Determination of N-Carbamylglutamate in Feeds and Animal Products by High Performance Liquid Chromatography Tandem Mass Spectrometry

Abstract

N-carbamylglutamate (NCG), a synthetic analogue of N-acetylglutamate, is an activator of blood ammonia conversion and endogenous arginine synthesis. Here, we established an accurate quantitative determination of NCG in feeds, animal tissues, and body fluids using the high performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). The sample pretreatment procedures included extraction with 0.5% of formic acid in water/methanol (80/20, v/v), and purification using an anionic solid phase extraction cartridge. Satisfactory separation of NCG was achieved in 20 min with the application of an Atlantis T3 column, and a confirmative detection of NCG was ensured by multiple reaction monitoring of positive ions. NCG spiked in feeds, tissues, and body fluids were evaluated in regard to linearity, sensitivity, recovery, and repeatability. Recoveries for different sample matrices were in the range of 88.12% to 110.21% with relative standard deviations (RSDs) less than 8.8%. Limits of quantification were within the range of 0.012 to 0.073 mg kg−1 and 0.047 to 0.077 μg mL−1 for solid and liquid samples, respectively. This study will provide a solid foundation for the evaluation of availability and metabolic mechanism of NCG in animals.

Effect of L‐citrulline supplementation on sperm characteristics and hormonal and antioxidant levels in blood and seminal plasma of rams

“…After feeding L ‐Cit to rams, our findings showed that serum levels of GnRH, FSH, LH and T, and seminal plasma increased significantly or very significantly, which was positively correlated with an increase in semen level, indicating that L ‐Cit can promote the release of reproductive hormone and regulate the semen quality of rams. L ‐Cit has two main metabolic pathways in the body, namely the NO and polyamine pathways (Ma et al., 2019). NO plays an important role in reproductive function and can participate in gonadotropin secretion, sexual behaviour, E 2 synthesis and ovulation (Dixit & Parvizi, 2001).…”

Effects of dietary N-carbamylglutamate supplementation on milk production performance, nutrient digestibility and blood metabolomics of lactating Holstein cows under heat stress

“…N-carbamylglutamate (NCG) is an analogue of NAG, which can enter the mitochondria to activate carbamylphosphate synthase-1 (CPS-1) and therefore promote the synthesis of arginine. In addition, NCG is stable in the ruminal environment and dose not antagonize other amino acids such as lysine and histidine [5]. Thus, oral supplementation of NCG can increase the endogenous synthesis of arginine.…”

Effects of dietary N-carbamylglutamate on rumen fermentation parameters, and bacterial community diversity of Holstein dairy cows in Tibet

“…Although Arg (Arginine) could attenuate inflammatory stress and significantly improve lactating performance of dairy cows ( Wang, 2016 ; Zhao et al, 2018 ; Ding et al, 2019 ), and improve the altitude reaction in high altitude animals by increasing blood oxygen saturation ( Schneider et al, 2001 ), the rapid degradation in the rumen ( Chacher et al, 2012 ) and the high price hindered its utilization in ruminant production. In the meanwhile, N-carbamoyl glutamate (NCG), a structural analog of N-acetylglutamate acts as an allosteric activator of essential cofactor aminophosphate synthase in endogenous arginine synthesis ( Gessler et al, 2010 ), is much more stable than Arg in the rumen ( Ma et al, 2019 ). The NCG is non-toxic to livestock ( Guffon et al, 1995 ), and previous studies consistently proven that supplementation of NCG to ruminant diets could improve milk yield and production performance for dairy cows ( Chacher et al, 2014 ; Ma et al, 2020 ; Gu et al, 2021 ), and improve reproduction of gestation Hu sheep ( Sun et al, 2017 ) and improved the health and growth of suckling Hu lambs ( Zhang et al, 2018 ).…”