2. The Energetic Importance of Fat in Ruminant Nutrition
The intensification of the rearing of polygastric animals, mainly dairy cattle, has contributed to an increased number of scientific reports on the impact of inadequately balanced rations in terms of animal energy requirements and the impact of supplementing ruminant diets with various natural energy sources
[23]. The energy balance associated with the transition into lactation is a challenge for many breeders. The problem of energy deficiencies in dairy cows intensifies especially during the transition period
[24]. During the periparturient period, cows reduce feed intake, which, along with increased energy requirements (necessary for lactogenesis and fetal growth) that exceed the available amount for the animal, contributes to a state of negative energy balance
[25]. Dairy cows that do not have an adequate adaptive response to the negative energy balance may experience the onset of metabolic disorders, including subclinical ketosis. Moreover, energy deficits can also extend over a much longer period of milk production in cows
[26]. The most common reason for worsening a negative energy balance is the insufficient energy value of feed intake compared to the requirement
[27]. Ruminant animals living in the wild are also affected by this issue, as their daily dietary intake is characterized by a relatively low amount of fats, which also contributes to the energy deficit. In addition, during the transition period, there are changes in the hormonal economy in the body, which contribute to changes in the activity of body tissues by increasing lipolysis and decreasing lipogenesis. Lipolysis at the beginning of lactation is a genetically determined process (homeorhesis), while the enzymes involved in lipogenesis are regulated by energy intake (homeostasis)
[28]. The result of the changes that have occurred is the metabolic mobilization of adipose tissue activated by, among others, catecholamines, cytokines, and growth hormones, leading to the breakdown of triacylglycerols present in adipocytes (fat cells) into glycerol and non-esterified fatty acids (NEFAs) to be released into the bloodstream (
Figure 2)
[29]. They are transported by various lipids, which include neutral lipids (NLs) and phospholipids (PLs)
[25].
Figure 2.
The process of lipolysis involved in fat mobilization.
Neutral lipids consist of triglycerides, diglycerides, monoglycerides, and cholesterol esters. The fatty acid profile of individual lipids varies, e.g., linolenic acid is found in neutral lipids, while polyunsaturated fatty acids (PUFAs) are found in phospholipids. During lipolysis, circulating NEFAs released from adipose tissue enter the liver and can be utilized in four pathways: (1) they can be completely oxidized for energy in the Krebs cycle, (2) they are converted to beta-hydroxybutyrate (BHB), (3) they can be re-synthesized to triglycerides (TG), where they can either be exported by very low-density lipoproteins (VLDL) or stored in the liver, or (4) they can be used to synthesize milk fatty acids in the mammary gland
[30]. The beta-oxidation of fatty acids occurs in the mitochondria of liver cells, and the resulting Acetyl CoA releases energy in the tricarboxylic fatty acid cycle
[31][32]. Carnitine palmitoyltransferase-I (CPT-I) is a limiting enzyme for the transport of fatty acids into the mitochondria where beta-oxidation occurs
[33][34]. In cows experiencing a state of negative energy balance, an increase in the activity of carnitine palmitoyltransferase I (CPT-I) is observed, as well as an upregulation of beta-oxidation
[34]. When the mobilization of fatty acids from body tissues is very intense, the acetyl CoA generated through B-oxidation is converted into ketone compounds such as acetoacetate and B-hydroxybutyric acid
[35]. The rate of formation of ketone bodies is not as energy-efficient as B-oxidation. In early lactation, however, ketogenesis is an important process because oxidized ketone compounds provide energy for vital organs
[36][37]. Free fatty acids that do not undergo hepatic B-oxidation are re-esterified into triglycerides and enter the bloodstream as very low-density lipoproteins (VLDLs). The intensity of the re-esterification process increases in a negative energy balance (NEB) state. However, the rate of VLDL transport from the liver of cows is low due to the low synthesis capacity of proteins (apoprotein B) that control the rate of VLDL synthesis and secretion
[38][39]. These mechanisms result in the accumulation of triglycerides in hepatocytes, leading to fat cow syndrome. Furthermore, cows with excessively high body condition scores (BCSs) in the early stages of the drying period mobilize fat tissue to a greater extent before parturition than those with normal conditions or low BCSs
[40]. Reducing the severity of NEB and triglyceride accumulation in the liver is possible, for example, by shortening the dry (nonlactating) period. Elevated concentrations of NEFA result in a reduction in cell number, chemotactic capacity, phagocytic activity, and the oxidative burst activity of polymorphonuclear cells
[41]. In addition, NEFA can lead to an increase in the number of reactive oxygen species and the breakdown of neutrophil granulocytes, a decrease in leukocyte proliferation and stimulation, and a decrease in the secretion of IgM and IFN-γ
[41]. Higher concentrations of those metabolites are typical in dairy cows in a transition period; however, the low capacity of the animal body to break them down results in the risk of production parameter disorders and the health status of cattle.
During the periparturient period, a negative energy balance is, to some extent, a biological necessity due to high energy requirements with insufficient energy intake from feed. At the same time, hypoglycemia occurs as a result of the increased demand for glucose—the primary precursor for the synthesis of milk lactose, which controls milk volume by maintaining the osmolarity of milk. In addition, during the resulting negative energy balance (NEB), cows use the release of nutrients from their body tissues as a key mechanism to maintain lactation as compensation for reduced nutrient intake from their diet
[42]. As a result of adipose tissue mobilization in animals, an increase in saturated and monounsaturated fatty acids can contribute to the compensation of the negative energy balance.
An increased ratio of unsaturated fatty acids (UFAs) to saturated fatty acids (SFAs) and the ratio of n-3 to n-6 fatty acids (PUFAs) can lead to biological membrane dysfunction and the onset of oxidative stress, which, in turn, can affect the health of dairy cows
[42]. This is extremely important because fat mobilization can result in too much lipid peroxidation in the liver and free radical formation, which can escalate oxidative stress in the animal body
[43]. Consequently, there is a great need for nutritional strategies to reduce the adverse effects of NEFA by mitigating fat mobilization and accelerating NEFA removal from the liver. An adequate supply of energy with the feed ration can reduce the severity of body fat mobilization to some extent
[27]. Among these, fatty acid supplementation, which has the effect of not only increasing the energy density of the diet but also mobilizing immune cells and the inflammatory response
[44]. In addition, polyunsaturated free fatty acids (FFAs), such as n-3 fatty acids and conjugated linoleic acids, are fed to reduce "de novo" fatty acid synthesis in the udder and thus the TAG content of milk, which can have modest benefits for the overall energy balance.
Furthermore, an excessive proportion of SFA caused by fat mobilization processes in the cell membrane of white blood cells significantly affects immune dysfunction and the formation of an unregulated inflammatory response in early lactation. This is due to a change in the profile of fatty acids in cell membranes, resulting in a change in the synthesis of oxylipids (also known as oxylipins or eicosanoids), which are responsible for anti-inflammatory and antimicrobial responses
[45]. Ling et al.
[46] demonstrated the impact of the lipid mobilization process occurring during the transition period on the fetus. The offspring of cows excessively exposed to non-esterified fatty acids were characterized by an increased index of the body’s oxidative status, which is expressed as the ratio of reactive oxygen and nitrogen species to the total antioxidant potential. The mechanism explaining the relationship between prenatal exposure to high maternal NEFA concentrations in late gestation and increased oxidative stress in born calves is unknown. However, in vitro studies indicate that fatty acids can directly induce the production of reactive oxygen and nitrogen species (RONS) in cells and indirectly affect oxidative stress in the organism through the activation of metabolic inflammatory pathways
[47]. Therefore, increasing the energy value of the diet, on the other hand, may have a beneficial effect in reducing the duration and severity of a negative energy balance condition while maintaining high milk yields in high-yielding dairy cows
[48].
3. Lipid Metabolism in Rumen—Impact of Rumen Microbiota on FA Biohydrogenation and Transformation and the Role of By-Pass Fat
The diet of ruminant animals contains a small amount of fat. The daily rations contain no more than 6% lipids, half of which comes from forage and the rest is supplemental fat
[22]. Unlike monogastric animals, due to chewing and rumen microbial activity, fatty acids reach further sections of the digestive tract in an altered form
[49]. The intensive metabolic transformation of feed fats results in changes in the absorption of fatty acids and their synthesis in the body. The two main metabolic processes are the hydrolysis of esters to release free fatty acids (lipolysis) and the biohydrogenation of unsaturated fatty acids (
Figure 3).
Figure 3. Lipid transformations in the rumen of dairy cows (based on Bauman et al.
[50]). Abbreviations: TG—triglycerides; GL—glycolipids; FA—fatty acids; VFA—volatile fatty acids; PL—phospholipids; SFA—saturated fatty acid; MC—Microbial cells.
Several types of rumen bacteria are involved in this process
[51]. The first step in rumen lipid metabolism is the lipolysis of feed lipids, which is the source of glyceride esters, consisting mainly of triacylglycerols from concentrates and galactolipids and phospholipids from forages, except for ensiled forages, from which free FAs are released by plant lipases. This process is determined by several factors, and the most important are rumen pH and the amount of fat in the daily ration, as too much of it decreases the activity of the lipolysis process
[52][53]. As a result of this transformation, lipids entering the rumen are first converted by microbial lipases. Among the most active bacteria involved in lipolysis is
Anaerovibrio lipolytica. However, their metabolites only affect triacylglycerols. In the case of galacto- and phospholipids, the most involved are lipases derived mainly from the bacteria of the genus
Butyrivibrio. As a result of their action, ester bonds of complex lipids are hydrolyzed so that unsaturated fatty acids such as linoleic acid (cis-9, cis-12 C18:2), linolenic acid (cis-9, cis-12, cis-15 C18:3), or oleic acid (cis-9 C18:1) are released along with glycerol. The rate of their transformation increases as the degree of unsaturation increases
[54]. FAs released by lipolysis are rapidly hydrogenated by the action of bacterial isomerases, followed by reductases. The isomerization process contributes to a change in the geometric configuration of the acids. Due to the displacement of the double bond and its transformation from the
cis to the
trans form, conjugated linoleic acids are formed. In addition, the formation of several isomers is also observed, among which the predominant fatty acid trans-11 stands out. The next step is the reduction in double bonds, which leads to the hydrogenation of the
trans double bonds formed during isomerization. Those fatty acids are partially incorporated into the bacteria responsible for the biohydrogenation process, during which polyunsaturated fatty acids (n-3 or n-6) are converted to monounsaturated (MUFA) or saturated (SFA) acids. The feed-derived acids, i.e., linoleic acid: C18:2, n-6 (LA) and C 18:3, n-3 linolenic acid (LNA) undergo biohydrogenation to stearic acid: C18:0. This process also results in the formation of trans-11-vaccenic acid (TVA, C18:1) and conjugated linoleic acid (CLA C18:2, cis-9, trans-11), along with isomers that are absorbed in the small intestine
[52]. Only a small proportion of unsaturated fatty acids can enter the lumen of the small intestine in an unchanged form, from which they are absorbed into the lymph in the jejunum
[52]. The amount of vaccenic acid (VA, trans-C18:1) entering the duodenum of cows’ ranges from 20 to 140 g, which is about 10–20% of the amount of PUFA fatty acids in the daily ration
[52].
The degree of biohydrogenation depends primarily on the type of fat, the length of incubation time in the rumen, as well as the composition of the rumen microbiota population. Bacteria such as
Butyrivibrio fibrisolvens,
Eubacterium spp.,
Ruminococcus albus,
Borrelia,
Micrococcus, and
Fusocillus spp. are involved in this process
[55]. The amount of long-chain unsaturated fatty acids affects the degree of the biohydrogenation of polyunsaturated fatty acids and decreases with an increase in saturated FAs
[55]. It is also dependent on the size of the rumen: in small ruminants (goats and sheep), the degree of total biohydrogenation is lower
[56]. The introduction of a fat supplement into the diet of ruminants, especially one containing polyunsaturated fatty acids, can, therefore, have a negative impact on the rumen fermentation process and ultimately affect total feed digestibility. The excessive supplementation of ruminant animals with fats may result in increased amounts of certain trans fatty acids in livestock products and a decrease in the fat and protein content of milk
[57][58]. Regarding growing ruminants, the dietary inclusion of lipids must be restricted (to ca. 60 g/kg DM consumed) to avoid the impairment of the rumen function
[59]. For this reason, the manipulation of ingredients in the ration abundant in PUFA fatty acids available to the rumen flora is limited
[60]. Their effect depends on the amount of additive introduced, its form, such as raw or processed (extruded, micronized) oilseeds, and the composition of the diet
[61][62].
Increasing the energy value of ruminant animal feed without reducing its digestibility is achieved through the use of appropriate additives in the form of specially formulated fats called rumen-protected fats, including calcium soaps, which provide an important safeguard against rumen biohydrogenation
[63]. One of the main advantages of using protected fat is its high digestibility rate, which is as high as 85%. Protected fats entering the rumen remain in their original form. In addition, they are inert to the rumen environment; therefore, they do not cause changes in their fermentation processes and lower the activity of cellulolytic microorganisms. Protected fat can be fed in the form of oilseeds that have undergone previous thermal processes, most often extrusion and micronization. Heat treatment in the form of an extrusion process of, for example, flax seeds, with optimally selected parameters
[64] can change the site of protein digestion from the rumen to the small intestine without adversely affecting total nutrient digestibility
[61], thus contributing to a faster fat absorption process
[65]. The addition of extruded flaxseed not only increases the energy concentration in the ration but contributes to improving reproductive rates in ruminant animals through the specific effects of n-3 fatty acids
[66], as well as increasing the proportion of polyunsaturated fatty acids contained in the milk of cows
[61]. Extrusion also increases the absorption rate of amino acids available to the intestine, increasing the pool of amino acids available for protein synthesis in the mammary glands
[67]. The complete digestion of protected fat takes place in the small intestine through the action of pancreatic lipase, and unchanged unsaturated fatty acids are absorbed in the small intestine
[68]. They provide a highly cost-effective alternative for starch-restricted feeds without reducing energy levels in the ration of ruminant animals and without compromising productivity. Behan et al.
[69] confirm this, claiming that the use of protected fats also plays a significant role in improving animal performance without affecting the rumen microbiome.
4. Regulatory Role of N-3 Fatty Acids in Reproduction of Ruminants
In the reproductive process, ovulation plays a key role, enabling the mature oocyte to have the ability to fertilize
[70]. The process of mammalian oocyte growth involves an important epigenetic process—reprogramming the maternal genome. This process involves changes in the cytoplasmic organelles, synthesis, and storage of mRNAs and proteins that are essential for the initial cell cycles of embryogenesis, the resumption and termination of meiosis, and epigenetic modifications
[71]. Striving for optimal metabolic health in dairy cows is a key strategy to ensure adequate ovarian physiology and high-quality oocytes and embryos
[72].
Fatty acids (FAs) play a key role in oocyte developmental programming
[73]. The concentration of n-3 fatty acids in an organism has an impact on the maturation process and developmental competence of oocytes
[74]. Introducing an appropriate diet containing n-3 PUFAs can have a positive impact on oocyte quality, maturation ability, and the probability of conception in cows. Studies conducted on Holstein heifers have confirmed the importance of fatty acids in oocytes and their impact on developmental programming
[71].
Polyunsaturated fatty acids (PUFAs), by altering hormone and metabolite levels, can affect various levels of the hypothalamic–pituitary–ovarian axis and locally stimulate follicle growth. Studies confirm that altering fatty acid (FA) intake in the diet of dairy cows can affect the concentration of circulating metabolic hormones and ovarian steroids, which, in turn, can affect the development and capacity of ovarian follicles and the composition and secretion of reproductive tissues
[75][76]. Acting as the endogenous stimulators of peroxisome proliferator-activated receptors (PPARs), n-3 fatty acids influence the regulation of sex steroid hormones involved in the processes of ovarian follicle growth and differentiation, oocyte maturation, and embryo development
[77].
During the ovulation process, the mature oocyte is surrounded by ovarian follicles that gradually grow
[78]. During the pre-ovulatory stage, the oocyte accumulates nutrients, mRNAs, proteins, and mitochondria and is surrounded by specialized hill cells and antral cavity fluid
[79]. The oocyte’s ability to develop into a viable embryo after fertilization is acquired mainly during terminal growth and meiotic maturation. The LH signal is transmitted through the granulosa and thalamus cells, which triggers the resumption of meiosis and the completion of the nuclear maturation of the oocyte into the metaphase II (MII) stage of meiosis, preparing it for fertilization
[80][81]. These rapid and synchronized events, such as the proliferation of granulosa layer cells, generation of the thalamic cell matrix, and segregation of chromosomes, are energy-intensive processes that require the production of sufficient ATP
[82]. Fatty acids play a key role as an energy source for the oocyte. The oocyte takes up both exogenous lipids and synthesizes and stores endogenous lipids in the ooplasm
[83]. Lipids have an important function, namely, signaling molecules in the regulatory mechanisms of oocyte maturation and competence acquisition
[84]. Free fatty acids (FAs), including unesterified FAs, are present in both the follicular fluid and the oocyte–cell complex (OCC) and are involved in the formation of the OCC follicle environment, which is directly related to oocyte quality
[85].
In vitro studies have confirmed the positive or neutral effects of n-3 PUFA treatment on oocyte maturation and developmental competence
[86]. Sharma et al.
[76] reported that increased follicle size in cows improves both oocyte quality and corpus luteum (CL) function. In cattle, on the other hand, PUFA supplementation during in vitro studies has led to disruptions in the expression of genes involved in the lipid metabolism of hill cells. These changes modify the molecular mechanisms that regulate oocyte maturation
[74]. Then, there are contradictory findings on the effect of n-3 PUFA on oocyte and embryo development
[87], suggesting that feeding cows flaxseed has no effect, which showed a reduction in oocyte fertilization and embryo quality. A study by Moallem et al.
[88] suggested a direct beneficial effect of n-3 PUFA fatty acids on the ovaries. In fact, in vitro embryo production performed after transvaginal ultrasound-guided oocyte retrieval (oocyte retrieval) without ovarian stimulation was increased after n-3 PUFA supplementation compared to a control consisting of saturated FA supplementation.
Dietary n-3 PUFA supplementation may be particularly recommended in cows with high genetic potential and unpredictable oocyte quality
[89]. Studies on the effects of n-3 fatty acids (N-3 PUFAs) on reproduction in ruminants mainly focus on the study of biomarkers that regulate ovarian function, ovarian processes such as steroidogenesis, and oocyte competence
[90].
The modification of fatty acids in animal diets can affect the concentration of circulating metabolic hormones and steroid hormones in the ovaries. This can affect the development and capacity of ovarian follicles and the composition and secretion of reproductive tissues. One of the parameters determining the reproductive success of animals is prostaglandin (PG) concentrations. This allows to assess the effect of n-3 PUFAs on processes in the uterus that regulate the estrous cycle and pregnancy formation
[91]. Prostaglandins (PG) play a key role as local signaling molecules. Their precursor is arachidonic acid (AA; C20:4), which is found in cell membranes as a component of the phospholipid bilayer
[92]. The process of AA release from cell membranes occurs through the action of the enzyme phospholipase A2 (prostaglandin-endoperoxidase 2—PTGS2), previously known as cyclooxygenase, which converts AA into the prostaglandin PGH2
[93]. PGH2 is a precursor for various series 2 prostaglandins, such as PGF2α, PGE2, PGD2, PGI2, and thromboxane A2 (TXA2)
[94].
An increased proportion of n-6 fatty acid-rich components (e.g., corn, soybeans) in the ration has been shown to contribute to the synthesis of PG series 1 and 2, while n-3 supplementation induces the production of PG series 3 (
Figure 4)
[95].
Figure 4. Metabolism of prostaglandins series 1, 2, and 3 (based on Park et al.
[95]). Explanations: C18:2 n-6—linoleic acid; C18:3 n-6—γ-linolenic acid; C20:3 n-6—dihomo- γ linolenic acid; C20:4 n-6—arachidonic acid; C18:3 n-3—α-linolenic acid; C18:4 n-3—stearidonic acid; C20:4 n-3—eicosatetraenoic acid; C20:5 n-3—eicosapentaenoic acid; PG1, PG2, PG3—prostaglandin 1 or 2 or 3.
During diestrus, there is an increase in the expression of the cyclooxygenase enzyme PTGS2, which leads to the synthesis and release of prostaglandin PGF2α, which is associated with the process of luteolysis or yellow body atrophy, which is crucial in regulating the estrous cycle
[44][96]. In contrast, early in pregnancy, there is an increased production of prostaglandin PGE2 under the influence of type I interferon, which regulates the expression of the enzyme PTGS2 in the endometrium
[97]. PGE2 helps maintain the luteal phase, which is essential for maintaining pregnancy and promotes embryo elongation and implantation. N-3 PUFA, especially docosahexaenoic acid (DHA), is a potent inhibitor of the activity of the enzyme PGHS (prostaglandin endonadoxide synthase) involved in the production of eicosanoids. Reducing the synthesis of PGE2s is a beneficial process as the excess of prostaglandin E-2s contributes to abnormalities in oocyte meiosis and fertilization
[83]. In addition, a significant proportion of bovine embryos are believed to fade due to the insufficient inhibition of uterine PGF2α. There are several mechanisms contributing to the reduction in prostaglandin F2α activity. The first is a reduction in the expression of genes involved in uterine PG synthesis. A study by Dirandeh et al.
[77] showed reduced PGFM concentrations in cows whose rations were based on elevated α-linolenic acid, indicating a significant reduction in the amount of PGF2α secreted in the uterus. Cows subjected to the test diet also had a reduced expression in PGFS, which converts prostaglandin H2 (PGH2)—an unstable intermediate compound in the PG biosynthesis pathway—into prostaglandin F2α. Research by Plewes et al.
[98] illustrated another important effect of changing the fatty acid composition of membrane phospholipids on reducing PGF2α secretion. Cows in early pregnancy fed for 60 days on feed supplemented with n-3-rich fishmeal were characterized by, among other things, a modified membrane lipid microdomain, resulting in reduced prostaglandin F2α signaling. As the researchers explained, the mechanism of the PUFA’s effect on membrane lipid microdomains involves altering the distribution of protein molecules, and, subsequently, transforming the protein–lipid complex or protein–protein interactions. Moreover, n-3 family fatty acids can affect the availability of arachidonic acid (AA) in endometrial cells
[99]. Studies have shown that the endometrium of lactating dairy cows contains a significant amount of AA acid on day 17 of the estrous cycle (about 15.3 g/100 g FA) when PGF 2α secretion is much lower than during the peripartum period
[100]. N-3 fatty acids have been shown to effectively reduce AA concentrations in animals. Giller et al.
[91], after supplementing the diets of heifers with 450 g of rumen-protected fish oil (omega 3 FA), observed a reduction in the concentration of AA in their bodies. This effect is a result of the specific metabolism of n-3, in which the process of competitive inhibition of Δ6 desaturase takes place. This is to prevent the synthesis of arachidonic acid (AA), excluding it from the phospholipid bilayer and competitively inhibiting the enzyme cyclooxygenase-2 (COX-2)
[101].
The dietary supplementation of dairy cows with n-3 PUFA has been associated with a larger corpus luteum (CL) size and higher blood progesterone levels, which, in turn, increases the number or size of ovarian follicles
[102]. Freret et al.
[103] also reported that cows receiving a diet supplemented with flaxseed showed an increase in the size of pre-ovulatory follicles and small follicles. This relationship was confirmed by a study by Ulfina et al.
[104] in which cows received either a supplement of crushed flaxseed at 750 g/day/head, 250 g/day/head of butyric acid, or a diet without additives. They found that cows given flaxseed, which is a source of n-3 fatty acids, had a significantly larger dominant ovarian follicle size (12.78 ± 0.62 vs. 10.54 ± 0.57) and larger corpus luteum size. In contrast, in a study where Sinedino et al.
[105] used algae supplementation in the diets of dairy cows, at 0 or 100 g/cow of algae product containing 10% DHA, from day 27 to 147 postpartum, it was shown that supplementation had no effect on either ovarian follicle diameter or progesterone concentration. The effect of feeding hemp seed cake on reproductive parameters in cattle was studied by Windersen et al.
[106]. Angus heifers whose diets contained 20% hemp cake as a substitute for 20% dried corn distillers’ broth were characterized by the regulated concentrations of the microorganisms present in the vagina and uterus. According to the authors, the achieved effect is important from the point of view of the reproductive parameters, since the bacterial infection of the uterus can contribute to disorders in the development of ovarian follicles. This is due to the secretion of lipopolysaccharides (LPSs) and tumor necrosis factor-α (TNF-α), which can limit ovarian follicle growth and estradiol production. As a result, the probability of maturation and ovulation of the first dominant ovarian follicle in infected cows is lower. The results obtained may be determined by the antimicrobial components contained in hemp cake, such as cannabinoid derivatives, which affect a set of Gram-positive and Gram-negative pathogens
[107].
Reproductive performance in ruminants plays a key role in the profitability of animal husbandry. However, reproductive disorders in dairy cattle are major problems contributing to infertility and low dairy productivity. Insufficient or unbalanced nutrient intake can cause reproductive failure through the malfunction of reproductive organs
[108]. Major metabolic changes are required to meet production demands. Dietary manipulation to improve fertility has shown great promise. A proper supply of n-3 polyunsaturated fatty acids (PUFA n-3) in the diet of dairy cows can increase the chances of conception
[102] and reduce the risk of pregnancy loss
[77]. Dietary supplements can be useful in improving the reproductive performance of cows by helping to reduce the negative energy balance that occurs after parturition, leading to an earlier return to the estrous cycle and improved fertility
[99]. As a result, providing an adequate diet and supplementation can improve the reproductive performance of dairy cows, which is key to increasing milk production efficiency and sustainability in the livestock industry
[109]. The results of the observed effect of n-3 FA supplementation on the reproductive performance in cows are summarized in
Table 1.
The use of flaxseed in various forms in ruminant nutrition during the breeding season contributes to improved reproductive performance and better nutrient utilization, which determines measurable health benefits
[110]. Swanepoel and Robinson
[111] reported that the use of a flaxseed component characterized by a high content of alpha-lipoic acid (ALA) in the feeding of dairy cows has a significant effect on conception rates and reduces the number of abortions. A study conducted by Sinedino et al.
[105] showed that supplementing the diet of dairy cows with seaweed rich in docosahexaenoic acid (DHA) at 100 g per cow, from 27 to 147 days postpartum, had a positive effect on animal reproduction. Increased levels of interferon (IFN)-stimulated genes in peripheral blood leukocytes (PBLs) were observed, indicating better reproductive fitness. In addition, more pregnancies were observed in algae-fed cows, both primiparous and multiparous, which is associated with enhanced embryonic development due to increased interferon-τ (IFN-τ) production. Daily dietary supplementation with DHA-rich algae improved estrous cyclicity and pregnancy rates at first artificial insemination in primiparous cows, as well as overall pregnancy rates, and reduced time to pregnancy in all cows. An examination of peripheral blood leukocytes from pregnant cows fed algae also showed increased expression of the RTP4 gene, which was associated with the amount of interferon-tau secreted by the fetus compared to the control group
[105]. Elis et al.
[102] analyzed the effects of n-3 fatty acids on the reproductive parameters of 46 Holstein cows. Animals that received a basal diet along with the addition of protected fish oil at 1% of dry matter between calving and 2 months after calving were characterized by a reduced rate of failure to fertilize and/or early embryo mortality. The study by Gonzalez et al.
[112] was conducted on 112 animals: 61 heifers and 51 Holstein Friesian cows. The animals were randomly divided into two experimental groups. The first group consisted of 66 animals (30 cows and 36 heifers), which received a commercial concentrate with added fats from extruded flaxseed (source of linolenic acid) and soybean oil (source of linoleic acid). The second group, consisting of 46 animals (21 cows and 25 heifers), received no fat supplements but were given the same diet as the animals in the first group without fat supplements. The supplementation of the first group lasted from 3 weeks before calving until the second month of lactation. The results suggest that supplementation had a positive effect on conception rates in heifers, reducing the days from calving to the first artificial insemination. In contrast, no significant differences were observed between treatments in reproductive parameters in cows. Velazquez et al.
[113] showed in their study that supplying n-3 PUFAs to the diet of small ruminants during pregnancy alters the physiology of the offspring. An analysis of the fetuses of animals fed 10 g/kg Ca salt (a source of n-3 PUFAs) from day 100 to 145 of gestation showed increased liver weight compared to the control group. In addition, beneficial effects on the development and growth of the offspring were observed, as well as the modification of the expression of immune mediators, enzymes involved in DNA methylation, and an acceleration in fetal growth.
Table 1.
Impact of n-3 fatty acid supplementation on reproductive performance—research overview.