**3. Dietary lipid composition**

Dietary triacylglycerol composition plays a major role in determining adipose tissue composition. Monogastric animals incorporate dietary fatty acids directly into tissue lipid deposits [21, 22] and, therefore, to manipulate carcass lipid quality, it is important to understand the interactions of dietary lipids with carcass lipid. Carcass fatty acid profiles closely mimic dietary fatty acid profile [21, 23], and therefore, potential exists to modify carcass lipid properties (i.e., firmness, fatty acid profile, etc.) by altering dietary lipid composition.

One of the strongest determinants of carcass fat quality in pigs is the level and composition of lipids in the diet [24]. Because the utilization efficiency of dietary fat is 90% in pigs fed above maintenance [24] and the transfer coefficient of dietary fat to carcass lipid is as high as 31-40% [25] the carcass lipid composition is a reflection of dietary fat. The impact of dietary lipids on carcass lipid may differ depending on the timing of feeding relative to growth and finishing, levels included in the diet, and interactions with other stressors.

### **3.1. Dietary fat**

214 Lipid Metabolism

acids into the cell mitochondria, and oxidation.

regulated [1] it is not pertinent to this discussion.

temperature, pH, and lipids [13, 14].

and acetyl CoA synthase [9].

levels as well as improving CPT-I mRNA stability [14].

**2.4. Regulation of lipid metabolism by transcription factors** 

components: activation of fatty acids in the cytosol of the cell, transport of activated fatty

The mechanism of the carnitine pathway is an ordered reaction where the binding of acyl-CoA begins the transport action [13]. Long chain fatty acids are converted to acyl-CoAs by acyl-CoA synthetase [14]. Acyl-CoAs are converted to acyl-carnitine molecules and transferred across the outer mitochondrial membrane by carnitine palmitoyltransferase-I (CPT; EC 2.3.1.21) [14]. Carnitine palmitoyltransferase-II is located on the inner mitochondrial membrane and liberates the carnitine from the acylcarnitine after transfer across the inner mitochondrial membrane [15]. After liberation, the acyl-CoA units are available for β-oxidation within the mitochondrial matrix [14]. Because CPT-II is not

Carnitine palmitoyltransferase-I is located on the outer mitochondrial membrane and limits the rate of fatty acid oxidation by controlling the transportion of fatty acyl-CoA to the mitochondrial matrix where β-oxidation occurs [13, 1]. Two transmembrane domains anchor CPT-I to the outer mitochondrial membrane [13]. There are three isoforms of CPT-I [1]. In liver, kidney, lung, and heart tissue, CPT-Ia is present; CPT-Ib is present in skeletal

Regulation of β-oxidation occurs during the initial transport step. The main route of CPT-I regulation is by malonyl-CoA, the first product of lipogenesis, which inhibits CPT-I and aids to prevent simultaneous oxidation and synthesis [1, 16]. Regulation of CPT-I allows βoxidation to be regulated by controlling the availability of acyl-CoA in the mitochondrial matrix [1, 14]. Though the sensivitiy of the CTP-Ia and CPT-Ib to malonyl-CoA are different, they both contain binding sites on the same side of the membrane as the active site [13]. The N-terminus of the enzyme, which is not required for catalytic activity, controls the response to malonyl-CoA [13]. The kinetics of inhibition by malonyl-CoA are responsive to

Fasting and glucagon increases CPT-I gene expression while hypothyroidism decreases expression by regulating the transcription level [13]. The insulin growth factor I receptor also controls CPT-I expression by mediating the inhibitory effects of insulin [13, 14]. Expression of CPT-I is also transcriptionally upregulated by PPARα [13]. Long chain fatty acids increase CPT-Ia mRNA expression in liver tissue by both increasing transcription

Sterol regulatory element binding proteins (SREBP) are helix loop helix proteins that are within the leucine zipper family of transcription factors [9]. The SREBPs are present as two isoforms, SREBP-1 (a and c subforms) and SREBP-2 [9]. While SREBP-2 is primarily involved in activation of cholesterol synthesis and metabolism, SREBP-1c is involved solely in regulation of fatty acid synthesis and SREBP-1a is capable of inducing both synthesis of cholesterol and fatty acids [9]. In the liver, SREBP-1c increases expression of SCD, ACC, FAS

muscle, heart, and adipose tissue; and CPT-Ic is brain tissue specific [1].

Dietary triacylglycerols alter carcass lipid composition at the level of the fatty acid profile [21]. Saturated fatty acids lack double bonds and have melting temperatures above 40ºC. Mono-, di- and poly-unsaturated fatty acids have one, two, or many double bonds, respectively and as the level of unsaturation increases, the melting point decreases [21]. The ratio of saturated to unsaturated fatty acids is a way of describing the relative saturation of a fatty acid profile [21]. Iodine value, a measure of double bonds in a lipid, is a method used to composite characteristics of lipids in regard to fluidity [21, 26]. Saturated to unsaturated ratios and iodine values can be utilized to describe the composition of lipids in both feedstuffs, total rations, and animal tissue.

Fat is commonly added in swine diets from 0.5% up to 7% of the ration and increases growth rate, reduces feed intake, and improves feed efficiency [21]. Because of the previously mentioned utilization efficiency and transfer coefficients, the level of saturation and iodine value of the feed lipid source will be strongly reflected in the carcass fatty acid profile and therefore, sources of dietary fats play a critical role in final carcass lipid quality. Vegetable oils are typically high in linoleic acid, have an unsaturated to saturated fatty acid ratio of 12:1 [22] and an iodine value greater than 100 [21]. Diets high in these unsaturated vegetable oils will result in oily, soft carcass fat [21]. Conversely, tallow, which is high in palmitate and stearate, has a saturated to unsaturated fatty acid ratio of 1:1 [22], an iodine value between 40 and 45 [21] and will result in firmer carcass fat when fed in the diet. Greater saturated:unsaturated fatty acid ratio in fat contained in pig carcasses results in fewer difficulties during processing [27] due to increased firmness at typical processing temperatures (2 to 4°C). Due to differences in calculation of these indices, some variations in fatty acid profile are captured with one ration but not the other, as seen in Figure 1. For this reason, it is best to utilize both the IV and saturated:unsaturated indices when characterizing fat quality, in order to identify all variations in fatty acid profile.

Impacts of Nutrition and Environmental Stressors on Lipid Metabolism 217

Dried distillers grains with solubles (DDGS) is the by-product of yeast fermentation of grains such as corn for ethanol production [28]. During fermentation, corn starch is converted into alcohol and the remaining grain components, protein, fat, fiber, minerals, and vitamins are concentrated in the fermentative co-product approximately 3-times that of corn [28]. The nutritional value of corn DDGS is variable and a function of DDGS processing

There are two processes by which ethanol can be extracted from corn, wet milling and dry grinding. Dry grinding is more commonly used and accounts for 70% of ethanol production processes [31]. Dry grinding yields the maximum ethanol from corn while wet milling yields other products including corn oil and corn gluten meal [31, 32]. The dry grind process begins by grinding the corn and mixing it with water (Figure 2). The resulting mash is then heated with enzymes to convert the starches to sugars which can be fermented by yeast. The product contains particulates and solubles which are distilled and dehydrated, producing ethanol and wet distiller's grains. The distiller's grains are then dried in order to increase

Mill

Corn

Liquefy

Saccharify & Ferment

Distill

Dehydrate

Dry

CO2

**Dried Distillers Grains with Solubles**

**Ethanol**

**Figure 2.** Dry grind processing of corn to produce ethanol. Progression of processing steps are shown in ovals and gray arrows, with inputs and outputs indicated by black arrows. The major byproduct of

ethanol production is dried distillers grains with solubles.

Enzyme

Enzyme

**3.2. Dried Distillers Grains with Solubles** 

[28, 29, 30].

shelf life [31, 32].

**Figure 1.** Differences in carcass lipid quality alter final product characteristics. Higher iodine values (IV) are associated with fat that is softer, resulting in increased difficulty slicing and processing. Panel *a* is backfat with an IV of 69 which represents fat that is firm and maintains shape and structure, while panel *b* is backfat with an IV of 79 which represents fat that will lack the firmness required for processing. Saturated to unsaturated fatty acid ratios are also used to characterize fatty acid profiles. While sausages (bottom panels) made from different animals have the same IV (59), the differences in saturated:unsaturated fatty acids results in a higher quality, firmer product in panel *c* (0.62) that has less smearing compared with panel *d* (0.59).

#### **3.2. Dried Distillers Grains with Solubles**

216 Lipid Metabolism

Fat is commonly added in swine diets from 0.5% up to 7% of the ration and increases growth rate, reduces feed intake, and improves feed efficiency [21]. Because of the previously mentioned utilization efficiency and transfer coefficients, the level of saturation and iodine value of the feed lipid source will be strongly reflected in the carcass fatty acid profile and therefore, sources of dietary fats play a critical role in final carcass lipid quality. Vegetable oils are typically high in linoleic acid, have an unsaturated to saturated fatty acid ratio of 12:1 [22] and an iodine value greater than 100 [21]. Diets high in these unsaturated vegetable oils will result in oily, soft carcass fat [21]. Conversely, tallow, which is high in palmitate and stearate, has a saturated to unsaturated fatty acid ratio of 1:1 [22], an iodine value between 40 and 45 [21] and will result in firmer carcass fat when fed in the diet. Greater saturated:unsaturated fatty acid ratio in fat contained in pig carcasses results in fewer difficulties during processing [27] due to increased firmness at typical processing temperatures (2 to 4°C). Due to differences in calculation of these indices, some variations in fatty acid profile are captured with one ration but not the other, as seen in Figure 1. For this reason, it is best to utilize both the IV and saturated:unsaturated indices when

characterizing fat quality, in order to identify all variations in fatty acid profile.

a b

c d

**Figure 1.** Differences in carcass lipid quality alter final product characteristics. Higher iodine values (IV) are associated with fat that is softer, resulting in increased difficulty slicing and processing. Panel *a* is backfat with an IV of 69 which represents fat that is firm and maintains shape and structure, while panel *b* is backfat with an IV of 79 which represents fat that will lack the firmness required for processing. Saturated to unsaturated fatty acid ratios are also used to characterize fatty acid profiles. While sausages (bottom panels) made from different animals have the same IV (59), the differences in saturated:unsaturated fatty acids results in a higher quality, firmer product in panel *c* (0.62) that has less

smearing compared with panel *d* (0.59).

Dried distillers grains with solubles (DDGS) is the by-product of yeast fermentation of grains such as corn for ethanol production [28]. During fermentation, corn starch is converted into alcohol and the remaining grain components, protein, fat, fiber, minerals, and vitamins are concentrated in the fermentative co-product approximately 3-times that of corn [28]. The nutritional value of corn DDGS is variable and a function of DDGS processing [28, 29, 30].

There are two processes by which ethanol can be extracted from corn, wet milling and dry grinding. Dry grinding is more commonly used and accounts for 70% of ethanol production processes [31]. Dry grinding yields the maximum ethanol from corn while wet milling yields other products including corn oil and corn gluten meal [31, 32]. The dry grind process begins by grinding the corn and mixing it with water (Figure 2). The resulting mash is then heated with enzymes to convert the starches to sugars which can be fermented by yeast. The product contains particulates and solubles which are distilled and dehydrated, producing ethanol and wet distiller's grains. The distiller's grains are then dried in order to increase shelf life [31, 32].

**Figure 2.** Dry grind processing of corn to produce ethanol. Progression of processing steps are shown in ovals and gray arrows, with inputs and outputs indicated by black arrows. The major byproduct of ethanol production is dried distillers grains with solubles.

#### 218 Lipid Metabolism

The nutritional value of DDGS for pigs is influenced by the processing procedure and production plant equipment and techniques [33, 34]. The nutrient profile of DDGS remains highly variable even within the same production site [30, 35]. The NRC published content for DDGS is 93% dry matter, 2.82 Mcal/kg metabolizable energy, 27.7% crude protein, 8.4% crude fat and 34.6% neutral detergent fiber [36], however there is significant plant to plant variation, as noted above.

Impacts of Nutrition and Environmental Stressors on Lipid Metabolism 219

**Figure 3.** Synthesis pathways for omega-3, -6, and -9 fatty acids in mammals. MUFA, monounsaturated fatty acid; HUFA, highly unsaturated fatty acid; EPA, eicosapentaenoic acid; DHA, docosahexaenoic

The ideal ratio of omega-6 to omega-3 fatty acids in human diets is between 4 to 6:1, although the average American diet is between 10 to 30:1 [42]. The change in this ratio is due to the increase in omega-6 intake relative to the level of omega-3 fatty acids [41]. The need to increase dietary intake of omega-3 fatty acids, specifically EPA and DHA, has increased demand for products with a ratio of omega-6 to -3 fatty acids more closely related to

The many health benefits of omega-3 fatty acids, such as lowering serum cholesterol and triacylglycerol concentrations, reduce platelet aggregation, reduction of blood pressure, and decreasing very-low-density and low-density lipoproteins, make dietary inclusion important [40]. The overall anti-inflammatory effects of omega-3 fatty acids have shown beneficial effects for arthritis and joint health in rats and humans [40]. Though it has not been directly studied in swine, omega-3 fatty acids could decrease the prevalence of lameness in sows if they result in the same joint and anti-inflammatory benefits noted in humans and rats. In Canada, 8-11% of sows culled were due to lameness [43] and in the United States lameness accounts for about 10% of culled sows during parity zero, one, and two or more, respectively [44]. Lameness results in the removal of sows at a younger age

than other culling reasons, thus decreasing breeding herd productivity [44].

American Heart Association (AHA) recommendations.

acid [9].

Two limiting factors for including DDGS in swine diets are the high level of unsaturation in the dietary fatty acid profile and the high fiber content [28, 31]. As discussed above, the composition of these fat sources is important when considering the carcass fat firmness [21]. Dietary fiber has also shown beneficial effects in swine diets including reduction of gastric ulceration and restriction of pathogenic bacteria in the intestinal tract; however, when fiber content of the diet exceeds 7%, growth is inhibited [37]. The high level of fat and fiber in DDGS have been shown to result in both decreased feed intake and increased unsaturated content of adipose tissue. In a trial utilizing 0, 10, 20, and 30% DDGS in grow-finish diets, pigs fed 20 or 30% DDGS had decreased growth performance and increased IV when compared to control fed pigs [38]. Incorporation of 0, 20, or 40% DDGS in diets during the final 30 days of the finishing phase resulted in reduced percent lean in bacon and decreased carcass firmness (based on IV and saturation); however, no effect on growth performance was observed [39].

The future direction of DDGS as a feed ingredient will likely be defined by the final use in global energy needs and not how it might be valued as a feed ingredient; that is, DDGS still contains a considerable amount of oil, a highly valued potential energy source. Today, DDGS is well suited for non-ruminants in terms of energy and protein content, price, and availability; however, the high linoleic acid content known to alter fat quality must be considered when determining dietary inclusions. As refiners investigate new approaches to removing the oil and protein, which may be of more value extracted, the future product could resemble a more fiber-like product, which would have wide range implications on non-ruminant animals and likely reduce it's future use in swine diets.

#### **3.3. Omega-3 and -6 fatty acids**

The levels of omega-3 and omega-6 fatty acids in the human diet are important for optimal health. Animals, including humans, lack the enzymes required to add double bonds between the methyl group and ninth carbon and therefore cannot synthesize omega-3 and -6 fatty acids, making these fatty acids essential in the diet [22]. Fatty acids in the omega-6 family, linoleic (LA; 18:2n-6) and arachidonic (AA; 20:4n-6), and those in the omega-3 family, α-linolenic (ALA; 18:3n-3) and subsequently eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3), must be supplied in animal diets [16, 21, 22). The synthesis pathways of omega-3 and -6 fatty acids and the parallel omega-9 pathways are shown in Figure 3. The omega-3 and -6 pathways compete for the ∆5 and ∆6 desaturases though both enzymes preferentially catalyze the reactions of the omega-3 pathway [40, 41].

Impacts of Nutrition and Environmental Stressors on Lipid Metabolism 219

218 Lipid Metabolism

variation, as noted above.

was observed [39].

**3.3. Omega-3 and -6 fatty acids** 

The nutritional value of DDGS for pigs is influenced by the processing procedure and production plant equipment and techniques [33, 34]. The nutrient profile of DDGS remains highly variable even within the same production site [30, 35]. The NRC published content for DDGS is 93% dry matter, 2.82 Mcal/kg metabolizable energy, 27.7% crude protein, 8.4% crude fat and 34.6% neutral detergent fiber [36], however there is significant plant to plant

Two limiting factors for including DDGS in swine diets are the high level of unsaturation in the dietary fatty acid profile and the high fiber content [28, 31]. As discussed above, the composition of these fat sources is important when considering the carcass fat firmness [21]. Dietary fiber has also shown beneficial effects in swine diets including reduction of gastric ulceration and restriction of pathogenic bacteria in the intestinal tract; however, when fiber content of the diet exceeds 7%, growth is inhibited [37]. The high level of fat and fiber in DDGS have been shown to result in both decreased feed intake and increased unsaturated content of adipose tissue. In a trial utilizing 0, 10, 20, and 30% DDGS in grow-finish diets, pigs fed 20 or 30% DDGS had decreased growth performance and increased IV when compared to control fed pigs [38]. Incorporation of 0, 20, or 40% DDGS in diets during the final 30 days of the finishing phase resulted in reduced percent lean in bacon and decreased carcass firmness (based on IV and saturation); however, no effect on growth performance

The future direction of DDGS as a feed ingredient will likely be defined by the final use in global energy needs and not how it might be valued as a feed ingredient; that is, DDGS still contains a considerable amount of oil, a highly valued potential energy source. Today, DDGS is well suited for non-ruminants in terms of energy and protein content, price, and availability; however, the high linoleic acid content known to alter fat quality must be considered when determining dietary inclusions. As refiners investigate new approaches to removing the oil and protein, which may be of more value extracted, the future product could resemble a more fiber-like product, which would have wide range implications on

The levels of omega-3 and omega-6 fatty acids in the human diet are important for optimal health. Animals, including humans, lack the enzymes required to add double bonds between the methyl group and ninth carbon and therefore cannot synthesize omega-3 and -6 fatty acids, making these fatty acids essential in the diet [22]. Fatty acids in the omega-6 family, linoleic (LA; 18:2n-6) and arachidonic (AA; 20:4n-6), and those in the omega-3 family, α-linolenic (ALA; 18:3n-3) and subsequently eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3), must be supplied in animal diets [16, 21, 22). The synthesis pathways of omega-3 and -6 fatty acids and the parallel omega-9 pathways are shown in Figure 3. The omega-3 and -6 pathways compete for the ∆5 and ∆6 desaturases though both enzymes preferentially catalyze the reactions of the omega-3 pathway [40, 41].

non-ruminant animals and likely reduce it's future use in swine diets.

**Figure 3.** Synthesis pathways for omega-3, -6, and -9 fatty acids in mammals. MUFA, monounsaturated fatty acid; HUFA, highly unsaturated fatty acid; EPA, eicosapentaenoic acid; DHA, docosahexaenoic acid [9].

The ideal ratio of omega-6 to omega-3 fatty acids in human diets is between 4 to 6:1, although the average American diet is between 10 to 30:1 [42]. The change in this ratio is due to the increase in omega-6 intake relative to the level of omega-3 fatty acids [41]. The need to increase dietary intake of omega-3 fatty acids, specifically EPA and DHA, has increased demand for products with a ratio of omega-6 to -3 fatty acids more closely related to American Heart Association (AHA) recommendations.

The many health benefits of omega-3 fatty acids, such as lowering serum cholesterol and triacylglycerol concentrations, reduce platelet aggregation, reduction of blood pressure, and decreasing very-low-density and low-density lipoproteins, make dietary inclusion important [40]. The overall anti-inflammatory effects of omega-3 fatty acids have shown beneficial effects for arthritis and joint health in rats and humans [40]. Though it has not been directly studied in swine, omega-3 fatty acids could decrease the prevalence of lameness in sows if they result in the same joint and anti-inflammatory benefits noted in humans and rats. In Canada, 8-11% of sows culled were due to lameness [43] and in the United States lameness accounts for about 10% of culled sows during parity zero, one, and two or more, respectively [44]. Lameness results in the removal of sows at a younger age than other culling reasons, thus decreasing breeding herd productivity [44].

Omega-6 fatty acids are the precursors of eicosanoids which include prostaglandins, thromboxanes and leukotrienes. These metabolites of n-6 fatty acids exhibit inflammatory effects [45]. Omega-3 fatty acids inhibit eicosanoid synthesis by decreasing the available arachidonic acid available for eicosanoid production [18, 45]. In addition to decreasing eicosanoid production, omega-3 fatty acids also decrease other inflammatory cytokines such as interleukin-1 and -6, and tumour necrosis factor [18, 45].

Impacts of Nutrition and Environmental Stressors on Lipid Metabolism 221

Atherosclerotic plaque formation is reduced by CLA [48]. Inclusion of 0.5 g/day in hypercholesterolemic diets fed to rabbits for 12 weeks resulted in significantly reduced serum triacylglycerols, low density lipoprotein (LDL) cholesterol levels and atherosclerotic plaque formation in the aorta [53]. The reduction of plaque deposits by CLA was proposed

Effects of CLA on the onset of diabetes and insulin resistance are inconsistant. Rats fed CLA have shown significantly reduced fasting glucose, insulinemia, triglyceridemia, free fatty acids, and leptinemia [48]. Butter enriched with c9t11 CLA failed to reduce glucose tolerance, lower adipose tissue or enhance glucose uptake leading to the conclusion that perhaps it is the t10c12 isomer which is responsible for the antidiabetogenic responses [48]. Insulin tolerance testing on CLA-fed mice showed marked insulin resistance without changes to blood glucose concentrations after oral glucose tolerance testing [54]. Other studies have examined the reduction of plasma leptin by CLA and the concomitant changes in blood glucose level due to regulation by leptin [46]. Feeding male mice high-fat diets with 1% CLA has resulted in reduced plasma leptin levels in one study [55] while resulting in no

The effects of feeding CLA to pigs have been evaluated in regard to fat quality [57]. Gilts fed 1% CLA for seven weeks had firmer bellies, higher levels of saturated fatty acids, lower levels of unsaturated fatty acids and decreased IV when compared to controls [58]. Barrows fed CLA had improved feed efficiency, decreased backfat, and improved loin marbling and firmness when CLA was included at 0.75% of grow-finish diets [51]. When CLA was fed to genetically lean gilts for eight weeks, an increase in average daily gain and gain:feed was observed [59]. The same study also noted an increase in saturated fatty acids, decrease in unsaturated fatty acids, and an increased level of saturation of the belly tissue [59]. Several studies have shown that CLA feeding increases fatty acid saturation, and firmness in back fat and belly fat [60, 61, 62]. Additionally, use of CLA when feeding by-products may alleviate some or all of the negative impact on carcass quality. When feeding 0, 20, or 40% DDGS during the final 30 days of the finishing period, the addition of 0.6% CLA minimized the negative impact of 20% DDGS inclusion on carcass lipid quality but was unable to

Dietary CLA in several species alters the activity of SCD-1, FAS, and ACC in adipose and liver. Conjugated linoleic acids decrease mRNA for FAS and ACC to significantly inhibit the capacity for *de novo* lipogenesis [47, 60]. In barrows and gilts fed 0.25 or 0.5% CLA for the finishing diet from 97 to 172 kg, ACC activity was significantly reduced compared to control pigs [64]. Alleviation of negative impacts of nutritional stress of lipid quality, such as during DDGS feeding, is likely through altered lipid metabolism as adipose mRNA expression of ACC was decreased with CLA supplementation with all inclusion levels of DDGS [39].

to be due to changes in LDL oxidative susceptibility [48].

change in plasma leptin or glucose levels in another [56].

overcome the negative effects of feeding 40% DDGS [63].

*3.4.2. Mechanism of CLA to alter lipid metabolism* 

*3.4.1. Feeding CLA to pigs* 

#### **3.4. Conjugated linoleic acid**

Conjugated linoleic acids (CLA) are a group of polyunsaturated fatty acids that are positional and geometric isomers of linoleic acid (C18:2). Because CLA and its precursor, *trans* vaccenic acid, are naturally produced during bacterial fermentation in the rumen of ruminant animals, the main sources of CLA in human nutrition are ruminant milk and meats [46, 47]. The main isomers of CLA are *cis*-9, *trans*-11(c9t11) and *trans*-10, *cis*-12 (t10c12; Figure 4). Though the main isomer produced by ruminants is c9t11, commercially available products commonly contain equal proportions of c9t11 and t10c12 [46, 47]. Research in rodents, pigs, and humans has been conducted on the effects of CLA and has shown beneficial effects of CLA against obesity, cancer, atherosclerosis, and diabetes, some of which are isomer specific [46, 47, 48].

**Figure 4.** Structure of linoleic acid compared with *cis*-9, *trans*-11 and *trans*-10, *cis*-12 conjugated linoleic acid (CLA).

Many studies have shown CLA mixtures are able to reduce adipose tissue depots in rodents, pigs, and humans and that this effect is specific to the t10c12 isomer or a mixture containing greater than 50% t10c12 [46, 48]. Postweanling mice fed 1% CLA for 28-30 d had a 50% reduction in total adipose tissue compared to control mice [49]. In pigs, CLA inclusion in feed has resulted in decreased backfat thickness at finishing [50, 51]. Overweight or obese humans supplemented with CLA for 12 weeks also demonstrated reduced body fat mass, although their body mass index remained unchanged [52].

Another noted effect of CLA is the inhibition of cancer, specifically, mammary, prostate, skin, colon, and stomach cancers [48]. The anti-carcinogenic effects of CLA have been mainly attributed to the c9t11 isomer [46]. In studies of mammary and prostate cancer cell lines, feeding 1% CLA significantly reduced growth of the cancerous cells. Other studies of the same cell lines have not demonstrated these effects of CLA [48].

Atherosclerotic plaque formation is reduced by CLA [48]. Inclusion of 0.5 g/day in hypercholesterolemic diets fed to rabbits for 12 weeks resulted in significantly reduced serum triacylglycerols, low density lipoprotein (LDL) cholesterol levels and atherosclerotic plaque formation in the aorta [53]. The reduction of plaque deposits by CLA was proposed to be due to changes in LDL oxidative susceptibility [48].

Effects of CLA on the onset of diabetes and insulin resistance are inconsistant. Rats fed CLA have shown significantly reduced fasting glucose, insulinemia, triglyceridemia, free fatty acids, and leptinemia [48]. Butter enriched with c9t11 CLA failed to reduce glucose tolerance, lower adipose tissue or enhance glucose uptake leading to the conclusion that perhaps it is the t10c12 isomer which is responsible for the antidiabetogenic responses [48]. Insulin tolerance testing on CLA-fed mice showed marked insulin resistance without changes to blood glucose concentrations after oral glucose tolerance testing [54]. Other studies have examined the reduction of plasma leptin by CLA and the concomitant changes in blood glucose level due to regulation by leptin [46]. Feeding male mice high-fat diets with 1% CLA has resulted in reduced plasma leptin levels in one study [55] while resulting in no change in plasma leptin or glucose levels in another [56].

#### *3.4.1. Feeding CLA to pigs*

220 Lipid Metabolism

acid (CLA).

Omega-6 fatty acids are the precursors of eicosanoids which include prostaglandins, thromboxanes and leukotrienes. These metabolites of n-6 fatty acids exhibit inflammatory effects [45]. Omega-3 fatty acids inhibit eicosanoid synthesis by decreasing the available arachidonic acid available for eicosanoid production [18, 45]. In addition to decreasing eicosanoid production, omega-3 fatty acids also decrease other inflammatory cytokines such

Conjugated linoleic acids (CLA) are a group of polyunsaturated fatty acids that are positional and geometric isomers of linoleic acid (C18:2). Because CLA and its precursor, *trans* vaccenic acid, are naturally produced during bacterial fermentation in the rumen of ruminant animals, the main sources of CLA in human nutrition are ruminant milk and meats [46, 47]. The main isomers of CLA are *cis*-9, *trans*-11(c9t11) and *trans*-10, *cis*-12 (t10c12; Figure 4). Though the main isomer produced by ruminants is c9t11, commercially available products commonly contain equal proportions of c9t11 and t10c12 [46, 47]. Research in rodents, pigs, and humans has been conducted on the effects of CLA and has shown beneficial effects of CLA against obesity, cancer, atherosclerosis, and diabetes, some of

**Figure 4.** Structure of linoleic acid compared with *cis*-9, *trans*-11 and *trans*-10, *cis*-12 conjugated linoleic

COOH

COOH

COOH

Many studies have shown CLA mixtures are able to reduce adipose tissue depots in rodents, pigs, and humans and that this effect is specific to the t10c12 isomer or a mixture containing greater than 50% t10c12 [46, 48]. Postweanling mice fed 1% CLA for 28-30 d had a 50% reduction in total adipose tissue compared to control mice [49]. In pigs, CLA inclusion in feed has resulted in decreased backfat thickness at finishing [50, 51]. Overweight or obese humans supplemented with CLA for 12 weeks also demonstrated reduced body fat mass,

Another noted effect of CLA is the inhibition of cancer, specifically, mammary, prostate, skin, colon, and stomach cancers [48]. The anti-carcinogenic effects of CLA have been mainly attributed to the c9t11 isomer [46]. In studies of mammary and prostate cancer cell lines, feeding 1% CLA significantly reduced growth of the cancerous cells. Other studies of

as interleukin-1 and -6, and tumour necrosis factor [18, 45].

although their body mass index remained unchanged [52].

the same cell lines have not demonstrated these effects of CLA [48].

**3.4. Conjugated linoleic acid** 

which are isomer specific [46, 47, 48].

Linoleic Acid, 18:2 n-6

CLA, 18:2 c-9, t-11

CLA, 18:2 t-10, c-12

The effects of feeding CLA to pigs have been evaluated in regard to fat quality [57]. Gilts fed 1% CLA for seven weeks had firmer bellies, higher levels of saturated fatty acids, lower levels of unsaturated fatty acids and decreased IV when compared to controls [58]. Barrows fed CLA had improved feed efficiency, decreased backfat, and improved loin marbling and firmness when CLA was included at 0.75% of grow-finish diets [51]. When CLA was fed to genetically lean gilts for eight weeks, an increase in average daily gain and gain:feed was observed [59]. The same study also noted an increase in saturated fatty acids, decrease in unsaturated fatty acids, and an increased level of saturation of the belly tissue [59]. Several studies have shown that CLA feeding increases fatty acid saturation, and firmness in back fat and belly fat [60, 61, 62]. Additionally, use of CLA when feeding by-products may alleviate some or all of the negative impact on carcass quality. When feeding 0, 20, or 40% DDGS during the final 30 days of the finishing period, the addition of 0.6% CLA minimized the negative impact of 20% DDGS inclusion on carcass lipid quality but was unable to overcome the negative effects of feeding 40% DDGS [63].

#### *3.4.2. Mechanism of CLA to alter lipid metabolism*

Dietary CLA in several species alters the activity of SCD-1, FAS, and ACC in adipose and liver. Conjugated linoleic acids decrease mRNA for FAS and ACC to significantly inhibit the capacity for *de novo* lipogenesis [47, 60]. In barrows and gilts fed 0.25 or 0.5% CLA for the finishing diet from 97 to 172 kg, ACC activity was significantly reduced compared to control pigs [64]. Alleviation of negative impacts of nutritional stress of lipid quality, such as during DDGS feeding, is likely through altered lipid metabolism as adipose mRNA expression of ACC was decreased with CLA supplementation with all inclusion levels of DDGS [39].

#### 222 Lipid Metabolism

Reductions in SCD-1 expression were observed with CLA feeding in both mouse liver and cultured preadipocytes [48]. Previous studies indicate that CLA tends to decrease both SCD-1 [65] and decreases the ∆9 desaturase index in pigs [65, 66]. Decreasing SCD-1 mRNA expression, and thereby decreasing the amount of saturated fatty acids being converted to unsaturated fatty acids, may be responsible for the increased levels of saturated fatty acids observed after feeding CLA [65, 66].

Impacts of Nutrition and Environmental Stressors on Lipid Metabolism 223

Within the thermoneutral zone of mammals, core body temperature is maintained without expending additional energy to warm or cool basal body temperature [72, 73]. If the environmental conditions are below this zone, additional energy of metabolism is devoted to generating heat to maintain the desired core temperature [72]. Conversely, at temperatures above this zone, the animal must dissipate energy to maintain core body temperature through additional heat loss mechanisms such as evaporative heat loss, convection, and conduction [72]. When environmental temperature rises above the point where heat production and heat loss are balanced, the animal is in a state of heat stress [74]. In swine, evaporative heat loss is limited due to their inability to sweat; therefore, heat loss is primarily by respiration, evaporation, and exposure to cool air and wet surfaces for convection and conduction, respectively [72, 73, 75]. As an adaptive mechanism to further cool the body and maintain a homeothermic temperature, the animal decreases feed intake

The optimum temperature for a finishing pig between 54.5 and 118.2 kg of body weight is 18.3°C, with a desirable temperature range between 10°C to 23.9°C [76]. The heat stress index (HSI; Figure 5), published by Iowa State University [77], is a practical guideline outlining temperature and humidity ranges for growing pigs. The HSI classifies environmental temperature and humidity conditions into three zones: alert, danger, and emergency. Within the alert range, producers are advised to monitor animal behavior, increase ventilation, and ensure that water is readily available. The danger range requires additional cooling by spraying or misting with water and increasing air flow. Under emergency conditions, producers are advised to avoid transporting animals, withdraw feed during the hottest part of the day, and reduce light levels. For example, when relative humidity is between 45 and 60%, 25.6°C is the alert threshold, 27.2°C is the danger threshold

For grow-finish swine, housing temperatures above 23.9°C decrease voluntary feed intake and growth rate compared to optimum housing temperatures [76, 78]. Voluntary decreases in feed intake decrease metabolic heat production to help maintain homeothermy [79]. Pigs challenged with heat-stress will have decreased feed consumption and average daily gain;

Nienaber et al. [81] noted that elevated temperature decreased daily feed consumption in both cattle and swine, through decreases in meal size and frequency. Feed intake was reduced by 55 g per degree of temperature increase above 22ºC [82]. A similar decrease in feed intake observed by Collin et al. [75] was coupled to reductions in thermic effect of

Humidity is also influential in the animal's ability to dissipate heat by evaporative heat loss [74]. In a study comparing 50, 65, and 80% humidity levels, respiration rate and rectal temperatures were increased at lower temperatures and 80% humidity compared to when humidity was 50 and 65% [74]. Increasing environmental humidity decreases the efficiency of evaporative cooling, resulting in symptoms of heat-stress occurring at lower

however, feed efficiency is maintained when compared to control animals [80].

in order to decrease the thermal effect of feeding [74].

and 30°C is the emergency threshold.

feeding and heat production.

temperatures.

The c10 t12 isomer of CLA decreases the expression of PPARγ in adipose tissue and increases the expression of PPARα in liver tissue [67, 68, 69]. By acting as a PPARγ modulator, CLA is able to prevent lipid accumulation as shown in cultured adipocytes [70]. Conjugated linoleic acid also acts as a PPARα activator and induces accumulation of PPARresponsive mRNAs in hepatic cells [67] serving to upregulate PPAR-responsive pathways.
