The impact of excess dietary leucine (Leu) was studied in two growth assays with pigs (8-25 kg). In each trial, forty-eight pigs were allotted to one of six dietary groups. The dietary Leu supply increased from treatment L100 to L200 (three increments). To guarantee that interactions between the branched-chain amino acids (BCAA) were not cushioned either surpluses of isoleucine (Ile, expt 1) or valine (Val; expt 2) were avoided. In the fifth treatment, the effects of a simultaneous excess of Leu and Val (expt 1), or of Leu and Ile (expt 2) were investigated. The sixth treatment was a positive control. An increase in dietary Leu decreased growth performance, and increased plasma Leu and serum a-keto-isocaproate levels in a linear, dose-dependent manner. Levels of plasma Ile and Val, and of serum a-keto-b-methylvalerate and a-keto-isovalerate, indicated increased catabolism. Linear increases in the activity of basal branched-chain a-keto acid dehydrogenase in the liver confirmed these findings. No major alterations occurred in the mRNA of branched-chain amino acid catabolism genes. In liver tissue from expt 2, however, the mRNA levels of growth hormone receptor, insulin-like growth factor acid labile subunit and insulin-like growth factor 1 decreased significantly with increasing dietary Leu. In conclusion, excess dietary Leu increased the catabolism of BCAA mainly through posttranscriptional mechanisms. The impact of excess Leu on the growth hormone -insulin-like growth factor-1 axis requires further investigation.Leucine excess: Amino acid interactions: Branched-chain a-keto acid dehydrogenase: PigsInteractions among the branched-chain amino acids (BCAA), such as the performance depressing effects of excess dietary leucine (Leu), are known in several species (1) . The impact of high dietary Leu levels needs to be elucidated in order to make correct estimates of adequate supplies and requirements for isoleucine (Ile) and valine (Val). Interactions among the BCAA include their catabolism, because all three compete for the same enzymes that catalyse the first two catabolic steps. The first step is a reversible transamination catalysed by the branched-chain amino acid transaminase (BCAT) isoenzymes, yielding branched-chain a-keto-acids (BCKA) that, in the second step, are oxidatively decarboxylated by a mitochondrial, multienzyme branchedchain a-keto acid dehydrogenase (BCKDH) complex. This step is irreversible, highly regulated and rate limiting for BCAA catabolism. The BCKDH complex consists of three catalytic components. The E1 subunit, a heterotetramer of a and b subunits, is a branched-chain a-keto acid decarboxylase. The E2 subunit is a dihydrolipoamide acyltransferase and the E3 subunit is a dihydrolipoamide dehydrogenase (2) . In contrast to the other subunits, the E3 is not BCKDHspecific and its expression is not analysed in the present work. BCKDH complex activity is regulated by covalent modification. Phosphorylation of its E1a subunits by a specific BCKDH kinase (BDKDK) causes inactivation, and dephospho...

The control of Maillard reaction (MR) is a key point to ensure processed foods quality. Due to the presence of a primary amino group on its side chain, lysine is particularly prone to chemical modifications with the formation of Amadori products (AP), Nε-(Carboxymethyl)-L-lysine (CML), Nε-(Carboxyethyl)-L-lysine (CEL). A new analytical strategy was proposed which allowed to simultaneously quantify lysine, CML, CEL and the Nε-(2-Furoylmethyl)-L-lysine (furosine), the indirect marker of AP. The procedure is based on stable isotope dilution assay followed by liquid chromatography tandem mass spectrometry. It showed high sensitivity and good reproducibility and repeatability in different foods. The limit of detection and the RSD% were lower than 5 ppb and below 8%, respectively. Results obtained with the new procedure not only improved the knowledge about the reliability of thermal treatment markers, but also defined new insights in the relationship between Maillard reaction products and their precursors.

Background Bacillus spp. seem to be an alternative to antimicrobial growth promoters for improving animals’ health and performance. However, there is little information on the effect of Bacillus spp. in combination with different dietary crude protein (CP) levels on the ileal digestibility and microbiota composition. Therefore, the objective of this study was to determine the effect of Bacillus spp. supplementation to low- (LP) and high-protein diets (HP) on ileal CP and amino acid (AA) digestibility and intestinal microbiota composition.MethodsEight ileally cannulated pigs with an initial body weight of 28.5 kg were randomly allocated to a row-column design with 8 pigs and 3 periods of 16 d each. The assay diets were based on wheat-barley-soybean meal with two protein levels: LP (14% CP, as-fed) and HP diet (18% CP, as-fed). The LP and HP diets were supplemented with or without Bacillus spp. at a level of 0.04% (as-fed). The apparent ileal digestibility (AID) and standardized ileal digestibility (SID) of CP and AA was determined. Bacterial community composition from ileal digesta was analyzed by Illumina amplicon sequencing and quantitative real-time PCR. Data were analyzed as a 2 × 2 factorial design using the GLIMMIX procedures of SAS.ResultsThe supplementation with Bacillus spp. did not affect both AID and SID of CP and AA in growing pigs. Moreover, there was no difference in AID of CP and AA between HP and LP diets, but SID of cystine, glutamic acid, glycine, and proline was lower (P < 0.05) in pigs fed the HP diets. The HP diets increased abundance of Bifidobacterium spp. and Lactobacillus spp., (P < 0.05) and by amplicon sequencing the latter was identified as predominant genus in microbiota from HP with Bacillus spp., whereas dietary supplementation of Bacillus spp. increased (P < 0.05) abundance of Roseburia spp..ConclusionsThe HP diet increased abundance of Lactobacillus spp. and Bifidobacterium spp.. The supplementation of Bacillus spp. resulted in a higher abundance of healthy gut associated bacteria without affecting ileal CP and AA digestibility, whereas LP diet may reduce the flow of undigested protein to the large intestine of pigs.

Valine is among the limiting AA in low protein diets for pigs, but data are currently lacking for empirical requirement studies. Therefore, 2 growth assays and 1 N-balance trial were conducted to determine the Val requirement of weaned pigs (8 to 25 kg of BW) expressed as the ratio of standardized ileal digestible (SID) Val to Lys. In Exp. 1, 48 pigs (individual housing; equal proportion of castrates and females; initial BW = 8.0 kg) were randomly subdivided into groups receiving 6 dietary treatments. Dietary SID Val:Lys ratio was increased by adding L-Val from 0.56 to 0.84% (planned increments of 0.06%) at an average SID Lys level of 0.99%. Other crystalline AA were supplied to meet optimum SID AA:Lys ratios based on the concept of an ideal protein. All diets were isocaloric (13.6 MJ of ME/kg) and isonitrogenous (17.8% CP). For the 35-d period, ADG and ADFI increased linearly (P< 0.01) and quadratically (P < 0.01) with increasing SID Val:Lys. However, the data were not suitable for estimation of Val requirement. Therefore, in Exp. 2 the dietary basal level of Val was reduced to 0.49% SID Val and 4 increments of 0.04% and 1 increment of 0.08% L-Val were used to give 6 dietary treatments (48 pigs; individual housing; equal proportion of castrates and females; initial BW = 7.9 kg). The SID Lys level averaged 0.99%, and SID Val:Lys ratios ranged from 0.49 to 74%. Other crystalline AA were supplied to meet optimum SID AA:Lys ratios. All diets were isocaloric (13.7 MJ of ME/kg) and isonitrogenous (17.7% CP). For the 34-d period, ADG, ADFI, and G:F increased linearly (P < 0.01), and ADG and G:F quadratically (P < 0.01) as the SID Val:Lys ratio increased. Estimates of optimum SID Val:Lys were 66, 67, and 61% for ADG, ADFI, and G:F, respectively. These estimates were confirmed by minimized concentrations of plasma urea and Lys and by markedly increased concentrations of plasma Val. To verify the estimates of the growth trials, a total of 24 N balances were conducted with 12 pigs using the dietary treatments of Exp. 2 (Exp. 3; equal proportion of males and females; average BW = 14.1 kg; 2 balances per pig). Pigs were restrictively fed 3 times daily. Precollection and collection periods lasted 7 d each and were repeated after rearranging the animals to treatments. Increasing the dietary SID Val:Lys ratio linearly increased N retention (P < 0.04) and quadratically decreased plasma concentrations of urea (P < 0.01). Optimal SID Val:Lys ratios of 65 and 62% were estimated for N retention and N utilization, respectively. We conclude that an optimum SID Val:Lys ratio of 65 to 67% is needed for optimal performance in 8- to 25-kg pigs.

Two growth assays and 1 N balance trial were conducted to determine the standardized ileal digestible (SID) Ile:Lys ratio in 8- to 25-kg pigs using spray-dried blood cells or corn gluten feed as a protein source. In Exp. 1, 48 individually penned pigs (initial BW = 7.7 kg) were used in a 6-point SID Ile titration study (analyzed SID Ile of 0.36, 0.43, 0.50, 0.57, 0.64, and 0.72%) by addition of graded levels of L-Ile. The basal diet contained 1.00% SID Lys, 18.4% CP, and 13.6 MJ of ME/kg. Diets were based on wheat, barley, corn, and 7.5% spray-dried blood cells as a protein source. Dietary SID Leu and Val levels were 1.61 and 1.02%, respectively. For the 35-d period, ADG, ADFI, and G:F increased linearly (P < 0.01) and quadratically (P < 0.04) with increasing SID Ile:Lys. Estimates of optimal SID Ile:Lys ratios were 59% for ADG and ADFI. In Exp. 2, 24 N balances were conducted using the Exp. 1 diets (12 pigs; individually penned; average BW = 11.5 kg). Pigs were fed 3 times daily with an amount equal to 1.0 MJ of ME/kg of BW(0.75). Preparation and collection periods (7 d each) were repeated after rearranging the animals to treatments. Increasing the dietary SID Ile:Lys ratio increased N retention linearly (P < 0.01), and N utilization linearly (P < 0.01) and quadratically (P < 0.01). An optimal SID Ile:Lys ratio of 54% was estimated for N retention. In Exp. 3, 48 individually penned pigs (initial BW = 8.0 kg) were fed grain-based diets in a 6-point SID Ile titration (analyzed SID Ile of 0.35, 0.41, 0.49, 0.56, 0.62, and 0.69%). Dietary SID Ile was increased by graded addition of L-Ile. The basal diet contained 0.97% SID Lys, 16.8% CP, and 13.6 MJ of ME/kg. In contrast to Exp. 1 and 2, spray-dried blood cells were excluded and corn gluten feed was used as a protein source. Dietary SID Leu and Val were set to 1.05 and 0.66%. For the 42-d period, ADG, ADFI, and G:F increased linearly (P < 0.01) and quadratically (P < 0.01) with increasing SID Ile:Lys. Estimated optimal SID Ile:Lys ratios were 54, 54, and 49 for ADG, ADFI, and G:F, respectively. These experiments suggest that the optimal SID Ile:Lys ratio depends on diet composition. In Exp. 1, AA imbalances because of increased Leu contents may have led to increased Ile nutritional needs. For ADG and ADFI, an optimum SID Ile:Lys ratio of 54% was estimated for 8- to 25-kg pigs in diets without Leu excess.