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BACKGROUND: Insufficient endogenous nutrients in the broiler embryo can lead to muscle gluconeogenesis, which ultimately affects the post‐hatching performance of chicks. This study investigated the effects of in ovo feeding (IOF) of N‐carbamylglutamate (NCG) on the growth hormones, carcass yield, and meat quality in broilers. Fertile eggs from a 30‐week‐old Ross 308 breeder flock were divided into three treatment groups: NC (non‐injection), SC (100 μL saline‐injection), and NCG (2 mg NCG injection). Each group had six replicates, with 70 eggs per replicate during incubation. Injections were administered on the 17.5th day of embryonic development. After hatching, 270 chicks were selected for 42‐day rearing for further sampling. RESULTS: Chicks in the NCG group had significantly higher body weight (BW) and average daily gain (ADG) at the growing phase, increased growth and testosterone hormone in both feeding phases (21 and 42 days), and improved average daily gain (ADG) and food conversion ratio (FCR) in both grower and entire feeding phases (P < 0.05). Triiodothyronine (T3) and tetraiodothyronine (T4) levels, carcass yield, dressing, drum weight, breast muscle weight, drumstick weights, thighs, pectoralis major, and their part percentage of carcass were improved in the NCG group (P < 0.05), these effects were varied along feeding phases. Moreover, IOF of the NCG also improved pectoralis breast muscle color values at 24 h post mortem (P < 0.05). CONCLUSION: These results suggest that NCG injection at the late embryonic age of broiler enhances growth performance and meat quality throughout the lifespan and this can probably be attributed to an increase in thyroid and testosterone hormones, indicating potential involvement in metabolic and nutrient partitioning pathway regulation. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

The present work focused on examining the function of rumen-protected L-arginine (RP-Arg) or N-carbamylglutamate (NCG) in jejunal oxidative resistance, integrity and immune function in the ovine foetal model of intrauterine growth restriction (IUGR). Thirty-two twin-bearing Hu ewes at d 35 of gestation were randomised as 4 treatment groups (n = 8 each): Control (CON), received 100% of the recommended National Research Council (NRC) for pregnancy; Restricted (RES), received 50% of the recommended NRC for pregnancy; RES + ARG, RES ewes added with 20 g/d of RP-Arg; or RES + NCG treatment, RES ewes added with 5 g/d of NCG. Foetal jejunal samples were collected on d 110 of pregnancy and were assayed for biomarkers of oxidative damage, integrity and immune function. The villus height was elevated (p

Simple Summary: Improvement of fetal growth in twin pregnancies is one of the major goals of extensive sheep production systems, as lambs' birth weight is correlated with their survival. This is especially relevant in harsh environments where natural maternal nutrient restriction is common. Maternal arginine (Arg) supplementation improves fetal growth, but Arg is degraded in the rumen, has a short biological half-life, and the high cost of protected forms limits its use in extensive sheep systems. N-carbamylglutamate (NCG) is not degraded in the rumen, increases arginine synthesis, and improves urea recycling, suggesting it may be an option as an alternative to Arg. The aim of this study was to evaluate the effect of oral NCG supplementation from 100 days of gestation (dga) to term in naturally nutrient-restricted grazing twin-bearing ewes, on maternal, placental, and fetal variables. The serum concentrations of NCG were increased 15-fold, and the plasma urea, albumin and phosphorus concentrations were reduced in supplemented ewes compared to controls, but there was no major effect on the dam or fetal body or organ weights nor the antioxidant markers and other blood biochemical parameters. These results indicate that NCG supplementation in mid-to-late gestation in grazing ewes was unable to rescue the negative production effects of severe natural nutritional restriction on both the dam and fetuses. The challenge of meeting the nutrient requirements of pregnant ewes in harsh environmental conditions reinforces the need for future research to identify novel strategies to improve lamb survival in such environments around the world. N-carbamylglutamate (NCG) is postulated to improve fetal growth in nutrient-restricted gestations when supplemented from day 35 to 110 of gestation, but the effects of supplementation from 100 days of gestation to birth have not been evaluated. The aim of this study was to evaluate the effect of oral NCG supplementation from 100 days of gestation (dga) to term in naturally nutrient-restricted grazing twin-bearing ewes, on the maternal body weight (BW), body condition score (BCS), placental morphology, fetal body and organ weights and blood biochemistry and antioxidant status in the ewe and fetuses. Eighteen twin-bearing ewes maintained under grazing management were randomly allocated to either a treatment group (NCG; n = 10), orally dosed once daily with 60 mg/kg of NCG from day 100 until 140 dga, or an unsupplemented control group (CON; n = 8). At 140 dga, blood gases, redox status, maternal and fetal plasma and fetal biometrics were obtained after caesarian section. The serum concentration of NCG was increased 15-fold in the NCG ewes compared to the CON. No major effects on dam or fetal body weight nor on blood biochemistry or antioxidant parameters were observed. These results indicate that NCG supplementation in mid-to-late gestation to grazing ewes was unable to rescue the negative production effects of severe natural nutritional restriction on both the dam and fetuses. [ABSTRACT FROM AUTHOR]

Background: This study aimed to evaluate the effects of N‐carbamylglutamate (NCG) on piglets' growth performance and immune response, and to unravel the mechanisms of such effects. In a 2 × 2 factorial design including diet (with or without NCG) and immunological challenge (saline or lipopolysaccharide (LPS)), 24 piglets were randomly distributed into four groups. After being fed a basic diet or a NCG‐supplemented diet for 21 days, piglets were administered LPS or saline intraperitoneally. Results: The results showed that NCG increased the average daily gain and average daily feed intake, and the feed conversion ratio of piglets, and alleviated the adverse effects of LPS stimulation on intestinal morphology. At the phylum level, NCG reversed the increase in the abundance of Firmicutes and the reduction in that of Actinomycete caused by LPS stimulation. At the genus level, NCG increased the abundance of Lactobacillus, Blautia, norank_Butyricicoccaceae, Subdoligranulum, and Ruminococcus_gauvreauii_group, and LPS decreased the abundance of Escherichia‐Shigella and Ruminococcus_gauvreauii_group. The short‐chain fatty acid content was increased by NCG, but LPS reduced its content. N‐Carbamylglutamate also inhibited significantly the LPS‐induced increase in the relative expression of signal transducer and activator of transcription (STAT) 3, related orphan receptor (RAR) c, and pro‐inflammatory cytokines, and the decrease in the relative expression of STAT5, forkhead box P3, IL‐10 and transforming growth factor beta 1 mRNA. A significant correlation was found between intestinal microbiota and inflammatory cytokines and short‐chain fatty acids. Conclusion: N‐Carbamylglutamate can improve piglets' growth performance. It can also attenuate LPS‐induced intestinal inflammation by modulating microbiota and Th17/Treg balance‐related immune signaling pathways. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the effects of dietary supplementation of underfed Hu ewes from d 35 to 110 of gestation with either rumen-protected L-arginine (RP-Arg) or N-carbamylglutamate (NCG) on placental amino acid (AA) transport, angiogenic gene expression, and steroid anabolism. On d 35 of gestation, 32 Hu ewes carrying twin fetuses were randomly divided into four treatment groups, each consisting of eight ewes, and were fed the following diets: A diet providing 100% of NRC's nutrient requirements for pregnant ewes (CON); A diet providing 50% of NRC's nutrient requirements for pregnant ewes (RES); RES diet plus 5 g/d NCG (RES + NCG); or RES diet plus 20 g/d RP-Arg (RES + ARG). On the d 110 of pregnancy, blood samples were taken from the mother, and samples were collected from type A cotyledons (COT; the fetal portions of the placenta). The levels of 17β-estradiol and progesterone in the maternal serum and both the capillary area density (CAD) and capillary surface density (CSD) in type A COT were decreased in response to Arg or NCG supplementation when compared to the RES group. The concentrations of arginine, leucine, putrescine and spermidine in type A COT were higher (P

ABSTRACT: Eggs rich in polyunsaturated fatty acids (PUFA), known as functional eggs, are animal products deemed beneficial to human health and possess high economic value. The production of functional eggs involves supplementing exogenous additives with the ability to regulate lipid metabolism. As N-Carbamylglutamate (NCG) serves as an endogenous arginine synthesizer, and arginine acts as the substrate for the formation of nitric oxide (NO), the biological function of NCG is partially mediated by NO. NO is a key regulatory molecule in lipid metabolism, suggesting that NCG may also have the ability to modulate lipid metabolism. In order to assess the capacity of NCG in regulating liver lipid metabolism and its potential application in producing functional eggs, we conducted a study to investigate the effects of dietary supplementation of NCG on production performance, serum, and liver NO levels, yolk fatty acid composition, and the liver transcriptome of layers. In this study, we utilized 30 layers of the Jinghong No.1 breed, all aged 45 wk. All the birds were randomly divided into 2 groups. Each group had 5 replicates, and each replicate had 3 birds. We provided them with different diets: one group received the basic diet, and the other group's diet was supplemented with 0.08% NCG. The experiment lasted for 14 wk. The results did not reveal any positive impact of NCG on production performance. However, NCG supplementation elevated NO levels in serum and liver, along with an increase in yolk PUFA, ω-3, and ω-6 fatty acids. Liver transcriptome analysis identified 124 upregulated differentially expressed genes (DEGs) and 43 downregulated DEGs due to NCG supplementation. Functional annotation using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database highlighted 3 upregulated DEGs (CPT1A, MOGAT1, and CHKA) and 2 downregulated DEGs (FASN and ETNPPL) associated with lipid metabolism. Pathway enrichment analysis revealed that CPT1A was enriched in the AMPK signaling pathway and the PPAR signaling pathway, while FASN was enriched in the AMPK signaling pathway. Thus, CPT1A and FASN are potential functional genes related to lipid metabolism facilitated by NCG supplementation. In summary, our study suggests that NCG supplementation modulates liver lipid metabolism, leading to the production of functional eggs in layers.

Simple Summary: NCG is a structural analogue of NAG, and can activate the rate-limiting enzyme of arginine synthesis and promote the synthesis of endogenous arginine in the intestine. Recent studies have found that NCG plays an important role in improving growth performance and maintaining intestinal health. However, there are few reports on the effects of NCG on the intestinal development and antioxidant capacity of piglets. The aim of this study was to investigate the effects of dietary NCG supplementation on the growth performance, apparent nutrient digestibility, jejunal morphology, amino acid transporters, and antioxidant capacity of weanling piglets. Studies have shown that dietary NCG supplementation can improve the growth performance and intestinal antioxidant capacity of weaned piglets. Weaning is an important period that affects the performance of piglets. However, the regulation of dietary amino acid levels is considered to be an effective way to alleviate the weaning stress of piglets. N-carbamylglutamate (NCG) plays an important role in improving the growth performance and antioxidant capacity of animals. A total of 36 weaned piglets were randomly assigned to two treatment groups, a control group (CON) and a 500 mg/kg NCG group (NCG), and the experiment lasted for 28 days. The results show that the NCG treatment group showed an increased 0–28 days average weight gain and average daily feed intake, and also increased contents of GLU and HDL, and lower SUN in serum, and an upregulation of the expression of the amino acid transporters SNAT2, EAAC1, SLC3A1, and SLC3A2 mRNA in the jejunum (p < 0.05), as well as an increased villus length and VH:CD ratio, and claudin-1, occludin, and ZO-1 mRNA expression in the jejunum (p < 0.05). The NCG treatment group showed an increased content of GSH-Px in serum and T-AOC and SOD in the jejunum, and a lower content of MDA (p < 0.05); and the upregulation of the mRNA expression related to antioxidant enzymes (CAT, SOD1, Gpx4, GCLC, GCLM and Nrf2, AhR, CYP1A1) in the jejunal mucosa (p < 0.05). In addition, compared with the control group, the NCG treatment group saw an upregulation in the mRNA expression of IL-10 and a decrease in the expression of IL-1β and IL-4 in the jejunal mucosa (p < 0.05). In summary, the results of this study suggest that NCG improved growth performance and jejunal morphology, improved the jejunal transport of amino acids related to the ornithine cycle, and improved the antioxidant capacity in weaned pigs. [ABSTRACT FROM AUTHOR]

The experiment was to investigate the effect of N-carbamylglutamate on reproductive performance, serum indexes and hormone secretion of cows. Sixty Angus cows with similar weight, parity and expected delivery were randomly divided into four groups with 15 cows in each group, one cow per replicate. The control group was basal diet, and each cow in test group 1, group 2 and group 3 were fed with basal diet supplemented with 10, 20 and 30 g/(head·d) of NCG, respectively. The experiment lasted for 120 d. The results showed that after cows delivery, the content of GLU and NO of cows in test group 3 were significantly higher than control group (P<0.05), the content of Arg of cows in test group 2 and group 3 were significantly higher than control group (P<0.05), and the content of P of cows in test group 2 were significantly higher than control group (P<0.05),while the content of E2 of cows in test group 3 were significantly lower than control group (P<0.05). Before cows breeding, the content of serum BUN of cows in test group 2 and group 3 were significantly lower than control group (P<0.05), while the content of GLU in test group 2 was significantly higher than control group (P<0.05), the content of Arg and NO in all the test groups were significantly higher than control group (P<0.05), the content of serum E2 in test group 3 was significantly higher than control group (P<0.05). The content of serum P in test group 2 and group 3 were significantly lower than control group (P<0.05). The estrus mating rates of cows in test group 2 and group 3 were higher than control group, and the conception rates of experimental groups were higher than that in control group, but the first postpartum mating days in test group 2 and group 3 were significantly lower than control group (P<0.05). The results indicates that the NCG can improve body metabolism, promote the secretion of reproduction-related hormones, and improve the reproductive performance of breeding cows, and the appropriate supplemental level is 20 and 30 g/(head·d). [ABSTRACT FROM AUTHOR]

N-carbamylglutamate (NCG) is postulated to improve fetal growth in nutrient-restricted gestations when supplemented from day 35 to 110 of gestation, but the effects of supplementation from 100 days of gestation to birth have not been evaluated. The aim of this study was to evaluate the effect of oral NCG supplementation from 100 days of gestation (dga) to term in naturally nutrient-restricted grazing twin-bearing ewes, on the maternal body weight (BW), body condition score (BCS), placental morphology, fetal body and organ weights and blood biochemistry and antioxidant status in the ewe and fetuses. Eighteen twin-bearing ewes maintained under grazing management were randomly allocated to either a treatment group (NCG; n = 10), orally dosed once daily with 60 mg/kg of NCG from day 100 until 140 dga, or an unsupplemented control group (CON; n = 8). At 140 dga, blood gases, redox status, maternal and fetal plasma and fetal biometrics were obtained after caesarian section. The serum concentration of NCG was increased 15-fold in the NCG ewes compared to the CON. No major effects on dam or fetal body weight nor on blood biochemistry or antioxidant parameters were observed. These results indicate that NCG supplementation in mid-to-late gestation to grazing ewes was unable to rescue the negative production effects of severe natural nutritional restriction on both the dam and fetuses.

IntroductionThe objective of this study was to investigate the effects of N-carbamylglutamate (NCG) supplementation on the growth performance, hindgut microbiota composition, and short-chain fatty acid (SCFA) contents in Charollais and Small Tail Han crossbred sheep.MethodsA total of 16 female crossbred mutton sheep (Charollais × Small Tail Han), aged 4 months, with an initial body weight of 30.03 ± 0.08 kg, were utilized in a 60 days experiment. The sheep were divided into two groups based on their initial body weight. Each group consisted of 8 replicates, with each individual sheep considered as a replicate. The dietary treatments comprised a basal diet supplemented with either 0.00% or 0.12% NCG.Results and discussionOur findings indicate that NCG supplementation did not have a significant effect on the growth performance of mutton sheep. However, it did lead to changes in hindgut SCFA contents. Specifically, NCG supplementation increased the content of propanoic acid while decreasing acetic acid and hexanoic acid in the hindgut. Through microbiota analysis using the 16S rRNA technique, we identified Lachnospiraceae_NK3A20_group and Parasutterella as biomarkers for the hindgut microbiota in mutton sheep fed a diet containing NCG. Further analysis of the microbiota composition revealed that NCG supplementation significantly increased the abundance of Lachnospiraceae_NK3A20_group and Parasutterella, while decreasing unclassified_f_Lachnospiraceae and Lachnoclostridium. Correlation analysis between hindgut SCFA contents and microbiota composition revealed that the abundance of Lachnoclostridium was positively correlated with the contents of acetic acid and hexanoic acid, but negatively correlated with propanoic acid. Additionally, the abundance of Lachnospiraceae_NK3A20_group and Parasutterella was positively correlated with the content of propanoic acid, while being negatively correlated with acetic acid and hexanoic acid. Based on these findings, we conclude that dietary supplementation of 0.12% NCG can modulate hindgut SCFA contents in mutton sheep by regulating the composition of the hindgut microbiota.

The experiment was conducted to investigate the effect of dietary NCG supplementation on growth performance, serum biochemical indexes and immune indexes of calves in Angus cows during late gestation. Sixty cows in late gestation period with similar weight parity and expected perinatal period were randomly divided into four groups with 15 cows in each group. Cows in control group were fed with a basal diet, cows in experimental groups were fed wtih basal diet supplemented with 10, 20 and 30 g/(head·d) NCG, respectively. The corresponding calves in each group were denoted as group 1, group 2 and group 3, respectively. The experiment starts from 4 w before delivery of cows and ends at 90 d of age of calves. The results showed that compared with control group, the birth weight of calves in group 3 was significantly higher than that in control group (P<0.05). Compared with control group, the contents of TP, ALB and NO in serum of newborn calves in group 3 were significantly increased (P<0.05), and the content of IgA, IgG and IgM in serum were significantly increased (P<0.05). The experiment indicates that the addition of NCG of 30 g/(head·d) to diet of cows in late gestation period can significantly increase birth weight, increase erum protein content of newborn calves, improve immune function, but these effects were lost at 90 days of age. [ABSTRACT FROM AUTHOR]

[Objectives] The purpose of this experiment was to investigate the effects of dietary supplementation of N-carbamylglutamate(NCG)or taurine(Tau)on reproductive performance, serum antioxidant indices, immunoglobulin levels of sows and the growth performance of piglets. [Methods] Forty-eight Danbred Landrace sows with similar body condition and parity were assigned randomly into 3 groups:control group(basal diet), NCG group(basal diet with 500 mg·kg-1 NCG)and Tau group(basal diet with 600 mg·kg-1 Tau), respectively. The experiment was carried on sows from 35 days of gestation to weaning at 26-day-age of piglets. [Results]1)Compared with the control group, both dietary NCG and Tau supplementation increased the number of heavier piglets(P<0.05), however, no significant difference was found between NCG group and Tau group(P>0.05). 2)Supplementation with dietary NCG and Tau showed a significant increase in piglet vitality(P<0.05); meanwhile, NCG group showed a remarkable decrease in duration of parturition(P<0.05). 3)Compared with the control group and NCG group, dietary supplemented with Tau significantly increased the average daily feed intake(ADFI)of the sows 5 days after delivery(P<0.05), while compared with the control group and Tau group, dietary NCG supplementation significantly increased the ADFI of sows during lactation(P<0.05), and enhanced the activity of superoxide dismutase(SOD)(P<0.05). Meanwhile, compared with control group, the activity of serum glutathione peroxidase(GPx)in NCG and Tau groups increased by 0.41% and 0.03% respectively, the malondialdehyde(MDA)content decreased by 0.43% and 1.71%respectively, but there was no statistical difference(P>0.05). 4)Both NCG and Tau supplementation tended to increase serum IgG(P=0.06)and IgM levels(P=0.07). 5)Compared with the control group and the Tau group, the supplementation with NCG significantly increased the average daily weight gain and average body weight of the weaned piglets(P<0.05); compared with the control group, the supplementation with Tau had no significant effect on the growth performance of the piglets(P>0.05). [Conclusions] Dietary supplemented with 500 mg·kg-1 NCG or 600 mg·kg-1 Tau could increase the number of heavier piglets and enhance the piglets' vitality, and NCG group showed a better efficiency than Tau group. Meanwhile, the supplementation with NCG can also shorten the duration of parturition, improve the feed intake of the sow during lactation period and the growth performance of weaned piglets. The mechanism of this action might be related to the fact that NCG could improve the antioxidant level and immune function of sows. [ABSTRACT FROM AUTHOR]

Previous studies have revealed that dietary N-carbamylglutamate (NCG) or L-arginine (Arg) improves small intestinal integrity and immune function in suckling Hu lambs that have experienced intrauterine growth retardation (IUGR). Whether these nutrients alter redox status and apoptosis in the colon of IUGR lambs is still unknown. This study, therefore, aimed at investigating whether dietary supplementation of Arg or NCG alters colonic redox status, apoptosis and endoplasmic reticulum (ER) stress and the underlying mechanism of these alterations in IUGR suckling Hu lambs. Forty-eight 7-d old Hu lambs, including 12 with normal birth weight (4.25 ± 0.14 kg) and 36 with IUGR (3.01 ± 0.12 kg), were assigned to 4 treatment groups (n = 12 each; 6 males and 6 females) for 3 weeks. The treatment groups were control (CON), IUGR, IUGR + Arg and IUGR + NCG. Relative to IUGR lambs, superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) content, as well as proliferation index, were higher (P

The aim of this study was as follows: 1) to investigate the effects of graded levels of N-carbamylglutamate (NCG) on performance, blood biochemical indexes, carcass traits and related indicators in growing-finishing pigs, and 2) to determine the optimal supplemental level. The toxicity of high-dose (much higher than recommended levels) NCG was assessed by routine blood tests and blood biochemical and histopathologic examinations of the heart, liver, spleen, lung, kidney and stomach. One hundred and forty-four growing-finishing pigs (Duroc × Large White × Landrace, 32.24 ± 1.03 kg) were used in a 74-d experiment and each treatment was replicated 6 times with 4 pigs (2 barrows and 2 gilts) per replicate. The dietary treatments were a corn-soybean meal basal diet supplemented with 0% (control), 0.05%, 0.1%, 0.15%, 0.2% or 1% NCG. The first 5 groups were used to explore the optimal supplemental level of NCG, while the control, 0.1% and 1% NCG groups were used to explore the safety of high-dose NCG. Compared with the normal control group, the final body weight and average daily gain tended to be higher in the 0.1% group (P = 0.08), the lean percentage tended to be higher in the 0.05% group (P = 0.07), the levels of free amino acids in the blood significantly increased in the 0.1% group (P

Introduction: Ammonia has been of concern for its high toxicity to animals. Ncarbamylglutamate (NCG) can reduce blood ammonia levels in mammals, but studies on ammonia tolerance in fish are insufficient. Methods: Juvenile yellow catfish were fed two levels of NCG (0.00% and 0.05%) for 84 days under three ammonia levels (0.00, 0.08, and 0.16 mg/L NH3). Results and Discussion: The results showed that survival rate (SUR), final body weight (FBW), weight gain (WG), and serum total protein (TP), triglycerides (TG), glucose (Glu), ornithine (Orn), citrulline (Cit) contents, and liver superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), arginase (ARG), ornithine transcarbamylase (OTC) activities decreased with the increase of ammonia levels, on the contrary, feed conversion ratio (FCR), hepatosomatic index (HSI), and serum ammonia, urea, alanine aminotransferase (ALT), aspartate aminotransferase (AST), glutamine (Gln), arginine (Arg) contents, and liver malondialdehyde (MDA), tumor necrosis factor (TNF), interleukin (IL) 1, IL 8 contents, and mRNA expressions of cu/zn sod, cat, gpx, gr, tnf ɑ, il 1, and il 8 were significantly increased. Dietary 0.05% NCG supplementation had higher SUR, FBW, WG, feed intake (FI), whole-body protein, and serum TP, total cholesterol (TC), Glu, citrulline (Cit) contents, and liver SOD, GPx, argininosuccinate synthetase (ASS), argininosuccinate lyase (ASL), inducible nitric oxide synthase (iNOS) activities compared to 0.00% NCG group, but had lower serum ammonia, urea, ALT, AST, Gln, Arg contents, and liver MDA, TNF, IL 1, IL 8 contents, and neuronal nitric oxide synthase activity. At the end of bacterial challenge, cumulative mortality (CM) increased with ammonia levels increased, but serum antibody titer (AT), lysozyme (LYZ) activity, 50% hemolytic complement, immunoglobulin (Ig) contents, respiratory burst (RB), phagocytic indices decreased with ammonia levels increased. CM in 0.05% NCG group was lower than that in 0.00% NCG group, but serum AT, LYZ activity, Ig content, RB in 0.05% NCG group were significantly higher. The correlation analysis found that iNOS was positively correlated with ASS activity. This study indicates that dietary NCG supplementation can improve the ammonia tolerance of yellow catfish, and ASS may also be the target of NCG to activate the urea cycle. [ABSTRACT FROM AUTHOR]

Introduction: The Tibetan Plateau is characterized by low temperature and hypoxia. N-carbamylglutamic acid (NCG) can increase blood oxygen saturation, and have the potential to be used to prevent the high-altitude hypoxia stress state of cows. However, its beneficial effect on the rumen microbiota of Holstein dairy cows remains unclear. Methods: Hence, the experiments 12 multiparous (parity ranged from 2 to 7) Holstein dairy cows (413.0 ± 42 kg) were randomly assigned to 2 treatments with 6 replicates in each treatment: basal diet (CON, control group) and basal diet plus 20 g/d/cow of NCG (NCG, experiment group), respectively. To study the effects of dietary NCG supplementation on rumen microbiota of Holstein dairy cows in Tibet. The experiment lasted for 45 days, with 15 days of pre-feeding and 30 days of formal trail period. Results: The results showed that ruminal NH3-N concentration in NCG group was lower (p < 0.05) than that in the CON group, while molar proportion of acetic acid and total volatile fatty acid (VFA) concentration were increased (p < 0.05) with the addition of NCG. Microbial diversity increased (p < 0.05) in NCG group, with Bacteroidetes, Firmicutes, and Patescibacteria as the most abundant phyla. The KEGG pathway analysis showed that the potential function of ruminal bacteria was mainly enriched in metabolism (carbohydrates, amino acids, lipids, energy, and nucleotides) and genetic information processing (replication, repair, and translation). Conclusion: In conclusion, NCG can improve rumen nitrogen utilization, total VFA and acetic acid production, and increase rumen microbial diversity, all of which could make the introduced Holstein dairy cows to better adapt to the harsh environment in Tibet and improve their production performance. [ABSTRACT FROM AUTHOR]

A total of 288 11‐week‐age roosters were used to evaluate the effects of dietary supplementation of N‐carbamylglutamate (NCG) on reproductive traits and testicular gene expression. The experimental periods were 12 weeks. All birds were randomly assigned to 4 treatments with 6 replicates per treatment and 12 birds per replicate. Dietary conditions were based on a basal diet and supplemented with 0%, 0.08%, 0.12%, or 0.16% NCG to form C, N1, N2 and N3 groups respectively. Dietary supplementation of NCG had positive effects on the seminiferous tubule parameters, serum gonadotropin‐releasing hormone and testosterone levels and the secondary sexual characteristics. Transcriptomics analysis was performed on the testicular tissues between C and N3 groups at the 16‐week‐age. Genes were mainly enriched in nine pathways, such as cytochrome P450 exogenous metabolism, drug metabolism, steroid hormone synthesis and glutathione metabolism, in which the ZP4 gene, cytochrome P450 family member 11A1 and other genes involved in the maintenance of gonadal function, steroid hormone biosynthesis and metabolism, and so forth, exist differences in expression levels. In summary, dietary supplementation of NCG had positive effects on the reproductive traits of roosters. NCG supplementation improved the development of reproductive traits of roosters by regulating the genes expression in testicular tissues and thus improved the synthesis of reproductive hormones in vivo. [ABSTRACT FROM AUTHOR]

The effect of feed additive N- carbamylglutamate (NCG) with different feeding levels on some biochemical parameters and amino acids in the serum of goats was investigated. Thirty male goats aged 10-11 months were distributed into three levels of concentrate 70%, 50%, and 30% with or without N- carbamylglutamate (NCG) in a 2×3 factorial experiment/Completely Randomized Design (CRD). After 77 days of individual feeding, jugular blood was sampled before morning feeding. The results showed that additive NCG with high concentrate (70%) led to a decrease (P<0.01) in total blood protein, triglycerides, and globulins, and a decrease (P<0.05) for creatinine, uric acid, and VLDL. Blood magnesium increased (P<0.01) with NCG of low concentrate (30%) treatment, while phosphor increased with NCG of 50% concentrate treatment. Calcium increased with all NCG treatments. The results of adding N- carbamylglutamate with high roughage led to a negative effect on blood amino acids. In conclusion, increasing roughage feeding without NCG is the best for blood homeostasis, and don’t use N- carbamylglutamate without non-protein nitrogen additives. [ABSTRACT FROM AUTHOR]

Simple Summary: Largemouth bass (Micropterus salmoides) has been widely cultured in China, and ammonia stress has become one of the important reasons limiting the development of largemouth bass' farming. However, the response of largemouth bass to ammonia stress is still to be investigated. In addition, N-carbamylglutamate has been considered as a potential anti-stress additive in terrestrial animals. Currently, it remains to be explored whether N-carbamylglutamate can enhance fish resistance under environmental stress. Our findings in this study could enrich our understanding of the responses in largemouth bass under ammonia stress and help us solve the existing problem of environmental stress in largemouth bass culture. Ammonia is a common environmental limiting factor in aquaculture. To investigate the effects of ammonia stress and explore the protective effect of N-carbamylglutamate (NCG) on Micropterus salmoides (M. salmoides), tissue sections and parameters related to oxidative stress and the inflammatory response in M. salmoides were carried out during the ammonia stress test and feeding test. The results demonstrated that the LC50 for 24 h, 48 h, 72 h, and 96 h under ammonia stress in M. salmoides were 25.78 mg/L, 24.40 mg/L, 21.90 mg/L, and 19.61 mg/L, respectively. Under ammonia stress, the structures of the tissues were damaged, and the GSH content decreased, while the MDA content increased with the increase in stress time and ammonia concentration. The NO content fluctuated significantly after the ammonia nitrogen stress. In the 15-day feeding test, with the increased NCG addition amount and feeding time, the GSH content increased while the MDA and NO contents decreased gradually in the NCG addition groups (NL group: 150 mg/kg; NM group: 450 mg/kg; NH group: 750 mg/kg) when compared with their control group (CK group: 0 mg/kg). In the ammonia toxicology test after feeding, the damage to each tissue was alleviated in the NL, NM, and NH groups, and the contents of GSH, MDA, and NO in most tissues of the NH group were significantly different from those in the CK group. The results suggested that ammonia stress caused tissue damage in M. salmoides, provoking oxidative stress and inflammatory response. The addition of NCG to the feed enhances the anti-ammonia ability of M. salmoides. Moreover, the gill and liver might be the target organs of ammonia toxicity, and the brain and kidney might be the primary sites where NCG exerts its effects. Our findings could help us to find feasible ways to solve the existing problem of environmental stress in M. salmoides culture. [ABSTRACT FROM AUTHOR]

Background: Methylmalonic acidemia (MMA) is an inborn error of metabolism whose optimal management, especially in the long‐term remains to be established. Methods: We describe the case of a child with MMA mut0 who was in a cycle of episodes of decompensation and hospitalization when we started to use carglumic acid (CA), a well‐known adjunctive therapy to standard care for the treatment of acute hyperammonemia due to MMA. Results: Using the lowest effective therapeutic dose of CA and adjusting the patient's diet with caloric and protein intake adequate for her age and pathology, we managed to keep ammonium levels within the normal range, and to ensure a normal growth pattern. Conclusion: The present case adds further confirmation of the long‐term management of MMA using CA, focusing on the long duration of follow up and on the use of a lower dose of CA in real life settings. [ABSTRACT FROM AUTHOR]

To study the effect of feed additive N- carbamylglutamate (NCG) with different levels of feeding on amino acids profile and some analytes in serum of goats, thirty male goats aged 7-8 months were distributed into three levels of concentrate 2%, 3%, 4% with or without NCG in a 2×3 factorial experiment/Completely Randomized Design (CRD). After 72 days of individual feeding, the blood from jugular vein was obtained before feeding in the morning. Results showed that additive NCG with 3% concentrate led to an increase (P<0.01) in total protein and albumin (P<0.05), globulins increased (P<0.01) with increasing concentrate without NCG. Creatinine decreased with increasing concentrate without NCG. Additive NCG led to a linear decrease in uric acid with increasing feed intake (P<0.05) and a linear increase without NCG. Blood magnesium, zinc, and copper increased with NCG. No difference in calcium between treatments. The level of feeding without NCG didn’t affect fat parameters, while LDL, HDL, and cholesterol increased with NCG and 4% concentrate. Additive N- carbamylglutamate affected negatively blood amino acids, especially with 4% concentrate. In conclusion, 3% concentrate was the best for blood homeostasis with or without NCG, and preferred to use NCG with urea. [ABSTRACT FROM AUTHOR]

Our previous studies demonstrated that prenatal in utero growth restriction impairs postnatal intestinal function. Thus, improving postpartal intestinal absorption capacity and growth by manipulating the maternal diet prepartum is of importance. This work was conducted to determine whether supplementation of N-carbamylglutamate (NCG) or rumen-protected L-arginine (RP-Arg) increased fetal intestinal amino acid (AA) profiles in intrauterine growth retardation (IUGR) fetuses. On d 35 of gestation, Hu ewes (n = 32) carrying twin fetuses were randomized into 4 groups (8 ewes and 16 fetuses in each group), where diets were as follows: 100% of nutrient requirements recommended by National Research Council (NRC, 2007) (CON); 50% of nutrient requirements recommended by NRC (2007) (RES); RES + RP-Arg (20 g/d), (RES + ARG); and RES + NCG (5 g/d), (RES + NCG). On d 110 of gestation, both fetal and maternal tissues were collected and weighed. Compared with RES, solute carrier family 1, member 5 (SLC1A5) was upregulated (P

Maternal nutrient restriction in twin- bearing ewes reduces the colostrum protein content, resulting in reduced postnatal growth and greater lamb mortality in twin- compared with single-bearing ewes. N-carbamylglutamate (NCG), an analogue of N-acetylglutamate increases endogenous arginine syn- thesis and is postulated to influence fetal growth and development and potentially lamb survival. The effect of maternal supplementation with NCG in mid-late gestation on colostrum quality and nutrient status of nutrient-restricted ewes has not been reported. The aim of this study was to evaluate the effect of oral maternal supplementation with NCG during mid–late gestation in nutrient restricted twin-bearing ewes on maternal body weight (BW) gain, body condition score (BCS), nutritional status and colostrum composition. Twin- bearing ewes (n = 20), managed in individual pens, were orally dosed once-daily with 60 mg/kg BW NCG in a water carrier from 100 d of gestation (dg) to lambing and were compared with unsupplemented controls (CON; n = 21). The ewes received a ration based on alfalfa (12.12% CP and 2.10 Mcal/kg ME) and pas- ture (6.85% CP and 2.13 Mcal/kg ME) hay, covering 50% of National Research Council nutritional require- ments for twin-bearing ewes to reflect commercial con- ditions. Maternal BW and BCS were recorded every 10 d, from 100 dg until lambing. Maternal blood samples were collected at 130 dg and 30 min after lambing and plasma beta-hydroxybutyrate (BHB), total protein, albumin and urea were evaluated. Colostrum was collected 30 min after the second lamb was delivered and fat, non-fat solids, density and protein contents were determined. The effect of treatment on ewe BW, BCS were analyzed using REML analysis in R (lme4 R package) with time (dg) and treatment (and their inter- action) as fixed effects. Plasma and colostrum metab- olites were analyzed using REML with treatment as the fixed effect. Ewe BW increased (P < 0.0001) and BCS decreased (P < 0.0001) with advancing gestation, but there was no effect of treatment (P > 0.05). At 130 dg plasma urea was a 12% greater (P = 0.02) and al- bumin tended to be 4% greater (P = 0.08) in NCG com- pared with CON ewes, but no treatment effect on BHB or total protein content was observed (P > 0.05). No treatment effect on maternal plasma metabolites were observed post lambing (P > 0.05). Colostrum protein content was 21% greater in NCG compared with CON (P = 0.04) ewes, but fat, non-fat solids or density (P > 0.05) did not differ between groups. Collectively, these results suggest that supplementation of nutritionally restricted ewes with 60 mg/kg BW from 100 dg to lambing may alter maternal nitrogen metabolism and colostrum protein content. The potential impact of these changes on lamb growth and/or survival, warrants further investigation. *Funded by FONDECYT Project N°1221042; AgResearch Strategic Science Investment Fund SSIF-A25765 and INIA Kampenaike. [ABSTRACT FROM AUTHOR]

Weaning is an important period that affects the performance of piglets. However, the regulation of dietary amino acid levels is considered to be an effective way to alleviate the weaning stress of piglets. N-carbamylglutamate (NCG) plays an important role in improving the growth performance and antioxidant capacity of animals. A total of 36 weaned piglets were randomly assigned to two treatment groups, a control group (CON) and a 500 mg/kg NCG group (NCG), and the experiment lasted for 28 days. The results show that the NCG treatment group showed an increased 0–28 days average weight gain and average daily feed intake, and also increased contents of GLU and HDL, and lower SUN in serum, and an upregulation of the expression of the amino acid transporters SNAT2, EAAC1, SLC3A1, and SLC3A2 mRNA in the jejunum (p < 0.05), as well as an increased villus length and VH:CD ratio, and claudin-1, occludin, and ZO-1 mRNA expression in the jejunum (p < 0.05). The NCG treatment group showed an increased content of GSH-Px in serum and T-AOC and SOD in the jejunum, and a lower content of MDA (p < 0.05); and the upregulation of the mRNA expression related to antioxidant enzymes (CAT, SOD1, Gpx4, GCLC, GCLM and Nrf2, AhR, CYP1A1) in the jejunal mucosa (p < 0.05). In addition, compared with the control group, the NCG treatment group saw an upregulation in the mRNA expression of IL-10 and a decrease in the expression of IL-1β and IL-4 in the jejunal mucosa (p < 0.05). In summary, the results of this study suggest that NCG improved growth performance and jejunal morphology, improved the jejunal transport of amino acids related to the ornithine cycle, and improved the antioxidant capacity in weaned pigs.

IntroductionSupplementation of exogenous additives is a strategy to improve laying performance of layers by regulating uterine function. N-Carbamylglutamate (NCG) as an activator for endogenous arginine synthesis has the potential to regulate the laying performance of layers, but its effects have not been fully understood.MethodsThis study investigated the effects of dietary supplementation of NCG on production performance, egg quality, and uterine gene expression in layers. A total of 360 45-week-old layers with a genetic line of Jinghong No. 1 were used in this study. The experimental period was 14 weeks. All birds were divided into 4 treatments with 6 replicates per treatment and 15 birds per replicate. Dietary treatments were based on a basal diet and supplemented with 0, 0.08, 0.12, or 0.16% NCG to form C, N1, N2, and N3 groups.Results and discussionWe found that layers in group N1 had higher egg production rate than those in group C. Egg weight was significantly reduced, while eggshell thickness was significantly improved, by treatment. However, the albumen height and Haugh unit were the lowest in group N3. Based on the above results, groups C and N1 were selected for further transcriptomics analysis of uterine tissue by RNA-seq. More than 7.4 Gb clean reads and 19,882 tentative genes were obtained using the Gallus gallus genome as a reference. Transcriptomics analysis in uterus tissue revealed that 95 differentially expressed genes (DEGs) were upregulated and 127 DEGs were downregulated. Functional annotation and pathway enrichment analysis showed that DEGs in uterine tissue were mainly enriched in glutathione metabolism, cholesterol metabolism, and glycerolipid metabolism, etc. Vitamin A metabolism-related gene, RBP1, nutrient transport-related gene, ALB, protein synthesis-related gene, METTL21C, and calcium transport-related gene, RYR2, CACNB2, RAMP3, and STAC, were significantly regulated by 0.08% NCG supplementation. Therefore, we concluded that NCG supplementation at a dose of 0.08% improved production performance and egg quality of layers by regulating uterus function.

Abstract Background Methylmalonic acidemia (MMA) is an inborn error of metabolism whose optimal management, especially in the long‐term remains to be established. Methods We describe the case of a child with MMA mut0 who was in a cycle of episodes of decompensation and hospitalization when we started to use carglumic acid (CA), a well‐known adjunctive therapy to standard care for the treatment of acute hyperammonemia due to MMA. Results Using the lowest effective therapeutic dose of CA and adjusting the patient's diet with caloric and protein intake adequate for her age and pathology, we managed to keep ammonium levels within the normal range, and to ensure a normal growth pattern. Conclusion The present case adds further confirmation of the long‐term management of MMA using CA, focusing on the long duration of follow up and on the use of a lower dose of CA in real life settings.

This study aimed to explore whether dietary rumen-protected L-arginine (RP-Arg) or N-carbamylglutamate (NCG) supplementation to feed-restricted pregnant ewes counteracts fetal hepatic inflammation and innate immune dysfunction associated with intrauterine growth retardation (IUGR) in ovine fetuses. On d 35 of pregnancy, twin-bearing Hu ewes (n = 32) were randomly assigned to 4 treatment groups (8 ewes and 16 fetuses per group) and fed diets containing 100% of the NRC requirements (CON), 50% of the NRC requirements (RES), RES + RP-Arg (20 g/d) (RESA), or RES + NCG (5 g/d) (RESN). At 08:00 on d 110 of gestation, fetal blood and liver tissue samples were collected. The levels of triglyceride, free fatty acid, cholesterol and β-hydroxybutyrate in the fetal blood of RESA and RESN groups were lower (P

ur previous studies have revealed that dietary N-carbamylglutamate (NCG) and L-arginine (Arg) supplementation improves redox status and suppresses apoptosis in the colon of suckling Hu lambs with intrauterine growth retardation (IUGR). However, no studies have reported the function of Arg or NCG in the colonic microbial communities, barrier function, and inflammation in IUGR-suckling lambs. This work aimed to further investigate how dietary Arg or NCG influences the microbiota, barrier function, and inflammation in the colon of IUGR lambs. Forty-eight newborn Hu lambs of 7 d old were assigned to four treatment groups (n = 12 per group; six male, six female) as follows: CON (normal birth weight, 4.25 ± 0.14 kg), IUGR (3.01 ± 0.12 kg), IUGR + Arg (2.99 ± 0.13 kg), and IUGR + NCG (3.03 ± 0.11 kg). A total of 1% Arg or 0.1% NCG was supplemented in a basal diet of milk replacer, respectively. Lambs were fed the milk replacer for 21 d until 28 d after birth. Compared to the non-supplemented IUGR lambs, the transepithelial electrical resistance (TER) was higher, while fluorescein isothiocyanate dextran 4 kDa (FD4) was lower in the colon of the NCG- or Arg-supplemented IUGR lambs (p < 0.05). The IUGR lambs exhibited higher (p < 0.05) colonic interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)-α, reactive oxygen species (ROS), and malondialdehyde (MDA) levels than the CON lambs; the detrimental effects of IUGR on colonic proinflammatory cytokine concentrations and redox status were counteracted by dietary Arg or NCG supplementation. Both IUGR + Arg and IUGR + NCG lambs exhibited an elevated protein and mRNA expression of Occludin, Claudin-1, and zonula occludens-1 (ZO-1) compared to the IUGR lambs (p < 0.05). Additionally, the lipopolysaccharide (LPS) concentration was decreased while the levels of acetate, butyrate, and propionate were increased in IUGR + Arg and IUGR + NCG lambs compared to the IUGR lambs (p < 0.05). The relative abundance of Clostridium, Lactobacillus, and Streptococcus was lower in the colonic mucosa of the IUGR lambs than in the CON lambs (p < 0.05) but was restored upon the dietary supplementation of Arg or NCG to the IUGR lambs (p < 0.05). Both Arg and NCG can alleviate colonic barrier injury, oxidative stress (OS), and inflammation by the modulation of colonic microbiota in IUGR-suckling lambs. This work contributes to improving knowledge about the crosstalk among gut microbiota, immunity, OS, and barrier function and emphasizes the potential of Arg or NCG in health enhancement as feed additives in the early life nutrition of ruminants. [ABSTRACT FROM AUTHOR]

Simple Summary: Larger litter size (born alive) and higher uniformity of piglets' birth weight are the most desired characteristics in pig production. Application of N-carbamylglutamate (NCG) was shown to increase litter size and decrease the coefficient of variation of piglets' birth weight from sows after conventional artificial insemination. In this study, we report that, in sows after fixed-time artificial insemination (FTAI), NCG improved the number of piglets born alive and the coefficient of variation of birth weight through direct effects (increased serum NCG levels) and indirect effects (altered intestinal microbiome and serum metabolites). These findings indicate that NCG can improve reproductive performance by improving both the number of piglets born alive and the uniformity of piglets' birth weight in sows after FTAI. N-carbamylglutamate (NCG) supplementation during gestation improves reproductive performance in sows after conventional artificial insemination. However, whether NCG can improve reproductive performance and change fecal microbiota and serum metabolite levels during pregnancy in sows after fixed-time artificial insemination (FTAI) remains unclear. Two hundred multiparous sows were assigned a diet from mating until farrowing: control (corn–soybean meal) or NCG supplementation (0.05% NCG). At days 30, 70, and 110 of gestation and after farrowing, maternal microbial diversity and serum metabolites were studied. Supplementation of NCG increased the number of piglets born alive and the litter weight (all p < 0.05) and altered the fetal microbial community during gestation. Some genera were particularly abundant at different time points during gestation and after farrowing, but none were commonly abundant across all four time points. Metabolic analysis revealed that NCG supplementation significantly increased the serum concentrations of NCG, ferulic acid, cinnamoylglycine, 3-phenyllactic acid, and gamma-glutamylglutamic acid in the NCG group compared with levels in the control group. Our results reveal that NCG supplementation during gestation improves reproductive performance in sows after FTAI, exerting both direct (increased serum NCG levels) and indirect effects (altered intestinal microbiome and serum metabolites) on sow reproduction and, ultimately, improving placental and fetal development. [ABSTRACT FROM AUTHOR]

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.

The objective of this study was to investigate the effects of dietary administration of l-arginine (Arg) or N-carbamylglutamate (NCG) on hepatic energy status and mitochondrial functions in suckling Hu lambs with intrauterine growth retardation (IUGR). Forty-eight newborn Hu lambs of 7 d old were allocated into 4 treatment groups of 12 lambs each, in triplicate with 4 lambs per replicate (2 males and 2 females) as follows: CON (lambs of normal birth weight, 4.25 ± 0.14 kg), IUGR (3.01 ± 0.12 kg), IUGR + 1% Arg (2.99 ± 0.13 kg), or IUGR + 0.1% NCG (3.03 ± 0.11 kg). The experiment lasted for 21 d, until d 28 after birth, and all lambs were fed milk replacer as a basal diet. Compared with IUGR lambs, NCG or Arg administration increased (P

The objective of the present study was to investigate the effects of N-carbamylglutamate (NCG) supplementation on metabolic profile and microbiota in ruminal content and feces of lactating dairy cows under heat stress (HS). Forty-eight lactating Holstein cows (154 ± 13.6 days in milk) were assigned randomly to four treatments (n = 12), to receive 0, 15, 20, or 25 g/day of commercial NCG (proportion: 97.7%) for the period of 60 days. The recorded ambient temperature–humidity index (THI) suggested that the cows were exposed to HS for almost the entire experimental period (average THI: 80.6). Samples of ruminal content and feces were collected at the end of the trial (day 60) to determine the biological effects of NCG supplementation on metabolome and microbiota using mass spectrometry-based metabolomics and 16S rRNA gene sequencing techniques, respectively. Results showed that NCG supplementation enhanced the levels of ruminal microbial protein, total volatile fatty acids (VFAs), and the molar proportion of propionate in the rumen, but lowered the ruminal pH, ammonia nitrogen (NH3-N), and the ratio of acetate to propionate. NCG at doses of 20 and 25 g/day reduced the community richness and diversity of ruminal microbiota with the decrease of Shannon and Simpson diversity. Compositions of ruminal and fecal microbiotas were altered by NCG, and the PICRUSt results revealed that metabolic pathways of the bacteria, such as amino acid metabolism, energy metabolism, and pyruvate metabolism, were enriched in NCG groups. Distinct changes in the metabolomic profile of ruminal fluid were observed between the control and NCG groups. Changes of 26 metabolites mainly involved in arginine metabolism, glutamate metabolism, and nitrogen metabolism were observed associated with NCG supplementation. These results provided new insights into the effects of NCG on metabolomic profile and microbiota in ruminal content and feces, and the optimal dose of NCG supplemented to dairy cows was 20 g/hd/day, which contributed to understanding the effects of NCG on HS in lactating dairy cows.

This study investigated the effects of dietary supplementation with N‐carbamylglutamate (NCG) on maternal endometrium and fetal development during early pregnancy of Inner Mongolia white cashmere goats. Forty‐eight pregnant Inner Mongolia white cashmere goats (average age 3 years old, average lactation parity 2, and average body weight 43.81 ± 2.66 kg) were randomly allocated to three groups: a basal diet (control group, n = 16), a basal diet plus 0.30‐g NCG/d (NCG1 group, n = 16), and a basal diet plus 0.40‐g NCG/d (NCG2 group, n = 16). All of the does were housed in individual pens and the NCG treatment was conducted from Days 0 to 90 of pregnancy. At Days 17 and 90 of pregnancy, six representative pregnant does in each group were slaughtered. The current study results demonstrated that maternal NCG administration during early pregnancy effectively increased the arginine family of amino acids and the glucogenic amino acids concentrations and promoted the mRNA expression of osteopontin (OPN), αv and β3 integrins, and endometrial development of Inner Mongolia white cashmere goats. The supplementation improved the fetal brown adipose tissue (BAT) stores and the mRNA expression of UCP‐1 and BMP7, thereby helping to the fetal early development. [ABSTRACT FROM AUTHOR]

Egg quality defects seriously reduce the quality grade and increase egg breakage in egg marketing activities. In this study, the effect of N-carbamylglutamate (NCG) on eggshell quality was investigated by evaluating calcium absorption and calcification in laying hens. A total of 30 newly hatched female Hy-Line chicks were randomly assigned to the control group (basal diet) and treatment group (basal diet supplemented with 1% NCG). At 25 wk, eggs from each group were obtained to assess egg quality parameters. Blood samples were collected for analysis of mineral, hormone, and amino acids levels at 16 h after laying egg. Uterine tissues were removed and fixed in 4% neutral paraformaldehyde solution or kept in liquid nitrogen for mineral determination, quantitative PCR, and Western blot. Results showed that the egg quality (eggshell thickness, strength and percentage, egg specific gravity, and eggshell effective thickness) was significantly increased while effective thickness of mastoid layer, width of mastoid gap, and mammillary knobs were significantly decreased by dietary NCG supplementation (P

ABSTRACT: N-carbamylglutamate (NCG), an analogue of N-acetyl-L-glutamate (NAG), can increase arginine synthesis in mammals and improve the reproductive performance. However, the effect of NCG on poultry laying performance is still unclear. This study investigated the effect of dietary NCG on development of chicken ovarian follicles. The dosage and timing for NCG administration were evaluated for its effect on follicular development. Results showed that supplementation with 1% NCG in the diet for 14 D led to accelerated development of growing follicles (over 60 μm in oocyte diameter) and significantly increased feed intake and feed efficiency. Plasma amino acids (AA) analysis showed that feeding with 1% NCG significantly increased of plasma AA levels. RNA-seq analysis revealed that NCG supplementation upregulated expression of genes related to angiogenesis and cell proliferation, but downregulated expression of apoptosis-related genes. Meanwhile, RT-qPCR and Western blot analysis validated the RNA-seq results. Moreover, NCG enhanced plasma NO level; upregulated expression of PKG-I, Raf1, and p-p38; and increased angiogenesis of the ovaries. In conclusion, dietary NCG (1% for 14 D) can promote development of ovarian follicles by increasing angiogenesis in ovaries of the chicken.

Ammonia is a common environmental limiting factor in aquaculture. To investigate the effects of ammonia stress and explore the protective effect of N-carbamylglutamate (NCG) on Micropterus salmoides (M. salmoides), tissue sections and parameters related to oxidative stress and the inflammatory response in M. salmoides were carried out during the ammonia stress test and feeding test. The results demonstrated that the LC50 for 24 h, 48 h, 72 h, and 96 h under ammonia stress in M. salmoides were 25.78 mg/L, 24.40 mg/L, 21.90 mg/L, and 19.61 mg/L, respectively. Under ammonia stress, the structures of the tissues were damaged, and the GSH content decreased, while the MDA content increased with the increase in stress time and ammonia concentration. The NO content fluctuated significantly after the ammonia nitrogen stress. In the 15-day feeding test, with the increased NCG addition amount and feeding time, the GSH content increased while the MDA and NO contents decreased gradually in the NCG addition groups (NL group: 150 mg/kg; NM group: 450 mg/kg; NH group: 750 mg/kg) when compared with their control group (CK group: 0 mg/kg). In the ammonia toxicology test after feeding, the damage to each tissue was alleviated in the NL, NM, and NH groups, and the contents of GSH, MDA, and NO in most tissues of the NH group were significantly different from those in the CK group. The results suggested that ammonia stress caused tissue damage in M. salmoides, provoking oxidative stress and inflammatory response. The addition of NCG to the feed enhances the anti-ammonia ability of M. salmoides. Moreover, the gill and liver might be the target organs of ammonia toxicity, and the brain and kidney might be the primary sites where NCG exerts its effects. Our findings could help us to find feasible ways to solve the existing problem of environmental stress in M. salmoides culture.

N-carbamylglutamate (NCG) supplementation during gestation improves reproductive performance in sows after conventional artificial insemination. However, whether NCG can improve reproductive performance and change fecal microbiota and serum metabolite levels during pregnancy in sows after fixed-time artificial insemination (FTAI) remains unclear. Two hundred multiparous sows were assigned a diet from mating until farrowing: control (corn–soybean meal) or NCG supplementation (0.05% NCG). At days 30, 70, and 110 of gestation and after farrowing, maternal microbial diversity and serum metabolites were studied. Supplementation of NCG increased the number of piglets born alive and the litter weight (all p < 0.05) and altered the fetal microbial community during gestation. Some genera were particularly abundant at different time points during gestation and after farrowing, but none were commonly abundant across all four time points. Metabolic analysis revealed that NCG supplementation significantly increased the serum concentrations of NCG, ferulic acid, cinnamoylglycine, 3-phenyllactic acid, and gamma-glutamylglutamic acid in the NCG group compared with levels in the control group. Our results reveal that NCG supplementation during gestation improves reproductive performance in sows after FTAI, exerting both direct (increased serum NCG levels) and indirect effects (altered intestinal microbiome and serum metabolites) on sow reproduction and, ultimately, improving placental and fetal development.

N-carbamylglutamate (NCG) supplementation promotes the synthesis of arginine, improves physiological parameters and modulates gut function in fish. However, its beneficial effect on the fish gut microbiota remains unclear. Thus, we investigated the beneficial effect of NCG on homeostasis of gut microbiota in tilapia. Tilapia were fed with diets supplemented with different levels of dietary NCG (Ctrl, NCGI and NCGII) for 8 weeks. The weight gain and the specific growth rate were significantly higher (p < 0.05) in NCG-supplemented groups compared to the group Ctrl. Moreover, the V3-V4 region of 16S rDNA was sequenced, and the bioinformatic analysis suggested that dietary NCG supplementation had a limited impact on the α-diversity of gut microbiota, while Non-metric multidimensional scaling analysis (NMDS) showed that dietary NCG leads to a clear separation on gut microbiota among the three groups. Furthermore, NCG supplementation reshaped the tilapia gut microbiota community, according to co-occurrence analysis and metabolic pathways prediction via PICRUSt analysis, leading to an enrichment of nutritional metabolism related functions, such as amino acid and lipid metabolisms (p < 0.05). Subsequently, the redundancy analysis (RDA) demonstrated that the structural changes of gut microbiota were determined by NCG concentration. In addition, the linear discriminant analysis effect size (LEfSe) analysis revealed 65 differentially abundant taxonomic clades were dominant in tilapia supplemented with NCG. The cross-validation and Random Forest model identified 12 key bacterial taxa in response to the NCG supplementation. Among them, four key bacterial taxa played a fundamental role in nitrogen-fixing, which may indicate that NCG increases the amino acid biosynthesis of gut microbiota and contributes to the growth promotion of tilapia.

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Background: Classic organic acidurias (OAs) usually characterized by recurrent episodes of acidemia, ketonuria, and hyperammonemia leading to coma and even death if left untreated. Acute hyperammonemia episodes can be treated effectively with N-carbamylglutamate (NCG). The effect of the long-term efficacy of N-carbamylglutamate is little known. Material-Methods: This retrospective study was conducted at Istanbul University-Cerrahpasa, Cerrahpasa Medical Faculty, Pediatric Nutrition and Metabolism Clinic between January 2012 to January 2018. Patients with classic OAs were enrolled in the study. Patients' ammonia levels, hospitalization needs, hyperammonemia episodes, and management of hyperammonemia were recorded. NCG usage for more than consecutively 15 days was considered as a long-term treatment. Results: Twenty-one patients, consisting of eleven patients with methylmalonic acidemia (MMA) and ten patients with propionic acidemia (PA) were eligible for the study. N-carbamylglutamate was used as ammonia scavenger for a total of 484 months with a median period of 23 months (min-max: 3–51 months) in all patients. A significant decrease in plasma ammonia levels was detected during long term NCG treatment (55.31 ± 13.762 μmol/L) in comparison with pre NCG treatment period (69.64 ± 17.828 μmol/L) (p = 0.021). Hospitalization required hyperammonemia episodes decreased with NCG treatment (p = 0.013). In addition, hyperammonemia episodes were also successfully treated with NCG (p = 0.000). Mean initial and final ammonia levels at the time of hyperammonemia episodes were 142 ± 46.495 μmol/L and 42.739 ± 12.120 μmol/L, respectively. The average NCG dosage was 85 mg/kg/day (range 12.5–250 mg/kg/day). No apparent side effects were observed. Conclusion: N-Carbamylglutamate may be deemed an effective and safe treatment modality in the chronic management of hyperammonemia in patients with PA and MMA.

The detoxification of ammonia to urea requires a functional hepatic urea cycle, which consists of six enzymes and two mitochondrial membrane transporters. The initial step of the urea cycle is catalyzed by carbamyl phosphate synthetase 1 (CPS1). CPS1 deficiency (CPS1D) is a rare autosomal recessive disorder. N-Carbamylglutamate (NCG), a deacylase-resistant analogue of N-acetylglutamate, can activate CPS1. We describe the therapeutic course of a patient suffering from neonatal onset CPS1D with compound heterozygosity for the c.2359C > T (p.Arg787*) and c.3559G > T (p.Val1187Phe) variants in CPS1, treated with NCG. She presented with hyperammonemia, which reached 944 μmol/L at the age of 2 days. The ammonia concentration decreased after treatment with continuous hemodiafiltration, NCG, sodium benzoate, sodium phenylbutyrate, L-arginine, vitamin cocktail (vitamin B1, vitamin B12, vitamin C, vitamin E, biotin), l-carnitine, coenzyme Q10, and parenteral nutrition. Her ammonia and glutamine levels remained low; thus, protein intake was increased to 1.2 g/kg/day. Furthermore, the amount of sodium benzoate and sodium phenylbutyrate were reduced. She remained metabolically stable and experienced no metabolic crisis following treatment with oral NCG, sodium benzoate, sodium phenylbutyrate, citrulline, vitamin cocktail, l-carnitine, and coenzyme Q10 until she underwent liver transplantation at 207 days of age. She had no neurological complications at the age of 15 months. Ammonia and glutamine levels of the patient were successfully maintained at a low level via NCG treatment with increased protein intake, which led to normal neurological development. Thus, undiagnosed urea cycle disorders should be treated rapidly with acute therapy including NCG, which should be maintained until a genetic diagnosis is reached. It is essential to prevent metabolic crises in patients with CPS1D until liver transplantation to improve their prognoses.

Abstract Background High-altitude pulmonary hypertension (HAPH) is a life-threating condition for animals in high altitude, and disturbance of endothelial nitric oxide (NO) synthesis contributes to its pathogenesis. N-carbamylglutamate (NCG), which enhances arginine synthesis, promotes endogenous synthesis of NO. In this study, we determined the effects of NCG on alleviating HAPH in Holstein heifers that ascended to Tibet (Lhasa, 3,658 m). Methods Exp. 1, 2,000 Holstein heifers were transported from low elevation (1,027 m) to Lhasa. After being exposed to hypoxia for 1 yr, Holstein heifers were assigned to a healthy group (Control, n = 6) with mean pulmonary hypertension (mPAP) 49 mmHg. Lung tissues were collected to evaluate histopathological changes and the expression of endothelial nitric oxide synthase (eNOS). Exp. 2, ten healthy heifers and 10 HAPH affected heifers were supplemented with NCG (20 g/d per heifer) for 4 wk. Physiological parameters were determined and blood samples were collected on d − 1 and d 28 of the feeding trial. Results Expression of eNOS in small pulmonary arteriole intima was higher in the healthy than HAPH group (P = 0.006), whereas HAPH group had significantly thicker media and adventitia than healthy group (all P

Abstract Background Propionic acidemia is a rare autosomal recessive inherited metabolic disorder that can inhibit the synthesis of N-acetylglutamate, the obligatory activator in urea synthesis, leading to hyperammonemia. N-carbamylglutamate ameliorates hyperammonemia in decompensated propionic acidemia. The effects of long-term continuous N-acetylglutamate administration in such patients are unknown. We report our clinical experience with continuous administration of N-acetylglutamate for 6 years in a patient with propionic acidemia frequently presenting with hyperammonemia. Case presentation A male Caucasian patient with frequently decompensated propionic acidemia and hyperammonemia was admitted 78 times for acute attacks during the first 9 years of his life. Continuous daily treatment with oral N-carbamylglutamate 100 mg/kg (50 mg/kg after 6 months) was initiated. During 6 years of treatment, he had a significant decrease in his mean plasma ammonia levels (75.7 μmol/L vs. 140.3 μmol/L before N-carbamylglutamate therapy, p < 0.005 [normal range 50–80 μmol/L]) and fewer acute episodes (two in 6 years). Conclusion Our results suggest a benefit of N-acetylglutamate administration outside the emergency setting. If this observation is confirmed, future studies should aim to optimize the dosage and explore effects of the dosage requirements on other drugs and on protein tolerance.

The objectives of this experiment were to investigate the effects of N-carbamylglutamate (NCG) on growth and slaughter performance, meat quality, nitrogen utilization, plasma antioxidant and amino acids of Holstein bulls. In this case, 24 Holstein bulls (490 ± 29.0 kg of body weights and 540 ± 6.1 d of age) were blocked by body weights and age and randomly assigned to 1 of 4 groups: (1) CON group: bulls were fed the control diet, (2) CON + NCG group: bulls were fed the control diet with 40 mg/kg BW NCG, (3) Urea group: bulls were fed the urea diet, and (4) Urea + NCG group: bulls were fed the urea diet with 40 mg/kg BW NCG. Feeding NCG significantly improved ADG, FCR, DM and CP digestibility, carcass weight, slaughter weight, DOP, eye muscle area, shear force (p = 0.001) and reduced L* of color, drip loss and cooking loss. Concurrently, feeding the urea diet induced a decreased ADG, carcass weight and slaughter weight, DOP, eye muscle area and shear force. NCG decreased contents of fecal N and urinary N, plasma urea in bulls and ammonia but increased N retention and utilization, plasma NO, plasma Arg, Leu, Ile and Tyr. On the other hand, feeding the urea diet increased urinary N, plasma urea and ammonia. Thus the study efficiently demonstrates that beef benefited from being fed a NCG product in the urea diet by enhancing its growth and slaughter performance, meat quality, nitrogen metabolism and plasma amino acids.

Egg quality defects seriously reduce the quality grade and increase egg breakage in egg marketing activities. In this study, the effect of N-carbamylglutamate (NCG) on eggshell quality was investigated by evaluating calcium absorption and calcification in laying hens. A total of 30 newly hatched female Hy-Line chicks were randomly assigned to the control group (basal diet) and treatment group (basal diet supplemented with 1% NCG). At 25 wk, eggs from each group were obtained to assess egg quality parameters. Blood samples were collected for analysis of mineral, hormone, and amino acids levels at 16 h after laying egg. Uterine tissues were removed and fixed in 4% neutral paraformaldehyde solution or kept in liquid nitrogen for mineral determination, quantitative PCR, and Western blot. Results showed that the egg quality (eggshell thickness, strength and percentage, egg specific gravity, and eggshell effective thickness) was significantly increased while effective thickness of mastoid layer, width of mastoid gap, and mammillary knobs were significantly decreased by dietary NCG supplementation (P < 0.05). The levels of minerals (Ca, P, Fe, Mg, Na, K) in eggshell, plasma, and uterus were remarkably elevated (P < 0.05). Meanwhile, the concentrations of calcium metabolism–related hormones (17β-estradiol, parathyroid hormone, and calcitonin) were increased in the NCG group (P < 0.05). Moreover, expression of calbindin 1, carbonic anhydrase 2, ovalbumin, ovotransferrin, ovocleidin-17, ovocleidin-116, and clusterin mRNAs, as well as calbindin 1 and ATP2A1 proteins in uterus, duodenum, and kidney, was all upregulated in hens fed with NCG (P < 0.05). In addition, the number of blood vessels in the uterus, height of uterine mucosal folds, villus length in endometrium, and areas of uterine mucosal folds were significantly increased in the NCG group (P < 0.05). In conclusion, dietary 1% NCG supplementation during 0 to 25 wk can improve eggshell quality through changes in endometrial morphology, expression of calcium metabolism–related genes, and secretion of related hormones to promote eggshell formation in the laying hens. [ABSTRACT FROM AUTHOR]

N-Carbamylglutamate (NCG) has been shown to enhance arginine synthesis and improve growth performance in animals. However, the effect of NCG on body fat deposition remains unknown. This study examined the effects of NCG on body fat deposition and evaluated the potential mechanisms involved. Rex rabbits (3 months old) were assigned to one of four dietary groups and supplemented with NCG at the following different concentrations in a feeding trial that lasted 67 d: 0 (control), 0·04, 0·08, and 0·12 %. NCG supplementation increased serum concentrations of arginine and proline by activating intestinal carbamoylphosphate synthase-І at the posttranscriptional level. Final body weights and growth performance were not affected by dietary NCG levels. However, NCG-treated rabbits had lower perirenal and subcutaneous fat percentages, serum TAG content, and hepatic fatty acid synthase (FAS) activity and increased NO synthase activity and serum levels of NO, growth hormone (GH), and insulin-like growth factor 1 (IGF-1). There were significant positive correlations between TAG content and perirenal fat percentage, as well as FAS activity and perirenal fat percentage, but significant negative correlations between TAG and NO levels, and FAS activity and IGF-1 level in rabbits after NCG treatment. NCG supplementation did not affect hepatic health indicators, except for serum ammonia concentrations, which were decreased in NCG-treated rabbits. Our results suggest that NCG can serve as a dietary supplement to reduce unfavourable fat deposition through inhibiting hepatic lipogenesis in animals since it appears to have no negative effects on growth performance or hepatic health. [ABSTRACT FROM AUTHOR]

The effects of N‐carbamylglutamate (NCG) on growth, intestinal enzyme activities, immunological and antioxidant parameters were evaluated by a 56‐d feeding trial in Pelteobagrus fulvidraco fed diets containing NCG with 0, 250, 500, 1,000 or 2,000 mg/kg, respectively. The results showed that 250 mg/kg of NCG resulted in significantly higher weight gain, intestine fold height, intestine lipase, serum lysozyme, glutathione peroxidase and total antioxidant capacity than control (p < 0.05). However, higher intestine trypsin, arginase, arginine decarboxylase, ornithine decarboxylase, diamine oxidase activities and serum nitric oxide content were observed in 500 mg/kg NCG group compared to control or 2,000 mg/kg (p < 0.05). The survival rate, intestine muscular layer thickness, serum lysozyme and superoxide dismutase activities in 2,000 mg/kg NCG group were significantly lower than those in control and 250 mg/kg group, accompanied by the higher feed conversion ratio in the same group (p < 0.05). Together, dietary NCG level at 250 or 500 mg/kg improved growth, intestinal enzyme activities, immunological and antioxidant abilities, while high NCG level of 2,000 mg/kg had a negative effect. Quadratic regression analysis on weigh growth, diamine oxidase and lysozyme activities indicated that the recommended optimum dietary NCG level was 213.48–314.50 mg/kg of the dry diet. [ABSTRACT FROM AUTHOR]

Eggs rich in polyunsaturated fatty acids (PUFA), known as functional eggs, are animal products deemed beneficial to human health and possess high economic value. The production of functional eggs involves supplementing exogenous additives with the ability to regulate lipid metabolism. As N-Carbamylglutamate (NCG) serves as an endogenous arginine synthesizer, and arginine acts as the substrate for the formation of nitric oxide (NO), the biological function of NCG is partially mediated by NO. NO is a key regulatory molecule in lipid metabolism, suggesting that NCG may also have the ability to modulate lipid metabolism. In order to assess the capacity of NCG in regulating liver lipid metabolism and its potential application in producing functional eggs, we conducted a study to investigate the effects of dietary supplementation of NCG on production performance, serum, and liver NO levels, yolk fatty acid composition, and the liver transcriptome of layers. In this study, we utilized 30 layers of the Jinghong No.1 breed, all aged 45 wk. All the birds were randomly divided into 2 groups. Each group had 5 replicates, and each replicate had 3 birds. We provided them with different diets: one group received the basic diet, and the other group's diet was supplemented with 0.08% NCG. The experiment lasted for 14 wk. The results did not reveal any positive impact of NCG on production performance. However, NCG supplementation elevated NO levels in serum and liver, along with an increase in yolk PUFA, ω-3, and ω-6 fatty acids. Liver transcriptome analysis identified 124 upregulated differentially expressed genes (DEGs) and 43 downregulated DEGs due to NCG supplementation. Functional annotation using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database highlighted 3 upregulated DEGs (CPT1A, MOGAT1, and CHKA) and 2 downregulated DEGs (FASN and ETNPPL) associated with lipid metabolism. Pathway enrichment analysis revealed that CPT1A was enriched in the AMPK signaling pathway and the PPAR signaling pathway, while FASN was enriched in the AMPK signaling pathway. Thus, CPT1A and FASN are potential functional genes related to lipid metabolism facilitated by NCG supplementation. In summary, our study suggests that NCG supplementation modulates liver lipid metabolism, leading to the production of functional eggs in layers. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the effects of N-carbamylglutamate (NCG) supplementation on the follicular development of yaks to identify potential mechanisms essential for fertility in yaks. Twelve multiparous anoestrous female yaks were randomly assigned to two groups—Control (fed with a basal diet, n = 6) and NCG (basal diet supplemented with 6.0 g day−1 NCG, n = 6). Yaks in the NCG group had higher numbers of large follicles (>5 mm in diameter) than those in the Control group. An RNA-sequencing analysis of yak ovaries revealed a total of 765 genes were differentially expressed between experimental groups, of which 181 genes were upregulated and 584 genes were downregulated following NCG supplementation. The results of a transcriptome functional analysis, qRT-PCR validation, and immunohistochemistry revealed that NCG supplementation increased angiogenesis and de novo synthesis of cholesterol in yak ovaries. NCG was also found to upregulate the gene expression of steroidogenic enzymes. Based on this, it was concluded that NCG supplementation promotes the follicular development of yaks mainly by affecting cholesterol metabolism to initiate steroidogenesis in ovaries. The results provide evidence for understanding the mechanisms responsible for NCG promoting follicular development of female yaks, which may contribute to the development and application of NCG in animal reproduction.

N-Carbamylglutamate (NCG), an analogue of N-acetylglutamate (NAG), can promote the synthesis of endogenous Arginine (Arg) in mammals, but not well studied in fish. This study was conducted to investigate the capacity of Arg endogenous synthesis by NCG, and the effects of various dietary NCG doses on growth performance, hepatic health and underlying nutrient regulation metabolism on ERK1/2-mTOR-S6K1 signaling pathway in Japanese seabass (Lateolabrax japonicus). Four experimental diets were prepared with NCG supplement levels of 0 (N0), 360 (N360), 720 (N720) and 3600 (N3600) mg/kg, in which N360 was at the maximum recommended level authorized by MOA, China in fish feed, and the N720 and N3600 levels were 2 and 10-fold of N360, respectively. Each diet was fed to 6 replicates with 30 Japanese seabass (initial body weight, IBW = 11.67 ± 0.02 g) in each tank. The results showed that the dietary NCG supplementation had no significant effects on the SGR and morphometric parameters of Japanese seabass, but 360–720 mg/kg NCG inclusion promoted PPV, while the 10-fold (3600 mg/kg) overdose of NCG had remarkably negative effects with significantly reduced feed efficiency, PPV and LPV. We found that Japanese seabass can utilize 360–720 mg/kg NCG to synthesis Arg to improve the amino acid metabolism by increasing plasma Arg and up-regulating intestinal ASL gene expression. Increased plasma GST and decreased MDA indicated the improved antioxidant response. Dietary NCG inclusion decreased plasma IgM and down-regulated the mRNA levels of inflammation (TNF-α and IL8), apoptosis (caspase family) and fibrosis (TGF-β1) related genes in the liver. The immunofluorescence examination revealed significantly decreased hepatic apoptosis and necrosis signals in the NCG groups. The ameliorated liver function and histological structure were closely related to the improved lipid metabolism parameters with decreased plasma VLDL and hepatic TG and NEFA accumulation, down-regulated fatty acid and cholesterol synthesis and simultaneously increased lipolysis gene mRNA levels, which regulated by inhibiting phosphorylation of ERK1/2-mTOR-S6K1 signaling pathway. Consuming 3600 mg/kg of dietary NCG is not safe for Japanese seabass culturing with the significantly increased FCR and decreased protein and lipid retention, and reduced plasma ALB. Accordingly, the observed efficacy and safety level of dietary NCG in the diet of Japanese seabass is 720 mg/kg. Arginine deficiency induces dysfunction of lipid metabolism in liver and highly expressed pro-inflammatory cytokines, which induced hepatocyte apoptosis. Dietary NCG alleviates the fatty liver and hepatocyte apoptosis by suppresses ERK1/2-mTOR-S6K1 protein phosphorylation to inhibiting lipogenesis and up-regulating lipolysis gene expression via promoting endogenous arginine synthesis in Japanese seabass. Image 1 • Japanese seabass can utilize NCG to synthesis endogenous arginine in vivo. • Dietary NCG attenuated hepatic inflammation and apoptosis of Japanese seabass. • NCG alleviated liver metabolic disease by improving lipid metabolism, which regulated by inhibiting ERK1/2-mTOR-S6K1 pathway; • The observed efficacy and safety level of dietary NCG in the diet of Japanese seabass is 720 mg/kg. • Overdose of dietary NCG (3600 mg/kg) decreased protein and lipid retention and plasma ALB level. [ABSTRACT FROM AUTHOR]