Protein-bound Amino Acids Research Articles

Nitrogen topdressing at panicle initiation modulated nitrogen allocation between storage proteins and free nitrogenous compounds in grains of japonica rice

Nitrogen (N) topdressing often leads to a decline in cooking and eating quality due to alterations in grain chemical composition. Investigation into the response of N compounds in grains, such as protein-bound amino acids (PAA), free amino acids (FAA), and other residue N compounds (Nresidue), would be beneficial for understanding the impact of N on rice quality. The present study selected five japonica rice cultivars and conducted a two-year field experiment employing two N fertilization treatments: the CK treatment, where all N fertilizer was applied as basal fertilizer, and the topdressing treatment, where 50% of the total N was applied as topdressing at the panicle initiation stage. Averaging across years and cultivars, N topdressing resulted in a 16.3% increase in PAA and an 8.65% decrease in FAA, with no significant impact on Nresidue. N topdressing resulted in a decrease in the proportion of essential amino acids found in prolamins, as well as a reduction in the proportion of amino acids belonging to the aspartate family and arginine. Significant genotypic variations were observed in the response of nitrogen components to topdressing. Furthermore, elite cultivars with premium quality exhibited greater stability in amylose content compared to protein content.

A simplified small-scale workflow for determination of complete protein-bound amino acids using pre-column derivatization HPLC method

Existing quantification methods of complete protein-bound amino acids profile of a food or feed often involve complicated hydrolysis setup and multiple hydrolysate treatment steps. Herein, we present a simplified workflow requiring only two steps: hydrolysis (in milligram scale) and direct sample dilution (in microliter scale), well adapted for automated pre-column O-phthaldialdehyde (OPA)/ 9-fluorenylmethyl chloroformate (FMOC) derivatized reverse phase high performance liquid chromatography (HPLC) method (the concentration range was established between 0.005 and 0.50 mM). This new workflow significantly reduces the analysis time (by up to 12 h for every 36 samples) while achieving over 90 % recovery for total protein-bound amino acids, including tryptophan, cysteine, and methionine. The easy hydrolysis setup eliminates the need for special hydrolysis tube or explosive reagents and substitutes laborious sample preparation steps like dry-down, reconstitution, neutralization with voluminous diluents and pH adjustment, with straightforward dilution using 0.1 N hydrochloric acid and deionized water. This method accommodates a small sample load, addressing concerns of limited sample availability. Robustness of this established protocol was validated by bovine serum albumin, and further confirmed by several plant proteins and casein standard. Good correlation of our results with an ISO accredited laboratory was verified (r = 0.992).

Microwave pretreatment effects on the aroma precursors, sensory characteristics and flavor profiles of fragrant rapeseed oil

Microwave technology offers a rapid and uniform heating method. This study investigated how microwave pretreatment affects the aroma precursors and flavor of fragrant rapeseed oils (FROs). Microwave pretreatment led to decreased levels of polyunsaturated fatty acids, sugars, protein-bound amino acids, and glucosinolates. Using gas chromatography–mass spectrometry, we identified 66 volatile compounds in the oil samples. Among these, based on odor activity values (OAV ≥ 1), we found 9 aldehydes, 1 ketone, 6 pyrazines, 1 isothiocyanate, and 7 nitriles as the key aroma-active compounds, contributing fatty-like, nutty-like, and pungent-like odors, respectively. The electronic nose results highlighted W5S and W1W as primary sensors for determining the flavor profiles of FROs. Notably, aroma-active pyrazines exhibited strong negative correlations with sucrose, cysteine, lysine, and isoleucine. This research provides essential insights for enhancing the aroma of FROs.

Increased Accumulation of Ginsenosides in Panax ginseng Sprouts Cultivated with Kelp Fermentates

Currently, new agri-tech has been developed and adapted for the cultivation of crops using smart farming technologies, e.g., plant factories and hydroponics. Kelp (Laminaria japonica), which has a high industrial value, was considered as an alternative to chemicals for its eco-friendly and sustainably wide use in crop cultivation. In this study, a fermented kelp (FK) was developed for use in hydroponics. The FK contained various free and protein-bound amino acid compositions produced by fermenting the kelp with Saccharomyces cerevisiae. Supplementing FK as an aeroponic medium when cultivating ginseng sprouts (GSs) elevated the total phenolic and flavonoid contents. Additionally, seven ginsenosides (Rg1, Re, Rb1, Rc, Rg2, Rb2, and Rd) in GSs cultivated with FK in a smart-farm system were identified and quantified by a high-performance liquid chromatography-evaporative light scattering detector/mass spectrometry analysis. Administering FK significantly increased the ginsenosides in the GSs compared to the control group, which was cultivated with tap water. These results indicate the FK administration contributed to the increased accumulation of ginsenosides in the GSs. Overall, this study suggests that FK, which contains abundant nutrients for plant growth, can be used as a novel nutrient solution to enhance the ginsenoside content in GSs during hydroponic cultivation.

Maternal stress is associated with higher protein-bound amino acid concentrations in human milk.

Maternal stress in the postpartum period affects not only the mother but also her newborn child, who is at increased risk of developing metabolic and mental disorders later in life. The mechanisms by which stress is transmitted to the infant are not yet fully understood. Human milk (HM) is a potential candidate as maternal stress affects various components of HM, e.g., fat and immunoglobulin concentrations. To date, it is unknown whether maternal stress also affects the amino acids (AAs) in HM, even though this nutrient is of extreme importance to child health and development. This study aimed to investigate whether and how maternal stress is associated with the AA composition of HM. In this observational cohort study (Amsterdam, The Netherlands), lactating women were recruited in two study groups: a high-stress (HS) group; women whose child was hospitalized (n = 24), and a control (CTL) group; women who gave birth to a healthy child (n = 73). HM was collected three times a day, on postpartum days 10, 17, and 24. Perceived psychological stress was measured using validated questionnaires, while biological stress measures were based on hair, saliva, and HM cortisol concentrations. HM protein-bound and free AAs were analyzed by liquid chromatography and compared between groups. Maternal perceived stress scores were higher in the HS group (p < 0.01). The concentrations of protein-bound AAs in HM were higher in the HS group compared to the CTL group (p = 0.028) and were positively associated with HM cortisol concentrations (p = 0.024). The concentrations of free AAs did not differ between study groups and were unrelated to cortisol concentrations. Findings from this prospective cohort study suggest that maternal stress in the postpartum period is associated with an altered human milk amino acid composition, which could play a role in the transmission of maternal stress effects to her child. The physiological implications of these stress-induced changes for infant development await future research.

Improvement of flesh quality, muscle growth and protein deposition in adult grass carp (Ctenopharyngodon idella): The role of tryptophan

The muscle is the main edible part of the animal trunk. Previous studies have shown that tryptophan is the limiting amino acid for protein synthesis. This study investigated whether dietary tryptophan could improve growth performance, flesh quality, muscle growth, and protein deposition in adult grass carp (Ctenopharyngodon idella). 450 grass carp (671.31 ± 1.66 g) were fed with six diets (iso‑nitrogenous) with different concentrations of tryptophan (0.25, 1.66, 3.12, 4.69, 5.95, and 7.61 g/kg diet) for 9 weeks. The results revealed that the optimal tryptophan levels (3.12 or 4.69 g/kg diet) increased feed intake (FI), feed efficiency (FE), percent weight gain (PWG), and specific growth rate (SGR) compared with the deficient group (0.25 g/kg diet), indicating that tryptophan improved the growth performance of adult grass carp. The optimal tryptophan levels increased the muscle contents of protein, lipid, protein-bound amino acids, free amino acids (FAA), pH24h, and shear force; improved the fatty acid (FA) profile in muscle; and reduced cooking loss and lactate content, suggesting that tryptophan improved the flesh quality of adult grass carp. The optimal tryptophan levels increased the myofiber diameter, frequency of >50 μm diameter fiber, mRNA levels of MRFs (MyoD, MyoG, Mrf4, and Myf5), and decreased the mRNA levels of MSTN in muscle, indicating that tryptophan promoted muscle growth in adult grass carp. We further found that the promotion of muscle protein deposition by dietary tryptophan may be related to protein synthesis, ubiquitin-proteasome, and autophagic lysosomal pathways. We discovered that dietary tryptophan increased the mRNA abundances of IGF-1, PI3K, AKT, TOR, and S6K1 and decreased the mRNA abundances of 4EBP1, FOXO1a, FOXO3a, MAFBX, MuRF1, UB, ULK1, Beclin1, ATG5, ATG12, p62, LC3–1, and LC3–2, thereby promoting muscle protein deposition in adult grass carp. The above results showed that dietary tryptophan enhanced growth performance, flesh quality, muscle growth, and muscle protein deposition in adult grass carp. Moreover, using broken line regression, the tryptophan requirement of adult grass carp was estimated to be 3.02 g/kg diet (11.64 g/kg protein) based on PWG.

9-Oxononanoic Acid and Its Lysine Schiff Base Adduct as a Novel Lipation Product in Peanuts.

9-Oxononanoic acid (9-ONA) was quantitated in peanuts roasted at 170 °C by GC-MS (EI). After roasting peanuts for 40 min, 9-ONA decreased from 1010 μmol/kg protein in the unheated sample to 722 μmol/kg protein, most likely due to modifications of nucleophilic side chains of protein-bound amino acids (lipation). After heating Nα-acetyl-l-lysine and 9-ONA in model experiments, a Schiff base in its reduced form, namely, Nε-carboxyoctyl-acetyl lysine, as well as two isomeric pyridinium derivatives, namely, dicarboxyhexylcarboxyheptylpyridinium-acetyl lysine 1 and 2, were tentatively identified by HPLC-ESI-MS/MS. Based on the identified lipation products of 9-ONA, it can be assumed that lipation reactions represent a mirror-image reaction. For quantitation of Nε-carboxyoctyllysine (COL) in roasted peanuts by means of HPLC-ESI-MS/MS, samples were reduced with sodium borohydride and acid hydrolyzed. For the first time, COL was quantitated after reduction in roasted peanuts. Furthermore, after prolonged roasting of peanuts for 40 min, COL decreased from 139.8 to 22.5 μmol/kg protein, which provides initial evidence for lipation of nucleophilic side chains of protein-bound amino acids by glycerol-bound oxidized fatty acids (GOFAs, e.g., 9-ONA) with the formation of neo-lipoproteins.

Intrinsic labelling of common beans with 2H2O to enable estimates of protein digestibility

ABSTRACT Background: The use of plant protein intrinsically labelled with stable isotopes provides an innovative solution to assess the efficiency of protein intake by humans. Here, the incorporation of 2H has been applied to intrinsically labelled plant protein in the common bean. This study aimed to evaluate which is the best phenological phase of seed maturation to incorporate the heavy hydrogen isotope 2H into seed amino acids. Common beans (Phaseolus vulgaris L.) were grown in pots, then, after 50 days sowing, 2H2O dissolved in irrigation water was applied, then again at an interval of either 3, 6, 9, and 12 days. Results: Applications of 2H2O at 6, 9, and 12 days after the first application, in the full-flowering stage, were the best treatments for enriching protein-bound amino acids in the bean seed with 2H. Conclusion: All treatments resulted in enrichment above 500 ppm, so the treatments (quantity and timing of 2H2O addition) were deemed successful for enriching bean seeds. This makes the intrinsically labelled seeds suitable for preparing test meals to assess the digestion and essential amino acid absorption of common bean amino acids in human subjects.

A novel dynamic proteomics approach for the measurement of broiler chicken protein fractional synthesis rate.

The study of protein synthesis in farm animals is uncommon despite its potential to increase knowledge about metabolism and discover new biomarkers of health and growth status. The present study describes a novel dynamic proteomics approach for the measurement of protein fractional synthesis rate (FSR) in broiler chickens. Chickens received a 10g/kg oral dose of 2 H2 O at day 21 of their life. Body water 2 H abundance was measured in plasma samples using a portable Fourier transform infrared spectrometer. Free and protein-bound amino acids (AAs) were isolated and had their 2 H enrichment measured by gas chromatography with mass spectrometry (GC/MS). Peptide 2 H enrichment was measured by proteomics analysis of plasma and muscle samples. Albumin, fibrinogen and muscle protein FSR were calculated from GC/MS and proteomics data. Ala appeared to be more enriched at the site of protein synthesis than in the AA free pools. Glu was found to be the AA closest to isotopic equilibrium between the different AA pools. Glu was used as an anchor to calculate n(AA) values necessary for chicken protein FSR calculation in dynamic proteomics studies. FSR values calculated using proteomics data and GC/MS data showed good agreement as evidenced by a Bland-Altman residual plot. A new dynamic proteomics approach for the measurement of broiler chicken individual protein FSR based on the administration of a single 2 H2 O oral bolus has been developed and validated. The proposed approach could facilitate new immunological and nutritional studies on free-living animals.

Free Amino Acids Can Replace Protein-Bound Amino Acids in Test Diets for Studies in Rainbow Trout (Oncorhynchus mykiss)

Effects of reducing dietary concentration of protein-bound amino acids on growth, feed intake and composition of gain in rainbow trout were studied in four experiments. Average initial body weights ranged between 29 ± 0.7 and 55 ± 0.5 g per trout. Diets contained ∼20 MJ digestible energy/kg dry matter. Each diet was fed to satiation to four replicate groups of 20 trout. Feed intake and growth rates were recorded for each group. Body composition was analyzed in representative groups at the start of each experiment and in all experimental groups at the end of each experiment. Reduction of dietary protein concentration to <380 g/kg dry matter caused significantly lower growth rates and reduced protein concentrations of gain, but these reductions in growth could be offset by the addition of 10 crystalline essential amino acids. Fish meal was completely replaced by a mixture of wheat gluten and crystalline amino acids without negative influences on growth. In the absence of fish meal, almost half the wheat gluten could be replaced by crystalline amino acids in diets containing about 32 g N × 6.25/kg dry matter without significant influences on growth. In such diets, concentrations of individual amino acids may be varied widely with no variation in other amino acids or nutrients.

The challenge to reduce crude protein contents of wheat-based broiler diets

The challenge to reduce crude protein (CP) contents of wheat-based broiler diets is both justified and formidable because the performance of broiler chickens offered reduced-CP, wheat-based diets is usually compromised. Moreover, broiler chickens offered wheat-based diets do not accommodate CP reductions as well as do those offered maize-based diets; this appears to stem from the higher protein concentrations and more rapid starch digestion rates of wheat. The higher protein concentrations of wheat than maize result in elevated inclusion levels of non-bound (synthetic, crystalline) amino acids (NBAA). This may be an impediment, because non-bound and protein-bound amino acids are not bioequivalent and intestinal uptakes of NBAA are more rapid than their protein-bound counterparts. This leads to post-enteral amino acid imbalances and the deamination of surplus amino acids, which generates ammonia (NH3). Because NH3 is inherently detrimental, it must be detoxified and eliminated as uric acid, which attracts metabolic costs. Moreover, inadequate NH3 detoxification may seriously compromise broiler growth performance. Also, consideration is given to some intrinsic wheat factors, including soluble non-starch polysaccharides, amylase–trypsin inhibitors and gluten, that may hold relevance. Several strategies are proposed that may enhance the performance of birds offered reduced-CP, wheat-based diets, including capping dietary starch:protein ratios, blending wheat with sorghum, whole-grain feeding in association with phytase, dietary inclusions of L-carnitine and the use of protected or slow-release amino acids. In future research, it should prove instructive to compare different wheats with a wide range of protein contents that, importantly, have been fully characterised for relevant parameters, to ascertain the most appropriate properties. The successful development and adoption of reduced-CP, wheat-based diets would be an enormous advantage for the Australian chicken-meat industry as it would diminish the huge dependence on imported, expensive soybean meal.

Effects of oral glutamine supplementation on jejunal morphology, development, and amino acid profiles in male low birth weight suckling piglets.

BackgroundIt has been shown that small intestine development in low birth weight (LBW) piglets is impaired. Glutamine (Gln) has been reported to improve piglet health and intestinal function in weaned piglets, but data is scarce in suckling piglets. This study was conducted to investigate the effects of oral Gln supplementation compared to Alanine (Ala) on jejunal development and function in 5 and 12 d old male LBW and normal birth weight (NBW) suckling piglets.ResultsGln had no effect on the jejunal morphology, development, tissue and digesta amino acid profiles and mRNA abundance of genes involved in amino acid transport, metabolism, glutathione synthesis in LBW piglets when compared to Ala supplementation and birth weight controls at 5 and 12 d. Only the concentration of Gln in jejunal tissue was higher in NBW piglets supplemented with Gln compared to Ala at 5 d (P < 0.05). A comparison of the birth weight groups showed no differences between LBW and NBW piglets at 5 and 12 d in any parameter. Jejunal crypt depth, villus height / width, tunica muscularis thickness, number of goblet and IgA positive cells, the ratio of jejunal RNA to DNA and the concentration of DNA, protein and RNA changed (P < 0.05) from 5 compared to 12 d. The concentrations of several free, and protein bound amino acids as well as amino metabolites differed between age groups in jejunal tissue but the digesta concentrations were affected to a lesser extent.ConclusionsOral Gln supplementation to suckling male piglets over the first 12 d of life was not associated with changes in jejunal parameters measured in this study. The absence of effects may indicate that Gln is absorbed as well as metabolized in the upper intestinal tract and thus could benefit intestinal development at a more proximal location.

The Impact of Digestive Dynamics on the Bioequivalence of Amino Acids in Broiler Chickens.

The purpose of this review is to consider the distinct possibility that dietary non-bound and protein-bound amino acids are not bioequivalent in broiler chickens. Usually, with conservative inclusions of a limited number of non-bound (synthetic, crystalline, feed-grade) amino acids in standard broiler diets, bioequivalency would not be an issue. However, reduced-crude protein (CP) broiler diets demand substantial inclusions of an extended range of non-bound amino acids to meet amino acid requirements. A standard diet may contain 5.0 g/kg non-bound amino acids, but a reduced-CP diet may contain up to 50 g/kg and this relative abundance skews the balance of non-bound to protein-bound amino acids and substantial proportions of certain amino acids are present in diets as non-bound entities. Importantly, tangible reductions in dietary CP, for example from 210 to 160 g/kg, usually both compromise broiler growth performance and increase fat deposition. Compromised growth performance is more evident in wheat- than maize-based diets but, paradoxically, fat deposition is more apparent in maize-based diets. The inability of birds to accommodate tangible dietary CP reductions appears to stem partially from the lack of bioequivalency between non-bound and protein-bound amino acids because of the differentials in intestinal uptake rates. Also, reduced-CP broiler diets generate perturbations in apparent amino acid digestibility coefficients which compound the fact that intestinal uptakes of non-bound acid acids are more rapid, and occur more anteriorly in the small intestine, than protein-bound amino acids. The likelihood is that greater proportions of non-bound amino acids transit the enterocytes of the gut mucosa without entering anabolic and/or catabolic pathways to gain entry to the portal circulation. This culminates in post-enteral amino acid imbalances and postprandial oxidation of surplus amino acids which involves deamination of amino acids and elevations in plasma ammonia (NH3) concentrations, but NH3 is inherently toxic and demands detoxification. Excessive deamination coupled with inadequate detoxification could result in 'ammonia overload' which would be expected to compromise growth performance. Thus, the hypothesis is that non-bound and protein-bound amino acids are not bioequivalent; moreover, it may be argued that this distinction is being overlooked and is thwarting the development and acceptance of reduced-CP broiler diets.

CCK and GLP-1 release in response to proteinogenic amino acids using a small intestine ex vivo model in pigs.

The impact of individual amino acids (AA) on gut hormone secretion and appetite regulation in pigs remains largely unknown. The aim of the present study was to determine the effect of the 20 proteinogenic AA on the release of the anorexigenic hormones cholecystokinin (CCK) and glucagon-like peptide 1 (GLP-1) in postweaning pigs. Six 25-d-old male piglets (Domestic Landrace × Large White; body weight = 6.94 ± 0.29 kg) were humanely killed for the collection of intestinal segments from the duodenum, jejunum, and ileum. Tissue samples from the three intestinal segments were used to determine which of the regions were more relevant for the analysis of gut peptides. Only the segments with the highest CCK and GLP-1 secretion and expression levels were evaluated with the 20 individual AA. Tissue segments were cut open, cleaned, and stripped of their muscle layer before identical circular samples were collected and incubated in 24-well plates for 1 h (37 °C, 5% v/v CO2). The culture broth consisted of a glucose-free KRB buffer containing no added AA (control) or with the addition of 10 mM of 1 of the 20 proteinogenic AA. Following incubation, tissues and supernatant were collected for gene expression and secretion analysis of CCK and GLP-1 levels. CCK secretion and mRNA expression were higher (P < 0.05) in duodenum when compared with proximal jejunum or ileum, whereas GLP-1/proglucagon levels were higher in ileum vs. duodenum (P < 0.05) and jejunum (P < 0.05, for GLP-1 only) in postweaning pigs. Based on these results, the effect of AA on CCK and GLP-1 secretion was studied in the duodenum and ileum, respectively. None of the AA tested stimulated both anorexigenic hormones. Of all the essential AA, Ile, Leu, Met, and Trp significantly (P < 0.05) stimulated GLP-1 from the ileum, while only Phe stimulated CCK from the duodenum. Of the nonessential AA, amide AA (Gln and Asn) caused the release of CCK, while Glu and Arg increased the release of GLP-1 from the ileum. Interpreting the results in the context of the digestion and absorption dynamics, non-bound AA are quickly absorbed and have their effect on gut peptide secretion limited to the proximal small intestine (i.e., duodenum), thus, mainly CCK. In contrast, protein-bound AA would only stimulate CCK release from the duodenum through feedback mechanisms (such as through GLP-1 secreted mainly in the ileum).

Replacement of fishmeal by yellow mealworm meal on the growth performance, feed utilisation and quality of large yellow croaker

An 80-day feeding trial was conducted to evaluate the effects of yellow mealworm (Tenebrio molitor, TM) meal as substitute for dietary fishmeal on the growth performance, feed utilisation and flesh quality of large yellow croaker (initial body weight: 189.18±0.13 g). The control diet (TM0) was designed to contain 56% of fishmeal. Based on the TM0, graded levels of TM meal (15, 30, 45, 60, 75 and 100%, respectively) were used to replace fishmeal to formulate the other six experimental diets (TM15, TM30, TM45, TM60, TM75 and TM100), respectively. The results showed that the survival was not significantly affected by dietary TM meal levels (P&gt;0.05). Compared with control group, the final body weight, weight gain rate and protein efficiency ratio decreased significantly when the replacement level over 30%, while feed conversion ratio increased significantly as replacement level over 45% (P&lt;0.05). The total protein-bound amino acid content in muscle was significantly increased with the increase of dietary TM meal inclusion (P&lt;0.05). With replacement level increasing, the percentage of eicosapentaenoic acid (EPA) and Σn-3/Σn-6 poly-unsaturated fatty acids (PUFA) in muscle significantly decreased (P&lt;0.05). Meanwhile, the skin redness (a*) and yellowness (b*) values in the ventral and bottom of ventral regions showed a decreasing and increasing trend, respectively (P&lt;0.05). The TM100 group showed a higher myofibre diameter and lower myofibre density compared to the control group (P&lt;0.05). Total replacement of fishmeal with TM meal significantly down-regulated and up-regulated the expression of myf6 and mstn, respectively (P&lt;0.05). The contents of inosine-5′-monophosphate and total free amino acids were significantly decreased with the increase of TM meal inclusion (P&lt;0.05). In conclusion, TM meal can replace at least 30% of dietary fishmeal protein without negative effects on the growth, feed utilisation and flesh quality of large yellow croaker.

Assessment of polyethersulfone and polyacrylonitrile hemodialysis clinical membranes: In situ synchrotron-based imaging of human serum proteins adsorption, interaction analyses, molecular docking and clinical inflammatory biomarkers investigations

While hemodialysis augments the physiological functions of the kidney in end stage renal disease (ESRD) patients, inherent membrane properties initiate life-threating biological episodes upon contact with blood. Membrane materials have different morphological and chemical properties that may become complement, leukocyte and even coagulation activators leading to several cardiovascular diseases due to biocompatibility issues upon contact with blood. The present study offers an in-depth understanding into two commonly used clinical polyethersulfone (PES) and polyacrylonitrile (PAN) membranes in Canadian hospitals and the effects of blood-membrane interactions on their biocompatibility, in terms of the initiation of blood activation and pro-inflammatory cytokine, from both experimental and theoretical standpoints. PES and PAN were thoroughly investigated to explore their susceptibility toward interaction with three human serum proteins. The extent of protein adsorption, and subsequent interactions on the membrane surfaces were investigated after ultrafiltration with human serum albumin (ALB), fibrinogen (FB) and transferrin (TR) proteins. The buildup of protein microparticles across different stratified membrane layers was probed using Synchrotron-based X-ray microtomography conducted with a biomedical imaging and therapy (BMIT 05ID-2) beamline at the Canada's national synchrotron light source facility. Furthermore, evidence of protein adhesion has been exhaustively investigated by surface chemical analyses and high-resolution surface imaging. The difference tendencies toward protein membrane-surface adhesion for both materials contributed to biological activations when studying in vitro clinical tests with inflammatory biomarkers using Human Magnetic Luminex assays of Serpin/ Antithrombin-III, Properdin, C5a, 1L-1α, 1L-1β, TNF-α, IL6, and vWF. Experimental results have been complemented with Molecular Modeling Docking analyses in the assessment of interaction patterns of human serum proteins in a view to understanding the relationship between functional group chemistry and membrane-surface bonding. Our models reveal inherent binding between PAN nitrogen atoms and protein-bound amino acids while the PES model proposes a contribution with its sulfone chemical groups as a prerequisite to membrane-surface binding resulting in high levels of complement and inflammation factors. The differences in protein adhesion between both clinical membrane modules are attributed to varying degrees of surface hydrophilicity as a direct factor of protein/membrane biocompatibility. Both membranes interacted differently with ALB, FB and TR, however, PAN membrane exhibited less affinity for protein adsorption and membrane-surface fouling.

The Metabolic Flux Probe (MFP)-Secreted Protein as a Non-Disruptive Information Carrier for 13C-Based Metabolic Flux Analysis.

Novel cultivation technologies demand the adaptation of existing analytical concepts. Metabolic flux analysis (MFA) requires stable-isotope labeling of biomass-bound protein as the primary information source. Obtaining the required protein in cultivation set-ups where biomass is inaccessible due to low cell densities and cell immobilization is difficult to date. We developed a non-disruptive analytical concept for 13C-based metabolic flux analysis based on secreted protein as an information carrier for isotope mapping in the protein-bound amino acids. This “metabolic flux probe” (MFP) concept was investigated in different cultivation set-ups with a recombinant, protein-secreting yeast strain. The obtained results grant insight into intracellular protein turnover dynamics. Experiments under metabolic but isotopically nonstationary conditions in continuous glucose-limited chemostats at high dilution rates demonstrated faster incorporation of isotope information from labeled glucose into the recombinant reporter protein than in biomass-bound protein. Our results suggest that the reporter protein was polymerized from intracellular amino acid pools with higher turnover rates than biomass-bound protein. The latter aspect might be vital for 13C-flux analyses under isotopically nonstationary conditions for analyzing fast metabolic dynamics.

Impact of parboiling and cultivars on the free and total amino acid composition of rice ( Oryza sativa L.)

Parboiling imparts significant changes to nutritional composition of rice. The study presents the variations in amino acid (AA) composition of rice observed during parboiling process, observed in selected Sri Lankan rice cultivars. Upon parboiling, the change in levels of free and protein bound amino acids inconsistently varied across the rice cultivars and upon the parboiling technique. In comparison to non-parboiled rice, parboiling resulted in pronounced increase in nutritional gamma amino butyric acid (GABA) levels ranging from 2 times up to 11 times rendering parboiled rice a functional food. Rice parboiled after soaking for 48 hr using steam reported the highest free AA and GABA contents, while parboiling with steeping in boiling water after 24 hr soaking accounted for higher total amino acid contents in rice. Hence, for production of parboiled rice with increased nutritive value in respect of AA levels, the significance of optimization of post-processing conditions in parboiling is highlighted. Novelty impact statement Parboiling is a post harvesting technique which improves the cooking, storage, and nutritional quality of rice. In the approach to investigate the variations in amino acid composition of rice upon parboiling, the present study discusses the significant impact caused by the cultivar and the technique being used on the amino acid composition. Hence, an insight is provided into the utilization of optimum conditions during the parboiling process to produce parboiled rice enriched with high amino acid levels including gamma amino butyric acid.

Functional Amino Acids in Pigs and Chickens: Implication for Gut Health.

In pigs and broiler chickens, the gastrointestinal tract or gut is subjected to many challenges which alter performance, animal health, welfare and livability. Preventive strategies are needed to mitigate the impacts of these challenges on gut health while reducing the need to use antimicrobials. In the first part of the review, we propose a common definition of gut health for pig and chickens relying on four pillars, which correspond to the main functions of the digestive tract: (i) epithelial barrier and digestion, (ii) immune fitness, (iii) microbiota balance and (iv) oxidative stress homeostasis. For each pillar, we describe the most commonly associated indicators. In the second part of the review, we present the potential of functional amino acid supplementation to preserve and improve gut health in piglets and chickens. We highlight that amino acid supplementation strategies, based on their roles as precursors of energy and functional molecules, as signaling molecules and as microbiota modulators can positively contribute to gut health by supporting or restoring its four intertwined pillars. Additional work is still needed in order to determine the effective dose of supplementation and mode of administration that ensure the full benefits of amino acids. For this purpose, synergy between amino acids, effects of amino acid-derived metabolites and differences in the metabolic fate between free and protein-bound amino acids are research topics that need to be furtherly investigated.

Growth performance, amino acid retention and mRNA levels of mTORC1 signaling pathway genes in Nile tilapia fingerlings fed protein-bound and crystalline amino acids

Recent research has provided new mechanistic insights that amino acids (AA) coordinate protein synthesis by expressing the evolutionarily conserved target of rapamycin complex 1 (mTORC1) signaling pathway genes. An 8-week feeding trial was conducted to evaluate the impacts of experimental diets (420 g kg−1 crude protein; 15 MJ g kg−1) with protein-bound AA (Control), crystalline amino acids (CAA) supplemented at 60 g kg−1 and CAA at 120 g kg−1 on growth performance, AA retention, and mTORC1 signaling pathway (mTOR, mLST8, and Raptor) genes in Nile tilapia, Oreochromis niloticus fingerling (n = 360; 4.3 ± 0.1). At the end of feeding trial, the time-course postprandial plasmatic lysine and methionine were determined by collecting blood samples of fish immediately after feeding and at 60, 120, 240, 480 and 960 min. Each diet was randomly distributed into eight replicates groups of 12 fish and hand-fed, six times a day until apparent satiety for eight weeks. Compared with fish fed CA120 diet, body weight gain (BWG), feed intake (FI), feed efficiency (FE), protein retention efficiency (PRE) and retention of six essential and five non-essential AA parameters were improved in fish fed Control and CA60 diets. Furthermore, fish fed CA60 diet showed significantly lower whole-body lipids and higher crude protein than those fed other diets, whereas humidity and ash were not affected by dietary treatments. Plasmatic lysine and methionine linearly increased in fish fed Control diet. However, time-course plasmatic lysine and methionine were affected along with quadratic effect, and plasmatic lysine and methionine peaked at 461 and 496 min in fish fed CA60 diet, being reduced at 364 and 405 min in fish fed CA120 diet, respectively. The mechanistic insights of well-balanced AA coordinating protein synthesis in fish fed Control and CA60 diets was identified through a similar response on relative gene expression of mLST8 and mTOR genes, but higher than that in fish fed CA120 diet. Overall, this study validated CAA up to 60 g kg−1 without adverse effects on fish growth performance. However, a high CAA level at 120 g kg−1 depressed growth performance and may be considered to produce industrial-scale feeds for precision tilapia feeding. Statement of relevanceThis study revealed new mechanistic insights that amino acids regulate protein synthesis through genomic approaches by regulating relative gene expression of mTORC1 signaling pathway. The data is relevant to the formulation of well-balanced amino acids precision tilapia feeding.