N-carbamoylglutamate Research Articles

Supplementation with N-carbamoylglutamate during the transition period improves the function of neutrophils and reduces inflammation and oxidative stress in dairy cows

The aim of this study was to investigate the effects of N-carbamoylglutamate (NCG) supplementation during the transition period on the functions of blood polymorphonuclear neutrophils (PMN), inflammation, and oxidative stress in dairy cows. Thirty multiparous Chinese Holstein dairy cows at wk 4 before parturition were blocked into 2 groups by parity, body weight, and milk yield of previous lactation, and randomly allocated to 2 dietary treatments of basal diet supplemented without (control, n = 15) or with 20 g/d per cow of NCG (NCG, n = 15). The supplementation was carried out from d -21 to 21 relative to calving. Health incidents (mastitis, retained placenta, and lameness) were recorded, and blood samples were collected at d -21, -7, 0 (the calving date), 7, and 21 relative to parturition and analyzed for variables related to inflammation and oxidative stress. In addition, whole blood was collected at d 7 to isolate PMN and used for analysis of the expression of functional genes and from d -21 to 21 for determination of weekly hematological parameters. The number of lymphocytes was greater at d 7 in the blood of NCG cows. The plasma level of malondialdehyde was lower in the NCG group, and blood reactive oxygen species were lower at d 7, whereas total antioxidant capacity tended to be greater in the NCG group and glutathione peroxidase tended to be higher at d 21 in cows fed NCG, suggesting that NCG supplementation improved antioxidation in cows. In addition, the concentration of serum amyloid A was lower in NCG-fed animals during the postpartum stage. Blood concentrations of IL6 and tumor necrosis factor-α were lower and tended to be lower in NCG-fed animals at d 7, respectively. Meanwhile, the concentrations of IL6 tended to be lower in NCG-fed animals at d 21. Furthermore, the expression of S100A9 and MMP9 in the PMN was lower and tended to be lower, respectively, whereas the expression of ITGB2, XBP1 tended to be higher and expression of CLEC6A was higher in NCG-fed cows. Overall, our results indicated that supplementation with NCG during the transition period showed the beneficial effects on animal health, by improving PMN functions and alleviating inflammation status and oxidative stress in dairy cows.

N-Carbamoylglutamate Supplementation on the Digestibility, Rumen Fermentation, Milk Quality, Antioxidant Parameters, and Metabolites of Jersey Cattle in High-Altitude Areas.

This study aimed to assess the impact of the dietary supplementation of N-carbamoylglutamate (NCG) on nutrient digestibility, rumen fermentation, milk quality, oxidative stress, and metabolites in the plasma and feces of Jersey cattle under high altitude with the hypoxic condition. A total of 14 healthy lactating Jersey dairy cows with similar body conditions were selected and randomly divided into 2 groups. The control group (CON group, N = 6 replicates) was fed with a conventional complete diet, whereas the experimental group (NCG group, N = 8 replicates) received 20 g/d per head NCG supplementation. The experiment lasted for 60 days, the adaptation period was 12 days, and the formal experiment period was 48 days. Except that the NCG group showed an upward trend in dry matter intake (DMI) (p = 0.09) and the fermentation parameters, the molar proportion of butyric acid tended to decrease (p = 0.08); the two groups had no significant differences (p > 0.05) in nutrients digestibility, plasma immunity, and antioxidant ability. However, compared with the CON group, the milk fat rate and blood oxygen saturation of the NCG group showed an upward trend (p = 0.09). For indexes associated with altitude stress, the contents of thyroxine, transferrin, and endothelin both decreased significantly (p < 0.05) in the NCG group. Meanwhile, heat shock protein (p = 0.07) and aldosterone (p = 0.06) also showed a downward trend. A total of 114 different metabolites were identified from feces and plasma, 42 metabolites were derived from plasma that mainly included 5 kinds of Super Class, and 72 metabolites were derived from feces that mainly included 9 kinds of Super Class. The significantly increased plasma differential metabolites were 2,5-dihydroxybenzoate and salicyluric acid, and the significantly increased fecal differential metabolites were Butenafine (fold change > 2). Pathway analysis showed that after applying NCG as a feed additive, the changes of the Jersey dairy cows mainly focused on amino acid metabolism and lipid metabolism. These results indicated that adding NCG to the diet can prevent the hypoxic stress state of lactating Jersey cows in high-altitude areas and has a tendency to improve milk quality.

Dietary provision of N-carbamoylglutamate to Holstein cows: A strategy to enhance the productive and reproductive efficiency during summer

The experiment was designed to investigate the effect of N-carbamoylglutamate (NCG) supplementation on productive, reproductive, and metabolic activity of Holstein cows (n = 169) during summer. The pregnant cows were supplemented NCG (20 g/day/cow) during last 20 days of pregnancy and extended for a period of 35 (NCG-35, n = 42), 50 (NCG-50, n = 44) or 70 days (NCG-70, n = 40), whereas, control group (NCG-0, n = 43) did not receive any supplement during the trial. Milk production, milk composition (total solids, lactose, fat, protein, somatic cell count, and urea nitrogen), and metabolic variables (glucose, alanine aminotransferase; ALT, aspartate aminotransferase; AST, arginine, blood urea nitrogen; BUN and nitric oxide; NO) were recorded at various intervals of NCG supplementation. After NCG supplementation, the cows were subjected to estrus synchronization at day 50 of calving using a modified Ovsynch protocol (GPGMH) and observed for reproductive variables. A significantly higher (P < 0.05) milk yield with improved milk lactose, milk protein and milk fat was found in NCG-70 group compared to NCG-0 group. Besides, the estrus response, ovulation rate, ovulatory follicle diameter, and pregnancy rate increased (P < 0.05) in NCG-70 group compared to NCG-0 or NCG-35 group. The serum metabolites (glucose, AST, arginine, BUN, and NO) showed variations at different time points (synchronization, AI, or post-AI) among the treatment groups. In conclusion, NCG supplementation improves the milk yield, milk composition, and serum metabolites in heat stressed dairy cows. The supplementation of NCG prior to GPGMH protocol application enhance the reproductive traits in dairy cows under summer conditions.

Supplementing N-carbamoylglutamate in late gestation increases newborn calf weight by enhanced placental expression of mTOR and angiogenesis factor genes in dairy cows.

The objective of this study was to investigate whether supplementation with N-carbamoylglutamate (NCG) to cows during late gestation alters uteroplacental tissue nutrient transporters, calf metabolism and newborn weight. Thirty multiparous Chinese Holstein cows were used in a randomized complete block design experiment. During the last 28 d of pregnancy, cows were fed a diet without (CON) or with NCG (20 g/d per cow). The body weight of calves was weighed immediately after birth. Placentome samples were collected at parturition and used to assess mRNA expression of genes involved in transport of arginine, glucose, fatty acid and angiogenesis factors, as well as the mammalian target of rapamycin (mTOR) pathway. Blood samples of calves before colostrum consumption were also collected for the detection of plasma parameters, amino acids (AA) and metabolomics analysis. The newborn weight (P = 0.02) and plasma Arg concentration of NCG-calves was significantly higher (P = 0.05) than that of CON-calves, and the plasma concentrations of urea nitrogen tended to be lower (P = 0.10) in the NCG group. The mRNA abundance of genes involved in glucose transport (solute carrier family 2 member 3 [SLC2A3], P < 0.01), angiogenesis (nitric oxide synthase 3 [NOS3], P = 0.02), and mTOR pathway (serine/threonine-protein kinase 1 [AKT1], P = 0.10; eukaryotic translation initiation factor 4B pseudogene 1 [EIF4BP1], P = 0.08; EIF4EBP2, P = 0.04; and E74-like factor 2 [ELF2], P = 0.03) was upregulated in the placentome of NCG-supplemented cows. In addition, 17 metabolites were significantly different in the placentome of NCG-supplemented cows compared to non-supplemented cows, and these metabolites are mainly involved in arginine and proline metabolism, alanine, aspartate and glutamate metabolism, and citrate cycle. In summary, the increased body weight of newborn calves from the NCG supplemented dairy cows may be attributed to the increased angiogenesis and uteroplacental nutrient transport and to the activated mTOR signal pathway, which may result in the increased nutrient supply to the fetus, and improved AA metabolism and urea cycle of the fetus.

N-carbamoylglutamate improves lipid metabolism, inflammation, and apoptosis responses in visceral adipocytes of Japanese seabass (Lateolabrax japonicus), invivo and invitro.

This study applied in vivo and in vitro methods to investigate the effect of dietary N-carbamoylglutamate (NCG) on lipid metabolism, inflammation and apoptosis related-gene expression in visceral adipose tissue and isolated adipocytes of Japanese seabass (Lateolabrax japonicus). A basal diet and a test diet supplemented with 720 mg/kg NCG were fed to the fish for 10 weeks. During the growth trial, no mortality and no significant differences in growth performance were observed in fish between the 2 groups (P > 0.05). Plasma Arg content and mRNA level of argininosuccinate synthetase (ASS) in adipose tissue were significantly increased, which indicated that NCG inclusion promoted endogenous Arg synthesis. Thereafter, the potential effects of NCG treatment on lipid metabolism-related genes expression were studied through in vivo and in vitro methods. In the present study, we successfully established a primary adipocytes culture system and isolated pre-adipocytes in vitro of Japanese seabass for the first time. Both the results in vivo and in vitro showed that NCG treatment decreased the mRNA levels of genes related to adipogenesis (fatty acid synthase, FASN), cholesterol synthesis (3-hydroxy-3-methylglutaryl-CoA reductase, HMGCR) and fat deposition (lipoprotein lipase [LPL] and leptin), which revealed the underlying mechanism of NCG on reducing fat deposition. The results of this study demonstrated that NCG inclusion reduced the expression of inflammatory and apoptosis cytokines markedly in vivo and in vitro. In conclusion, NCG did exert beneficial effects on ameliorating adipogenesis, inflammation and apoptosis via promoting Arg endogenous synthesis in Japanese seabass.

Dietary supplementation with N-carbamoylglutamate initiated from the prepartum stage improves lactation performance of postpartum dairy cows

The objective of this study was to investigate the effects of supplementing N-carbamoylglutamate (NCG), an Arg enhancer, on amino acid (AA) supply and utilization and productive performance of early-lactating dairy cows. Thirty multiparous Chinese Holstein dairy cows were randomly divided into control (CON, n = 15) and NCG (CON diet supplemented with NCG at 20 g/d per cow, n = 15) groups at 4 wk before calving. Diets were offered individually in tie-stalls, and NCG was supplemented by top-dress feeding onto total mixed ration for the NCG group. The experiment lasted until wk 10 after calving. Dry matter intake tended to be higher (P = 0.06), and yields of milk (P < 0.01), milk protein (P < 0.01), and milk fat (P < 0.01) were higher in the NCG-cows than in the CON-cows. Plasma activities of aspartate aminotransferase (P < 0.01), alanine aminotransferase (P = 0.03), and plasma level of β-hydroxybutyrate (P = 0.04) were lower in the NCG-cows than in the CON-cows, whereas plasma glucose (P = 0.05) and nitric oxide (NO, P < 0.01) concentrations were higher. Coccygeal vein concentrations of Cys (P < 0.01), Pro (P < 0.01), Tyr (P = 0.05), most essential AA except Thr and His (P < 0.01), total essential AA (P < 0.01), and total AA (P < 0.01) were higher in the NCG-cows than in the CON-cows. The arterial supply of all AA was greater in the NCG-cows than in the CON-cows. The NCG-cows had higher mammary plasma flow of AA (P = 0.04) and clearance rate of Cys (P < 0.01), Pro (P < 0.01) and Asp (P < 0.01), and higher ratios of uptake to output of Met (P = 0.05), Lys (P < 0.01), Cys (P = 0.01), Pro (P = 0.03), and Asp (P = 0.01). In summary, addition of NCG initiated from the prepartum period improved the lactation performance of postpartum dairy cows, which might attribute to greater Arg and NO concentrations, as well as improved AA supply and utilization, liver function, and feed intake in these cows.

Short communication: Effects of dietary N-carbamoylglutamate supplementation on the milk amino acid profile and mozzarella cheese quality in mid-lactating dairy cows

N-Carbamoylglutamate (NCG) is an enhancer of Arg, which is a functional AA and could prevent cardiovascular disease and improve immunity. The present study was conducted to investigate the effects of supplementing NCG in diets of lactating cattle on the NCG concentration and AA composition of raw milk and on mozzarella cheese quality. Thirty multiparous cows with a mean body weight of 669 kg (standard deviation = 71) and days in milk of 176 (standard deviation = 55) were blocked based on parity and milk production and randomly assigned to 1 of 2 treatments: basal diet (CON) and basal diet supplemented with 40 g of NCG per day per cow (NCG). After 8 wk of treatment, raw milk samples were collected from the 2 groups for AA analysis and mozzarella cheese-making. Furthermore, the NCG concentration and distribution in milk and mozzarella cheese were detected. The AA concentration in milk was greater and the NCG concentration in raw milk was approximately 6 times greater in the NCG group than in the CON group. No NCG was detected in cheese from the CON group, and very little NCG (<1.0 μg/kg) of cheese was detected in the NCG group. Most of the dietary NCG was transferred into whey, stretch water, and brine during cheese production. No significant difference was found between the 2 groups on cheese texture and color except that hardness was lower in the NCG group. Overall, the results indicated that dietary supplementation of NCG could improve the NCG and AA concentrations in raw milk without affecting the quality of cheeses such as mozzarella.

The suppressed autophagy induced by carbon disulfide could be rescued by N-carbamoyl glutamate during the window of embryo implantation in mice

ObjectivesTo explore the effects of carbon disulfide (CS2) and N-carbamoyl glutamate (NCG) on autophagy during the window of embryo implantation in mice and whether dietary NCG supplementation can promote embryo implantation in case of CS2 exposure. MethodsPregnant mice that received single intraperitoneal injection of CS2 on Gestational day (GD)4 were fed basal diet with or without NCG supplementation from GD1 to endpoints. The control mice were injected solvents. There were four endpoints (GD5, GD6, GD7 and GD9 endpoints) in each group. The uterus was collected on endpoints to detect autophagy-related markers by using the methods of transmission electron microscopy (TEM), immunohistochemistry (IHC), quantitative real-time polymerase chain reaction (qRT-PCR) and ELISA. ResultsThe P62 brown punctate staining increased in CS2 exposure group and reduced after dietary NCG supplementation, which was opposite with LC3B, Beclin1 and ATG5 on GD5 endpoint. Simultaneously, P62 protein expression raised 43.33% on GD5 endpoint (p < 0.01) when exposed to CS2 and descended to the control level after NCG supplementation. The rate of decline of LC3B and Beclin1 proteins were 27.04% (p < 0.01) and 23.27% (p < 0.05) on GD5 endpoint, 20.20% (p < 0.05) and 11.30% on GD7 endpoint in CS2 exposure group, respectively, then NCG supplementation caused the LC3B and Beclin1 protein expression to rise in different degrees. Comparatively, the mRNA expression of all autophagy-related gene changed more apparently on three endpoints than the protein expression. The images of TEM showed that nearly no autophagosome could be seen in CS2 exposure group, while dietary NCG supplementation increased the number of autophagosome obviously on GD5 endpoint. The number of implanted embryos which declined due to CS2 exposure returned to normal in NCG supplementation group. ConclusionsDietary NCG supplementation could rescue the suppressed autophagy induced by CS2 in the window of implantation and increase the number of implanted embryos.

Determination of N-Carbamoylglutamate in Rumen Fluid

N-carbamoylglutamate (NCG) is a feed additives obtained synthetically from sodium glutamate,which is a structural analogue of N-acetylglutamate. Current study is aimed at developing ion chromatographic method for the determination of NCG in rumen fluid. Successful determination was performed on cation-exchange column and non-suppressed conductivity detection with total rune time of 11 minutes. Simple one-step cleanup using anion exchange cartridge was used to remove interfering compounds. Method was applied to determine NCG in rumen fluid, which was obtained from three rumen fistulated dairy cows. Incubated NCG sample in rumen fluid was assessed at various time intervals (0, 2, 4, 6, 12 and 24 h at 39 oC). Results showed that NCG was disappeared at the rate of 7.8, 12.8, 13.3, 14.2, and 17.8 % at 2, 4, 6, 12 and 24 h, respectively. It is concluded from the results that the procedure is quick and uncomplicated. NCG can be detected in rumen fluid without lengthy separation using cation-exchange chromatography. This technique can be applied for determination of NCG in rumen fluid as well as for practical use in animals feed.

Effectiveness of citrulline and N-carbamoyl glutamate as arginine precursors on reproductive performance in mammals: A systematic review.

The use of functional nutrients has been proposed to reduce the occurrence of intrauterine growth retardation in animals at birth in several mammalian species. The objective of this study was to verify the effectiveness of citrulline and N-carbamylglutamate (NCG) dietary supplementation as arginine precursors for mammalian species, and the effects on fetal development through a systematic review. The search for studies was performed during August 2018 in the PubMed, ISI Web of Science, Science Direct, and Scopus databases. The literature search was conducted using “arginine precursor”, “citrulline”, or “N-carbamylglutamate” as keywords, combined with “gestation”, “pregnancy”, “fetus”, “newborn”, or “reproduction”. Studies in which arginine precursors were evaluated in gestating mammals and their effects on parameters related to the intrauterine development of the conceptus were selected. Of 1,379 articles, 18 were selected, primarily based on the title and the abstract. Supplementation with NCG (0.5 g to 2 g/kg of feed) increased maternal plasma arginine concentrations in all studies that evaluated this variable. Fetal number increased in 55.56% of the studies that evaluated it, and fetal weight increased in the majority (62.5%) of the studies evaluating this variable. By supplementing citrulline, only fetal weight was improved, with an increase in maternal plasma arginine in 40% of the studies. In conclusion, N-carbamoyl glutamate seems to be an arginine precursor more effective than L-citrulline during gestation; however, both precursors, beside L-Arginine, should be evaluated in similar conditions to confirm the existence of specific particularities such as periods and levels of supplementation, which need to be considered for different species of animals. The supplementation of NCG increases arginine concentrations in maternal plasma, thus improving mammalian reproductive efficiency and fetal development, mainly by promoting higher birth weight.

Effect of N-carbamoylglutamate supplementation on the growth performance, antioxidant status and immune response of mirror carp (Cyprinus carpio) fed an arginine-deficient diet

The present study was conducted to determine the effect of dietary N-carbamoylglutamate (NCG) supplementation on the growth performance, antioxidant capability and immune responses of mirror carp (Cyprinus carpio) fed an arginine (Arg)-deficient diet. A total of 630 mirror carp (41.65 ± 0.14 g) were fed diets (Arg 1.24% of the diet) that were supplemented with 0.50% Arg (control diet) or graded levels of NCG at 0 (Arg deficiency diet), 0.04%, 0.08%, 0.12%, 0.16% and 0.20% for 8 weeks. The results showed that, compared with the control diet, the Arg-deficient diet supplementation with 0 NCG (1) decreased the final body weight (FWB), the weight gain rate (WGR) or the protein efficiency ratio (PER) and increased the feed conversion ratio (FCR); (2) decreased the concentration of Arg and nitric oxide (NO) and the activity of total nitric oxide synthetase (T-NOS) in the plasma; (3) decreased the activities of superoxide dismutase (SOD) in the proximal intestine (PI), catalase (CAT) in the PI and distal intestine (DI), and glutathione peroxidase (GPx) in PI and mid-intestine (MI) and increased the concentration of malondialdehyde (MDA) in the PI, MI and DI; and (4) decreased the activity of lysozyme in the plasma, increased the relative mRNA expression of tumor necrosis factor-α (TNF-α), interleukin1β (IL-1β) and interleukin 8 (IL-8) in the PI, MI and DI, and decreased the relative mRNA expression of interleukin 10 (IL-10) in the PI and MI, and transforming growth factor β2 (TGF-β2) in the PI, MI and DI. Compared with the Arg deficient-diet supplementation with 0 NCG, (1) 0.12% or 0.16% NCG increased the FBW, WGR and PER, and 0.16% NCG increased the FCR; (2) 0.08%–0.20% NCG increased the concentration of Arg, NO and the activity of T-NOS; (3) 0.08% NCG increased the activities of SOD in the PI and MI, and 0.12% NCG increased activities of CAT and GPx in the PI, MI and DI; and (4) 0.04%–0.20% NCG increased the activity of lysozyme, 0.04%–0.20% NCG decreased the relative mRNA expression of TNF-α, IL-1β and IL-8 in the PI and MI, and 0.04%–0.20% NCG increased the relative mRNA expression of IL-10 and TGF-β2 in the PI and MI. The present results indicated that dietary 0.12% or 0.16% NCG improved the growth performance, feed utilization, intestinal antioxidant capacity and immune response of mirror carp fed an Arg-deficient diet.

Short communication: Effects of dietary addition of N-carbamoylglutamate on milk composition in mid-lactating dairy cows

The present study was conducted to investigate the effect of N-carbamoylglutamate (NCG) on milk production and composition in mid-lactating Holstein dairy cows. Sixty multiparous cows with a mean body weight of 669 kg (standard deviation = 71) and 176 days in milk (standard deviation = 55) were blocked based on parity and milk production and randomly assigned into 4 treatments, a basal diet supplemented with 0, 10, 20, or 40 g of NCG/d per cow. Milk yield and composition were recorded weekly, whereas dry matter intake and plasma variables were determined every 2 wk. The results showed that the addition of NCG had no effect on the dry matter intake and milk yield of the cows. Milk fat content and yield increased linearly with NCG addition. The contents of milk protein and total solid also increased linearly in the cows fed NCG, whereas the yield of protein was not affected by the treatments. Conversely, dietary addition of NCG increased the plasma nitric oxide content in a quadratic manner. Moreover, addition of NCG linearly increased the plasma Arg content. Overall, the results indicate that dietary NCG addition increased the milk protein and fat contents, which improved the milk quality of lactating dairy cows.

Brain MRS glutamine as a biomarker to guide therapy of hyperammonemic coma

Acute idiopathic hyperammonemia in an adult patient is a life-threatening condition often resulting in a rapid progression to irreversible cerebral edema and death. While ammonia-scavenging therapies lower blood ammonia levels, in comparison, clearance of waste nitrogen from the brain may be delayed. Therefore, we used magnetic resonance spectroscopy (MRS) to monitor cerebral glutamine levels, the major reservoir of ammonia, in a gastric bypass patient with hyperammonemic coma undergoing therapy with N-carbamoyl glutamate and the ammonia-scavenging agents, sodium phenylacetate and sodium benzoate. Improvement in mental status mirrored brain glutamine levels, as coma persisted for 48h after plasma ammonia normalized. We hypothesize that the slower clearance for brain glutamine levels accounts for the delay in improvement following initiation of treatment in cases of chronic hyperammonemia. We propose MRS to monitor brain glutamine as a noninvasive approach to be utilized for diagnostic and therapeutic monitoring purposes in adult patients presenting with idiopathic hyperammonemia.

Interaction of Excitatory Amino Acid Transporters 1 – 3 (EAAT1, EAAT2, EAAT3) with N-Carbamoylglutamate and N-Acetylglutamate

Background/Aims: Inborn deficiency of the N-acetylglutamate synthase (NAGS) impairs the urea cycle and causes neurotoxic hyperammonemia. Oral administration of N-carbamoylglutamate (NCG), a synthetic analog of N-acetylglutamate (NAG), successfully decreases plasma ammonia levels in the affected children. Due to structural similarities to glutamate, NCG may be absorbed in the intestine and taken up into the liver by excitatory amino acid transporters (EAATs). Methods: Using Xenopus laevis oocytes expressing either human EAAT1, 2, or 3, or human sodium-dependent dicarboxylate transporter 3 (NaDC3), transport-associated currents of NAG, NCG, and related dicarboxylates were assayed. Results: L-aspartate and L-glutamate produced saturable inward currents with K_m values below 30 µM. Whereas NCG induced a small inward current only in EAAT3 expressing oocytes, NAG was accepted by all EAATs. With EAAT3, the NAG-induced current was sodium-dependent and saturable (K_m 409 µM). Oxaloacetate was found as an additional substrate of EAAT3. In NaDC3-expressing oocytes, all dicarboxylates induced much larger inward currents than did L-aspartate and L-glutamate. Conclusion: EAAT3 may contribute to intestinal absorption and hepatic uptake of NCG. With respect to transport of amino acids and dicarboxylates, EAAT3 and NaDC3 can complement each other.

Effect of dietary N-carbamoylglutamate on milk production and nitrogen utilization in high-yielding dairy cows

The objective of the current study was to investigate the effect of N-carbamoylglutamate (NCG) supplementation on milk production and nitrogen (N) utilization in Chinese Holstein dairy cows. Sixty multiparous cows (78±17.3 d in milk, 635±61.00kg of body weight, and 41.9±7.9kg/d milk yield; mean ± SD) were blocked by parity, days in milk, and milk yield and randomly allocated to 1 of 4 groups, each of which was fed a dietary treatment containing 0 (control), 10, 20, or 30g of NCG/d. Milk yield was recorded weekly. Dry matter intake, milk composition, plasma variables, and urea N contents in plasma, urine, and milk were determined every other week. Blood samples were collected from the coccygeal vein. Rumen microbial protein synthesis was estimated based on the purine derivatives in the urine. Dry matter intake was found to be similar between the treatments. Addition of 20g of NCG/d tended to increase milk yield (40.2 vs. 38.1kg/d) and increased the content (2.83 vs. 2.74%) and yield (1.12 vs. 1.02kg/d) of milk protein compared with the control. The yield and content of milk fat were similar between the treatments, whereas the contents of lactose and total solids increased linearly with an increase in NCG. Dietary supplementation of NCG linearly increased the plasma nitric oxide level and decreased the plasma ammonia N level. Compared with the control, the plasma Arg concentration in cows fed 10, 20, and 30g of NCG/d was increased by 1.1, 10.4, and 16.0%, respectively. The urea N concentrations in the milk, plasma, and urine decreased with the addition of NCG, although the lowest urea N concentrations were observed with the addition of 20g of NCG/d. The conversion of dietary crude protein to milk protein exhibited quadratic trends of improvement by NCG supplementation, with a peak at 20g of NCG/d. The rumen microbial protein synthesis was not altered by NCG supplementation, but the metabolizable protein tended to show a quadratic increase in cows fed 20g of NCG/d. In conclusion, supplementation of 20g of NVG/d may alter the plasma metabolites, optimize the AA profile, increase the metabolizable protein utilization, and thereby improve the lactation performance and N utilization of high-yielding dairy cows.

Potential role of N-carbamoyl glutamate in biosynthesis of arginine and its significance in production of ruminant animals.

Arginine (ARG) exerts many beneficial effects on animal body and enhanced angiogenesis, lactogenesis, which finally leads to the improvement in nitrogen (N) metabolism, reproduction, lactation, immunity and growth. Unfortunately, unprotected ARG will be degraded in the rumen and its price is high, thus feeding rumen-protected ARG seems to be uneconomical. Alternatively, N-carbamoyl glutamate (NCG) is structural analogue of N-acetyl glutamate, cofactor of cabamoyl phosphate synthetase1, is lower in rumen degradation compared to ARG. Additionally, rumen epithelial and duodenal cells have potentially utilized the NCG for ureagenesis. Supplementation of NCG to high yielding dairy cows increased plasma concentration of ARG and nitric oxide, decreased the plasma ammonia N and improved lactation performance and N utilization. Supplementation of NCG enhanced pregnancy rates in rats, improved litter size and fetal survival rate, thereby improved the reproductive performance of sows. Oral NCG supplementation increases plasma ARG and somatotropin levels, and increased growth rate and muscle protein synthesis in nursing piglets. The NCG is potential a relatively cheaper source of feed additive to offer vital compensation over oral administration of ARG, resulting in improved ruminant animal health and production. In this article, we reviewed the mechanism of ARG biosynthesis by NCG and their significance in growth, reproduction, milk production and N utilization in ruminant animals.

Effect of dietary arginine and N-carbamoylglutamate supplementation on reproduction and gene expression of eNOS, VEGFA and PlGF1 in placenta in late pregnancy of sows

The objectives of this study were to investigate the potential mechanisms of dietary arginine (Arg) and N-carbamoylglutamate (NCG) supplementation on reproductive performance of sows. Twenty-seven crossbred (Landrace×Large White) sows with similar body weight and parity at day (90±1) of gestation were assigned randomly into 3 groups (n=9) control group, Arg group, NCG group, and fed with the following diets: a control diet, and the control diet supplemented with 1.0% Arg or 0.1% NCG. Litter size was recorded. Blood samples were obtained for biochemical analyses. Placenta chorioallantoic membrane tissue collected immediately after birth to preserve in RNA stabilizer for mRNA analysis of endothelial nitric oxide synthase (eNOS), endothelial growth factor a (VEGFA) and placenta growth factor 1 (PlGF1) by real time-PCR. The results showed that compared with the control group, the average birth weight of all piglets born alive were 16.2% and 14.3% higher in the Arg and NCG groups (P<0.05), respectively; plasma VEGFA was higher in the Arg group (P<0.05). The expression of VEGFA in the allantochorion tissue of the NCG-supplemented group was higher (P<0.01), and tended to be higher in the Arg-supplemented group (0.05<P<0.1). NCG significantly increased protein concentration of PlGF1 in plasma (P<0.05), and enhanced PlGF1mRNA expression in allantochorion tissue of placenta (P<0.01). The results suggested that dietary Arg and NCG supplementation play important roles in meliorating placental vascular function and promoting the nutrients supply to fetus.

Degradation of L-arginine and N-carbamoyl glutamate and their effect on rumen fermentation in vitro

The objective of this experiment was to determine the degradation of L-arginine (ARG) and N-carbamoyl glutamate (NCG) and to examine their effect on rumen fermentation. Rumen fluids were collected from 3 rumen-fistulated cows and then incubated with ARG or NCG at 1 mmol/L in a glass syringe system at 39°C for 24 h. The control treatment was given neither ARG nor NCG. Gas production (GP) was recorded, and pH at 2, 4, 6, 12, and 24 h was also determined. At 12 and 24 h, the measurements were also made for ammonia-nitrogen (N), volatile fatty acids (VFA) and microbial crude protein (MCP) yield on purine quantification basis. At 24 h, the proportion of ARG and NCG degradation in rumen fluid was 100.0 and 17.8%, respectively. Gas production and the acetate to propionate ratio increased in groups treated with ARG and NCG, compared with the control (P<0.01). Ammonia nitrogen concentration was higher (P<0.01) in the ARG group than in the NCG and control groups. Microbial crude protein concentration diminished in ARG and NCG groups, in comparison with the control (P<0.01). In conclusion, the effects of ARG and NCG on rumen fermentation were numerically relatively similar. Rapid degradation of ARG in rumen is a nutritionally wasteful process. Thus, ARG should be spared from rumen degradation, while NCG could be fed to ruminant without need for coating.

Metabolic Fates of Ammonia–N in Ruminal Epithelial and Duodenal Mucosal Cells Isolated from Growing Sheep

The objective of this experiment was to determine the capability of ruminant gut tissues to detoxify ammonia-N using short-term incubations of isolated cells in vitro. Ruminal epithelial cells (REC) and duodenal mucosal cells (DMC) were isolated from growing Texel-Polypay ram lambs (n=4) fed a pelleted forage:concentrate-based diet. Immediately after isolation, primary cells were incubated for 60min with glucose (1mM), glutamate (1mM), [15N]ammonium chloride (5, 10, 20, or 40mM), and 1 of 4 combinations of substrates (1mM each) that could support urea synthesis [control, N-carbamoylglutamate (NCG); NCG + ornithine (ONCG); and ONCG + aspartate (AONCG)]. Treatments were arranged in a 4×4 factorial design. Incorporation of ammonia-15N into alanine, citrulline, arginine, and urea was determined by gas chromatography-mass spectrometry. For both cell types, ammonia-N transfer to alanine was lower when incubation medium contained NCG compared with control, whereas use of ammonia-N for net alanine synthesis increased quadratically with ammonia concentration regardless of substrate treatment. For REC, ammonia-N was not incorporated into citrulline, arginine, or urea, nor into arginine or urea by DMC. Ammonia-N use for net citrulline synthesis exhibited an inverse relationship with ammonia concentration, decreasing linearly as media ammonia concentration increased. Thus, ala-nine synthesis may be a significant metabolic pathway for ruminant gut tissues to detoxify ammonia-N when it is presented luminally at high concentrations as compared with detoxification by the ornithine-urea cycle. Furthermore, DMC do exhibit a metabolic capability to incorporate ammonia-N into citrulline, but low or absent activity of downstream enzymes of the ornithine-urea cycle appears to limit ammonia-N transfers to urea.