The acute postprandial response of homocysteine to multivitamin and mineral supplementation with a standard meal is not impaired in older compared to younger adults.

PURPOSE: B vitamins are required for the complex regulation of homocysteine and one-carbon (1C) metabolism. Nutritional supplements are frequently used by older adults to counter nutritional inadequacies. However, the postprandial use of B vitamins from supplements in 1C metabolism may be altered with age owing to impaired nutrient absorption and metabolic regulation. Despite implications for health and nutritional status, postprandial 1C metabolite responses have not been characterised in older adults. METHODS: Healthy older (n = 20, 65-76 years) and younger (n = 20, 19-30 years) participants were recruited through online and printed advertisements in Auckland, New Zealand. Participants consumed a multivitamin and mineral supplement with a standard breakfast meal. Blood samples were collected at baseline and hourly for 4 h following ingestion. Plasma 1C metabolites (betaine, choline, cysteine, dimethylglycine, glycine, methionine, serine) were quantified using liquid chromatography coupled with mass spectrometry. Serum homocysteine, folate and vitamin B12 were quantified on a Cobas e411 autoanalyzer. RESULTS: Older adults had higher fasting homocysteine concentrations (older: 14.0 ± 2.9 µmol/L; younger: 12.2 ± 2.5 µmol/L; p = 0.036) despite higher folate (older: 36.7 ± 17.4 nmol/L; younger: 21.6 ± 7.6 nmol/L; p < 0.001) and similar vitamin B12 concentrations (p = 0.143) to younger adults. However, a similar postprandial decline in homocysteine was found in older and younger subjects in response to the combined meal and supplement. Except for a faster decline of cystathionine in older adults (p = 0.003), the postprandial response of other 1C metabolites was similar between young and older adults. CONCLUSION: Healthy older adults appear to maintain postprandial responsiveness of 1C metabolism to younger adults, supported by a similar postprandial decline in homocysteine concentrations.

The impact of heat treatment of bovine milk on gastric emptying and nutrient appearance in peripheral circulation in healthy females: a randomized controlled trial comparing pasteurized and ultra-high temperature milk.

BACKGROUND: Heat treatments of dairy, including pasteurization and ultra-high temperature (UHT) processing, alter milk macromolecular structures, and ultimately affect digestion. In vitro, animal, and human studies show faster nutrient release or circulating appearance after consuming UHT milk (UHT-M) compared with pasteurized milk (PAST-M), with a faster gastric emptying (GE) rate proposed as a possible mechanism. OBJECTIVES: To investigate the impact of milk heat treatment on GE as a mechanism of faster nutrient appearance in blood. We hypothesized that GE and circulating nutrient delivery following consumption would be faster for UHT-M than PAST-M. METHODS: In this double-blind randomized controlled cross-over trial, healthy female (n = 20; 27.3 ± 1.4 y, mean ± SD) habitual dairy consumers, consumed 500 mL of either homogenized bovine UHT-M or PAST-M (1340 compared with 1320 kJ). Gastric content volume (GCV) emptying half-time (T50) was assessed over 3 h by magnetic resonance imaging subjective digestive symptoms, plasma amino acid, lipid and B vitamin concentrations, and gastric myoelectrical activity were measured over 5 h. RESULTS: Although GCV T50 did not differ (102 ± 7 min compared with 89 ± 8 min, mean ± SEM, UHT-M and PAST-M, respectively; P = 0.051), GCV time to emptying 25% of the volume was 31% longer following UHT-M compared with PAST-M (42 ± 2 compared with 32 ± 4 min, P = 0.004). Although GCV remained larger for a longer duration following UHT-M (treatment × time interaction, P = 0.002), plasma essential amino acid AUC was greater following UHT-M than PAST-M (55,324 ± 3809 compared with 36,598 ± 5673 µmol·min·L-1, P = 0.006). Heat treatment did not impact gastric myoelectrical activity, plasma appetite hormone markers or subjective appetite scores. CONCLUSIONS: Contrary to expectations, GE was slower with UHT-M, yet, as anticipated, aminoacidemia was greater. The larger GCV following UHT-M suggests that gastric volume may poorly predict circulating nutrient appearance from complex food matrices. Dairy heat treatment may be an effective tool to modify nutrient release by impacting digestion kinetics. CLINICAL TRIAL REGISTRY: www.anzctr.org.au (ACTRN12620000172909).

Comparing Response of Sheep and Cow Milk on Acute Digestive Comfort and Lactose Malabsorption: A Randomized Controlled Trial in Female Dairy Avoiders.

Background: Sheep milk (SM) is a possible alternate dairy source for those who experience digestive symptoms with cow milk (CM). While both the milks contain lactose, one of the causes for self-reported intolerance to CM, the composition of SM and CM also differs across proteins and fats, which have been shown to impact digestive processes. Objective: To compare the acute digestive comfort and lactose malabsorption of SM to CM in female dairy avoiders. Method: In a double-blinded, randomized cross over trial, 30 dairy-avoiding females (aged 20-30 years) drank 650 mL of SM or CM (each reconstituted from spray dried powder) following an overnight fast, on two separate occasions at least 1 week apart. Blood samples were collected for glucose and insulin assessment, and single nucleotide polymorphisms of the lactase (LCT) gene (C/T13910 and G/A22018). Breath H2 and visual analog scale (VAS) digestive symptom scores were recorded at fasting and regular intervals over 4 h after ingestion. Results: Eighty percentage of study participants were lactase non-persistent (LNP; CC13910 and GG22018 genotype). Digestive symptoms, including abdominal cramps, distension, rumbling, bloating, belching, diarrhea, flatulence, vomiting, and nausea, were similar in response to SM and CM ingestion (milk × time, P > 0.05). Breath H2 was greater after CM than SM (72 ± 10 vs. 43 ± 6 ppm at 240 min, P < 0.001), which may be due to greater lactose content in CM (33 vs. 25 g). Accordingly, when corrected for the lactose content breath H2 did not differ between the two milks. The response remained similar when analyzed in the LNP subset alone (n = 20). Conclusions: Despite a higher energy and nutrient content, SM did not increase adverse digestive symptoms after ingestion, relative to CM, although there was a reduced breath H2 response, which could be attributed to the lower lactose content in SM. The tolerability of SM should be explored in populations without lactose intolerance for whom underlying trigger for intolerance is unknown.

Responsiveness of one-carbon metabolites to a high-protein diet in older men: Results from a 10-wk randomized controlled trial.

OBJECTIVES: Dietary strategies to promote successful aging are divergent. Higher-protein diets are recommended to preserve skeletal muscle mass and physical function. Conversely, increased B-vitamin intake, supporting one-carbon (1C) metabolism, reduces the risk of cognitive decline and cardiovascular disease. On the hypothesis that higher protein intake through animal-based sources will benefit 1C regulation by the supply of B vitamins (folate, riboflavin, and vitamins B6 and B12) and methyl donors (choline) despite higher methionine intake, this study explored the effect of a higher-protein diet on 1C metabolite status in older men compared to current protein recommendations. METHODS: Older men (age, 74 ± 3 y) were randomized to receive a diet for 10 wk containing either the recommended dietary allowance (RDA) of protein (0.8 g/kg body weight/d, n = 14), or double that amount (2RDA, n = 15), with differences in protein accounted for by modifying carbohydrate intake. Intervention diets were matched to each individual's energy requirements based on the Harris-Benedict equation and adjusted fortnightly as required depending on physical activity and satiety. Fasting plasma 1C metabolite concentrations were quantified by liquid chromatography coupled with mass spectrometry at baseline and after 10 wk of intervention. RESULTS: Plasma homocysteine concentrations were reduced from baseline to follow-up with both diets. Changes in metabolite ratios reflective of betaine-dependent homocysteine remethylation were specific to the RDA diet, with an increase in the betaine-to-choline ratio and a decrease in the dimethylglycine-to-betaine ratio. Comparatively, increasing folate intake was positively associated with a change in choline concentration and inversely with the betaine-to-choline ratio for the 2RDA group. CONCLUSIONS: Adding to the known benefits of higher protein intake in older people, this study supports a reduction of homocysteine with increased consumption of animal-based protein, although the health effects of differential response of choline metabolites to a higher-protein diet remain uncertain.

The Effect of Elevated Protein Intake on DNA Damage in Older People: Comparative Secondary Analysis of Two Randomized Controlled Trials.

A high protein intake at old age is important for muscle protein synthesis, however, this could also trigger protein oxidation with the potential risk for DNA damage. The aim of this study was to investigate whether an increased protein intake at recommended level or well above would affect DNA damage or change levels of reduced (GSH) and oxidised glutathione (GSSG) in community-dwelling elderly subjects. These analyses were performed in two randomized intervention studies, in Austria and in New Zealand. In both randomized control trials, the mean protein intake was increased with whole foods, in the New Zealand study (n = 29 males, 74.2 ± 3.6 years) to 1.7 g/kg body weight/d (10 weeks intervention; p < 0.001)) in the Austrian study (n = 119 males and females, 72.9 ± 4.8 years) to 1.54 g/kg body weight/d (6 weeks intervention; p < 0.001)). In both studies, single and double strand breaks and as formamidopyrimidine-DNA glycosylase-sensitive sites were investigated in peripheral blood mononuclear cells or whole blood. Further, resistance to H2O2 induced DNA damage, GSH, GSSG and CRP were measured. Increased dietary protein intake did not impact on DNA damage markers and GSH/GSSG levels. A seasonal-based time effect (p < 0.05), which led to a decrease in DNA damage and GSH was observed in the Austrian study. Therefore, increasing the protein intake to more than 20% of the total energy intake in community-dwelling seniors in Austria and New Zealand did not increase measures of DNA damage, change glutathione status or elevate plasma CRP.

Circulating Branched Chain Amino Acid Concentrations Are Higher in Dairy-Avoiding Females Following an Equal Volume of Sheep Milk Relative to Cow Milk: A Randomized Controlled Trial.

Background: Intolerances to bovine dairy are a motivating factor in consumers seeking alternate-or replacement-dairy beverages and foods. Sheep milk (SM) is an alternate dairy source, with greater protein, although similar amino acid composition compared to cow milk (CM). Studies are yet to address the appearance of circulating amino acids following consumption of SM, relative to CM, in humans. Objective: To clinically determine the appearance of branched chain amino acids, and other amino acids, in circulation in response to equal servings of SM and CM, in females who avoid dairy products. Design: In a double-blinded, randomized, cross-over trial, 30 self-described dairy avoiding females (20-40 years) drank 650 mL of SM or CM that were reconstituted from the spray dried powders (30 and 25 g in 180 mL water, respectively) on separate occasions, following an overnight fast. After reconstitution, the energy and protein provided by SM was higher than for CM (2,140 vs. 1,649 kJ; 29.9 vs. 19.4 g protein); content of branched chain amino acids (BCAAs) were 10.5 and 6.5 mg·mL-1, respectively. Blood samples were collected at fasting and at regular intervals over 5 h after milk consumption. Plasma amino acids were measured by HPLC. Results: 80% of subjects self-identified as lactose intolerant, and the majority (47%) "avoided drinking milk" "most of the time". SM resulted in greater plasma appearance of BCAAs at 60 min (641.1 ± 16.3 vs. 563.5 ± 14.4 µmol·L-1; p < 0.001) compared with CM. SM similarly resulted in elevated postprandial concentrations of the amino acids lysine, methionine, and proline, particularly at 240 min (time × milk interactions p = 0.011, 0.017, and p = 0.002, respectively). Postprandial increases in plasma alanine concentrations were sustained to 120 min after CM (time × milk interaction p = 0.001) but not after SM, despite greater quantities provided by SM. Conclusions: SM is a rich source of protein, and relative to CM, provides a greater quantity of BCAAs, with a corresponding elevation of the postprandial circulating BCAA response. SM is therefore a possible dairy alternative of benefit to those who need to increase total protein intake or for individuals with heightened protein requirements. Unique Identifier and Registry: https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=375324, identifier U1111-1209-7768.

Circulatory and Urinary B-Vitamin Responses to Multivitamin Supplement Ingestion Differ between Older and Younger Adults.

Multivitamin and mineral (MVM) supplements are frequently used amongst older populations to improve adequacy of micronutrients, including B-vitamins, but evidence for improved health outcomes are limited and deficiencies remain prevalent. Although this may indicate poor efficacy of supplements, this could also suggest the possibility for altered B-vitamin bioavailability and metabolism in older people. This open-label, single-arm acute parallel study, conducted at the Liggins Institute Clinical Research Unit in Auckland, compared circulatory and urinary B-vitamer responses to MVM supplementation in older (70.1 ± 2.7 y, n = 10 male, n = 10 female) compared to younger (24.2 ± 2.8 y, n = 10 male, n = 10 female) participants for 4 h after the ingestion of a single dose of a commercial MVM supplement and standardized breakfast. Older adults had a lower area under the curve (AUC) of postprandial plasma pyridoxine (p = 0.02) and pyridoxal-5'phosphate (p = 0.03) forms of vitamin B6 but greater 4-pyridoxic acid AUC (p = 0.009). Urinary pyridoxine and pyridoxal excretion were higher in younger females than in older females (time × age × sex interaction, p < 0.05). Older adults had a greater AUC increase in plasma thiamine (p = 0.01), riboflavin (p = 0.009), and pantothenic acid (p = 0.027). In older adults, there was decreased plasma responsiveness of the ingested (pyridoxine) and active (pyridoxal-5'phosphate) forms of vitamin B6, which indicated a previously undescribed alteration in either absorption or subsequent metabolic interconversion. While these findings cannot determine whether acute B6 responsiveness is adequate, this difference may have potential implications for B6 function in older adults. Although this may imply higher B vitamin substrate requirements for older people, further work is required to understand the implications of postprandial differences in availability.

Analysis of Human Faecal Host Proteins: Responsiveness to 10-Week Dietary Intervention Modifying Dietary Protein Intake in Elderly Males.

Faecal proteomics targeting biomarkers of immunity and inflammation have demonstrated clinical application for the identification of changes in gastrointestinal function. However, there are limited comprehensive analyses of the host faecal proteome and how it may be influenced by dietary factors. To examine this, the Homo sapiens post-diet proteome of older males was analysed at the completion of a 10-week dietary intervention, either meeting the minimum dietary protein recommendations (RDA; n = 9) or twice the recommended dietary allowance (2RDA, n = 10). The host faecal proteome differed markedly between individuals, with only a small subset of proteins present in ≥ 60% of subjects (14 and 44 proteins, RDA and 2RDA, respectively, with only 7 common to both groups). No differences were observed between the diet groups on the profiles of host faecal proteins. Faecal proteins were detected from a wide range of protein classes, with high inter-individual variation and absence of obvious impact in response to diets with markedly different protein intake. This suggests that well-matched whole food diets with two-fold variation in protein intake maintained for 10 weeks have minimal impact on human faecal host proteins.

Validity of a Portable Breath Analyser (AIRE) for the Assessment of Lactose Malabsorption.

Hydrogen (H2) measurement in exhaled breath is a reliable and non-invasive method to diagnose carbohydrate malabsorption. Currently, breath H2 measurement is typically limited to clinic-based equipment. A portable breath analyser (AIRE, FoodMarble Digestive Health Limited, Dublin, Ireland) is a personalised device marketed for the detection and self-management of food intolerances, including lactose malabsorption (LM). Currently, the validity of this device for breath H2 analysis is unknown. Individuals self-reporting dairy intolerance (six males and six females) undertook a lactose challenge and a further seven individuals (all females) underwent a milk challenge. Breath samples were collected prior to and at frequent intervals post-challenge for up to 5 h with analysis using both the AIRE and a calibrated breath hydrogen analyser (BreathTracker, QuinTron Instrument Company Inc., Milwaukee, WI, USA). A significant positive correlation (p < 0.001, r > 0.8) was demonstrated between AIRE and BreathTracker H2 values, after both lactose and milk challenges, although 26% of the AIRE readings demonstrated the maximum score of 10.0 AU. Based on our data, the cut-off value for LM diagnosis (25 ppm H2) using AIRE is 3.0 AU and it is effective for the identification of a response to lactose-containing foods in individuals experiencing LM, although its upper limit is only 81 ppm.

Comparison of the Acute Postprandial Circulating B-Vitamin and Vitamer Responses to Single Breakfast Meals in Young and Older Individuals: Preliminary Secondary Outcomes of a Randomized Controlled Trial.

B-vitamin deficiency is common in ageing populations either due to altered dietary habits or altered digestive and metabolic functions. There is limited data on the acute circulating concentrations of B-vitamins and their various forms (vitamers), following ingestion of realistic meals. This study compared the acute circulating B-vitamin and vitamer responses to either an energy-dense (ED) or a nutrient-dense (ND) breakfast meal, consumed in a randomized cross-over sequence, in older and younger adults (n = 15 and 15, aged 67.3 ± 1.5 and 22.7 ± 0.5 years (mean ± SEM), respectively). Eleven differing B-vitamins and vitamers were determined in plasma samples by ultra-high-performance liquid chromatography-tandem mass spectrometry, in the fasting and postprandial state (hourly for 5 h). While postprandial thiamine concentration increased following both meals, riboflavin increased only following a ND meal in both age groups. Many vitamins including nicotinic acid, pantothenic acid, pyridoxal, pyridoxamine, pyridoxal-5'phosphate, and 4-pyridoxic acid remained unaltered, and flavin mononucleotide (FMN), nicotinamide and nicotinuric acid concentrations reduced following both meals. Biological age and food composition had minimal impact on postprandial B-vitamin concentrations, yet the differences between the ED and ND meals for riboflavin highlight the importance of riboflavin intake to achieve adequacy.

Impact of a High Protein Intake on the Plasma Metabolome in Elderly Males: 10 Week Randomized Dietary Intervention.

High protein diets may improve the maintenance of skeletal muscle mass in the elderly, although it remains less clear what broader impact such diets have on whole body metabolic regulation in the elderly. Non-targeted polar metabolomics analysis using HILIC HPLC-MS was used to profile the circulating plasma metabolome of elderly men (n = 31; 74.7 ± 4.0 years) who were randomized to consume for 10 weeks a diet designed to achieve either protein (RDA; 0.8·g-1·kg-1) or that doubled this recommend intake (2RDA; 1.6.g.kg-1). A limited number of plasma metabolites (n = 24) were significantly differentially regulated by the diet. These included markers of protein anabolism, which increased by the 2RDA diet, including; urea, creatine, and glutarylcarnitine. Whilst in response to the RDA diet; glutamine, glutamic acid, and proline were increased, relative to the 2RDA diet (p < 0.05). Metaboanalyst identified six major metabolic pathways to be influenced by the quantity of protein intake, most notably the arginine and proline pathways. Doubling of the recommended protein intake in older males over 10 weeks exerted only a limited impact on circulating metabolites, as determined by LC-MS. This metabolomic response was almost entirely due to increased circulating abundances of metabolites potentially indicative of altered protein anabolism, without evidence of impact on pathways for metabolic health. Trial Registration: This trial was registered on 3rd March 2016 at the Australia New Zealand Clinical Trial Registry (www.anzctr.org.au) at ACTRN 12616000310460.

Protein Intake at Twice the RDA in Older Men Increases Circulatory Concentrations of the Microbiome Metabolite Trimethylamine-N-Oxide (TMAO).

Higher dietary protein intake is increasingly recommended for the elderly; however, high protein diets have also been linked to increased cardiovascular disease (CVD) risk. Trimethylamine-N-oxide (TMAO) is a bacterial metabolite derived from choline and carnitine abundant from animal protein-rich foods. TMAO may be a novel biomarker for heightened CVD risk. The purpose of this study was to assess the impact of a high protein diet on TMAO. Healthy men (74.2 ± 3.6 years, n = 29) were randomised to consume the recommended dietary allowance of protein (RDA: 0.8 g protein/kg bodyweight/day) or twice the RDA (2RDA) as part of a supplied diet for 10 weeks. Fasting blood samples were collected pre- and post-intervention for measurement of TMAO, blood lipids, glucose tolerance, insulin sensitivity, and inflammatory biomarkers. An oral glucose tolerance test was also performed. In comparison with RDA, the 2RDA diet increased circulatory TMAO (p = 0.002) but unexpectedly decreased renal excretion of TMAO (p = 0.003). LDL cholesterol was increased in 2RDA compared to RDA (p = 0.049), but no differences in other biomarkers of CVD risk and insulin sensitivity were evident between groups. In conclusion, circulatory TMAO is responsive to changes in dietary protein intake in older healthy males.

Comprehensive Profiling of the Circulatory miRNAome Response to a High Protein Diet in Elderly Men: A Potential Role in Inflammatory Response Modulation.

SCOPE: MicroRNA are critical to the coordinated post-transcriptional regulation of gene expression, yet few studies have addressed the influence of habitual diet on microRNA expression. High protein diets impact cardiometabolic health and body composition in the elderly suggesting the possibility of a complex systems response. Therefore, high-throughput small RNA sequencing technology is applied in response to doubling the protein recommended dietary allowance (RDA) over 10 weeks in older men to examine alterations in circulating miRNAome. METHODS AND RESULTS: Older men (n = 31; 74.1 ± 0.6 y) are randomized to consume either RDA (0.8 g kg-1  day-1 ) or 2RDA (1.6 g kg-1  day-1 ) of protein for 10 weeks. Downregulation of five microRNAs (miR-125b-5p, -100-5p, -99a-5p, -23b-3p, and -203a) is observed following 2RDA with no changes in the RDA. In silico functional analysis highlights target gene enrichment in inflammation-related pathways. qPCR quantification of predicted inflammatory genes (TNFα, IL-8, IL-6, pTEN, PPP1CB, and HOXA1) in peripheral blood mononuclear cells shows increased expression following 2RDA diet (p ≤ 0.05). CONCLUSION: The study findings suggest a possible selective alteration in the post-transcriptional regulation of the immune system following a high protein diet. However, very few microRNAs are altered despite a large change in the dietary protein.

The effects of dietary protein intake on appendicular lean mass and muscle function in elderly men: a 10-wk randomized controlled trial.

Background: The Recommended Daily Allowance (RDA) for protein intake in the adult population is widely promoted as 0.8 g · kg-1 · d-1 Aging may increase protein requirements, particularly to maintain muscle mass.Objective: We investigated whether controlled protein consumption at the current RDA or twice the RDA (2RDA) affects skeletal muscle mass and physical function in elderly men.Design: In this parallel-group randomized trial, 29 men aged >70 y [mean ± SD body mass index (in kg/m2): 28.3 ± 4.2] were provided with a complete diet containing either 0.8 (RDA) or 1.6 (2RDA) g protein · kg-1 · d-1, aimed to balance energy needs. Before treatment and after 10 wk of intervention, whole-body and appendicular lean mass were measured by using dual-energy X-ray absorptiometry. Knee-extension peak power was measured with dynamometry.Results: Both groups were found to have been in a moderate negative energy balance (mean ± SD RDA: 209 ± 213 kcal/d; 2RDA 145 ± 214 kcal/d; P= 0.427 for difference between the groups). In comparison with RDA, whole-body lean mass increased in 2RDA (P = 0.001; 1.49 ± 1.30 kg, P < 0.001 compared with -0.55 ± 1.49 kg, P = 0.149). This difference was mostly accounted for by an increase in trunk lean mass found in 2RDA (+1.39 ± 1.09 kg, P < 0.001). Appendicular lean mass also decreased in RDA compared with 2RDA (P = 0.022), driven by a reduction in RDA (-0.64 ± 0.91 kg, P = 0.005 compared with 0.11 ± 0.57 kg, P = 0.592). Adjusting for energy imbalances did not alter these findings. Knee-extension peak power was also differently affected (P = 0.012; 26.6 ± 47.7 W, P = 0.015 in 2RDA compared with -11.7 ± 31.0 W, P = 0.180 in RDA).Conclusions: Consumption of a diet providing 2RDA for protein compared with the current guidelines was found to have beneficial effects on lean body mass and leg power in elderly men. These effects were not explained by differences in energy balance. This trial was registered at the Australia New Zealand Clinical Trial Registry (www.anzctr.org.au) as ACTRN12616000310460.