comparisons between lipids of different cell types and under different conditions. Chapter Review KEY TERMS Terms in bold are deﬁned in the glossary. fatty acid polyunsaturated fatty acid (PUFA) omega-3 (ω -3) fatty acids triacylglycerol lipases phospholipid sterols glycerophospholipid ether lipid plasmalogen glycolipid galactolipid sphingolipid ceramide sphingomyelin glycosphingolipid cerebroside globoside ganglioside sterol cholesterol bile acids eicosanoid prostaglandin (PG) thromboxane (TX) leukotriene (LT) lipoxin (LX) vitamin vitamin D3 cholecalciferol vitamin A1 (all-trans-retinol) carotenoids vitamin E tocopherol vitamin K dolichol polyketide lipidome PROBLEMS 1. Operational Deﬁnition of Lipids How is the deﬁnition of “lipid” different from the types of deﬁnitions used for other biomolecules such as amino acids, nucleic acids, and proteins? 2. Structure of an Omega-3 Fatty Acid The omega-3 fatty acid docosahexaenoic acid (DHA, 22:6(Δ4,7,10,13,16,19)) is the most abundant omega-3 fatty acid in the brain and an important component of breast milk. Draw the structure of this fatty acid. 3. Melting Points of Lipids The melting points of a series of 18-carbon fatty acids are stearic acid, 69.6 °C; oleic acid, 13.4 °C; linoleic acid, −5 °C; and linolenic acid, −11 °C. a. What structural aspect of these 18-carbon fatty acids can be correlated with the melting point? b. Draw all the possible triacylglycerols that can be constructed from glycerol, palmitic acid, and oleic acid. Rank them in order of increasing melting point. c. Branched-chain fatty acids are found in some bacterial membrane lipids. Would their presence increase or decrease the ﬂuidity of the membrane (that is, give the lipids a lower or a higher melting point)? Why? 4. Catalytic Hydrogenation of Vegetable Oils Catalytic hydrogenation, used in the food industry, converts double bonds in the fatty acids of the oil triacylglycerols to — CH2— CH2— . How does this affect the physical properties of the oils? 5. Impermeability of Waxes What property of the waxy cuticles that cover plant leaves makes the cuticles impermeable to water? 6. Naming Lipid Stereoisomers Carvone, a member of the terpenoid family of chemicals, forms two enantiomers with quite different properties. One enantiomer, abundant in spearmint, smells sweet and minty. The other enantiomer, abundant in caraway seeds, smells spicy and of rye bread. Name the compounds abundant in spearmint and caraway seeds using the RS system. 7. Chemical Reactivity of Lipids Soaps are salts of fatty acids and can be made by mixing triacylglycerols with a strong base such as NaOH. This saponiﬁcation reaction produces glycerol and fatty acid salts. In a lab experiment, students saponify the triacylglycerol tristearin in the presence of 18O-labeled water. What saponiﬁcation reaction products will contain the 18O label? 8. Hydrophobic and Hydrophilic Components of Membrane Lipids A common structural feature of membrane lipids is their amphipathic nature. For example, in phosphatidylcholine, the two fatty acid chains are hydrophobic and the phosphocholine head group is hydrophilic. Name the components that serve as the hydrophobic and hydrophilic units for each membrane lipid: a. phosphatidylethanolamine b. sphingomyelin c. galactosylcerebroside d. ganglioside e. cholesterol. 9. Deducing Lipid Structure from Composition A biochemist completely digests a glycerophospholipid with a mixture of phospholipases A and D. HPLC and MS analysis reveals the presence of an amino acid of 105.09 Da, a saturated fatty acid of 256.43 Da, and an omega-3 monounsaturated fatty acid of 282.45 Da. Which amino acid does the glycerophospholipid contain? Draw the most likely structure of this glycerophospholipid. 10. Deducing Lipid Structure from Molar Ratio of Components Complete hydrolysis of a glycerophospholipid yields glycerol, two fatty acids (16:1(Δ9) and 16:0), phosphoric acid, and serine in the molar ratio 1:1:1:1:1. Name this lipid and draw its structure. 11. Lipids in Blood Group Determination We note in Figure 10-13 that the structure of glycosphingolipids determines the blood groups A, B, and O in humans. It is also true that glycoproteins determine blood groups. How can both statements be true? 12. The Action of Phospholipases The venom of the Eastern diamondback rattler and the Indian cobra contains phospholipase A2, which catalyzes the hydrolysis of fatty acids at the C-2 position of glycerophospholipids. The phospholipid breakdown product of this reaction is lysolecithin, which is derived from phosphatidylcholine. At high concentrations, this and other lysophospholipids act as detergents, dissolving the membranes of erythrocytes and lysing the cells. Extensive hemolysis may be life-threatening. a. All detergents are amphipathic. What are the hydrophilic and hydrophobic portions of lysolecithin? b. The pain and inﬂammation caused by a snake bite can be treated with certain steroids. What is the basis of this treatment? c. Though the high levels of phospholipase A2 in venom can be deadly, this enzyme is necessary for a variety of normal metabolic processes. What are these processes? 13. Intracellular Messengers from Phosphatidylinositols The hormone vasopressin is an extracellular signal that activates a speciﬁc phospholipase C in the membrane. Cleavage of PIP2 by phospholipase C generates two products. What are they? Compare their properties and their solubilities in water, and predict whether either would diffuse readily through the cytosol. 14. Isoprene Units in Isoprenoids Geraniol, farnesol, and squalene are called isoprenoids because they are synthesized from ﬁve-carbon isoprene units. In each compound, circle the ﬁve-carbon units representing isoprene units (see Fig. 10-22). 15. Hydrolysis of Lipids Name the products of mild hydrolysis with dilute NaOH of a. 1-stearoyl-2,3-dipalmitoylglycerol b. 1-palmitoyl-2-oleoylphosphatidylcholine. 16. Effect of Polarity on Solubility Rank a triacylglycerol, a diacylglycerol, and a monoacylglycerol in order of decreasing solubility in water. Assume that each acylglycerol contains only palmitic acid. 17. Chromatographic Separation of Lipids Suppose that you apply a mixture of lipids to a silica gel column and then wash the column with increasingly polar solvents. The mixture consists of phosphatidylserine, phosphatidylethanolamine, phosphatidylcholine, cholesteryl palmitate (a sterol ester), sphingomyelin, palmitate, n-tetradecanol, triacylglycerol, and cholesterol. In what order will the lipids elute from the column? Explain your reasoning. 18. Identiﬁcation of Unknown Lipids Johann Thudichum, who practiced medicine in London about 100 years ago, also dabbled in lipid chemistry in his spare time. He isolated a variety of lipids from neural tissue and characterized and named many of them. His carefully sealed and labeled vials of isolated lipids were rediscovered many years later. a. How would you conﬁrm, using techniques not available to Thudichum, that the vials labeled “sphingomyelin” and “cerebroside” actually contain these compounds? b. How would you distinguish sphingomyelin from phosphatidylcholine by chemical, physical, or enzymatic tests? BIOCHEMISTRY ONLINE 19. Using the LIPID MAPS Database to Find Solubility Information Lipidomics has identiﬁed thousands of cellular lipids. LIPID MAPS is an online database containing over 40,000 unique lipid structures, as well as information on the chemical and physical properties of each lipid (www.lipidmaps.org). One important parameter when working with lipids is log P, where P is the octanol:water partition coefﬁcient, an indicator of lipophilicity. a. Look up cholesterol, sphingosine, linoleic acid, and stearic acid in LIPID MAPS and use the reported log P values to place them in order of increasing solubility in octanol. b. Pharmacologists oen study log P values when developing new drugs. Why would knowing a drug’s log P value be informative? 20. Characteristics of Lipid Transport Proteins Oen when lipids are transported between different tissues, they are carried by proteins. In this exercise, you will explore the interactions between a lipid and a protein using the PDB (www.rcsb.org). Use the PDB identiﬁer 2YG2 and study the structure of the complex between HDL-associated apolipoprotein M and sphingosine-1-phosphate. Navigate to 3D View: Structure to answer the following questions. a. What protein motif is adopted by apolipoprotein M? b. Which amino acid residues do you ﬁnd lining the sphingosine binding pocket? What do they have in common? c. The phosphoryl group of sphingosine-1-phosphate is exposed on the surface of the protein. Why do you suppose it is important that the transport protein binds the hydrocarbon tail of sphingosine-1-phosphate but not necessarily the polar head group? DATA ANALYSIS PROBLEM 21. Determining the Structure of the Abnormal Lipid in Tay-Sachs Disease Box 10-1, Figure 1, shows the pathway of breakdown of gangliosides in healthy (normal) individuals and in individuals with certain genetic diseases. Some of the data on which the ﬁgure is based were presented in a paper by Lars Svennerholm (1962). Note that the sugar Neu5Ac, N- acetylneuraminic acid, represented in the Box 10-1 ﬁgure as , is a sialic acid. Svennerholm reported that “about 90% of the monosialogangliosides isolated from normal human brain” consisted of a compound with ceramide, hexose, N- acetylgalactosamine, and N-acetylneuraminic acid in the molar ratio 1:3:1:1. a. Which of the gangliosides (GM1 through GM3 and globoside) in Box 10-1, Figure 1, ﬁts this description? Explain your reasoning. b. Svennerholm reported that 90% of the gangliosides from a patient with Tay-Sachs disease had a molar ratio (of the same four components given above) of 1:2:1:1. Is this consistent with the Box 10-1 ﬁgure? Explain your reasoning. To determine the structure in more detail, Svennerholm treated the gangliosides with neuraminidase to remove the N-acetylneuraminic acid. This resulted in an asialoganglioside that was much easier to analyze. He hydrolyzed it with acid, collected the ceramide-containing products, and determined the molar ratio of the sugars in each product. He did this for both the normal gangliosides and the Tay-Sachs gangliosides. His results are shown below. Ganglioside Ceramide Glucose Galactose Galactosamine Normal Fragment 1 1 1 0 0 Fragment 2 1 1 1 0 Fragment 3 1 1 1 1 Fragment 4 1 1 2 1 Tay-Sachs Fragment 1 1 1 0 0 Fragment 2 1 1 1 0 Fragment 3 1 1 1 1 c. Based on these data, what can you conclude about the structure of the normal ganglioside? Is this consistent with the structure in Box 10-1? Explain your reasoning. d. What can you conclude about the structure of the Tay- Sachs ganglioside? Is this consistent with the structure in Box 10-1? Explain your reasoning. Svennerholm also reported the work of other researchers who “permethylated” the normal asialoganglioside. Permethylation is the same as exhaustive methylation: a methyl group is added to every free hydroxyl group on a sugar. They found the following permethylated sugars: 2,3,6- trimethylglycopyranose; 2,3,4,6- tetramethylgalactopyranose; 2,4,6- trimethylgalactopyranose; and 4,6-dimethyl-2-deoxy-2- aminogalactopyranose. e. To which sugar of GM1 does each of the permethylated sugars correspond? Explain your reasoning. f. Based on all the data presented so far, what pieces of information about normal ganglioside structure are missing? Reference Svennerholm, L. 1962. The chemical structure of normal human brain and Tay-Sachs gangliosides. Biochem. Biophys. Res. Comm. 9:436–441.

Stems are from the chapter Problems section; correct choices are drawn from Abbreviated Solutions to Problems (Appendix B) in the same edition.

Practice questions (from chapter Problems & Appendix B)Score: 0 / 25

1. Operational Deﬁnition of Lipids How is the deﬁnition of “lipid” different from the types of deﬁnitions used for other biomolecules such as amino acids, nucleic acids, and proteins?

AThe term “lipid” does not specify a particular chemical structure. Compounds are categorized as lipids based on their greater solubility in organic solvents than in water. BHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH) CFirst eluted to last eluted: cholesteryl palmitate and triacylglycerol; cholesterol and n- tetradecanol; phosphatidylcholine and phosphatidylethanolamine; sphingomyelin; phosphatidylserine and palmitate. The lipids elute from the silica gel column in order of polarity. The least polar lipid will elute ﬁrst and the most polar lipid will elute last. Da. Subject acid hydrolysates of each compound to chromatography (GC or silica gel TLC), and compare the result with known standards. Sphingomyelin hydrolysate: sphingosine, fatty acids, phosphocholine, choline, and phosphate; cerebroside hydrolysate: sphingosine, fatty acids, sugars, but no phosphate. b. Strong alkaline hydrolysis of sphingomyelin yields sphingosine; phosphatidylcholine yields glycerol. Detect hydro…

2. Structure of an Omega-3 Fatty Acid The omega-3 fatty acid docosahexaenoic acid (DHA, 22:6(Δ4,7,10,13,16,19)) is the most abundant omega-3 fatty acid in the brain and an important component of breast milk. Draw the structure of this fatty acid.

3. Melting Points of Lipids The melting points of a series of 18-carbon fatty acids are stearic acid, 69.6 °C; oleic acid, 13.4 °C; linoleic acid, −5 °C; and linolenic acid, −11 °C. a. What structural aspect of these 18-carbon fatty acids can be correlated with the melting point? b. Draw all the possible triacylglycerols that can be constructed from glycerol, palmitic acid, and oleic acid. Rank them in order of increasing melting point. c. Branched-chain fatty acids are found in some bacterial membrane lipids. Would their presence increase or decrease the ﬂuidity of the membrane (that is, give the lipids a lower or a higher melting point)? Why?

ASpearmint is (R)-carvone; caraway is (S)-carvone. Ba. Subject acid hydrolysates of each compound to chromatography (GC or silica gel TLC), and compare the result with known standards. Sphingomyelin hydrolysate: sphingosine, fatty acids, phosphocholine, choline, and phosphate; cerebroside hydrolysate: sphingosine, fatty acids, sugars, but no phosphate. b. Strong alkaline hydrolysis of sphingomyelin yields sphingosine; phosphatidylcholine yields glycerol. Detect hydro… Ca. The number of cis double bonds. Each cis double bond causes a bend in the hydrocarbon chain, lowering the melting temperature. b. Six different triacylglycerols can be constructed, in order of increasing melting points: OOO< OOP=OPO< PPO=POP< PPP c. where O = oleic and P = palmitic acid. The greater the content of saturated fatty acid, the higher the melting point. d. Branched-chain fatty acids increase the ﬂuidi… DDiacylglycerol is hydrophobic and remains in the membrane. Inositol 1,4,5-trisphosphate is highly polar, very soluble in water, and more readily diffusible in the cytosol. Both are second messengers.

4. Catalytic Hydrogenation of Vegetable Oils Catalytic hydrogenation, used in the food industry, converts double bonds in the fatty acids of the oil triacylglycerols to — CH2— CH2— . How does this affect the physical properties of the oils?

5. Impermeability of Waxes What property of the waxy cuticles that cover plant leaves makes the cuticles impermeable to water?

ASerine BFirst eluted to last eluted: cholesteryl palmitate and triacylglycerol; cholesterol and n- tetradecanol; phosphatidylcholine and phosphatidylethanolamine; sphingomyelin; phosphatidylserine and palmitate. The lipids elute from the silica gel column in order of polarity. The least polar lipid will elute ﬁrst and the most polar lipid will elute last. CLong, saturated acyl chains, nearly solid at air temperatures, form a hydrophobic layer in which a polar compound such as H2O cannot dissolve or diffuse. DThe part of the membrane lipid that determines blood type is the oligosaccharide in the head group of the membrane sphingolipids (see Fig. 10-13). This same oligosaccharide is attached to certain membrane glycoproteins, which also serve as points of recognition by the antibodies that distinguish blood groups.

6. Naming Lipid Stereoisomers Carvone, a member of the terpenoid family of chemicals, forms two enantiomers with quite different properties. One enantiomer, abundant in spearmint, smells sweet and minty. The other enantiomer, abundant in caraway seeds, smells spicy and of rye bread. Name the compounds abundant in spearmint and caraway seeds using the RS system.

Aa. β barrel b. Phe, Trp, Tyr, Leu. All are hydrophobic or have nonpolar R groups. c. The polar head group can hydrogen bond with water; the hydrocarbon tail cannot. Hydrophobic portions of the residues of the pocket protect the tail from contact with water as it moves through the bloodstream. Ba. GM1 and globoside. Both glucose and galactose are hexoses, so “hexose” in the molar ratio refers to glucose + galactose. The ratios for the four gangliosides are GM1, 1:3:1:1; GM2, 1:2:1:1; GM3, 1:2:0:1; globoside, 1:3:1:0. b. Yes. The ratio matches GM2, the ganglioside expected to build up in Tay-Sachs disease (see Box 10-1, Fig. 1). c. This analysis is similar to that used by Sanger to determine the amino acid … CSpearmint is (R)-carvone; caraway is (S)-carvone. DHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH)

7. Chemical Reactivity of Lipids Soaps are salts of fatty acids and can be made by mixing triacylglycerols with a strong base such as NaOH. This saponiﬁcation reaction produces glycerol and fatty acid salts. In a lab experiment, students saponify the triacylglycerol tristearin in the presence of 18O-labeled water. What saponiﬁcation reaction products will contain the 18O label?

AThe 18O label appears in the fatty acid salts. BSerine CLong, saturated acyl chains, nearly solid at air temperatures, form a hydrophobic layer in which a polar compound such as H2O cannot dissolve or diffuse. DIt reduces double bonds, which increases the melting point of lipids containing the fatty acids.

8. Hydrophobic and Hydrophilic Components of Membrane Lipids A common structural feature of membrane lipids is their amphipathic nature. For example, in phosphatidylcholine, the two fatty acid chains are hydrophobic and the phosphocholine head group is hydrophilic. Name the components that serve as the hydrophobic and hydrophilic units for each membrane lipid: a. phosphatidylethanolamine b. sphingomyelin c. galactosylcerebroside d. ganglioside e. cholesterol.

AFirst eluted to last eluted: cholesteryl palmitate and triacylglycerol; cholesterol and n- tetradecanol; phosphatidylcholine and phosphatidylethanolamine; sphingomyelin; phosphatidylserine and palmitate. The lipids elute from the silica gel column in order of polarity. The least polar lipid will elute ﬁrst and the most polar lipid will elute last. BHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH) Ca. The number of cis double bonds. Each cis double bond causes a bend in the hydrocarbon chain, lowering the melting temperature. b. Six different triacylglycerols can be constructed, in order of increasing melting points: OOO< OOP=OPO< PPO=POP< PPP c. where O = oleic and P = palmitic acid. The greater the content of saturated fatty acid, the higher the melting point. d. Branched-chain fatty acids increase the ﬂuidi… Da. The free —OH group on C-2 and the phosphocholine head group on C-3 are hydrophilic; the fatty acid on C-1 of lysolecithin is hydrophobic. b. Certain steroids, such as prednisone, inhibit the action of phospholipase A2, inhibiting the release of arachidonic acid from C-2. Arachidonic acid is converted to a variety of eicosanoids, some of which cause inﬂammation and pain. c. Phospholipase A2 releases arachidonic ac…

9. Deducing Lipid Structure from Composition A biochemist completely digests a glycerophospholipid with a mixture of phospholipases A and D. HPLC and MS analysis reveals the presence of an amino acid of 105.09 Da, a saturated fatty acid of 256.43 Da, and an omega-3 monounsaturated fatty acid of 282.45 Da. Which amino acid does the glycerophospholipid contain? Draw the most likely structure of this glycerophospholipid.

AHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH) BSerine CThe term “lipid” does not specify a particular chemical structure. Compounds are categorized as lipids based on their greater solubility in organic solvents than in water. Da. β barrel b. Phe, Trp, Tyr, Leu. All are hydrophobic or have nonpolar R groups. c. The polar head group can hydrogen bond with water; the hydrocarbon tail cannot. Hydrophobic portions of the residues of the pocket protect the tail from contact with water as it moves through the bloodstream.

10. Deducing Lipid Structure from Molar Ratio of Components Complete hydrolysis of a glycerophospholipid yields glycerol, two fatty acids (16:1(Δ9) and 16:0), phosphoric acid, and serine in the molar ratio 1:1:1:1:1. Name this lipid and draw its structure.

Aa. GM1 and globoside. Both glucose and galactose are hexoses, so “hexose” in the molar ratio refers to glucose + galactose. The ratios for the four gangliosides are GM1, 1:3:1:1; GM2, 1:2:1:1; GM3, 1:2:0:1; globoside, 1:3:1:0. b. Yes. The ratio matches GM2, the ganglioside expected to build up in Tay-Sachs disease (see Box 10-1, Fig. 1). c. This analysis is similar to that used by Sanger to determine the amino acid … Ba. Subject acid hydrolysates of each compound to chromatography (GC or silica gel TLC), and compare the result with known standards. Sphingomyelin hydrolysate: sphingosine, fatty acids, phosphocholine, choline, and phosphate; cerebroside hydrolysate: sphingosine, fatty acids, sugars, but no phosphate. b. Strong alkaline hydrolysis of sphingomyelin yields sphingosine; phosphatidylcholine yields glycerol. Detect hydro… C DFirst eluted to last eluted: cholesteryl palmitate and triacylglycerol; cholesterol and n- tetradecanol; phosphatidylcholine and phosphatidylethanolamine; sphingomyelin; phosphatidylserine and palmitate. The lipids elute from the silica gel column in order of polarity. The least polar lipid will elute ﬁrst and the most polar lipid will elute last.

11. Lipids in Blood Group Determination We note in Figure 10-13 that the structure of glycosphingolipids determines the blood groups A, B, and O in humans. It is also true that glycoproteins determine blood groups. How can both statements be true?

ASolubility in water: monoacylglycerol > diacylglycerol > triacylglycerol BSpearmint is (R)-carvone; caraway is (S)-carvone. CThe part of the membrane lipid that determines blood type is the oligosaccharide in the head group of the membrane sphingolipids (see Fig. 10-13). This same oligosaccharide is attached to certain membrane glycoproteins, which also serve as points of recognition by the antibodies that distinguish blood groups. DHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH)

12. The Action of Phospholipases The venom of the Eastern diamondback rattler and the Indian cobra contains phospholipase A2, which catalyzes the hydrolysis of fatty acids at the C-2 position of glycerophospholipids. The phospholipid breakdown product of this reaction is lysolecithin, which is derived from phosphatidylcholine. At high concentrations, this and other lysophospholipids act as detergents, dissolving the membranes of erythrocytes and lysing the cells. Extensive hemolysis may be life-threatening. a. All detergents are amphipathic. What are the hydrophilic and hydrophobic portions of lysolecithin? b. The pain and inﬂammation caused by a snake bite can be treated with certain steroids. What is the basis of this treatment? c. Though the high levels of phospholipase A2 in venom can be deadly, this enzyme is necessary for a variety of normal metabolic processes. What are these processes?

AFirst eluted to last eluted: cholesteryl palmitate and triacylglycerol; cholesterol and n- tetradecanol; phosphatidylcholine and phosphatidylethanolamine; sphingomyelin; phosphatidylserine and palmitate. The lipids elute from the silica gel column in order of polarity. The least polar lipid will elute ﬁrst and the most polar lipid will elute last. Ba. The free —OH group on C-2 and the phosphocholine head group on C-3 are hydrophilic; the fatty acid on C-1 of lysolecithin is hydrophobic. b. Certain steroids, such as prednisone, inhibit the action of phospholipase A2, inhibiting the release of arachidonic acid from C-2. Arachidonic acid is converted to a variety of eicosanoids, some of which cause inﬂammation and pain. c. Phospholipase A2 releases arachidonic ac… CHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH) DThe part of the membrane lipid that determines blood type is the oligosaccharide in the head group of the membrane sphingolipids (see Fig. 10-13). This same oligosaccharide is attached to certain membrane glycoproteins, which also serve as points of recognition by the antibodies that distinguish blood groups.

13. Intracellular Messengers from Phosphatidylinositols The hormone vasopressin is an extracellular signal that activates a speciﬁc phospholipase C in the membrane. Cleavage of PIP2 by phospholipase C generates two products. What are they? Compare their properties and their solubilities in water, and predict whether either would diffuse readily through the cytosol.

Aa. β barrel b. Phe, Trp, Tyr, Leu. All are hydrophobic or have nonpolar R groups. c. The polar head group can hydrogen bond with water; the hydrocarbon tail cannot. Hydrophobic portions of the residues of the pocket protect the tail from contact with water as it moves through the bloodstream. BDiacylglycerol is hydrophobic and remains in the membrane. Inositol 1,4,5-trisphosphate is highly polar, very soluble in water, and more readily diffusible in the cytosol. Both are second messengers. CThe term “lipid” does not specify a particular chemical structure. Compounds are categorized as lipids based on their greater solubility in organic solvents than in water. Da. GM1 and globoside. Both glucose and galactose are hexoses, so “hexose” in the molar ratio refers to glucose + galactose. The ratios for the four gangliosides are GM1, 1:3:1:1; GM2, 1:2:1:1; GM3, 1:2:0:1; globoside, 1:3:1:0. b. Yes. The ratio matches GM2, the ganglioside expected to build up in Tay-Sachs disease (see Box 10-1, Fig. 1). c. This analysis is similar to that used by Sanger to determine the amino acid …

14. Isoprene Units in Isoprenoids Geraniol, farnesol, and squalene are called isoprenoids because they are synthesized from ﬁve-carbon isoprene units. In each compound, circle the ﬁve-carbon units representing isoprene units (see Fig. 10-22).

A Ba. The free —OH group on C-2 and the phosphocholine head group on C-3 are hydrophilic; the fatty acid on C-1 of lysolecithin is hydrophobic. b. Certain steroids, such as prednisone, inhibit the action of phospholipase A2, inhibiting the release of arachidonic acid from C-2. Arachidonic acid is converted to a variety of eicosanoids, some of which cause inﬂammation and pain. c. Phospholipase A2 releases arachidonic ac… CThe term “lipid” does not specify a particular chemical structure. Compounds are categorized as lipids based on their greater solubility in organic solvents than in water. Da. GM1 and globoside. Both glucose and galactose are hexoses, so “hexose” in the molar ratio refers to glucose + galactose. The ratios for the four gangliosides are GM1, 1:3:1:1; GM2, 1:2:1:1; GM3, 1:2:0:1; globoside, 1:3:1:0. b. Yes. The ratio matches GM2, the ganglioside expected to build up in Tay-Sachs disease (see Box 10-1, Fig. 1). c. This analysis is similar to that used by Sanger to determine the amino acid …

15. Hydrolysis of Lipids Name the products of mild hydrolysis with dilute NaOH of a. 1-stearoyl-2,3-dipalmitoylglycerol b. 1-palmitoyl-2-oleoylphosphatidylcholine.

Aa. The number of cis double bonds. Each cis double bond causes a bend in the hydrocarbon chain, lowering the melting temperature. b. Six different triacylglycerols can be constructed, in order of increasing melting points: OOO< OOP=OPO< PPO=POP< PPP c. where O = oleic and P = palmitic acid. The greater the content of saturated fatty acid, the higher the melting point. d. Branched-chain fatty acids increase the ﬂuidi… Ba. Glycerol and the sodium salts of palmitic and stearic acids b. D-Glycerol 3-phosphocholine and the sodium salts of palmitic and oleic acids CThe term “lipid” does not specify a particular chemical structure. Compounds are categorized as lipids based on their greater solubility in organic solvents than in water. DFirst eluted to last eluted: cholesteryl palmitate and triacylglycerol; cholesterol and n- tetradecanol; phosphatidylcholine and phosphatidylethanolamine; sphingomyelin; phosphatidylserine and palmitate. The lipids elute from the silica gel column in order of polarity. The least polar lipid will elute ﬁrst and the most polar lipid will elute last.

16. Effect of Polarity on Solubility Rank a triacylglycerol, a diacylglycerol, and a monoacylglycerol in order of decreasing solubility in water. Assume that each acylglycerol contains only palmitic acid.

Aa. Glycerol and the sodium salts of palmitic and stearic acids b. D-Glycerol 3-phosphocholine and the sodium salts of palmitic and oleic acids BSolubility in water: monoacylglycerol > diacylglycerol > triacylglycerol Ca. The free —OH group on C-2 and the phosphocholine head group on C-3 are hydrophilic; the fatty acid on C-1 of lysolecithin is hydrophobic. b. Certain steroids, such as prednisone, inhibit the action of phospholipase A2, inhibiting the release of arachidonic acid from C-2. Arachidonic acid is converted to a variety of eicosanoids, some of which cause inﬂammation and pain. c. Phospholipase A2 releases arachidonic ac… DSerine

17. Chromatographic Separation of Lipids Suppose that you apply a mixture of lipids to a silica gel column and then wash the column with increasingly polar solvents. The mixture consists of phosphatidylserine, phosphatidylethanolamine, phosphatidylcholine, cholesteryl palmitate (a sterol ester), sphingomyelin, palmitate, n-tetradecanol, triacylglycerol, and cholesterol. In what order will the lipids elute from the column? Explain your reasoning.

Aa. Glycerol and the sodium salts of palmitic and stearic acids b. D-Glycerol 3-phosphocholine and the sodium salts of palmitic and oleic acids BFirst eluted to last eluted: cholesteryl palmitate and triacylglycerol; cholesterol and n- tetradecanol; phosphatidylcholine and phosphatidylethanolamine; sphingomyelin; phosphatidylserine and palmitate. The lipids elute from the silica gel column in order of polarity. The least polar lipid will elute ﬁrst and the most polar lipid will elute last. CSerine Da. Sphingosine (4.78); linoleic acid (5.88); stearic acid (6.33); cholesterol (7.68) b. Log P describes the lipophilicity of the drug, crucial for determining how to formulate the drug for transport through aqueous compartments of the body such as the gut and the bloodstream. Log P also determines the likelihood of a drug being absorbed by fats and fatty tissues, which can alter its effectiveness, half-life, and pot…

18. Identiﬁcation of Unknown Lipids Johann Thudichum, who practiced medicine in London about 100 years ago, also dabbled in lipid chemistry in his spare time. He isolated a variety of lipids from neural tissue and characterized and named many of them. His carefully sealed and labeled vials of isolated lipids were rediscovered many years later. a. How would you conﬁrm, using techniques not available to Thudichum, that the vials labeled “sphingomyelin” and “cerebroside” actually contain these compounds? b. How would you distinguish sphingomyelin from phosphatidylcholine by chemical, physical, or enzymatic tests? BIOCHEMISTRY ONLINE

AIt reduces double bonds, which increases the melting point of lipids containing the fatty acids. BDiacylglycerol is hydrophobic and remains in the membrane. Inositol 1,4,5-trisphosphate is highly polar, very soluble in water, and more readily diffusible in the cytosol. Both are second messengers. Ca. Subject acid hydrolysates of each compound to chromatography (GC or silica gel TLC), and compare the result with known standards. Sphingomyelin hydrolysate: sphingosine, fatty acids, phosphocholine, choline, and phosphate; cerebroside hydrolysate: sphingosine, fatty acids, sugars, but no phosphate. b. Strong alkaline hydrolysis of sphingomyelin yields sphingosine; phosphatidylcholine yields glycerol. Detect hydro… DSpearmint is (R)-carvone; caraway is (S)-carvone.

19. Using the LIPID MAPS Database to Find Solubility Information Lipidomics has identiﬁed thousands of cellular lipids. LIPID MAPS is an online database containing over 40,000 unique lipid structures, as well as information on the chemical and physical properties of each lipid (www.lipidmaps.org). One important parameter when working with lipids is log P, where P is the octanol:water partition coefﬁcient, an indicator of lipophilicity. a. Look up cholesterol, sphingosine, linoleic acid, and stearic acid in LIPID MAPS and use the reported log P values to place them in order of increasing solubility in octanol. b. Pharmacologists oen study log P values when developing new drugs. Why would knowing a drug’s log P value be informative?

Aa. Sphingosine (4.78); linoleic acid (5.88); stearic acid (6.33); cholesterol (7.68) b. Log P describes the lipophilicity of the drug, crucial for determining how to formulate the drug for transport through aqueous compartments of the body such as the gut and the bloodstream. Log P also determines the likelihood of a drug being absorbed by fats and fatty tissues, which can alter its effectiveness, half-life, and pot… BHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH) CSerine Da. β barrel b. Phe, Trp, Tyr, Leu. All are hydrophobic or have nonpolar R groups. c. The polar head group can hydrogen bond with water; the hydrocarbon tail cannot. Hydrophobic portions of the residues of the pocket protect the tail from contact with water as it moves through the bloodstream.

20. Characteristics of Lipid Transport Proteins Oen when lipids are transported between different tissues, they are carried by proteins. In this exercise, you will explore the interactions between a lipid and a protein using the PDB (www.rcsb.org). Use the PDB identiﬁer 2YG2 and study the structure of the complex between HDL-associated apolipoprotein M and sphingosine-1-phosphate. Navigate to 3D View: Structure to answer the following questions. a. What protein motif is adopted by apolipoprotein M? b. Which amino acid residues do you ﬁnd lining the sphingosine binding pocket? What do they have in common? c. The phosphoryl group of sphingosine-1-phosphate is exposed on the surface of the protein. Why do you suppose it is important that the transport protein binds the hydrocarbon tail of sphingosine-1-phosphate but not necessarily the polar head group? DATA ANALYSIS PROBLEM

AFirst eluted to last eluted: cholesteryl palmitate and triacylglycerol; cholesterol and n- tetradecanol; phosphatidylcholine and phosphatidylethanolamine; sphingomyelin; phosphatidylserine and palmitate. The lipids elute from the silica gel column in order of polarity. The least polar lipid will elute ﬁrst and the most polar lipid will elute last. Ba. β barrel b. Phe, Trp, Tyr, Leu. All are hydrophobic or have nonpolar R groups. c. The polar head group can hydrogen bond with water; the hydrocarbon tail cannot. Hydrophobic portions of the residues of the pocket protect the tail from contact with water as it moves through the bloodstream. Ca. Glycerol and the sodium salts of palmitic and stearic acids b. D-Glycerol 3-phosphocholine and the sodium salts of palmitic and oleic acids Da. GM1 and globoside. Both glucose and galactose are hexoses, so “hexose” in the molar ratio refers to glucose + galactose. The ratios for the four gangliosides are GM1, 1:3:1:1; GM2, 1:2:1:1; GM3, 1:2:0:1; globoside, 1:3:1:0. b. Yes. The ratio matches GM2, the ganglioside expected to build up in Tay-Sachs disease (see Box 10-1, Fig. 1). c. This analysis is similar to that used by Sanger to determine the amino acid …

21. Determining the Structure of the Abnormal Lipid in Tay-Sachs Disease Box 10-1, Figure 1, shows the pathway of breakdown of gangliosides in healthy (normal) individuals and in individuals with certain genetic diseases. Some of the data on which the ﬁgure is based were presented in a paper by Lars Svennerholm (1962). Note that the sugar Neu5Ac, N- acetylneuraminic acid, represented in the Box 10-1 ﬁgure as , is a sialic acid. Svennerholm reported that “about 90% of the monosialogangliosides isolated from normal human brain” consisted of a compound with ceramide, hexose, N- acetylgalactosamine, and N-acetylneuraminic acid in the molar ratio 1:3:1:1. a. Which of the gangliosides (GM1 through GM3 and globoside) in Box 10-1, Figure 1, ﬁts this description? Explain your reasoning. b. Svennerholm reported that 90% of the gangliosides from a patient with Tay-Sachs disease had a molar ratio (of the same four components given above) of 1:2:1:1. Is this consistent with the Box 10-1 ﬁgure? Explain your reasoning. To determine the structure in more detail, Svennerholm treated the gangliosides with neuraminidase to remove the N-acetylneuraminic acid. This resulted in an asialoganglioside that was much easier to analyze. He hydrolyzed it with acid, collected the ceramide-containing products, and determined the molar ratio of the sugars in each product. He did this for both the normal gangliosides and the Tay-Sachs gangliosides. His results are shown below. Ganglioside Ceramide Glucose Galactose Galactosamine Normal Fragment 1 1 1 0 0 Fragment 2 1 1 1 0 Fragment 3 1 1 1 1 Fragment 4 1 1 2 1 Tay-Sachs Fragment 1 1 1 0 0 Fragment 2 1 1 1 0 Fragment 3 1 1 1 1 c. Based on these data, what can you conclude about the structure of the normal ganglioside? Is this consistent with the structure in Box 10-1? Explain your reasoning. d. What can you conclude about the structure of the Tay- Sachs ganglioside? Is this consistent with the structure in Box 10-1? Explain your reasoning. Svennerholm also reported the work of other researchers who “permethylated” the normal asialoganglioside. Permethylation is the same as exhaustive methylation: a methyl group is added to every free hydroxyl group on a sugar. They found the following permethylated sugars: 2,3,6- trimethylglycopyranose; 2,3,4,6- tetramethylgalactopyranose; 2,4,6- trimethylgalactopyranose; and 4,6-dimethyl-2-deoxy-2- aminogalactopyranose. e. To which sugar of GM1 does each of the permethylated sugars correspond? Explain your reasoning. f. Based on all the data presented so far, what pieces of information about normal ganglioside structure are missing? Reference Svennerholm, L. 1962. The chemical structure of normal human brain and Tay-Sachs gangliosides. Biochem. Biophys. Res. Comm. 9:436–441.

ASpearmint is (R)-carvone; caraway is (S)-carvone. Ba. The number of cis double bonds. Each cis double bond causes a bend in the hydrocarbon chain, lowering the melting temperature. b. Six different triacylglycerols can be constructed, in order of increasing melting points: OOO< OOP=OPO< PPO=POP< PPP c. where O = oleic and P = palmitic acid. The greater the content of saturated fatty acid, the higher the melting point. d. Branched-chain fatty acids increase the ﬂuidi… CHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH) Da. GM1 and globoside. Both glucose and galactose are hexoses, so “hexose” in the molar ratio refers to glucose + galactose. The ratios for the four gangliosides are GM1, 1:3:1:1; GM2, 1:2:1:1; GM3, 1:2:0:1; globoside, 1:3:1:0. b. Yes. The ratio matches GM2, the ganglioside expected to build up in Tay-Sachs disease (see Box 10-1, Fig. 1). c. This analysis is similar to that used by Sanger to determine the amino acid …

22. Operational Deﬁnition of Lipids How is the deﬁnition of “lipid” different from the types of deﬁnitions used for other biomolecules such as amino acids, nucleic acids, and proteins?

Aa. Glycerol and the sodium salts of palmitic and stearic acids b. D-Glycerol 3-phosphocholine and the sodium salts of palmitic and oleic acids Ba. β barrel b. Phe, Trp, Tyr, Leu. All are hydrophobic or have nonpolar R groups. c. The polar head group can hydrogen bond with water; the hydrocarbon tail cannot. Hydrophobic portions of the residues of the pocket protect the tail from contact with water as it moves through the bloodstream. CThe term “lipid” does not specify a particular chemical structure. Compounds are categorized as lipids based on their greater solubility in organic solvents than in water. DSerine

23. Structure of an Omega-3 Fatty Acid The omega-3 fatty acid docosahexaenoic acid (DHA, 22:6(Δ4,7,10,13,16,19)) is the most abundant omega-3 fatty acid in the brain and an important component of breast milk. Draw the structure of this fatty acid.

Aa. Subject acid hydrolysates of each compound to chromatography (GC or silica gel TLC), and compare the result with known standards. Sphingomyelin hydrolysate: sphingosine, fatty acids, phosphocholine, choline, and phosphate; cerebroside hydrolysate: sphingosine, fatty acids, sugars, but no phosphate. b. Strong alkaline hydrolysis of sphingomyelin yields sphingosine; phosphatidylcholine yields glycerol. Detect hydro… Ba. The number of cis double bonds. Each cis double bond causes a bend in the hydrocarbon chain, lowering the melting temperature. b. Six different triacylglycerols can be constructed, in order of increasing melting points: OOO< OOP=OPO< PPO=POP< PPP c. where O = oleic and P = palmitic acid. The greater the content of saturated fatty acid, the higher the melting point. d. Branched-chain fatty acids increase the ﬂuidi… C DHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH)

24. Melting Points of Lipids The melting points of a series of 18-carbon fatty acids are stearic acid, 69.6 °C; oleic acid, 13.4 °C; linoleic acid, −5 °C; and linolenic acid, −11 °C. a. What structural aspect of these 18-carbon fatty acids can be correlated with the melting point? b. Draw all the possible triacylglycerols that can be constructed from glycerol, palmitic acid, and oleic acid. Rank them in order of increasing melting point. c. Branched-chain fatty acids are found in some bacterial membrane lipids. Would their presence increase or decrease the ﬂuidity of the membrane (that is, give the lipids a lower or a higher melting point)? Why?

AHydrophobic units: a. 2 fatty acids b., c., and d. 1 fatty acid and the hydrocarbon chain of sphingosine e. the steroid nucleus and acyl side chain. Hydrophilic units: a. phosphoethanolamine b. phosphocholine c. D-galactose d. several sugar molecules e. alcohol group (—OH) BThe 18O label appears in the fatty acid salts. CSpearmint is (R)-carvone; caraway is (S)-carvone. Da. The number of cis double bonds. Each cis double bond causes a bend in the hydrocarbon chain, lowering the melting temperature. b. Six different triacylglycerols can be constructed, in order of increasing melting points: OOO< OOP=OPO< PPO=POP< PPP c. where O = oleic and P = palmitic acid. The greater the content of saturated fatty acid, the higher the melting point. d. Branched-chain fatty acids increase the ﬂuidi…

25. Catalytic Hydrogenation of Vegetable Oils Catalytic hydrogenation, used in the food industry, converts double bonds in the fatty acids of the oil triacylglycerols to — CH2— CH2— . How does this affect the physical properties of the oils?

AThe part of the membrane lipid that determines blood type is the oligosaccharide in the head group of the membrane sphingolipids (see Fig. 10-13). This same oligosaccharide is attached to certain membrane glycoproteins, which also serve as points of recognition by the antibodies that distinguish blood groups. BIt reduces double bonds, which increases the melting point of lipids containing the fatty acids. Ca. GM1 and globoside. Both glucose and galactose are hexoses, so “hexose” in the molar ratio refers to glucose + galactose. The ratios for the four gangliosides are GM1, 1:3:1:1; GM2, 1:2:1:1; GM3, 1:2:0:1; globoside, 1:3:1:0. b. Yes. The ratio matches GM2, the ganglioside expected to build up in Tay-Sachs disease (see Box 10-1, Fig. 1). c. This analysis is similar to that used by Sanger to determine the amino acid … Da. The free —OH group on C-2 and the phosphocholine head group on C-3 are hydrophilic; the fatty acid on C-1 of lysolecithin is hydrophobic. b. Certain steroids, such as prednisone, inhibit the action of phospholipase A2, inhibiting the release of arachidonic acid from C-2. Arachidonic acid is converted to a variety of eicosanoids, some of which cause inﬂammation and pain. c. Phospholipase A2 releases arachidonic ac…