Protein intake and outcome of critically ill patients: analysis of a large international database using piece-wise exponential additive mixed models

Background Proteins are an essential part of medical nutrition therapy in critically ill patients. Guidelines almost universally recommend a high protein intake without robust evidence supporting its use. Methods Using a large international database, we modelled associations between the hazard rate of in-hospital death and live hospital discharge (competing risks) and three categories of protein intake (low: < 0.8 g/kg per day, standard: 0.8–1.2 g/kg per day, high: > 1.2 g/kg per day) during the first 11 days after ICU admission (acute phase). Time-varying cause-specific hazard ratios (HR) were calculated from piece-wise exponential additive mixed models. We used the estimated model to compare five different hypothetical protein diets (an exclusively low protein diet, a standard protein diet administered early (day 1 to 4) or late (day 5 to 11) after ICU admission, and an early or late high protein diet). Results Of 21,100 critically ill patients in the database, 16,489 fulfilled inclusion criteria for the analysis. By day 60, 11,360 (68.9%) patients had been discharged from hospital, 4,192 patients (25.4%) had died in hospital, and 937 patients (5.7%) were still hospitalized. Median daily low protein intake was 0.49 g/kg [IQR 0.27–0.66], standard intake 0.99 g/kg [IQR 0.89– 1.09], and high intake 1.41 g/kg [IQR 1.29–1.60]. In comparison with an exclusively low protein diet, a late standard protein diet was associated with a lower hazard of in-hospital death: minimum 0.75 (95% CI 0.64, 0.87), and a higher hazard of live hospital discharge: maximum HR 1.98 (95% CI 1.72, 2.28). Results on hospital discharge, however, were qualitatively changed by a sensitivity analysis. There was no evidence that an early standard or a high protein intake during the acute phase was associated with a further improvement of outcome. Conclusions Provision of a standard protein intake during the late acute phase may improve outcome compared to an exclusively low protein diet. In unselected critically ill patients, clinical outcome may not be improved by a high protein intake during the acute phase. Study registration ID number ISRCTN17829198

Hydrogen properties and charges in a sub-1.2 Å resolution cryo-EM structure revealed by a cold field emission beam

The cold field emission (CFE) beam produces the less-attenuated contrast transfer function of electron microscopy, thereby enhancing high-resolution signals and this particularly benefits higher-resolution single particle cryogenic electron microscopy. Here, we present a sub-1.2 Å resolution structure of a standard protein sample, apoferritin. Image data were collected with the CFE beam in a high-throughput scheme while minimizing beam tilt deviations from the coma-free axis. A difference map reveals positive densities for most hydrogen atoms in the core region of the protein complex including those in water molecules, while negative densities around acidic amino-acid side chains likely represent negative charges. The position of the hydrogen densities depends on parent bonded-atom type, which is validated by an estimated level of coordinate errors.

Branched Chain Amino Acid Supplementation to a Hypocaloric Diet Does Not Affect Resting Metabolic Rate but Increases Postprandial Fat Oxidation Response in Overweight and Obese Adults after Weight Loss Intervention

Background: Branched chain amino acids (BCAA) supplementation is reported to aid in lean mass preservation, which may in turn minimize the reduction in resting metabolic rate (RMR) during weight loss. Our study aimed to examine the effect of BCAA supplementation to a hypocaloric diet on RMR and substrate utilization during a weight loss intervention. Methods: A total of 111 Chinese subjects comprising 55 males and 56 females aged 21 to 45 years old with BMI between 25 and 36 kg/m2 were randomized into three hypocaloric diet groups: (1) standard-protein (14%) with placebo (CT), (2) standard-protein with BCAA, and (3) high-protein (27%) with placebo. Indirect calorimetry was used to measure RMR, carbohydrate, and fat oxidation before and after 16 weeks of dietary intervention. Results: RMR was reduced from 1600 ± 270 kcal/day to 1500 ± 264 kcal/day (p < 0.0005) after weight loss, but no significant differences in the change of RMR, respiratory quotient, and percentage of fat and carbohydrate oxidation were observed among the three diet groups. Subjects with BCAA supplementation had an increased postprandial fat (p = 0.021) and decreased postprandial carbohydrate (p = 0.044) oxidation responses compared to the CT group after dietary intervention. Conclusions: BCAA-supplemented standard-protein diet did not significantly attenuate reduction of RMR compared to standard-protein and high-protein diets. However, the postprandial fat oxidation response increased after BCAA-supplemented weight loss intervention.

Ruminal ammonia concentration and fermentation kinetics of commercial herbal feed additives with amino acids

The objective of this study was to characterize the chemical composition of rumen fermentation while estimating it’s in vitro protein degradation (from ruminal ammonia concentration) and kinetics regarding two herbal feed plant additives. The tested herbal mixtures were elaborated with Phaseolus mango and Linum usitatissimum, providing lysine (Lys) and Trigonella foenum-graecum and Allium sativa, providing Methionine (Met). They were compared to alfalfa (Medicago sativa) and solvent extracted soybean meal (Glicine max), as standard sources of protein using the in vitro gas production technique modified to estimate N-NH3, recording fermentation kinetics and dry matter digestibility (72 h), in a completely randomized design followed by Tukey test. Ruminal ammonia concentration in the herbal mixtures was lower (P<0.05) than in the standard protein sources, indicating that protein from herbal mixtures could resist ruminal degradation. Herbal additives with Lys or Met showed minimum N-NH3 concentration in the first 4 h of incubation. At 8 h, the concentration was 0.27 and 0.54 mg dL-1 for the herbal products with Lys and Met, significantly lower than solvent extracted soybean meal and alfalfa (1.15 and 2.24 mg dL-1 respectively, P<0.05). Highlights The tested herbal mixture elaborated with Phaseolus mango and Linum usitatissimum, provide bypass Lysine. The tested herbal mixture elaborated with Trigonella foenum-graecum and Allium sativa, provide bypass Methionine. Ruminal ammonia concentration in the herbal mixtures was lower than in the standard protein sources. Protein from herbal mixtures could resist ruminal degradation.

Increased Carvone Production in Escherichia Coli by Balancing Expression of Limonene Conversion Enzymes Through Targeted QconCAT Proteome Analysis

Background Natural mint flavor is produced by extraction from mint, which is not efficient enough to make it environment friendly process. (−)-Carvone is the monoterpenoid with key flavor of spearmint, and there has been an attempt to produce (−)-carvone by recombinant Escherichia coli. Although all enzymes in (−)-carvone biosynthesis have been functionally expressed in E. coli independently, the yield of (−)-carvone was low in the previous study.ResultsWe have found a by-product formation when cytochrome P450 limonene-6-hydroxylase (P450)/cytochrome P450 reductase (CPR) and carveol dehydrogenase (CDH) were expressed in single cell. These by-products were determined as dihydrocarveol and dihydrocarvone. We hypothesized that the enzymatic kinetics and the expression levels of P450 and CDH are quite different in E. coli. Therefore, two strains independently expressing P450/CPR and CDH were mixed with different mixing ratio, confirming increase in carvone production and decrease in by-product formation when CDH input was reduced. To determine the optimum balance of enzyme expressions, proteome analysis quantification concatemer (QconCAT) method to quantify P450, CPR, and CDH was developed. Using the QconCAT standard protein that was artificially created by concatenating the tryptic peptides, the ratio between P450 and CDH was calculated, and their optimum ratio to maximize (−)-carvone production was shown. Then, a single strain expressing both P450/CPR and CDH was constructed to imitate the superior expression ratio. The upgraded strain showed 15-fold improvement compared to the initial strain, showing 44 ± 6.3 mg/L of (−)-carvone production from 100 mg/L (−)-limonene as a starting substrate.ConclusionsImproved expression balance of P450 and CDH in recombinant E. coli increased the (−)-carvone production using (−)-limonene as the direct substrates by the whole-cell biocatalysis, showing approximately 150 times higher titer than previous report. Our study showed the usefulness of proteome analysis QconCAT method in the strain development for industrial biotechnology field.

Critical Aspects to Produce Low-Cost Protein Molecular Weight Marker: A Review

Proteomic studies usually begin with characterizing protein profile, like the molecular weight of protein, which can be done by the SDS-PAGE technique followed by Western Blot. These methods need a standard protein called molecular weight marker (MWM). In this review, the important, basic aspects of either recombinant or native MWM production was discussed, including the type and effect of dyes can be used for preparing a prestained MWM. Moreover, buffer and polyols used in the formulation can also affect the quality and stability. Also, another adjuvant may be needed to increase the robustness of MWM to lower the risk of protease contamination that can breakdown the protein inside the MWM during storage. Understanding those critical aspects will help to produce/formulate a good quality of MWM.

Effects of anti-nutritional factors on the nutritional potential of three varieties of lablab bean (Lablab purpureus ) in rats

Twenty five Wistar strain rats weighing between 68 to 80g were allotted to five dietary groups. Group I received standard protein diet which served as control, group II received a protein-free diet, which served as a negative control, while rats on III, IV and V were given rawRongai Brown (RB), rawRongaiWhite (RW) and rawHighworthBlack (HB) based diets as the main sources of protein, respectively. The rats were fed 10g experimental diets per day for seven days. a 3-day adaptation period, faecal samples were collected quantitatively for four days, bulked for each rat, weighed, oven-dried at 105 C for 24 hr, milled and stored for analyses. All the rats fed the three varieties of raw lablab beans had significant decrease (p

PSIV-6 Effect of a Phytogenic Water Additive on Growth Performance, Blood Metabolites and Gene Expression of Amino Acids Transporters in Nursery Pigs Fed with Low Protein Diets

Moderately low protein (MLP) diets can help decrease nutrient excretion from the swine production. However, MLP diets negatively impact growth performance. We hypothesized that supplementing MLP diets with phytogenics may reduce the negative effects of these diets on growth. The objective of this study was to investigate the effect of a phytogenic water additive (PWA; Herbanimal®) on growth performance, blood metabolite and gene expression of amino acids transporters in pigs fed with MLP diets. Forty-eight weaned barrows were allotted to six dietary treatments (n = 8) for 4 weeks: &gt;CON-NS: standard protein diet-no PWA; CON-LS: standard protein diet-low PWA dose (4 ml/L); CON-HS: standard protein diet-high PWA dose (8 ml/L); LP-NS: low protein diet-no PWA; LP-LS: low protein diet-low PWA dose (4 ml/L); LP-HS: low protein diet- high PWA dose (8 ml/L). Feed intake and body weight were recorded daily and weekly, respectively. At week 4, blood and tissue samples were collected and analyzed for metabolites using a chemistry analyzer and amino acid transporters using qPCR, respectively. The data were analyzed by univariate GLM (SPSS®) and the means were separated using paired Student’s t-test corrected by Benjamini-Hochberg. Pigs fed CON-HS improved the average daily gain and serum calcium and phosphorus concentrations compared to CON-NS. Pigs fed LP-LS had higher serum phosphorus and blood urea nitrogen compared to the pigs fed with LP-NS. The mRNA abundance of SLC7A11 in the jejunum was lower in CON-LS and CON-HS compared to CON-NS. Additionally, mRNA abundance of SLC6A19 in the jejunum of pigs fed with LP-LS was higher compared to LP-NS and lower in CON-HS relative to pigs fed with CON-LS. In conclusion, PWA improved the growth performance of pigs fed standard protein diets but not low protein diets. Further, the PWA improved the concentrations of blood calcium and phosphorous in pigs fed MLP diets. Funding: Agrivida and Animal Health and Production and Animal Products: Improved Nutritional Performance, Growth, and Lactation of Animals from the USDA-NIFA.