High Bioavailability from Ferric Pyrophosphate-Fortified Bouillon Cubes in Meals is Not Increased by Sodium Pyrophosphate: a Stable Iron Isotope Study in Young Nigerian Women.

BACKGROUND: It is challenging to find an iron compound that combines good bioavailability with minimal sensory changes when added to seasonings or condiments. Ferric pyrophosphate (FePP) is currently used to fortify bouillon cubes, but its bioavailability is generally low. Previously, the addition of a stabilizer, sodium pyrophosphate (NaPP), improved iron bioavailability from a bouillon drink. OBJECTIVE: We assessed whether there is a dose-response effect of added NaPP on iron bioavailability from local meals prepared with intrinsically labeled FePP-fortified bouillon cubes in young Nigerian women using iron stable isotope techniques. METHODS: In a double-blind, randomized, cross-over trial, women (n = 24; aged 18-40 y; mean BMI 20.5 kg/m2) consumed a Nigerian breakfast and lunch for 5 d prepared with bouillon cubes containing 2.5 mg 57Fe (as FePP) and 3 different molar ratios of NaPP: 57Fe (0:1, 3:1, and 6:1). Iron bioavailability was assessed by measuring 57Fe incorporation into erythrocytes 16 d after each 5 d NaPP: 57Fe feeding period. Data were analyzed using a linear regression model of log iron absorption on NaPP ratio, with body weight and baseline body iron stores as covariates and subject as a random intercept. RESULTS: Of the women included, 46% were anemic and 26% were iron deficient. Iron bioavailability was 10.8, 9.8, and 11.0% for the 0:1, 3:1, and 6:1 NaPP:57Fe treatments, respectively. There was no dose-response effect of an increasing NaPP:57Fe ratio (ß ± SE: 0.003 ± 0.028, P = 0.45). CONCLUSIONS: In this study, the addition of NaPP did not increase iron bioavailability from FePP-fortified bouillon cubes. However, iron bioavailability from the Nigerian meals prepared with FePP-fortified bouillon cubes was higher than expected. These results are encouraging for the potential of bouillon cubes as a fortification vehicle. Further studies are needed to assess the effect of FePP-fortified bouillon cubes on improving iron status in low-income populations. This trial was registered at clinicaltrials.gov as NCT02815449.

Intestinal phase-II metabolism of quercetin in HT29 cells, 3D human intestinal tissues and in healthy volunteers: a qualitative comparison using LC-IMS-MS and LC-HRMS.

Flavonoids are a large class of dietary molecules, among which quercetin is the most ubiquitous, which undergo an extensive intestinal phase-II metabolism. We compared the in vivo metabolism of quercetin in healthy volunteers with two in vitro models, HT29 cells and 3 D human intestinal tissues. Supernatants of the in vitro experiments and the human intestinal fluids (HIF) were analyzed by LC-IMS-MS and LC-HRMS in a qualitative way. Quercetin glucuronides, sulfates and their methyl conjugates were detected in all three systems. The metabolic profiles were found to be different, both in terms of the metabolites produced and their relative proportions. In particular, quercetin sulfates were almost absent in supernatants from HT29 cells incubations while they were a major metabolite in HIF and also found in 3 D intestinal tissues incubations. IMS provided structural information as well as a third dimension of characterization, while HRMS brought increased sensitivity and MS/MS confirmation. HT29 cells are a useful tool to generate phase-II metabolites but do not represent the in vivo situation. 3 D intestinal tissues appear as a more relevant tool to study the intestinal phase-II metabolism of flavonoids.

Identification of phase-II metabolites of flavonoids by liquid chromatography-ion-mobility spectrometry-mass spectrometry.

Flavonoids are a class of natural compounds with a broad range of potentially beneficial health properties. They are subjected to an extensive intestinal phase-II metabolism, i.e., conjugation to glucuronic acid, sulfate, and methyl groups. Flavonoids and their metabolites can interact with drug transporters and thus interfere with drug absorption, causing food-drug interactions. The site of metabolism plays a key role in the activity, but the identification of the various metabolites remains a challenge. Here, we developed an analytical method to identify the phase-II metabolites of structurally similar flavonoids. We used liquid chromatography-ion-mobility spectrometry-mass spectrometry (LC-IMS-MS) analysis to identify phase-II metabolites of flavonols, flavones, and catechins produced by HT29 cells. We showed that IMS could bring valuable structural information on the different positional isomers of the flavonols and flavones. The position of the glucuronide moiety had a strong influence on the collision cross section (CCS) of the metabolites, with only minor contribution of hydroxyl and methyl moieties. For the catechins, fragmentation data obtained from MS/MS analysis appeared more useful than IMS to determine the structure of the metabolites, mostly due to the high number of metabolites formed. Nevertheless, CCS information as a molecular fingerprint proved to be useful to identify peaks from complex mixtures. LC-IMS-MS thus appears as a valuable tool for the identification of phase-II metabolites of flavonoids. Graphical abstract Structural identification of phase-II metabolites of flavonoids using LC-IMS-MS.

Sodium pyrophosphate enhances iron bioavailability from bouillon cubes fortified with ferric pyrophosphate.

Fe fortification of centrally manufactured and frequently consumed condiments such as bouillon cubes could help prevent Fe deficiency in developing countries. However, Fe compounds that do not cause sensory changes in the fortified product, such as ferric pyrophosphate (FePP), exhibit low absorption in humans. Tetra sodium pyrophosphate (NaPP) can form soluble complexes with Fe, which could increase Fe bioavailability. Therefore, the aim of this study was to investigate Fe bioavailability from bouillon cubes fortified with either FePP only, FePP+NaPP, ferrous sulphate (FeSO4) only, or FeSO4+NaPP. We first conducted in vitro studies using a protocol of simulated digestion to assess the dialysable and ionic Fe, and the cellular ferritin response in a Caco-2 cell model. Second, Fe absorption from bouillon prepared from intrinsically labelled cubes (2·5 mg stable Fe isotopes/cube) was assessed in twenty-four Fe-deficient women, by measuring Fe incorporation into erythrocytes 2 weeks after consumption. Fe bioavailability in humans increased by 46 % (P<0·005) when comparing bouillons fortified with FePP only (4·4 %) and bouillons fortified with FePP+NaPP (6·4 %). Fe absorption from bouillons fortified with FeSO4 only and with FeSO4+NaPP was 33·8 and 27·8 %, respectively (NS). The outcome from the human study is in agreement with the dialysable Fe from the in vitro experiments. Our findings suggest that the addition of NaPP could be a promising strategy to increase Fe absorption from FePP-fortified bouillon cubes, and if confirmed by further research, for other fortified foods with complex food matrices as well.

How do metabolites differ from their parent molecules and how are they excreted?

Understanding which physicochemical properties, or property distributions, are favorable for successful design and development of drugs, nutritional supplements, cosmetics, and agrochemicals is of great importance. In this study we have analyzed molecules from three distinct chemical spaces (i) approved drugs, (ii) human metabolites, and (iii) traditional Chinese medicine (TCM) to investigate four aspects determining the disposition of small organic molecules. First, we examined the physicochemical properties of these three classes of molecules and identified characteristic features resulting from their distinctive biological functions. For example, human metabolites and TCM molecules can be larger and more hydrophobic than drugs, which makes them less likely to cross membranes. We then quantified the shifts in physicochemical property space induced by metabolism from a holistic perspective by analyzing a data set of several thousand experimentally observed metabolic trees. Results show how the metabolic system aims to retain nutrients/micronutrients while facilitating a rapid elimination of xenobiotics. In the third part we compared these global shifts with the contributions made by individual metabolic reactions. For better resolution, all reactions were classified into phase I and phase II biotransformations. Interestingly, not all metabolic reactions lead to more hydrophilic molecules. We were able to identify biotransformations leading to an increase of logP by more than one log unit, which could be used for the design of drugs with enhanced efficacy. The study closes with the analysis of the physicochemical properties of metabolites found in the bile, faeces, and urine. Metabolites in the bile can be large and are often negatively charged. Molecules with molecular weight >500 Da are rarely found in the urine, and most of these large molecules are charged phase II conjugates.

Comparable reduction in cholesterol absorption after two different ways of phytosterol administration in humans.

PURPOSE: Consumption of phytosterols is a nutritional strategy to reduce cholesterol absorption, but the efficacy of various phytosterol intake modalities remains uncertain. The main objective was to investigate the effects of phytosterol esters (PE) provided either as a spread (dispersed in fat) during a mixed meal or as a minidrink (micro-dispersed in liquid form) after a meal. METHODS: In a randomized, single-blinded crossover design, 12 healthy intubated volunteers tested three different liquid meal sequences with and without PE. The liquid meal (500 mL, Fortisip) contained an oral dose (80 mg) of deuterium-enriched cholesterol (D7C). The intubation was stopped at 240 min, and the fate of sterols was determined in the different phases of duodenal content samples as function of time. A second solid fat-containing meal without sterols was consumed at 270 min. D7C was quantified in chylomicrons and plasma for 8 h. The conditions tested were as follows: (1) no PE added (control), (2) PE in a spread added into a liquid meal (PE-spread meal) and (3) PE given 30 min after a liquid meal as 100-g yoghurt drink (PE-minidrink meal). RESULTS: Addition of PE decreased the incorporation of cholesterol into the duodenum aqueous phase including micelles. PE added as a spread or as a minidrink significantly and comparably lowered meal cholesterol occurrence in chylomicrons (-40 % for PE-spread and -54 % for PE-minidrink, p < 0.0001) compared with the control meal. CONCLUSIONS: PE either dispersed in fat during a meal or micro-dispersed in a liquid form after a meal resulted in a markedly reduced occurrence of meal-derived cholesterol in the circulation at a comparable extent.

Effect of mulberry fruit extract on glucose fluxes after a wheat porridge meal: a dual isotope study in healthy human subjects.

BACKGROUND: Previous research has shown the efficacy of mulberry extracts for lowering post-prandial glucose (PPG) responses. The postulated mechanism is slowing of glucose absorption, but effects on glucose disposal or endogenous production are also possible. This research assessed the effect of a specified mulberry fruit extract (MFE) on these three glucose flux parameters. METHODS: The study used a double-blind, randomized, controlled, full cross-over design. In 3 counter-balanced treatments, 12 healthy adult male subjects, mean (SD) age 24.9 (2.50) years and body mass index 22.5 (1.57) kg/m2, consumed porridge prepared from 13C-labelled wheat, with or without addition of 0.75 g MFE, or a solution of 13C-glucose in water. A co-administered 2H-glucose venous infusion allowed for assessment of glucose disposal. Glucose flux parameters, cumulative absorption (time to 50% absorption, T50%abs), and PPG positive incremental area under the curve from 0 to 120 min (+iAUC0-120) were determined from total and isotopically labelled glucose in plasma. As this exploratory study was not powered for formal inferential statistical tests, results are reported as the mean percent difference (or minutes for T50%abs) between treatments with 95% CI. RESULTS: MFE increased mean T50%abs by 10.2 min, (95% CI 3.9-16.5 min), and reduced mean 2 h post-meal rate of glucose appearance by 8.4% (95% CI -14.9 to -1.4%) and PPG + iAUC0-120 by 11% (95% CI -26.3 to -7.3%), with no significant changes in glucose disposal or endogenous production. CONCLUSIONS: The PPG-lowering effect of MFE is primarily mediated by a reduced rate of glucose uptake.

Absolute oral bioavailability and metabolic turnover of ß-sitosterol in healthy subjects.

The metabolic turnover, absolute oral bioavailability, clearance, and volume of distribution for ß-sitosterol were measured in healthy subjects. [(14)C]ß-Sitosterol was used as an isotopic tracer to distinguish pulse doses from dietary sources and was administered by both oral and intravenous routes. The administered doses of [(14)C]ß-sitosterol were in the region of 3 to 4 µg, sufficiently low as not to perturb the kinetics of ß-sitosterol derived from the diet. Because the plasma concentrations of [(14)C]ß-sitosterol arising from such low doses were anticipated to be very low, the ultrasensitive isotope ratio analytical method of accelerator mass spectrometry was used. The limit of quantification for [(14)C]ß-sitosterol was approximately 0.1 pg/ml, the oral absolute bioavailability was just 0.41%, clearance was 85 ml/h, volume of distribution was 46 L, and the turnover was 5.8 mg/day. Given the steady-state concentrations of ß-sitosterol (2.83 µg/ml), then the dietary load was calculated to be approximately 1400 mg/day.

Phytosterol ester processing in the small intestine: impact on cholesterol availability for absorption and chylomicron cholesterol incorporation in healthy humans.

Phytosterols (plant sterols and stanols) can lower intestinal cholesterol absorption, but the complex dynamics of the lipid digestion process in the presence of phytosterol esters (PEs) are not fully understood. We performed a clinical experiment in intubated healthy subjects to study the time course of changes in the distribution of all lipid moieties present in duodenal phases during 4 h of digestion of meals with 3.2 g PE (PE meal) or without (control meal) PE. In vitro experiments under simulated gastrointestinal conditions were also performed. The addition of PE did not alter triglyceride (TG) hydrolysis in the duodenum or subsequent chylomicron TG occurrence in the circulation. In contrast, cholesterol accumulation in the duodenum aqueous phase was markedly reduced in the presence of PE (-32%, P < 0.10). In vitro experiments confirmed that PE reduces cholesterol transfer into the aqueous phase. The addition of PE resulted in a markedly reduced presence of meal-derived hepta-deuterated cholesterol in the circulation, i.e., in chylomicrons (-43%, PE meal vs. control; P < 0.0001) and plasma (-54%, PE meal vs. control; P < 0.0001). The present data show that addition of PE to a meal does not alter TG hydrolysis but displaces cholesterol from the intestinal aqueous phase and lowers chylomicron cholesterol occurrence in humans.

Action of plant sterol intervention on sterol kinetics in hypercholesterolemic men with high versus low basal circulatory plant sterol concentrations.

BACKGROUND: The relationship between plant sterol (PS) absorption and circulatory concentrations with cholesterol absorption and biosynthesis during PS consumption has yet to be clearly elucidated in humans. It is therefore essential to examine campesterol, ß-sitosterol, and cholesterol absorption and cholesterol fractional synthesis rate (FSR) following PS consumption in individuals with high versus low basal circulatory PS concentrations. DESIGN: A randomized, crossover trial was conducted in 82 hypercholesterolemic men consuming spreads with or without 2 g/d of PS for two 4-week periods, each separated by a 4-week washout. Endpoint tracer enrichments after ingestion of (2)H-labeled campesterol or ß-sitosterol and (13)C-labeled cholesterol were determined by isotope ratio mass spectrometry. RESULTS: For both phases of dietary intervention, the endpoint cholesterol absorption index was positively correlated with campesterol (r = 0.5864, p < 0.0001) and ß-sitosterol (r = 0.4676, p < 0.0001) absorption indices; inversely, endpoint cholesterol FSR correlated negatively with the absorption indices of campesterol (r = -0.5004, p < 0.0009), ß-sitosterol (r = -0.4154, p < 0.05), and cholesterol (r = -0.4056, p < 0.0001). PS intervention reduced absorption indices of campesterol, ß-sitosterol, and cholesterol by 36.5% ± 2.7%, 39.3% ± 2.9%, and 34.3% ± 1.9%, respectively, but increased cholesterol FSR by 33.0% ± 3.3% relative to control. Endpoint circulatory PS levels (cholesterol adjusted) were positively associated with endpoint absorption indices of campesterol (r = 0.5586, p < 0.0001, for placebo; r = 0.6530, p < 0.0001, for PS intake) and cholesterol (r = 0.3683, p < 0.001 for placebo; r = 0.3469, p < 0.002, for PS intake) and were negatively associated with cholesterol FSR (r = -0.3551, p < 0.002, for placebo; r = -0.3643, p < 0.001, for PS intake). The cholesterol-lowering effect of PS was most pronounced among individuals falling within the 50th-75th percentiles of basal PS concentrations. CONCLUSION: These data suggest that basal PS concentrations indicate not only sterol absorption efficiency but also the extent of PS-induced cholesterol reduction and thus might be clinically useful to predict the extent of cholesterol response to PS intervention within a given individual.

Current in vitro testing of bioactive peptides is not valuable.

Dietary peptides have been suggested to possess biological activity in vivo and could affect cardiovascular disease parameters, based on data derived from in vitro experiments. Isolated peptides are often tested in in vitro cellular assays or on heterologously expressed molecular target proteins. The stimulatory or inhibitory effect on target proteins in vitro has often been used as the justification to test these compounds directly in vivo. Unfortunately, this research approach has an inherent flaw. It neglects the poor absorption, distribution, metabolism, and excretion (ADME) properties of peptides resulting in low peptide bioavailability. Because peptides are prone to extensive hydrolysis in the gastrointestinal tract by stomach, small intestinal, and brush border peptidases, most of them do not reach the absorption stage in the duodenum and jejunum. Therefore, a valid research approach should include the demonstration of stability of the peptide toward luminal and brush border peptidases and evaluate its ADME properties. Surprisingly, only very few animal and human studies determined absolute concentrations and kinetics of bioactive peptides. These studies have shown the presence of selected peptides in plasma samples at pico- and nanomolar concentrations with fast elimination kinetics in the minute range. For the correct interpretation of results, it is advised that researchers refer to the data currently available concerning bioavailability and ADME properties in humans. Two mandatory criteria for future in vitro studies investigating potential biological activities of peptides should be using physiologically relevant concentrations and times.

Continuous dose-response relationship of the LDL-cholesterol-lowering effect of phytosterol intake.

Phytosterols (plant sterols and stanols) are well known for their LDL-cholesterol (LDL-C)-lowering effect. A meta-analysis of randomized controlled trials in adults was performed to establish a continuous dose-response relationship that would allow predicting the LDL-C-lowering efficacy of different phytosterol doses. Eighty-four trials including 141 trial arms were included. A nonlinear equation comprising 2 parameters (the maximal LDL-C lowering and an incremental dose step) was used to describe the dose-response curve. The overall pooled absolute (mmol/L) and relative (%) LDL-C-lowering effects of phytosterols were also assessed with a random effects model. The pooled LDL-C reduction was 0.34 mmol/L (95% CI: -0.36, -0.31) or 8.8% (95% CI: -9.4, -8.3) for a mean daily dose of 2.15 g phytosterols. The impacts of subject baseline characteristics, food formats, type of phytosterols, and study quality on the continuous dose-response curve were determined by regression or subgroup analyses. Higher baseline LDL-C concentrations resulted in greater absolute LDL-C reductions. No significant differences were found between dose-response curves established for plant sterols vs. stanols, fat-based vs. non fat-based food formats and dairy vs. nondairy foods. A larger effect was observed with solid foods than with liquid foods only at high phytosterol doses (>2 g/d). There was a strong tendency (P = 0.054) towards a slightly lower efficacy of single vs. multiple daily intakes of phytosterols. In conclusion, the dose-dependent LDL-C-lowering efficacy of phytosterols incorporated in various food formats was confirmed and equations of the continuous relationship were established to predict the effect of a given phytosterol dose. Further investigations are warranted to investigate the impact of solid vs. liquid food formats and frequency of intake on phytosterol efficacy.

Postprandial changes in solubilizing capacity of human intestinal fluids for BCS class II drugs.

PURPOSE: To explore the effect of the nutritional state on the solubilizing properties of human intestinal fluids (HIF) on a time-after-food administration basis. METHODS: HIF were collected in fractions of 30 min from five volunteers in the fasted, fed and fat-enriched fed state. In vitro solubility of five BCS class II drugs (danazol, diazepam, nifedipine, ketoconazole, indomethacin) was assessed in the intestinal fractions and simulated intestinal fluids. RESULTS: Solubilities in intestinal fractions were characterized by high time- and subject-dependent variability. For the non-ionized drugs, solubility in early intestinal fractions was higher in both fed states compared to the fasted state, and in the fat-enriched fed state compared to the fed state. Solubility in simulated intestinal fluids did not sufficiently predict the solubilizing capacity of the early postprandial phase. Solubility in HIF was shown to be determined by a complex interplay of various intraluminal parameters. For the ionized drugs, pH played a significant role for indomethacin (R (2) = 0.86); for the partly ionized ketoconazole other intraluminal parameters were also important. CONCLUSIONS: Solubilizing capacity of HIF in the fed state is strongly time-dependent. Intraluminal dissolution may, therefore, vary with drug arrival time in the small intestine and constitute a source of variability in intestinal drug absorption.

Novel natural and synthetic inhibitors of solute carriers SGLT1 and SGLT2.

Selective analogs of the natural glycoside phloridzin are marketed drugs that reduce hyperglycemia in diabetes by inhibiting the active sodium glucose cotransporter SGLT2 in the kidneys. In addition, intestinal SGLT1 is now recognized as a target for glycemic control. To expand available type 2 diabetes remedies, we aimed to find novel SGLT1 inhibitors beyond the chemical space of glycosides. We screened a bioactive compound library for SGLT1 inhibitors and tested primary hits and additional structurally similar molecules on SGLT1 and SGLT2 (SGLT1/2). Novel SGLT1/2 inhibitors were discovered in separate chemical clusters of natural and synthetic compounds. These have IC50-values in the 10-100 µmol/L range. The most potent identified novel inhibitors from different chemical clusters are (SGLT1-IC50 Mean ± SD, SGLT2-IC50 Mean ± SD): (+)-pteryxin (12 ± 2 µmol/L, 9 ± 4 µmol/L), (+)-ε-viniferin (58 ± 18 µmol/L, 110 µmol/L), quinidine (62 µmol/L, 56 µmol/L), cloperastine (9 ± 3 µmol/L, 9 ± 7 µmol/L), bepridil (10 ± 5 µmol/L, 14 ± 12 µmol/L), trihexyphenidyl (12 ± 1 µmol/L, 20 ± 13 µmol/L) and bupivacaine (23 ± 14 µmol/L, 43 ± 29 µmol/L). The discovered natural inhibitors may be further investigated as new potential (prophylactic) agents for controlling dietary glucose uptake. The new diverse structure activity data can provide a starting point for the optimization of novel SGLT1/2 inhibitors and support the development of virtual SGLT1/2 inhibitor screening models.

The angiotensin converting enzyme inhibitory tripeptides Ile-Pro-Pro and Val-Pro-Pro show increasing permeabilities with increasing physiological relevance of absorption models.

Transepithelial transport of the ACE inhibitory peptides Ile-Pro-Pro and Val-Pro-Pro was studied in different models of absorption. Apparent permeability (P(app)) values for absorptive transport across Caco-2 monolayers were 1.0+/-0.9 x 10(-8) (Ile-Pro-Pro) and 0.5+/-0.1 x 10(-8)cms(-1) (Val-Pro-Pro). Ex vivo transport across jejunal segments in the Ussing chamber was 5-times (Ile-Pro-Pro) to 10-times (Val-Pro-Pro) higher with no significant differences (p>0.05) observed between both peptides. The peptidase inhibitor bestatin increased permeability for the absorptive direction for Ile-Pro-Pro by twofold. Neither a transepithelial pH gradient nor increased apical tripeptide concentration nor longitudinal localization of the intestinal segment influenced P(app) in the ex vivo experiments. Val-Pro-Pro transport across Peyer's patches, however, was 4-times higher (P(app)=21.0+/-9.3 x10(-8)cms(-1)) as compared to duodenum (P(app)=4.8+/-1.4 x 10(-8)cms(-1)). In the in situ perfusion experiments P(app) values varied greatly among different animals ranging from 0.5 to 24.0 x10(-8)cms(-1) (Ile-Pro-Pro) and from 1.0 to 15.6 x 10(-8)cms(-1) (Val-Pro-Pro). In summary, Caco-2 and ex vivo absorption models differ considerably regarding their peptide permeability. The in situ model seems to be less appropriate because of the observed large variability in peptide permeability. The results of this study demonstrate that the ACE inhibitory peptides Ile-Pro-Pro and Val-Pro-Pro are absorbed partially undegraded.

Pharmacokinetics of proline-rich tripeptides in the pig.

Tripeptides may possess bioactive properties. For instance, blood pressure lowering is attributed to the proline-rich tripeptides Ile-Pro-Pro (IPP), Leu-Pro-Pro (LPP), and Val-Pro-Pro (VPP). However, little is known about their absorption, distribution, and elimination characteristics. The aim of this study was to characterize the pharmacokinetic behavior of IPP, LPP, and VPP in a conscious pig model. Synthetic IPP, LPP, and VPP were administered intravenously or intragastrically (4.0 mg kg(-1) BW in saline) to 10 piglets (approximately 25 kg body weight) in the postabsorptive state. After intravenous dosing, the elimination half-life for IPP was significantly higher (P<0.001) than for LPP and VPP (2.5+/-0.1, 1.9+/-0.1, and 2.0+/-0.1 min, respectively). After intragastric dosing, however, the elimination half-lives were not significantly different between the peptides (9+/-1, 15+/-4, and 12+/-6 min, respectively). Maximum plasma concentrations were about 10 nmol l(-1) for the three tripeptides. The fraction dose absorbed was 0.077+/-0.010, 0.059+/-0.009, and 0.073+/-0.015%, for IPP, LPP, and VPP, respectively. Proline-rich tripeptides reach the blood circulation intact, with an absolute bioavailability of about 0.1% when administered via a saline solution. Because half-lives of absorption and elimination were maximally about 5 and 15 min, respectively, this suggests that under these conditions a bioactive effect of these tripeptides would be rather acute.

Genetic variation in ABC G5/G8 and NPC1L1 impact cholesterol response to plant sterols in hypercholesterolemic men.

ATP-binding cassette hetero-dimeric transporters G5 and G8 (ABCG5/G8) have been postulated to mediate intestinal cholesterol efflux, whereas Niemann-Pick C1 Like 1 (NPC1L1) protein is believed to be essential for intestinal cholesterol influx. The individual or combined genetic markers, such as single nuclear polymorphisms (SNPs), of these two transporter genes may explain inter-individual variations in plasma cholesterol response following plant sterol (PS) intervention. The present study was aimed at investigating the association between ABCG5/G8 and NPC1L1 genotype SNPs with sterol absorption and corresponding plasma concentrations. The study used a 4-week crossover design with 82 hypercholesterolemic men characterized by high vs. low basal plasma PS concentrations consuming spreads with or without 2 g/day of PS. For the ABCG8 1289 C > A (T400 K) polymorphism, the A allele carriers with high basal plasma PS concentrations demonstrated a 3.9-fold greater reduction (p < 0.05) in serum low density lipoprotein cholesterol (LDL-C) than their low basal plasma PS counterparts. For the NPC1L1 haplotype of 872 C > G (L272L) and 3929 G > A (Y1291Y), individuals carrying mutant alleles showed a 2.4-fold greater (p < 0.05) reduction in LDL-C levels, compared to wild type counterparts. Results suggest that genetic and metabolic biomarkers together may predict inter-individual lipid level responsiveness to PS-intervention, and thus could be useful in devising individualized cholesterol lowering strategies.

Intestinal disposition of quercetin and its phase-II metabolites after oral administration in healthy volunteers.

OBJECTIVES: Quercetin is one of the main dietary flavonoids and undergoes a substantial intestinal phase-II metabolism. Quercetin conjugates have been detected in plasma and in urine, but their presence in the small intestine has not been assessed. This study aimed to investigate the intestinal metabolism and metabolite excretion of quercetin by the human small intestinal wall after oral dosing. METHODS: Six healthy volunteers were given a capsule of 500 mg of quercetin with 240 ml of water. Duodenal fluids were collected using the intraluminal sampling technique for 4 h and analysed by LC-MS/MS. KEY FINDINGS: Phase-II metabolites of quercetin were detected and quantified in aspirated intestinal fluids. Metabolites appeared almost immediately after administration, indicating an intestinal metabolism and apical excretion into the lumen. Quercetin-3'-O-glucuronide was found to be the main intestinal metabolite. Our results could not conclude on the enterohepatic recycling of quercetin or its metabolites, although several individual profiles showed distinctive peaks. CONCLUSIONS: This study highlights the intestinal metabolism and excretion of quercetin and its conjugates in humans and gives insights into the relevant concentrations which should be used to investigate potential food-drug interactions in vitro.

Characterization of Human Duodenal Fluids in Fasted and Fed State Conditions.

This work provides an elaborate characterization of human intestinal fluids (HIF) collected in fasted- and fed-state conditions. HIF from 20 healthy volunteers (10 M/F) were aspirated by intubation near the ligament of Treitz in a time-dependent manner (10-min intervals) and characterized for pH, bile salts, phospholipids, cholesterol, triacylglycerides (TAG), diacylglycerides (DAG), monoacylglycerides (MAG), free fatty acids (FFA), pancreatic lipase, phospholipase A2, and nonspecific esterase activity. For almost all parameters, a food-induced effect was observed. Results were characterized by a high variability, as illustrated by the broad ranges observed for each parameter: pH (fasted: 3.4-8.3; fed: 4.7-7.1), bile salts (fasted: 0.03-36.18 mM; fed: 0.74-86.14 mM), phospholipids (fasted: 0.01-6.33 mM; fed: 0.16-14.39 mM), cholesterol (fasted: 0-0.48 mM; fed: 0-3.29 mM), TAG (fed: 0-6.76 mg/mL), DAG (fed: 0-3.64 mg/mL), MAG (fasted: 0-1.09 mg/mL; fed: 0-11.36 mg/mL), FFA (fasted: 0-3.86 mg/mL; fed: 0.53-15.0 mg/mL), pancreatic lipase (fasted: 26-86 g/mL; fed: 146-415 g/mL), phospholipase A2 (fasted: 3-6 ng/mL; fed: 4.3-27.7 ng/mL), and nonspecific esterase activity (fasted: 270-4900 U/mL; fed: 430-4655 U/mL). This comprehensive overview may serve as reference data for physiologically based pharmacokinetic modeling and the optimization of biorelevant simulated intestinal fluids for the use in in vitro dissolution, solubility, and permeability profiling.