Triglyceride-rich Lipoprotein Fractions Research Articles

Clustering analysis of lipoprotein profiles to identify subtypes of hypertriglyceridemia in Miniature Schnauzers.

Hypertriglyceridemia (HTG) is prevalent in Miniature Schnauzers, predisposing them to life-threatening diseases. Varied responses to management strategies suggest the possibility of multiple subtypes. To identify and characterize HTG subtypes in Miniature Schnauzers through cluster analysis of lipoprotein profiles. We hypothesize that multiple phenotypes of primary HTG exist in this breed. Twenty Miniature Schnauzers with normal serum triglyceride concentration (NTG), 25 with primary HTG, and 5 with secondary HTG. Cross-sectional study using archived samples. Lipoprotein profiles, generated using continuous lipoprotein density profiling, were clustered with hierarchical cluster analysis. Clinical data (age, sex, body condition score, and dietary fat content) was compared between clusters. Six clusters were identified. Dogs with primary HTG were dispersed among 4 clusters. One cluster showed the highest intensities for triglyceride-rich lipoprotein (TRL) and low-density lipoprotein (LDL) fractions and also included 4 dogs with secondary HTG. Two clusters had moderately high TRL fraction intensities and low-to-intermediate LDL intensities. The fourth cluster had high LDL but variable TRL fraction intensities with equal numbers of NTG and mild HTG dogs. The final 2 clusters comprised only NTG dogs with low TRL intensities and low-to-intermediate LDL intensities. The clusters did not appear to be driven by differences in the clinical data. The results of this study support a spectrum of lipoprotein phenotypes within Miniature Schnauzers that cannot be predicted by triglyceride concentration alone. Lipoprotein profiling might be useful to determine if subtypes have different origins, clinical consequences, and response to treatment.

Vitamin A and D Absorption in Adults with Metabolic Syndrome versus Healthy Controls: A Pilot Study Utilizing Targeted and Untargeted LC-MS Lipidomics.

Persons with metabolic syndrome (MetS) absorb less vitamin E than healthy controls. It is hypothesized that absorption of fat-soluble vitamins (FSV) A and D2 would also decrease with MetS status and that trends would be reflected in lipidomic responses between groups. Following soymilk consumption (501 IU vitamin A, 119 IU vitamin D2 ), the triglyceride-rich lipoprotein fractions (TRL) from MetS and healthy subjects (n = 10 age- and gender-matched subjects/group) are assessed using LC-MS/MS. Absorption is calculated using area under the time-concentration curves (AUC) from samples collected at 0, 3, and 6 h post-ingestion. MetS subjects have ≈6.4-fold higher median vitamin A AUC (retinyl palmitate) versus healthy controls (P = 0.07). Vitamin D2 AUC is unaffected by MetS status (P = 0.48). Untargeted LC-MS lipidomics reveals six phospholipids and one cholesterol ester with concentrations correlating (r = 0.53-0.68; P <0.001) with vitamin A concentration. The vitamin A-phospholipid association suggests increased hydrolysis by PLB, PLRP2, and/or PLA2 IB may be involved in the trend in higher vitamin A bioavailability in MetS subjects. Previously observed differences in circulating levels of these vitamins are likely not due to absorption. Alternate strategies should be investigated to improve FSV status in MetS.

37-LB: GLP-2 Regulation of Dietary Fat Absorption and Intestinal Lipoprotein Production: The Role of the Enteric Nervous System via nNOS Signaling

Objective: Postprandial dyslipidemia, a condition commonly presented in individuals with type 2 diabetes (T2D), is characterized by the overproduction of triglyceride (TG)-rich lipoproteins (TRL). This results in excess atherogenic chylomicron (CM) remnants in circulation, significantly increasing the risk of cardiovascular disease (CVD). Using a Syrian golden hamster model, our laboratory has revealed that intestinal hormone glucagon-like peptide-2 (GLP-2) regulates dietary fat uptake and increases CM production in insulin-resistant states. However, the mechanisms by which GLP-2 enhances CM production remain unclear. Previous studies have implicated nitric oxide (NO) in the actions of GLP-2, demonstrating the requirement of NO in CM secretion. Therefore, this study investigated the role of neuronal NO synthase (nNOS)-mediated NO generation in lipid uptake and CM formation. Methods: nNOS-/- and C57BL/6 (WT) mice received an intraduodenal injection of olive oil, followed by an acute intraperitoneal (i.p.) injection of GLP-2 (0.25mg/kg; n=7) or vehicle (PBS; n=8). Blood samples were collected over a 2-hour period to assess postprandial lipid accumulation, and TRL fractions were isolated via ultracentrifugation. Immunoblotting was used to measure plasma apolipoprotein B48 (apoB48) protein expression. Results: While GLP-2-treated WT mice showed characteristic rises in postprandial TG and cholesterol in whole plasma and TRL fractions, the effect of GLP-2 in nNOS-/- mice was abolished. The effect of GLP-2 on postprandial plasma apoB48 protein expression was also abolished in nNOS-/- mice relative to GLP-2-treated WT mice. Conclusions: These findings demonstrate a novel mechanism in which nNOS-generated NO is crucial for GLP-2-mediated lipid absorption and CM production in the postprandial state. Understanding this pathway will provide insight for future therapeutic interventions to treat lipid disorders in T2D. Disclosure E.M. Grande: None. K. Adeli: None. Funding Canadian Institutes of Health Research

131-OR: GLP-1R in the Nodose Ganglion Modulate Intestinal Lipoprotein Production via a Neuroendocrine Mechanism

Diabetic dyslipidemia is characterized by overproduction of ApoB48-containing chylomicron particles, which are an independent risk factor for the development of cardiovascular disease. Chylomicron production is regulated in-part by gut-derived hormones such as glucagon-like peptide (GLP)-1. Notably, we have shown that GLP-1 acts in a neuroendocrine manner; activating GLP-1 receptor (GLP-1R) containing vagal neurons in the portal vein thereby reducing postprandial lipemia. GLP-1Rs have also been identified in the vagal nodose ganglion (NG), however their relevance to lipoprotein metabolism is yet unknown. Syrian Golden hamsters received bilateral NG injections of either PBS vehicle, the GLP-1R agonist exendin-4, the GLP-1R antagonist exendin 9-39, or a combination of both. To investigate the propagation of vagal signalling from the portal bed hamsters received injections of GLP-1 or PBS vehicle into the portal vein, and concurrent NG injections of either exendin-4 or exendin 9-39. In addition, selective deafferination of GLP-1R-containing NG neurons was achieved via nodose injection of the cytotoxin saponin conjugated to exendin-4. After treatment, all animals received a gavage of olive oil, and an intravenous infusion of tyloxapol to prevent lipoprotein clearance. Postprandial accumulation of TG-rich lipoproteins was assessed via blood draws over a 6-hour period. Triglyceride-rich lipoprotein (TRL) fractions were then isolated via density ultracentrifugation for biochemical analysis. NG injection of exendin-4 significantly reduced postprandial plasma TG and TRL. The ability of portal GLP-1 to reduce postprandial lipemia was lost with 9-39 injection into the NG, suggesting that active GLP-1R in this ganglion are requisite to elicit this effect. Finally, selective deafferination of NG GLP-1R containing neurons caused significant rises in postprandial TG and TRL, underscoring the importance of vagal signalling in intestinal lipoprotein regulation. Disclosure S.S. Hoffman: None. K. Adeli: None. Funding Canadian Institutes of Health Research (967131)

Serum triglyceride and cholesterol concentrations and lipoprotein profiles in dogs with naturally occurring pancreatitis and healthy control dogs.

BackgroundPrevious studies have reported an association between hyperlipidemia and pancreatitis in dogs, but details of this association remain poorly defined.Hypothesis/ObjectivesTo compare serum triglyceride and cholesterol concentrations and lipoprotein profiles between dogs with naturally occurring pancreatitis and healthy dogs.AnimalsSeventeen dogs with a clinical diagnosis of pancreatitis (Group 1) and 53 healthy control dogs (Group 2).MethodsProspective case‐control study.ResultsIn Group 1, 3/17 dogs (18%) had hypertriglyceridemia whereas in Group 2, 4/53 dogs (7.5%) had hypertriglyceridemia (odds ratio [OR], 2.63; 95% confidence interval [CI], 0.52‐13.14; P = .35). A significant difference was found in serum triglyceride concentrations between Group 1 (median, 67 mg/dL) and Group 2 (median, 54 mg/dL; P = .002). In Group 1, 4/17 dogs (24%) had hypercholesterolemia, whereas 1/53 (1.9%) dogs in Group 2 had hypercholesterolemia (OR, 16; 95% CI, 1.64‐155.5; P = .01). No significant difference was found in serum cholesterol concentrations between Group 1 (median, 209 mg/dL) and Group 2 (median, 227 mg/dL; P = .56). Lipoprotein profiles were significantly different between Group 1 and Group 2 dogs (Eigenvalues, 0.6719; R 2 = 1.0; P = .001).Conclusions and Clinical ImportanceMost dogs with pancreatitis (>70%) had serum triglyceride and cholesterol concentrations within reference intervals. In the small percentage of dogs that had hypertriglyceridemia, hypercholesterolemia, or both, increases were mild. Important differences were identified in lipoprotein profiles between dogs with pancreatitis and healthy control dogs. Dogs with pancreatitis had higher low‐density lipoprotein fractions and lower triglyceride‐rich lipoprotein and high‐density lipoprotein fractions than healthy dogs.

Identification of molecular species of phosphatidylcholine hydroperoxides in native and copper-oxidized triglyceride-rich lipoproteins in humans.

Background Triglyceride-rich lipoproteins are considered to be independent predictors of atherosclerotic cardiovascular disease. The molecular basis of its atherogenicity is uncertain. Here, we aim to identify molecular species of phosphatidylcholine hydroperoxides (PCOOH) in triglyceride-rich lipoproteins. For comparison, copper-oxidized triglyceride-rich lipoproteins were investigated as well. Methods A fasting EDTA blood sample was collected from six healthy human volunteers to isolate two major triglyceride-rich lipoproteins fractions – very low-density lipoproteins (VLDL) and intermediate-density lipoproteins (IDL) using sequential ultracentrifugation. Triglyceride-rich lipoproteins and plasma samples were studied for PCOOH by liquid chromatography (LC) coupled with Orbitrap mass spectrometry. Results Twelve molecular species of PCOOH in triglyceride-rich lipoproteins and/or plasma were identified using the following criteria: (1) high-resolution mass spectrometry (MS) with mass accuracy within 5 ppm, (2) retention time in LC and (3) fragmentation pattern in MS2 and MS3. PC36:4-OOH was most often detected in VLDL, IDL and plasma. The ratio of total PCOOH to phosphatidylcholine progressively increased with the duration of oxidation in both VLDL and IDL. Conclusion This study demonstrated the presence of 12 molecular species of PCOOH in native triglyceride-rich lipoproteins. The frequent detection of PCOOH in triglyceride-rich lipoproteins provides a molecular basis of the atherogenicity of triglyceride-rich lipoproteins. PCOOH in triglyceride-rich lipoproteins might serve as a molecular basis of the atherogenicity of triglyceride-rich lipoproteins.

The Effect of an Iron Supplement on Lycopene Metabolism and Absorption During Digestion in Healthy Humans.

To investigate the formation and absorption of lycopene (LYC) metabolites in the human upper gastrointestinal lumen, in the absence and presence of iron. Healthy males (n = 7) consumed test meals that deliver ≈22mg LYC + ≈0.3mg apo-lycopenals from oleoresin without (-FeSO4 ) and with ferrous sulfate (160mg, +FeSO4 ). Subjects were intubated with a naso-gastric/naso-duodenal tube. Digesta, blood plasma, and the triglyceride-rich lipoprotein (TRL) fractions of plasma were analyzed using LC-MS/MS, to measure LYC and apo-lycopenoids. Digesta LYC concentrations increased with time (p = 1.2×10-7 ), decrease with time×iron (p = 1.1×10-5 ), and remain ≈200× higher than apo-lycopenals/lycopenone. Digesta apo-8'-, -10'-, -12'-, -14'-, -15-lycopenal, and apo-13-lycopenone concentrations increased with time (p <0.01), apo-12'-, -14'-, -15-lycopenal, apo-13-lycopenone increase with iron (p <0.05), and time×iron decrease apo-8'-, -10'-, -12'-, -14'-, -15-lycopenal, apo-13-lycopenone concentrations (p <0.01). A 1.9-fold decrease in LYC TRL area-under-the-time-concentration-curve is observed after the test meal +FeSO4 versus the test meal -FeSO4 (p = 0.02). Apo-lycopenals were detected in later TRL fractions, and no apo-lycopenols or apo-lycopenoic acids were observed in any samples. FeSO4 reduces LYC absorption. Apo-lycopenals appear to be absorbed from foods, and not made in significant quantities during digestion.

Synergistic Effects of DHA and Sucrose on Body Weight Gain in PUFA-Deficient Elovl2 -/- Mice.

The omega-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) is implicated in the regulation of both lipid and carbohydrate metabolism. Thus, we questioned whether dietary DHA and low or high content of sucrose impact on metabolism in mice deficient for elongation of very long-chain fatty acids 2 (ELOVL2), an enzyme involved in the endogenous DHA synthesis. We found that Elovl2 -/- mice fed a high-sucrose DHA-enriched diet followed by the high sucrose, high fat challenge significantly increased body weight. This diet affected the triglyceride rich lipoprotein fraction of plasma lipoproteins and changed the expression of several genes involved in lipid metabolism in a white adipose tissue. Our findings suggest that lipogenesis in mammals is synergistically influenced by DHA dietary and sucrose content.

Extracellular RNA Profile in Mesenteric Lymph from Exemplar Rat Models of Acute and Critical Illness.

Background: Mesenteric lymph (ML) has been implicated in the development of multiple organ dysfunction syndrome in critical illness. Extracellular RNAs play a role in cell-to-cell communication during physiological and disease processes but they are rarely studied in ML. We aimed at examining the RNA profiles of peripheral plasma, ML, and ML's extracellular vesicle (ML-EV) and triglyceride-rich lipoprotein (ML-TRL) fractions, obtained from rodent models of critical illness. Methods and Results: We collected ML for 5 hours from rodent models of critical illness [Acute Pancreatitis, Cecal Ligation and Incision (CLI), Gut Ischemia-Reperfusion (IR)] and matching Sham control rats. ML-EV and ML-TRL fractions were also isolated. RNA sequencing was performed on the RNA extracted from ML, ML-EV, ML-TRL, and plasma by using the Ion Torrent Personal Genome Machine platform. RNA sequences were searched using the Basic Local Alignment Search Tool against rat genome and RefSeq, microRNA (miRNA), genomic tRNA, functional RNA, and Genbank nucleotide databases, and the read counts were analyzed. Each sample type had a distinct RNA profile. ML contained more RNA per volume and a larger proportion of tRNA fragments than plasma. ML-EVs were enriched with miRNA, whereas ML-TRLs contained low absolute amounts of RNA. The RNA size profiles for CLI and Gut IR were different from Sham. ML carried intestinal RNAs and in a CLI model it was significantly enriched with bacterial RNA sequences. Conclusions: We found the distinct but diverse RNA profiles of ML and its compartments, and their different profiles in critical illness. Intestinal-derived small RNAs in ML may have a direct role in critical illness and utility as potential biomarkers.

Abstract P201: Separation of Postprandial Lipoproteins: Purification of Chylomicrons Using an ApoB100 Immunoaffinity Method

Introduction: Elevated fasting and postprandial triglyceride-rich lipoproteins (TRL) are associated with increased risk for atherosclerosis. In the postprandial state circulating lipids consist of dietary fat transported from the intestine, by chylomicrons (containing ApoB48) and fat transported from the liver, in very low-density lipoproteins (VLDL, containing ApoB100). Research into the roles of endogenous versus dietary fat has been hindered because current methods are inadequate to fully separate these particles. Specifically, chylomicron fractions have considerable contamination from VLDL. Hypothesis: We hypothesized that the generation of resin crosslinked to specific ApoB100 antibodies unique to VLDL would allow purification of chylomicron particles. Proof-of-principle experiments will determine: 1) individual contributions of newly synthesized fatty acids (de novo lipogenesis (DNL)), to triglycerides found in VLDL and chylomicrons; and 2) the proportion of labeled acetyl-CoA (precursor-pool value (PPV)) used to synthesize new palmitate. A difference in DNL and PPV for the purified VLDL and chylomicron fractions will confirm the successful separation of these particles. Methods: Polyclonal antibodies were produced against the C-terminus of the ApoB100 protein and crosslinked to resin generating ApoB100-specific affinity columns. To yield purified chylomicron and VLDL particles we used this technique in conjunction with stable-isotope tracer methodology in a human feeding study. Two healthy volunteers consumed a standardized diet accompanied by oral dosing of 1- 13 C-acetate every 30 minutes for 8 hours. Plasma collected hourly was used for the isolation of TRL, which was applied to the affinity resin and purified chylomicrons and VLDL collected. Triglycerides were isolated and derivatized. The enrichment of methyl-palmitate was measured by GC/MS and Mass Isotopomer Distribution Analysis was used to calculate DNL and PPV. Results: Western blots confirmed the separation of chylomicrons and VLDL by the described method. From the purified TRL fractions, the DNL-AUC (%DNL x 5 hours) values for subject 1 were chylomicron 4% and VLDL 34%, an 88% difference, and for subject 2 were chylomicron 8% and VLDL 83%, a 90% difference. The PPV calculated for each fraction in subject 1 were chylomicron 10% and VLDL 13% and in subject 2 were chylomicron 10%, VLDL 12%. On average, the PPV in the chylomicron fraction were 18% less than the VLDL fraction. Conclusions: In conclusion, results demonstrate that the immunoaffinity method effectively separated postprandial lipoproteins showing fractionated particles carry unique information regarding tissue of origin . Future applications include investigating the respective contribution of fat and sugar to atherosclerosis risk.

Production of asymmetric oxidative metabolites of [13C]-β-carotene during digestion in the gastrointestinal lumen of healthy men

Asymmetric β-apo-carotenoids (nonvitamin A-active metabolites) of provitamin A carotenoids have been observed in humans, but no study has investigated their formation during digestion. The aim of this study was to follow the formation and absorption of asymmetric β-apo-carotenoids during digestion. Healthy men were intragastrically and intraduodenally intubated, and randomly assigned to consume a lipid-rich control meal (n=3) or a lipid-rich test meal containing 20 mg [13C-10]-β-carotene (n=7). Digesta samples were collected over 5 h, and blood collected over 7 h. The triglyceride-rich lipoprotein (TRL) fractions of plasma were also isolated. Lipophilic extracts of digesta, plasma, and TRL were analyzed via a high-performance liquid chromatography-tandem mass spectrometry method developed to identify [13C]-labeled β-apo-carotenals/carotenone, [13C]-β-apo-carotenols, and [13C]-β-apo-carotenoic acids. Relative to [13C]-β-carotene, [13C]-β-apo-carotenal levels remained ∼3 orders of magnitude lower throughout digestion (no [13C]-β-apo-carotenols, or [13C]-β-apo-carotenoic acids were observed). A mixed model determined relative influence of digesta type and time on digesta metabolite level. Increasing time significantly increased the model levels of digesta [13C]-β-apo-10',12',14',15-carotenal and [13C]-β-apo-13-carotenone (P<0.05) and trended toward decreased [13C]-β-apo-8'-carotenal (P=0.0876). Gastric digesta were associated with a significantly higher level of [13C]-β-apo-8'-carotenal (P=0.0289), and lower levels of [13C]-β-apo-12',14',15-carotenal (P<0.05), relative to duodenal digesta. Anticipated retinoids, but no asymmetric [13C]-β-apo-carotenals, [13C]-β-apo-carotenols, or [13C]-β-apo-carotenoic acids, were observed in the blood or TRL samples. β-Carotene appears to be robust to digestion, with minor amounts of β-apo-carotenals/carotenone formed. Absence of asymmetric [13C]-β-apo-carotenals in plasma and TRL suggests lack of absorption, levels below the limit of detection, lack of stability, or further conversion during the digestive process to as-yet unidentified products. Lack of asymmetric [13C]-β-apo-carotenals in plasma also suggests a lack of postprandial intestinal BCO2 activity in healthy humans. This trial was registered at clinicaltrials.gov as NCT03492593.

Altered Metabolic Profile of Triglyceride-Rich Lipoproteins in Gut-Lymph of Rodent Models of Sepsis and Gut Ischemia-Reperfusion Injury.

Triglyceride-rich lipoproteins are important in dietary lipid absorption and subsequent energy distribution in the body. Their importance in the gut-lymph may have been overlooked in sepsis, the most common cause of critical illness, and in gut ischemia-reperfusion injury, a common feature of many critical illnesses. We aimed to undertake an exploratory study of triglyceride-rich lipoprotein fractions in gut-lymph using untargeted metabolic profiling to identify altered metabolites in sepsis or gut ischemia-reperfusion. The gut-lymph was collected from rodent sham, sepsis, and gut ischemia-reperfusion models. The triglyceride-rich lipoprotein-enriched fractions isolated from the gut-lymph were subjected to a dual metabolomics analysis approach: non-polar metabolite analysis by ultra-high performance liquid chromatography-mass spectrometry and polar metabolite analysis by gas chromatography-mass spectrometry. The metabolite analysis of gut-lymph triglyceride-rich lipoprotein fractions revealed a significant increase (FDR-adjusted P value < 0.05) in myo-inositol in the sepsis group and monoacylglycerols [(18:1) and (18:2)] in gut ischemia-reperfusion. There were no significantly increased specific metabolites in the lipoprotein-enriched fractions of both sepsis and gut ischemia-reperfusion. In contrast, there was a widespread decrease in multiple lipid species in sepsis (35 out of 190; adjusted P < 0.05), but not in the gut ischemia-reperfusion. Increased levels of myo-inositol and monoacylglycerols, and decreased multiple lipid species in the gut-lymph triglyceride-rich lipoprotein fraction could be candidates for new biomarkers and/or involved in the progression of sepsis and gut ischemia-reperfusion pathobiology.

Relative contribution of α-carotene to postprandial vitamin A concentrations in healthy humans after carrot consumption

Background: Asymmetric α-carotene, a provitamin A carotenoid, is cleaved to produce retinol (vitamin A) and α-retinol (with negligible vitamin A activity). The vitamin A activity of α-carotene-containing foods is likely overestimated because traditional analytic methods do not separate α-retinol derivatives from active retinol.Objective: This study aimed to accurately characterize intestinal α-carotene cleavage and its relative contribution to postprandial vitamin A in humans after consumption of raw carrots.Design: Healthy adults (n = 12) consumed a meal containing 300 g raw carrot (providing 27.3 mg β-carotene and 18.7 mg α-carotene). Triglyceride-rich lipoprotein fractions of plasma were isolated and extracted, and α-retinyl palmitate (αRP) and retinyl palmitate were measured over 12 h postprandially via high-performance liquid chromatography-tandem mass spectrometry. The complete profile of all α-retinyl esters and retinyl esters was measured at 6 h, and total absorption of α- and β-carotene was calculated.Results: αRP was identified and quantified in every subject. No difference in preference for absorption of β- over α-carotene was observed (adjusting for dose, 28% higher, P = 0.103). After absorption, β-carotene trended toward preferential cleavage compared with α-carotene (22% higher, P = 0.084). A large range of provitamin A carotenoid conversion efficiencies was observed, with α-carotene contributing 12-35% of newly converted vitamin A (predicted contribution = 25.5%). In all subjects, a majority of α-retinol was esterified to palmitic acid (as compared with other fatty acids).Conclusions: α-Retinol is esterified in the enterocyte and transported in the blood analogous to retinol. The percentage of absorption of α-carotene from raw carrots was not significantly different from β-carotene when adjusting for dose, although a trend toward higher cleavage of β-carotene was observed. The results demonstrate large interindividual variability in α-carotene conversion. The contribution of newly absorbed α-carotene to postprandial vitamin A should not be estimated but should be measured directly to accurately assess the vitamin A capacity of α-carotene-containing foods. This trial was registered at clinicaltrials.gov as NCT01432210.

Absorption and Excretion of Vitamin K Varies by Age and Triglycerides: A Metabolic Study in Older and Younger Adults Using Deuterium‐Labeled Collard Greens

Vitamin K is an essential fat‐soluble nutrient, with its most established role as a cofactor in coagulation. Emerging roles in soft tissue calcification have been proposed. However, further progress in understanding absorption and metabolism is needed, including knowledge of the factors influencing the physiologic response to vitamin K. Phylloquinone (PK) is the primary form of vitamin K in the diet and circulation. Large intra‐individual variances in circulating PK exist, and little is known about the dietary and non‐dietary factors that influence PK metabolism.The purpose of this study was to compare absorption efficiency and body retention of PK in older and younger adults following dietary vitamin K depletion and repletion.Forty‐two older (55–80 y; n=21) and younger (18–40 y; n=21) adults were maintained on sequential 28‐day vitamin K depletion (~10 ug PK/day) and vitamin K repletion (~500 ug PK/day) diets. At the end of each phase, subjects were fed deuterated collard greens (2H‐PK) and monitored over 72 hours. Liquid chromatography‐mass spectrometry (LC‐MS) was used to measure labeled and unlabeled PK in plasma and triglyceride‐rich lipoprotein (TRL) fractions, and labeled and unlabeled menadione (a PK metabolite) from 24‐hr urine collections.Absorption efficiency of 2H‐PK did not differ between PK depletion and repletion, but was significantly higher in older adults than in younger adults (p=0.02). 2H‐PK in TRL fractions was correlated with the total 2H‐PK in plasma (r=0.95, p&lt;&lt;0.001), corroborating previous data that PK is primarily transported in circulation associated with triglycerides. Age (p=0.015) and plasma triglycerides (p&lt;0.001) were significant linear predictors of 2H‐PK area under the curve (AUC). Both age and plasma triglycerides retained significance in the multivariate model, and together explained 31.6% of the variance in absorbed plasma 2H‐PK during the depletion period, and 29.9% in the repletion period. Analysis of urinary 2H‐menadione showed no differences in body retention of vitamin K by age, sex, or depletion or repletion.In conclusion, age and circulating triglycerides are determinants of vitamin K absorption efficiency and together explained about 30% of the variance observed in absorbed PK. Body retention of vitamin K did not vary by age, sex, or dietary intake. There is a need to elucidate non‐traditional factors, such as genetics, that may influence absorption and metabolism of vitamin K.Support or Funding InformationStudy supported by the USDA ARS under Cooperative Agreement 58‐1950‐7‐707 and the NIH (5R01DK69341‐4).

13C β‐Carotene stability during digestion in healthy humans

Non‐provitamin A metabolites of β‐carotene may be produced after the consumption of β‐carotene, but to date, no study has investigated their formation during digestion. Nine healthy men were recruited in total, with seven men consuming a liquid test meal containing 20 mg of 13C β‐Carotene, and two men consuming a control meal. Gastric and duodenal samples were taken postprandially over 5 hours using a nasoduodenal tube, while plasma samples, from which the triglyceride‐rich lipoprotein (TRL) fraction containing chylomicrons was isolated, were taken over 7 hours. An LC‐MS/MS method was developed to identify and quantitate 13C β‐apo‐carotenals, 13C β‐apo‐carotenols, and 13C β‐apo‐carotenoic acids in gastric and duodenal digesta, plasma, and TRL samples using mixtures of standards. Gastric and duodenal digesta 13C β‐apo‐carotenal concentrations (including13C retinal) increased over 5 hours relative to 13C β‐carotene, with the exception of 13C β‐apo‐8′‐carotenal which peaked at 2 hours. 13C β‐apo‐carotenols and 13C β‐apo‐carotenoic acids were not observed in the digesta samples. 13C retinyl esters and 13C β‐carotene were observed in the whole plasma and in the TRL fraction, and 13C retinol was observed in whole plasma. However, no non‐vitamin A 13C β‐apo‐carotenals, 13C β‐apo‐carotenols, or 13C β‐apo‐carotenoic acids were observed in the plasma or TRL samples. Taken together, our data suggests that the conditions of digestion favor β‐apo‐carotenal formation. However, it is not clear if these β‐apo‐carotenals are absorbed at levels too low for our limit of detection, or whether they are not absorbed at all. The biological effect(s) of these compounds in the digestive tract remains to be elucidated.Support or Funding InformationThis work was supported by a grant from the PACA (Provence‐Alpes‐Côte d'Azur) region. Rachel Kopec received support from the European Union, in the framework of the Marie‐Curie FP7 COFUND People Programme, through the award of an AgreenSkills fellowship (under grant agreement n° 267196), as well as the Foundation Olga Triballat.

Enhanced bioavailability of lycopene when consumed as cis-isomers from tangerine compared to red tomato juice, a randomized, cross-over clinical trial.

Tangerine tomatoes (Solanum lycopersicum) are rich in tetra-cis-lycopene resulting from natural variation in carotenoid isomerase. Our objective was to compare the bioavailability of lycopene from tangerine to red tomato juice, and elucidate physical deposition forms of these isomers in tomatoes by light and electron microscopy. Following a randomized cross-over design, subjects (n = 11, 6 M/5 F) consumed two meals delivering 10 mg lycopene from tangerine (94% cis) or red tomato juice (10% cis). Blood was sampled over 12 h and triglyceride-rich lipoprotein fractions of plasma were isolated and analyzed using HPLC-DAD-MS/MS. Lycopene was crystalline in red tomato chromoplasts and globular in tangerine tomatoes. With tangerine tomato juice we observed a marked 8.5-fold increase in lycopene bioavailability compared to red tomato juice (p < 0.001). Fractional absorption was 47.70 ± 8.81% from tangerine and 4.98 ± 1.92% from red tomato juices. Large heterogeneity was observed among subjects. Lycopene is markedly more bioavailable from tangerine than from red tomato juice, consistent with a predominance of cis-lycopene isomers and presence in chromoplasts in a lipid dissolved globular state. These results justify using tangerine tomatoes as a lycopene source in studies examining the potential health benefits of lycopene-rich foods.

Avocado Consumption Enhances Human Postprandial Provitamin A Absorption and Conversion from a Novel High–β-Carotene Tomato Sauce and from Carrots

Dietary lipids have been shown to increase bioavailability of provitamin A carotenoids from a single meal, but the effects of dietary lipids on conversion to vitamin A during absorption are essentially unknown. Based on previous animal studies, we hypothesized that the consumption of provitamin A carotenoids with dietary lipid would enhance conversion to vitamin A during absorption compared with the consumption of provitamin A carotenoids alone. Two separate sets of 12 healthy men and women were recruited for 2 randomized, 2-way crossover studies. One meal was served with fresh avocado (Persea americana Mill), cultivated variety Hass (delivering 23 g of lipid), and a second meal was served without avocado. In study 1, the source of provitamin A carotenoids was a tomato sauce made from a novel, high–β-carotene variety of tomatoes (delivering 33.7 mg of β-carotene). In study 2, the source of provitamin A carotenoids was raw carrots (delivering 27.3 mg of β-carotene and 18.7 mg of α-carotene). Postprandial blood samples were taken over 12 h, and provitamin A carotenoids and vitamin A were quantified in triglyceride-rich lipoprotein fractions to determine baseline-corrected area under the concentration-vs.-time curve. Consumption of lipid-rich avocado enhanced the absorption of β-carotene from study 1 by 2.4-fold (P < 0.0001). In study 2, the absorption of β-carotene and α-carotene increased by 6.6- and 4.8-fold, respectively (P < 0.0001 for both). Most notably, consumption of avocado enhanced the efficiency of conversion to vitamin A (as measured by retinyl esters) by 4.6-fold in study 1 (P < 0.0001) and 12.6-fold in study 2 (P = 0.0013). These observations highlight the importance of provitamin A carotenoid consumption with a lipid-rich food such as avocado for maximum absorption and conversion to vitamin A, especially in populations in which vitamin A deficiency is prevalent. This trial was registered at clinicaltrials.gov as NCT01432210.

Decrease in circulating fibroblast growth factor 21 after an oral fat load is related to postprandial triglyceride-rich lipoproteins and liver fat

Elevated levels of circulating fibroblast growth factor 21 (FGF21) are commonly encountered in type 2 diabetes, dyslipidemia, and non-alcoholic fatty liver disease, all of which share exaggerated postprandial lipemia as a common proatherogenic feature. How FGF21 responds to an oral fat load in man is unknown. We measured liver fat contents and subcutaneous and visceral fat volumes in 47 healthy subjects, who also underwent an oral fat load with measurements of plasma FGF21 and free fatty acid (FFA). Triglyceride (TG), apolipoprotein B-48 (apoB-48), and apoB-100 concentrations were measured in triglyceride-rich lipoprotein (TRL) fractions. When compared with fasting levels, the concentration of FGF21 decreased significantly at 4 h (P < 0.05) and tended to return to fasting levels at 8 h after an oral fat load. Fasting and postprandial FGF21 correlated significantly with liver fat as well as with TRLs in the chylomicron and especially in very low-density lipoprotein 1 (VLDL1) and VLDL2 fractions representing remnant particles, but not with FFA. Subjects with increased liver fat (>5%, n = 12) showed impaired suppression of FGF21 at 4 h (P < 0.05) and at 8 h (P=0.01) and demonstrated higher postprandial TG area under the curve in plasma and TRL fractions (P ≤ 0.032) compared with those with normal liver fat (≤ 5%, n = 35). We observed a significant decrease of FGF21 concentration after an oral fat load. Fasting and postprandial FGF21 levels were closely related to large VLDL and remnants, but not to plasma FFA. Our pilot findings suggest that the postprandial accumulation of TRL remnants and liver fat may modulate postprandial FGF21 levels.

Decrease in Brain Distribution of Fluvoxamine in Experimental Hyperlipidemic Rats

Many clinical reports and trials have suggested that fluvoxamine (FLV) reduces plasma lipoprotein levels. However, few studies have reported the effect of plasma lipoproteins on FLV pharmacokinetics. The aim of the present study was to investigate the affinities of FLV to plasma lipoproteins and the effect of plasma lipoproteins on the biodistribution of FLV using an experimental hyperlipidemic (HL) rat model. HL rats were prepared by intraperitoneal administration of Poloxamer-407 solution (1.0 g/kg). In vitro protein binding and distribution of FLV in plasma lipoproteins were determined in control and HL rats. In vivo pharmacokinetic study (intravenous administration of FLV, 5.0 mg/kg) and biodistribution analysis for brain and liver at a steady state (infusion, 1.5 mg/kg/hr, 6 hrs) were also performed. The plasma protein binding of FLV was around 83% and 95% in control and HL rats, respectively, whereas the FLV recoveries in triglyceride-rich lipoprotein fractions were increased in HL. Therefore, the elevation of lipoproteins was likely responsible for the increase in protein binding in HL. After intravenous administration, the area under the plasma concentration vs. time curve (AUC) in HL was 3.9-fold greater than that in control rats, whereas the distribution ratio of FLV plasma concentration to the brain at a steady state was decreased to approximately 20% of that of the control. FLV has an affinity to plasma lipoproteins, and their elevation might decrease the FLV biodistribution to brain; the plasma lipoprotein levels could not be found to correlate positively with the FLV pharmacokinetic effect in brain, but rather may attenuate it.

Postprandial lipemia in the elderly involves increased incorporation of ingested fat in plasma free fatty acids and small (Sf20–400) triglyceride-rich lipoproteins

In the elderly, the rise in postprandial plasma triglyceride (TG) concentrations is increased, contributing to their increased risk of cardiovascular disease. We sought to determine the incorporation of ingested fat (whipping cream enriched with [1,1,1-(13)C]triolein) into plasma lipids during the postprandial period in six healthy elderly (67 ± 1 yr old) and six healthy young (23 ± 2 yr old) subjects. Blood and expired air samples were taken before and at 2-h intervals during the 8-h postprandial period. As expected, the area under the curve of postprandial plasma TG concentrations was larger in the elderly compared with the young subjects (152 ± 38 vs. 66 ± 27 mg·dl(-1)·h, P < 0.05). The incorporation of [(13)C]oleate in plasma free fatty acids (FFAs) and TG of the small (S(f) = 20-400) triglyceride-rich lipoprotein (TRL) fraction was significantly higher in the elderly compared with the young subjects, resulting in increased postprandial contributions of the ingested lipid to plasma FFAs (41 ± 3 vs. 26 ± 6%, P < 0.05) and the small TRL fraction (36 ± 5 vs. 21 ± 3%, P < 0.05) in elderly. Plasma apoB-100 concentration was higher, whereas the rate of oxidation of the ingested lipid was lower (P < 0.05) in the elderly. We conclude that increased postprandial lipemia in the elderly involves increased contribution of ingested lipid to the plasma small TRLs. This appears to be driven at least in part by increased appearance of the ingested fat as plasma FFA and increased availability of apo B-100 lipoproteins in the elderly.