Dietary Supplementation with L-Citrulline Between Days 1 and 60 of Gestation Enhances Embryonic Survival in Lactating Beef Cows

Abstract

Low fertility limits reproductive efficiency in cattle. This study was conducted with multiparous Brangus cows receiving dietary supplementation with or without L-citrulline [Cit; an immediate precursor of L-arginine (Arg)]. During the entire experimental period, cows grazed green pasture and had free access to drinking water and mineral blocks. One hundred and seven (107) cows were assigned randomly to one of three treatment groups: dried distillers grains with solubles (DDGS) without Cit supplement (n = 36); DDGS top-dressed with rumen-protected Cit product (RPAA; n = 36); or unprotected Cit product (RUAA; n = 35). After 2 months of lactation, all cows were synchronized to estrus and were bred once via artificial insemination. From Day 1 to Day 60 of gestation, cows were individually fed once daily 0.84 kg of a supplement (DDGS; control), 0.56 kg of DDGS plus 0.28 kg of RUAA (containing 0.07 kg of unencapsulated Cit), or 0.56 kg of DDGS plus 0.28 kg of RPAA (containing 0.07 kg of rumen-protected Cit). The supplemental dose of Cit was equivalent to 0.5% of the estimated daily intake of 14 kg dry matter from pasture. On Days 40 and 60 of gestation, ultrasound was used to determine pregnancy rates. Each pregnant cow had a single conceptus. On Day 60 of gestation, blood samples were obtained from the jugular vein. All cows grazed normally and appeared healthy. Birth rates for live-born calves were 22% and 35% in cows receiving DDGS alone and Cit supplementation, respectively (p < 0.05). The beneficial effect of Cit was associated with increases in concentrations of Cit (+19%), Arg (+20%), ornithine (+19%), proline (+17%), and insulin (+82%) but decreases in concentrations of ammonia (-14%) in maternal plasma (p < 0.05). Thus, dietary supplementation with Cit is a simple, novel, and cost-effective nutritional method to increase the reproductive efficiency of lactating beef cows.

Keywords: arginine; beef cows; citrulline; conceptus; nutrition; pregnancy rate.

Figure 1

Timeline of the experiment. After 2 months of lactation, all beef cows were synchronized to estrus (Syn), followed (one week later) by artificial insemination (AI; Day 0 of gestation). One day after the AI services, cows in the control [dried distillers grains with solubles (DDGS) only], rumen-protected citrulline (RPAA), and unprotected citrulline (RUAA)] groups were fed their respective supplements once daily before grazing pasture for 60 days. Pregnancy (Preg) was checked on Days 40 and 60 of gestation for each cow using the transrectal ultrasonography method. Between Day 61 of gestation and parturition, cows fully grazed on pasture without any DDGS or citrulline (AA) supplementation. After the end of the 60-day period of AA supplementation, calves from the previous pregnancy continued to stay with their mothers until they were weaned at 6 months of age.

Figure 2

Proposed biochemical mechanisms responsible for the beneficial effect of dietary L-citrulline (Cit) supplementation in improving pregnancy outcomes in beef cows. Dietary Cit bypasses catabolism in the rumen to enter the small intestine, where it is absorbed into the portal circulation. Cit is not taken up by the liver and instead is effectively used for the synthesis of L-arginine (Arg) by extrahepatic tissues and cells (e.g., the kidneys, placentae, and endothelial cells) in gestating dams. Arg is the precursor for the formation of nitric oxide, polyamines, and creatine that are essential for placental angiogenesis and growth, uterine–umbilical blood flow, the transfer of oxygen and nutrients (e.g., amino acids, glucose, water, fatty acids, vitamins, and ions) from mother to fetus via aquaporins (AQPs), and conceptus energy metabolism. In addition, Arg stimulates the secretion of insulin from pancreatic β-cells and activates the mechanistic target of rapamycin (MTOR) signaling pathway in cells, thereby promoting the initiation and elongation of protein synthesis while inhibiting proteolysis. Furthermore, Arg increases the expression of genes for the synthesis of glutathione (GSH), as well as antioxidative and anti-inflammatory responses in maternal tissues and conceptuses. Finally, as an activator of N-acetylglutamate (NAG) synthase, which catalyzes the formation of NAG (an allosteric activator of carbamoyl phosphate synthase), Arg enhances the removal of ammonia (a metabolite that is highly toxic to the conceptus at elevated concentrations). Abbreviations: EHT, the conversion of citrulline into arginine in extra-hepatic tissues in the presence of aspartate via argininosuccinate synthase (ASS) and argininosuccinate lyase (ASL); ↑, increase or improve; ↓, decrease; “+”, activation.