Effects of different protein sources on amino acid absorption and plasma appearance of tryptophan, large neutral amino acids, and tryptophan metabolites in pigs.

BACKGROUND: Absorption of tryptophan (TRP) across the gut epithelium is potentially modulated by competing large neutral amino acids (LNAAs), which could affect the appearance of TRP and its metabolites in the bloodstream. OBJECTIVES: This study aimed to determine, in a growing pig model of an adult human, the absorption of TRP and other LNAAs from the gastrointestinal tract, and plasma appearance of TRP, LNAAs and TRP metabolites , in response to dietary proteins varying in TRP content. METHODS: Pigs were adapted for seven days to each of four diets that differed in their protein source and TRP content: 1) alpha-lactalbumin (AL; 9.95mg TRP/g diet DM), 2) whey protein (6.59mg TRP/g), 3) casein (3.73mg TRP/g), or 4) zein (0.14mg TRP/g). On day 8, after a 12h fast, pigs received a test meal consisting of 45g protein, or a protein-free meal, and were euthanized 0 (baseline), 1, 2, 3, 4, or 6h later (n=6 pigs at each time in each meal group). Tryptophan and LNAA absorption from the small intestine, and appearance of TRP, LNAAs and TRP metabolites (melatonin, serotonin, kynurenine pathway metabolites), in the portal vein and systemic circulation, were determined. RESULTS: AL intake resulted in sustained elevated plasma TRP concentrations after an overnight fast. The amount of TRP absorbed was dose-dependently related to protein TRP content (p=0.028), with fastest rates for pigs fed AL (371mg/h). Portal and systemic plasma TRP, TRP/LNAA and the TRP metabolites were highest (p≤0.05) after AL intake , and remained above baseline levels for ∼4h postprandially. Absorption rates of TRP correlated with postprandial plasma TRP and TRP metabolites(p≤0.05). CONCLUSIONS: In adult humans, postprandial plasma TRP and TRP metabolite concentrations can likely be modulated by the TRP content of the meal.

Effects of bovine whey protein on exercise-induced gut permeability in healthy adults: a randomised controlled trial.

PURPOSE: Intestinal permeability is a critical component of gut barrier function. Barrier dysfunction can be triggered by certain stressors such as exercise, and if left unmanaged can lead to local and systemic disorders. The aim of this study was to investigate the effects of a specific whey protein fraction in alleviating exercise-induced gut permeability as assessed by recovery of lactulose/rhamnose (L/R) and lactulose/mannitol (L/M) urinary probes. METHODS: Eight males and eight females (aged 18-50) completed two arms of a double-blind, placebo-controlled, crossover study. For each arm participants performed two baseline intestinal permeability assessments, following which they consumed the treatment (2 g/day of milk powder containing 200 mg of whey protein) or placebo (2 g/day of milk powder) for 14 days, before performing a post-exercise permeability assessment. The exercise protocol involved a 20-min run at 80% of maximal oxygen uptake on a 1% incline. RESULTS: Mixed model analysis revealed an increase in L/R (23%; P < 0.001) and L/M (20%; P < 0.01) recovery following exercise. However, there was no treatment or treatment × exercise effect. CONCLUSION: The exercise protocol utilised in our study induces gut permeability. However, consuming whey protein, at the dose and timing prescribed, is not able to mitigate this effect.

A metabolomics analysis of inter-species and seasonal trends in ruminant milk: the molecular difference between bovine, caprine and ovine milk.

Ruminant milk composition can be impacted by many factors, primarily inter-species differences, but also environmental factors (e.g., season, feeding system and feed composition). Pasture-based feeding systems are known to be influenced by seasonal effects on grass composition. Spring pasture is rich in protein and low in fiber compared with late-season pasture, potentially inducing variability in the composition of some milk metabolites across the season. This study aimed to investigate inter-species and seasonal differences in the milk metabolome across the 3 major commercial ruminant milk species from factories in New Zealand: bovine, caprine and ovine milk. Bovine and caprine raw milk samples were collected monthly for a period of 9 mo (August-April, 2016-2017; bovine n = 41, caprine n = 44 samples); while ovine milk samples were collected for a period of 5 mo (August-January, n = 20 samples). Milk samples were subjected to biphasic extraction, and untargeted metabolite profiling was performed using 2 separate liquid chromatography high-resolution mass spectrometry analytical methods (polar metabolites and lipids). Major differences in milk metabolome were observed between the 3-ruminant species, with 414 of 587 (71%) polar metabolite features and 210 of 233 (87%) lipid features significantly different between species. Significant seasonal trends were observed in the polar metabolite fraction for bovine, caprine and ovine milk (17, 24 and 32 metabolites, respectively), suggesting that the polar metabolite relative intensities of ovine and caprine milk were more susceptible to changes within seasons than bovine milk. There was no significant seasonal difference for the triglycerides (TG) species measured in bovine milk, while 3 and 52 TG species changed in caprine and ovine milk, respectively, across the seasons. Four phosphatidylcholines and 2 phosphatidylethanolamines varied in caprine milk within the season, and 8 diglycerides varied in ovine milk. The inter-species and seasonal metabolite differences reported here provide a knowledge base of components potentially linked to milk physiochemical properties, and potential health benefits of New Zealand pasture-fed dairy ingredients.

Comparative lipidomics analysis of in vitro lipid digestion of sheep milk: Influence of homogenization and heat treatment.

This study investigated the changes in sheep milk lipids during in vitro gastrointestinal digestion in response to heat treatment (75°C/15 s and 95°C/5 min) and homogenization (200/50 bar) using lipidomics. Homogenized and pasteurized sheep milk had higher levels of polar lipids in gastric digesta emptied at 20 min than raw sheep milk. Intense heat treatment of homogenized sheep milk resulted in a reduced level of polar lipids compared with homogenized-pasteurized sheep milk. The release rate of free fatty acids during small intestinal digestion for gastric digesta emptied at 20 min followed the order: raw ≤ pasteurized < homogenized-pasteurized ≤ homogenized-heated sheep milk; the rate for gastric digesta emptied at 180 min showed a reverse order. No differences in the lipolysis degree were observed among differently processed sheep milks. These results indicated that processing treatments affect the lipid composition of digesta and the lipolysis rate but not the lipolysis degree during small intestinal digestion.

Dissecting the relationship between plasma and tissue metabolome in a cohort of women with obesity: Analysis of subcutaneous and visceral adipose, muscle, and liver.

Untargeted metabolomics of blood samples has become widely applied to study metabolic alterations underpinning disease and to identify biomarkers. However, understanding the relevance of a blood metabolite marker can be challenging if it is unknown whether it reflects the concentration in relevant tissues. To explore this field, metabolomic and lipidomic profiles of plasma, four sites of adipose tissues (ATs) from peripheral or central depot, two sites of muscle tissue, and liver tissue from a group of nondiabetic women with obesity who were scheduled to undergo bariatric surgery (n = 21) or other upper GI surgery (n = 5), were measured by liquid chromatography coupled with mass spectrometry. Relationships between plasma and tissue profiles were examined using Pearson correlation analysis subject to Benjamini-Hochberg correction. Plasma metabolites and lipids showed the highest number of significantly positive correlations with their corresponding concentrations in liver tissue, including lipid species of ceramide, mono- and di-hexosylceramide, sphingomyelin, phosphatidylcholine (PC), phosphatidylethanolamine (PE), lysophosphatidylethanolamine, dimethyl phosphatidylethanolamine, ether-linked PC, ether-linked PE, free fatty acid, cholesteryl ester, diacylglycerol and triacylglycerol, and polar metabolites linked to several metabolic functions and gut microbial metabolism. Plasma also showed significantly positive correlations with muscle for several phospholipid species and polar metabolites linked to metabolic functions and gut microbial metabolism, and with AT for several triacylglycerol species. In conclusion, plasma metabolomic and lipidomic profiles were reflective more of the liver profile than any of the muscle or AT sites examined in the present study. Our findings highlighted the importance of taking into consideration the metabolomic relationship of various tissues with plasma when postulating plasma metabolites marker to underlying mechanisms occurring in a specific tissue.

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.

Plasma metabolomic response to high-carbohydrate meals of differing glycaemic load in overweight women.

BACKGROUND: Metabolomic dysregulation following a meal in overweight individuals with the Metabolic Syndrome (MetS) involves multiple pathways of nutrient storage and oxidation. OBJECTIVE: The aim of the current study was to perform an acute cross-over intervention to examine the interactive actions of meal glycaemic load (GL) on the dynamic responses of the plasma metabolome in overweight females. METHODS: Postmenopausal women [63 ± 1.23y; Healthy (n = 20) and MetS (n = 20)] ingested two differing high-carbohydrate test meals (73 g carbohydrate; 51% energy) composed of either low glycemic index (LGI) or high (HGI) foods in a randomised sequence. Plasma metabolome was analysed using liquid chromatography-mass spectrometry (LC-MS). RESULTS: In the overweight women with MetS, there were suppressed postprandial responses for several amino acids (AAs), including phenylalanine, leucine, valine, and tryptophan, p < 0.05), irrespective of the meal type. Meal GL exerted a limited impact on the overall metabolomic response, although the postprandial levels of alanine were higher with the low GL meal and uric acid was greater following the high GL meal (p < 0.05). CONCLUSIONS: MetS participants exhibited reduced differences in the concentrations of a small set of AAs and a limited group of metabolites implicated in energy metabolism following the meals. However, the manipulation of meal GL had minimal impact on the postprandial metabolome. This study suggests that the GL of a meal is not a major determinant of postprandial response, with a greater impact exerted by the metabolic health of the individual. Trial registration Australia New Zealand Clinical Trials Registry: ACTRN12615001108505 (21/10/2015).

pH-dependent sedimentation and protein interactions in ultra-high-temperature-treated sheep skim milk.

Sheep milk is considered unstable to UHT processing, but the instability mechanism has not been investigated. This study assessed the effect of UHT treatment (140°C/5 s) and milk pH values from 6.6 to 7.0 on the physical properties of sheep skim milk (SSM), including heat coagulation time, particle size, sedimentation, ionic calcium level, and changes in protein composition. Significant amounts of sediment were found in UHT-treated SSM at the natural pH (∼6.6) and pH 7.0, whereas lower amounts of sediment were observed at pH values of 6.7 to 6.9. The proteins in the sediment were mainly κ-casein (CN)-depleted casein micelles with low levels of whey proteins regardless of the pH. Both the pH and the ionic calcium level of the SSM at all pH values decreased after UHT treatment. The dissociation levels of κ-, ß-, and αS2-CN increased with increasing pH of the SSM before and after heating. The protein content, ionic calcium level, and dissociation level of κ-CN were higher in the SSM than values reported previously in cow skim milk. These differences may contribute to the high amounts of sediment in the UHT-treated SSM at natural pH (∼6.6). Significantly higher levels of κ-, ß-, and αS2-CN were detected in the serum phase after heating the SSM at pH 7.0, suggesting that less κ-CN was attached to the casein micelles and that more internal structures of the casein micelles may have been exposed during heating. This could, in turn, have destabilized the casein micelles, resulting in the formation of protein aggregates and high amounts of sediment after UHT treatment of the SSM at pH 7.0.

A Diet Enriched with Lacticaseibacillus rhamnosus HN001 and Milk Fat Globule Membrane Alters the Gut Microbiota and Decreases Amygdala GABA a Receptor Expression in Stress-Sensitive Rats.

Brain signalling pathways involved in subclinical anxiety and depressed mood can be modulated via the gut brain axis (GBA), providing the potential for diet and dietary components to affect mood. We investigated behavioural, physiological and gut microbiome responses to the Lacticaseibacillus rhamnosus strain HN001 (LactoB HN001™), which has been shown to reduce postpartum anxiety and depression, and a milk fat globule membrane-enriched product, Lipid 70 (SurestartTM MFGM Lipid 70), which has been implicated in memory in stress-susceptible Wistar Kyoto rats. We examined behaviour in the open field, elevated plus maze and novel object recognition tests in conjunction with the expression of host genes in neuro-signalling pathways, and we also assessed brain lipidomics. Treatment-induced alterations in the caecal microbiome and short-chain fatty acid (SCFA) profiles were also assessed. Neither ingredient induced behavioural changes or altered the brain lipidome (separately or when combined). However, with regard to brain gene expression, the L. rhamnosus HN001 + Lipid 70 combination produced a synergistic effect, reducing GABAA subunit expression in the amygdala (Gabre, Gat3, Gabrg1) and hippocampus (Gabrd). Treatment with L. rhamnosus HN001 alone altered expression of the metabotropic glutamate receptor (Grm4) in the amygdala but produced only minor changes in gut microbiota composition. In contrast, Lipid 70 alone did not alter brain gene expression but produced a significant shift in the gut microbiota profile. Under the conditions used, there was no observed effect on rat behaviour for the ingredient combination. However, the enhancement of brain gene expression by L. rhamnosus HN001 + Lipid 70 implicates synergistic actions on region-specific neural pathways associated with fear, anxiety, depression and memory. A significant shift in the gut microbiota profile also occurred that was mainly attributable to Lipid 70.

Kinetics of heat-induced interactions among whey proteins and casein micelles in sheep skim milk and aggregation of the casein micelles.

The interactions among the proteins in sheep skim milk (SSM) during heat treatments (67.5-90°C for 0.5-30 min) were characterized by the kinetics of the denaturation of the whey proteins and of the association of the denatured whey proteins with casein micelles, and changes in the size and structure of casein micelles. The relationship between the size of the casein micelles and the association of whey proteins with the casein micelles is discussed. The level of denaturation and association with the casein micelles for ß-lactoglobulin (ß-LG) and α-lactalbumin (α-LA) increased with increasing heating temperature and time; the rates of denaturation and association with the casein micelles were markedly higher for ß-LG than for α-LA in the temperature range 80 to 90°C; the Arrhenius critical temperature was 80°C for the denaturation of both ß-LG and α-LA. The casein micelle size increased by 7 to 120 nm, depending on the heating temperature and the holding time. For instance, the micelle size (about 293 nm) of SSM heated at 90°C for 30 min increased by about 70% compared with that (about 174.6 nm) of unheated SSM. The casein micelle size increased slowly by a maximum of about 65 nm until the level of association of the denatured whey proteins with casein micelles reached 95%, and then increased markedly by a maximum of about 120 nm when the association level was greater than about 95%. The marked increases in casein micelle size in heated SSM were due to aggregation of the casein micelles. Aggregation of the casein micelles and association of whey protein with the micelles occurred simultaneously in SSM during heating.

Untargeted metabolomics reveals plasma metabolites predictive of ectopic fat in pancreas and liver as assessed by magnetic resonance imaging: the TOFI_Asia study.

BACKGROUND: Excess visceral obesity and ectopic organ fat is associated with increased risk of cardiometabolic disease. However, circulating markers for early detection of ectopic fat, particularly pancreas and liver, are lacking. METHODS: Lipid storage in pancreas, liver, abdominal subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) from 68 healthy or pre-diabetic Caucasian and Chinese women enroled in the TOFI_Asia study was assessed by magnetic resonance imaging/spectroscopy (MRI/S). Plasma metabolites were measured with untargeted liquid chromatography-mass spectroscopy (LC-MS). Multivariate partial least squares (PLS) regression identified metabolites predictive of VAT/SAT and ectopic fat; univariate linear regression adjusting for potential covariates identified individual metabolites associated with VAT/SAT and ectopic fat; linear regression adjusted for ethnicity identified clinical and anthropometric correlates for each fat depot. RESULTS: PLS identified 56, 64 and 31 metabolites which jointly predicted pancreatic fat (R2Y = 0.81, Q2 = 0.69), liver fat (RY2 = 0.8, Q2 = 0.66) and VAT/SAT ((R2Y = 0.7, Q2 = 0.62)) respectively. Among the PLS-identified metabolites, none of them remained significantly associated with pancreatic fat after adjusting for all covariates. Dihydrosphingomyelin (dhSM(d36:0)), 3 phosphatidylethanolamines, 5 diacylglycerols (DG) and 40 triacylglycerols (TG) were associated with liver fat independent of covariates. Three DGs and 12 TGs were associated with VAT/SAT independent of covariates. Notably, comparison with clinical correlates showed better predictivity of ectopic fat by these PLS-identified plasma metabolite markers. CONCLUSIONS: Untargeted metabolomics identified candidate markers of visceral and ectopic fat that improved fat level prediction over clinical markers. Several plasma metabolites were associated with level of liver fat and VAT/SAT ratio independent of age, total and visceral adiposity, whereas pancreatic fat deposition was only associated with increased sulfolithocholic acid independent of adiposity-related parameters, but not age.

Plasma Biomarkers and Identification of Resilient Metabolic Disruptions in Patients With Venous Thromboembolism Using a Metabolic Systems Approach.

OBJECTIVE: Deep vein thrombosis and pulmonary embolism referred as venous thromboembolism (VTE) are a common cause of morbidity and mortality. Plasma from healthy controls or individuals who have experienced a VTE were analyzed using metabolomics to characterize biomarkers and metabolic systems of patients with VTE. Approach and Results: Polar metabolite and lipidomic profiles from plasma collected 3 months after an incident VTE were obtained using liquid chromatography mass spectrometry. Fasting-state plasma samples from 42 patients with VTE and 42 healthy controls were measured. Plasma metabolomic profiling identified 512 metabolites forming 62 biological clusters. Multivariate analysis revealed a panel of 21 metabolites altogether capable of predicting VTE status with an area under the curve of 0.92 (P=0.00174, selectivity=0.857, sensitivity=0.971). Multiblock systems analysis revealed 25 of the 62 functional biological groups as significantly affected in the VTE group (P<0.05 to control). Complementary correlation network analysis of the dysregulated functions highlighted a subset of the lipidome composed mainly of n-3 long-chain polyunsaturated fatty acids within the predominant triglycerides as a potential regulator of the post-VTE event biological response, possibly controlling oxidative and inflammatory defence systems, and metabolic disorder associated dysregulations. Of interest was microbiota metabolites including trimethylamine N-oxide that remained associated to post incident VTE patients, highlighting a possible involvement of gut microbiota on VTE risk and relapse. CONCLUSIONS: These findings show promise for the elucidation of underlying mechanisms and the design of a diagnostic test to assess the likely efficacy of clinical care in patients with VTE.

Gut Microbial Metabolites and Biochemical Pathways Involved in Irritable Bowel Syndrome: Effects of Diet and Nutrition on the Microbiome.

The food we consume and its interactions with the host and their gut microbiota affect normal gut function and health. Functional gut disorders (FGDs), including irritable bowel syndrome (IBS), can result from negative effects of these interactions, leading to a reduced quality of life. Certain foods exacerbate or reduce the severity and prevalence of FGD symptoms. IBS can be used as a model of perturbation from normal gut function with which to study the impact of foods and diets on the severity and symptoms of FGDs and understand how critical processes and biochemical mechanisms contribute to this impact. Analyzing the complex interactions between food, host, and microbial metabolites gives insights into the pathways and processes occurring in the gut which contribute to FGDs. The following review is a critical discussion of the literature regarding metabolic pathways and dietary interventions relevant to FGDs. Many metabolites, for example bile acids, SCFAs, vitamins, amino acids, and neurotransmitters, can be altered by dietary intake, and could be valuable for identifying perturbations in metabolic pathways that distinguish a "normal, healthy" gut from a "dysfunctional, unhealthy" gut. Dietary interventions for reducing symptoms of FGDs are becoming more prevalent, but studies investigating the underlying mechanisms linked to host, microbiome, and metabolite interactions are less common. Therefore, we aim to evaluate the recent literature to assist with further progression of research in this field.

Human milk and infant formula differentially alters the microbiota composition and functional gene relative abundance in the small and large intestines in weanling rats.

PURPOSE: Human breast milk is the optimal source of nutrients for growing infants. However, many circumstances can arise which preclude breast milk feeding, leading to the use of infant formula, including during the weaning period. Many diet-related effects are modulated by the gut microbiome. Therefore, we investigated the effect of human milk (HM) or infant formula (IF) on the gut microbiota in weanling rats. METHODS: The gut microbiota of weanling male Sprague-Dawley rats fed HM or IF for 28 days was analysed by shotgun metagenome sequencing. Caecal contents were analysed by liquid chromatography-mass spectrometry metabolomics. RESULTS: Numerous genera within the Proteobacteria phylum were relatively more abundant in the ileum, caecum, and colon of rats fed HM, including ileal Escherichia (HM = 9.6% ± 4.3 SEM; IF = 0.9% ± 0.3 SEM; P = 0.03). Other taxa that differed between HM- and IF-fed rats included Prevotella and Ruminococcus. Overall, more differences were observed in the ileum than the caecum and colon between rats fed HM and IF. For the rats fed IF, in the ileum, the relative abundance of Bifidobacterium was higher (HM = 1.7% ± 0.7 SEM; IF = 5.0% ± 1.5 SEM; P = 0.04) with gene functions related to carbohydrate and amino acid metabolism also decreased. In the caecum, metabolic features such as bile acids were elevated while amino sugars were also decreased. CONCLUSION: Our results show that HM and IF composition differences are reflected in the gut microbiome composition and function in both the small and large intestines.

Correction to: Effects of short- and long-term glucocorticoid-induced osteoporosis on plasma metabolome and lipidome of ovariectomized sheep.

An amendment to this paper has been published and can be accessed via the original article.

Effects of short- and long-term glucocorticoid-induced osteoporosis on plasma metabolome and lipidome of ovariectomized sheep.

BACKGROUND: Understanding the metabolic and lipidomic changes that accompany bone loss in osteoporosis might provide insights about the mechanisms behind molecular changes and facilitate developing new drugs or nutritional strategies for osteoporosis prevention. This study aimed to examine the effects of short- or long-term glucocorticoid-induced osteoporosis on plasma metabolites and lipids of ovariectomized (OVX) sheep. METHODS: Twenty-eight aged ewes were divided randomly into four groups: an OVX group, OVX in combination with glucocorticoids for two months (OVXG2), and OVX in combination with five doses of glucocorticoids (OVXG5) to induce bone loss, and a control group. Liquid chromatography-mass spectrometry untargeted metabolomic analysis was applied to monthly plasma samples to follow the progression of osteoporosis over five months. RESULTS: The metabolite profiles revealed significant differences in the plasma metabolome of OVX sheep and OVXG when compared with the control group by univariate analysis. Nine metabolites were altered, namely 5-methoxytryptophan, valine, methionine, tryptophan, glutaric acid, 2-pyrrolidone-5-carboxylic acid, indole-3-carboxaldehyde, 5-hydroxylysine and malic acid. Similarly, fifteen lipids were perturbed from multiple lipid classes such as lysophoslipids, phospholipids and ceramides. CONCLUSION: This study showed that OVX and glucocorticoid interventions altered the metabolite and lipid profiles of sheep, suggesting that amino acid and lipid metabolisms are potentially the main perturbed metabolic pathways regulating bone loss in OVX sheep.

Omics analysis reveals variations among commercial sources of bovine milk fat globule membrane.

Milk fat globule membrane (MFGM) is a glycosylated, protein-embedded, phospholipid fraction that surrounds triglycerides in milk. Commercial bovine sources have recently come to the market as a novel food ingredient and have been added to various products, including infant formula. Considering that MFGM is a heterogeneous mixture of fat, protein, and carbohydrate, it can be expected that variations among MFGM products exist. For this reason, our aim was to characterize the composition of commercial MFGM samples through a combination of proteomic and lipidomic analyses. Six bovine milk fractions, represented as MFGM fractions or phospholipid fractions, were obtained from various commercial sources. Additionally, the MFGM samples were compared with 2 infant formulas, a standard formula as well as a premium formula containing MFGM. For proteomic analysis, bottom-up data-dependent liquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed on each MFGM fraction, and nearly a thousand proteins were identified across all samples, with 364 of them having different abundance across the samples tested. One hundred twelve proteins differed by a fold-change of 10 or greater, 14 by a fold-change of 50, and 2 by a fold-change of 100 in at least 1 pair, suggesting large differences in the proteins present in these fractions. Even though the classical MFGM proteins were enriched in the MFGM fractions, the relative protein composition varied considerably, and all contain an abundance of milk (casein and whey) proteins. Lipidomic analysis identified a total of 393 lipid species across both positive and negative ionization modes, with the major classes detected being triglycerides, sphingomyelins, and several phospholipids. Across all samples, triglycerides comprised at least 50% of total lipids, with phosphatidylcholine and sphingomyelin being the second and third most abundant lipid classes, respectively. These findings demonstrate the heterogeneous nature of various bovine commercial MFGM fractions. This variation must be considered when evaluating and describing potential functional benefits of these products shown in clinical trials.

A combination of lipidomics, MS imaging, and PET scan imaging reveals differences in cerebral activity in rat pups according to the lipid quality of infant formulas.

We evaluated the effect of adding docosahexaenoic:arachidonic acids (3:2) (DHA+ARA) to 2 representative commercial infant formulas on brain activity and brain and eye lipids in an artificially reared rat pup model. The formula lipid background was either a pure plant oil blend, or dairy fat with a plant oil blend (1:1). Results at weaning were compared to breast milk-fed pups. Brain functional activity was determined by positron emission tomography scan imaging, the brain and eye fatty acid and lipid composition by targeted and untargeted lipidomics, and DHA brain regional location by mass-spectrometry imaging. The brain functional activity was normalized to controls with DHA+ARA added to the formulas. DHA in both brain and eyes was influenced by formula intake, but more than two-thirds of tissue DHA-glycerolipids remained insensitive to the dietary challenge. However, the DHA lipidome correlated better with brain function than sole DHA content ( r = 0.70 vs. r = 0.48; P < 0.05). Brain DHA regional distribution was more affected by the formula lipid background than the provision of PUFAs. Adding DHA+ARA to formulas alters the DHA content and lipidome of nervous tissue in the neonate, making it closer to dam milk-fed controls, and normalizes brain functional activity.-Aidoud, N., Delplanque, B., Baudry, C., Garcia, C., Moyon, A., Balasse, L., Guillet, B., Antona, C., Darmaun, D., Fraser, K., Ndiaye, S., Leruyet, P., Martin, J.-C. A combination of lipidomics, MS imaging, and PET scan imaging reveals differences in cerebral activity in rat pups according to the lipid quality of infant formulas.

Low pyrrolizidine alkaloid levels in perennial ryegrass is associated with the absence of a homospermidine synthase gene.

BACKGROUND: Pyrrolizidine alkaloids (PAs) are a class of secondary metabolites that function as feeding deterrents in a range of different plant species. In perennial ryegrass (Lolium perenne L.) the only PAs that have been identified are the thesinine-rhamnoside group, which displays significant genetic variation. Homospermidine synthase (HSS) has evolved from deoxyhypusine synthase (DHS) and catalyses the first step in the PA pathway, making it a key candidate for the investigation of genes influencing observed PA trait variation. RESULTS: During PCR amplification and sequence analysis of DHS we identified two putative HSS genes in perennial ryegrass. One of the genes (LpHSS1) was absent in some perennial ryegrass plants. Thesinine-rhamnoside levels were measured using liquid chromatography coupled with mass spectrometry in a diverse association mapping population, consisting of 693 plants free of fungal endophytic symbionts. Association tests that accounted for population structure identified a significant association of absence of the LpHSS1 gene with lower levels of thesinine-rhamnoside PAs. HSS-like gene sequences were identified for other grass species of the Poaceae, including tall fescue, wheat, maize and sorghum. CONCLUSION: HSS is situated at the crucial first step in the PA pathway making it an important candidate gene for investigation of involvement in PA phenotypic variation. In this study, PA level in perennial ryegrass was strongly associated with the presence or absence of the LpHSS1 gene. A genetic marker, developed for the presence/absence of LpHSS1, may be used for marker-assisted breeding to either lower or increase PAs in breeding populations of perennial or Italian ryegrass to investigate a potential role in the deterrence of herbivore pests. The presence of HSS-like genes in several other Poaceae species suggests that PA biosynthesis may occur in plant family members beyond perennial ryegrass and tall fescue and identifies a potential route for manipulating PA levels.

Gastroparesis and lipid metabolism-associated dysbiosis in Wistar-Kyoto rats.

Altered gastric accommodation and intestinal morphology suggest impaired gastrointestinal (GI) transit may occur in the Wistar-Kyoto (WKY) rat strain, as common in stress-associated functional GI disorders. Because changes in GI transit can alter microbiota composition, we investigated whether these are altered in WKY rats compared with the resilient Sprague-Dawley (SD) rats under basal conditions and characterized plasma lipid and metabolite differences. Bead transit was tracked by X-ray imaging to monitor gastric emptying (4 h), small intestine (SI) transit (9 h), and large intestine transit (12 h). Plasma extracts were analyzed by lipid and hydrophilic interaction liquid chromatography (HILIC) and liquid chromatography-mass spectrometry (LC-MS). Cecal microbial composition was determined by Illumina MiSeq 16S rRNA amplicon sequencing and analysis using the QIIME pipeline. Stomach retention of beads was 77% for WKY compared with 35% for SD rats. GI transit was decreased by 34% (9 h) and 21% (12 h) in WKY compared with SD rats. Excluding stomach retention, transiting beads moved 29% further along the SI over 4-9 h for WKY compared with SD rats. Cecal Ruminococcus, Roseburia, and unclassified Lachnospiraceae genera were less abundant in WKY rats, whereas the minor taxa Dorea, Turicibacter, and Lactobacillus were higher. Diglycerides, triglycerides, phosphatidyl-ethanolamines, and phosphatidylserine were lower in WKY rats, whereas cholesterol esters and taurocholic acids were higher. The unexpected WKY rat phenotype of delayed gastric emptying, yet rapid SI transit, was associated with altered lipid and metabolite profiles. The delayed gastric emptying of the WKY phenotype suggests this rat strain may be useful as a model for gastroparesis.NEW & NOTEWORTHY This study reveals that the stress-prone Wistar-Kyoto rat strain has a baseline physiology of gastroparesis and rapid small intestine transit, together with metabolic changes consistent with lipid metabolism-associated dysbiosis, compared with nonstress-prone rats. This suggests that the Wistar-Kyoto rat strain may be an appropriate animal model for gastroparesis.