The effects of branched-chain amino acid interactions on growth performance, blood metabolites, enzyme kinetics and transcriptomics in weaned pigs.

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 alpha-keto-isocaproate levels in a linear, dose-dependent manner. Levels of plasma Ile and Val, and of serum alpha-keto-beta-methylvalerate and alpha-keto-isovalerate, indicated increased catabolism. Linear increases in the activity of basal branched-chain alpha-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.

The quantification of free Amadori compounds and amino acids allows to model the bound Maillard reaction products formation in soybean products.

The quantification of protein bound Maillard reaction products (MRPs) is still a challenge in food chemistry. Protein hydrolysis is the bottleneck step: it is time consuming and the protein degradation is not always complete. In this study, the quantitation of free amino acids and Amadori products (APs) was compared to the percentage of blocked lysine by using chemometric tools. Eighty thermally treated soybean samples were analyzed by mass spectrometry to measure the concentration of free amino acids, free APs and the protein-bound markers of the Maillard reaction (furosine, Nε-(carboxymethyl)-l-lysine, Nε-(carboxyethyl)-l-lysine, total lysine). Results demonstrated that Discriminant Analysis (DA) and Correlated Component Regression (CCR) correctly estimated the percent of blocked lysine in a validation and prediction set. These findings indicate that the measure of free markers reflects the extent of protein damage in soybean samples and it suggests the possibility to obtain rapid information on the quality of the industrial processes.

Effect of Bacillus subtilis and Bacillus licheniformis supplementation in diets with low- and high-protein content on ileal crude protein and amino acid digestibility and intestinal microbiota composition of growing pigs.

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. METHODS: Eight 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. RESULTS: The 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.. CONCLUSIONS: The 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.

Quantification of Nε-(2-Furoylmethyl)-L-lysine (furosine), Nε-(Carboxymethyl)-L-lysine (CML), Nε-(Carboxyethyl)-L-lysine (CEL) and total lysine through stable isotope dilution assay and tandem mass spectrometry.

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.