European Journal of Nutrition (v.50, #7)

The impact of hepatitis B virus (HBV) or hepatitis C virus (HCV) infection upon B vitamins status and antioxidative defense in infected patients was examined.Dietary record and blood levels of B vitamins and oxidative stress–associated biomarkers were determined for 195 healthy controls, 132 HBV, and 114 HCV patients.HBV-infected patients had significantly higher levels of total cholesterol, free fatty acids (FFA), and lower ghrelin level (p < 0.05); and HCV-infected patients had significantly higher Ishak inflammation score and lactate dehydrogenase activity (p < 0.05). HBV patients had significantly lower red blood cell (RBC) vitamins B2 and B6 levels, and HCV infection significantly decreased vitamins B2, B6 and folate levels in RBC and/or plasma (p < 0.05). Correlation coefficients of RBC vitamin B2 versus serum FFA in HBV patients, RBC vitamins B2 and B6 versus HCV RNA and Ishak inflammation score, and plasma vitamin B6 vs Ishak inflammation score in HCV patients were <−0.5. HBV-infected patients had significantly higher oxidized glutathione level and lower glutathione peroxidase activity (p < 0.05), but HCV patients had significantly lower superoxide dismutase and catalase activities (p < 0.05).HBV or HCV infection enhanced oxidative stress and lowered B vitamins in circulation. In order to avoid other healthy risk, nutrition status should be monitored and limitation or supplementation of certain nutrients might be helpful for HBV- or HCV-infected patients.
Keywords: Hepatitis B virus; Hepatitis C virus; B vitamins; Oxidative stress; Lipid metabolism

Intestinal transit and systemic metabolism of apple polyphenols by Kathrin Kahle; Michael Kempf; Peter Schreier; Wolfgang Scheppach; Dieter Schrenk; Tanja Kautenburger; Dorothée Hecker; Wolfgang Huemmer; Matthias Ackermann; Elke Richling (507-522).
Apples are the most widely consumed fruits in Germany and various other countries. Positive health effects of apple-derived polyphenols in vivo depend on their absorption, metabolism, distribution, and elimination from the body after consumption. Data on the metabolism of these polyphenols in humans are scarce. In order to study the intestinal transit and metabolism of apple polyphenols in humans, a variety of experiments were carried out.Polyphenols were incubated with saliva (for 5 min), simulated gastric or duodenal juice (4 or 10 h, respectively), or rat hepatocytes (4 h) under aerobic conditions, and with ileostomy fluid under aerobic conditions for 10 h. The polyphenol profile in human serum (8 h later) and renal elimination in urine (24 h later) were also investigated after consumption of 1 L apple juice. Polyphenols and their metabolites were identified and quantified by high-performance liquid chromatography with diode array detection (HPLC–DAD), HPLC–electrospray ionization–tandem mass spectrometry (ESI-MS/MS), and gas chromatography (GC)-MS.In the presence of native saliva or ileostomy fluid, β-glycosides of phloretin and quercetin were hydrolyzed, to varying degrees depending on the sugar moiety, and to much lesser degrees in the presence of antibiotics. In the gastric milieu, almost complete degradation of procyanidin B2 to (−)-epicatechin was observed. In the presence of artificial duodenal juice flavan-3-ol epimerization occurred. Quercetin was completely converted to phloroglucinol, 3,4-dihydroxybenzoic acid, and 2,4,6-trihydroxybenzoic acid. Formation of isomeric products of hydroxycinnamic acid esters and their corresponding methyl esters was also observed, and similar results were obtained after incubation with rat hepatocytes. Products of phase II metabolism, two phloretin O-glucuronides and eight (methyl) quercetin O-glucuronides, were identified in the hepatocyte samples. Following enzymatic hydrolysis, 5-caffeoylquinic acid, 4-p-coumaroylquinic acid, caffeic acid, (−)-epicatechin, phloretin, and quercetin were recovered in both serum and urine (5.3% and 3.5% of the amounts consumed, respectively). In addition, 19.5% of the polyphenols consumed were identified in the urine in the form of hydroxylated phenolic and hippuric acids.The findings relating to the absorption, metabolism, and systemic availability of polyphenols in vivo should contribute to our understanding of their biological effects, and the characterization of newly formed metabolites should facilitate further studies.
Keywords: Metabolism; Bioavailability; Apple; Polyphenols; Intestinal transit; Urine; Plasma

The effects of PG102, a water-soluble extract from Actinidia arguta, on serum total IgE levels: a double-blind, randomized, placebo-controlled exploratory clinical study by Sae-Hoon Kim; Sunyoung Kim; So-Hee Lee; Heung-Woo Park; Yoon-Seok Chang; Kyung-Up Min; Sang-Heon Cho (523-529).
Recent studies have reported that blocking IgE has a potentially beneficial role in the treatment of various allergic diseases. Previously, we found that PG102, a water-soluble extract prepared from the edible fruits of Actinidia arguta, can effectively reduce IgE levels using murine models.To evaluate the efficacy of PG102 at lowering levels of total IgE in asymptomatic subjects with atopy.A total of 90 asymptomatic subjects with atopy were randomized equally to a PG102 group or a placebo control group and treated for 8 weeks in a double-blind manner. Total serum IgE, eosinophilic cation protein (ECP), eotaxin, thymus, and activation-regulated chemokine (TARC), IL-4, IL-5, and IL-13 levels were measured. Eosinophil counts were determined before and after treatment, and results were compared. In addition, possible adverse reactions were thoroughly checked in this first human trial.Levels of total IgE significantly increased in the control group but showed no change in the PG102 group, and change differences between the control and PG102 groups were significant (+12.9%, vs.−5.7%, p = 0.015). Levels of ECP and eotaxin and eosinophil counts produced similar results. However, the other variables showed no significant changes after treatment.In this exploratory clinical trial, it was found that 8 weeks of treatment with PG102 effectively reduced the levels of total IgE in apparently asymptomatic subjects with atopy.
Keywords: Actinidia arguta ; Allergy; IgE; PG102

Vitamin A (VA) deficiency is still a major health problem in the developing world. It affects various cellular functions and causes hypolipidemic effects in the body. β-Carotene (BC)-rich foods are promising sources of VA. Phospholipids are reported to improve BC bioefficacy in normal rats, but whether they show similar effects during VA deficiency is unknown.To compare the BC metabolism and plasma lipid responses in VA-sufficient (+VA) and VA-deficient (−VA) rats after a single oral dose of micellar BC containing phospholipids.Groups of rats were fed with a VA-free diet and when they attained the weight-plateau stage of deficiency, both +VA and −VA rats were divided into 2 groups (phosphatidylcholine, PC and lysophosphatidylcholine, LPC). Each group was further divided into 4 subgroups (1, 2, 3, and 6 h; n = 5 rats/time point) and determined the BC metabolism and plasma lipid responses to a post-dose of micellar BC with phospholipids.Maximal plasma BC (pmol/mL) levels were observed at 2 h in PC (1330 ± 124) and at 1 h in LPC (1576 ± 144) groups of +VA rats, and at 3 h in the PC (1621 ± 158) and LPC (2248 ± 675) groups of −VA rats. Liver BC (pmol/g) was maximum at 1 h in the PC (218 ± 32) and LPC (249 ± 24) groups of +VA rats, and at 2 h in PC (228 ± 23) and at 3 h in LPC (277 ± 18) groups of −VA rats. Plasma and liver BC levels were significantly (P < 0.05) higher in −VA rats than +VA rats. Plasma retinyl palmitate (pmol/mL) was maximum at 3 h in PC (97 ± 18) and at 2 h in LPC (126 ± 14) groups of +VA rats, and at 2 h in the PC (92 ± 13) and LPC (134 ± 27) groups of −VA rats. The higher (P < 0.05) BC monoxygenase activity in −VA rats compared to +VA rats supports the BC bioefficacy. Plasma retinol level was improved in the PC and LPC groups, but the effect of LPC was higher (P < 0.05) than PC. Micellar phospholipids mitigate the VA deficiency–induced hypolipidemic effects.Micellar phospholipids improved BC metabolism and reinstated the hypolipidemic effects, perhaps by modifying the fat-metabolizing enzymes and repairing the altered intestinal membrane structure.
Keywords: Bioavailability; β-Carotene; Mixed micelles; Phospholipids; Vitamin A deficiency

Impairment of cardiac insulin signaling in fructose-fed ovariectomized female Wistar rats by Zorica Zakula; Goran Koricanac; Snezana Tepavcevic; Mojca Stojiljkovic; Tijana Milosavljevic; Esma R. Isenovic (543-551).
Fructose consumption produces deleterious metabolic effects in animal models. The sites of fructose-induced insulin resistance are documented to be the liver, skeletal muscle, and adipose tissue, but effects of fructose-rich diet on cardiac insulin signaling and action were not investigated.In order to study the potential fructose effects on development of cardiac insulin resistance, we analyzed biochemical parameters relevant for insulin action and phosphorylation of insulin signaling molecules, plasma membrane glucose transporter type 4 (GLUT4) content, and phosphorylation of endothelial nitric oxide synthase (eNOS), in ovariectomized female rats on fructose-enriched diet, in basal and insulin-stimulated conditions.Fructose-fed rats (FFR) had increased content of visceral adipose tissue, but not body weight. Food intake was decreased, while fluid and caloric intake were increased in FFR. Additionally, fructose diet increased plasma insulin, blood triglycerides level, and HOMA index. Stimulation of protein kinase B (Akt) signaling pathway by insulin was reduced in rats on fructose-enriched diet, but effect of fructose on extracellular signal-regulated kinase (Erk 1/2) phosphorylation was not observed. Furthermore, insulin-induced GLUT4 presence in plasma membranes of cardiac cells was decreased by fructose diet, as well as insulin stimulation of eNOS phosphorylation at Ser1177.In summary, these results strongly support our hypothesis that fructose diet-induced changes of plasma lipid profile and insulin sensitivity are accompanied with decrease in cardiac insulin action in ovariectomized female rats.
Keywords: Fructose; Heart; Insulin resistance; Glucose transporter type 4; Nitric oxide synthase type III

Anti-platelet effects of olive oil extract: in vitro functional and proteomic studies by Baukje de Roos; Xuguang Zhang; Guillermo Rodriguez Gutierrez; Sharon Wood; Garry J. Rucklidge; Martin D. Reid; Gary J. Duncan; Louise L. Cantlay; Garry G. Duthie; Niamh O’Kennedy (553-562).
Platelets play a key role in haemostasis and wound healing, contributing to formation of vascular plugs. They are also involved in formation of atherosclerosic plaques. Some traditional diets, like the Mediterranean diet, are associated with a lower risk of cardiovascular disease. Components in these diets may have anti-platelet functions contributing to their health benefits.We studied the effects of alperujo extract, an olive oil production waste product containing the majority of polyphenols found in olive fruits, through measurement of effects on platelet aggregation and activation in isolated human platelets, and through identification of changes in the platelet proteome.Alperujo extract (40 mg/L) significantly decreased in vitro ADP- (p = 0.002) and TRAP- (p = 0.02) induced platelet activation as measured by the flow cytometry using the antibody for p-selectin (CD62p), but it did not affect the conformation of the fibrinogen receptor as measured by flow cytometry using the antibodies for anti-fibrinogen, CD42a and CD42b. Alperujo extract (100 mg/L) inhibited both collagen- and TRAP-induced platelet aggregation by 5% (p < 0.05), and a combination of hydroxytyrosol and 3,4-dihydroxyphenylglycol were, at least partly, responsible for this effect. Proteomic analysis identified nine proteins that were differentially regulated by the alperujo extract upon ADP-induced platelet aggregation. These proteins represent important mechanisms that may underlie the anti-platelet effects of this extract: regulation of platelet structure and aggregation, coagulation and apoptosis, and signalling by integrin αIIb/β3.Alperujo extract may protect against platelet activation, platelet adhesion and possibly have anti-inflammatory properties.
Keywords: Platelet function; Mediterranean diet; Alperujo extract; Proteomics

To study the effect of diet supplementation with polyphenols on several functions suffering age-related changes, in peritoneal leucocytes from mature and old mice.Five groups of female ICR mice were used. Four groups received a supplementation (20% wt/wt) of biscuits with different cereal fractions naturally rich in polyphenols (named CO49, CO50, CO52, CO53), containing different amounts of catechin, p-hydroxybenzoic acid, vanillic acid, p-coumaric acid, sinapic acid, ferulic acid, rutin and oryzanol. The control group received only standard maintenance diet. Peritoneal suspensions were obtained after 15 and 30 weeks of diet supplementation, when the age of the animals was 49 ± 2 (mature mice) and 64 ± 2 weeks (old mice), respectively. The functions analysed were: chemotaxis of macrophages and lymphocytes, phagocytosis of particles by macrophages, intracellular superoxide anion levels, lymphoproliferative response to mitogens (concanavalin A and lipopolysaccharide), interleukin-2 secretion and natural killer (NK) activity, as functions that decrease with age, and adherence of macrophages and lymphocytes and tumour necrosis factor-α secretion as functions with age-related increase.The supplementation, in general, increased the functions that decrease with age and decreased those that increase with age. There were differences in the effects shown by the four kinds of biscuits depending on the function studied and the number of weeks of supplementation.Since the immune system has been proposed as a good marker of health and predictor of longevity, diet supplementation with cereals naturally rich in polyphenols could be an important way for health preservation with age and reaching high longevity.
Keywords: Leucocyte functions; Polyphenols; Ageing; Mice

Plasma pharmacokinetics of catechin metabolite 4′-O-Me-EGC in healthy humans by Mathieu Renouf; Karine Redeuil; Karin Longet; Cynthia Marmet; Fabiola Dionisi; Martin Kussmann; Gary Williamson; Kornél Nagy (575-580).
Tea is an infusion of the leaves of the Camellia sinensis plant and is the most widely consumed beverage in the world after water. Green tea contains significant amounts of polyphenol catechins and represents a promising dietary component to maintain health and well-being. Epidemiological studies indicate that polyphenol intake may have potential health benefits, such as, reducing the incidence of coronary heart disease, diabetes and cancer. While bioavailability of green tea bioactives is fairly well understood, some gaps still remain to be filled, especially the identification and quantification of conjugated metabolites in plasma, such as, sulphated, glucuronidated or methylated compounds.In the present study, we aimed to quantify the appearance of green tea catechins in plasma with particular emphasis on their methylated forms.After feeding 400 mL of green tea, 1.25% infusion to 9 healthy subjects, we found significant amounts of EC, EGC and EGCg in plasma as expected. EGC was the most bioavailable catechin, and its methylated form (4′-O-Me-EGC) was also present in quantifiable amounts. Its kinetics followed that of its parent compound. However, the relative amount of the methylated form of EGC was lower than that of the parent compound, an important aspect which, in the literature, has been controversial so far. The quantitative results presented in our study were confirmed by co-chromatography and accurate mass analysis of the respective standards. We show that the relative abundance of 4′-O-Me-EGC is ~40% compared to the parent EGC.4′-O-Me-EGC is an important metabolite derived from catechin metabolism. Its presence in significant amounts should not be overlooked when assessing human bioavailability of green tea.
Keywords: Green tea; Bioavailability; Catechins

Relation of body mass index to blood folate and total homocysteine concentrations in Japanese adults by Mio Nakazato; Takahiro Maeda; Noboru Takamura; Mitsuhiro Wada; Hironori Yamasaki; Kelley E. Johnston; Tsunenobu Tamura (581-585).
Plasma folate concentrations are suggested to be negatively associated with body mass index (BMI, kg/m2), although these findings are controversial. Our objective was to evaluate the association of BMI with blood folate and total homocysteine (tHcy) concentrations.We measured plasma and erythrocyte folate and plasma tHcy concentrations in 434 healthy adults (343 women and 91 men; mean age of 63.8 ± 10.7 [SD, range 23–88] years), who participated in a 2007 population-based survey in western Japan.The overall mean plasma and erythrocyte folate and tHcy were 21.6 (±11.0, SD) nmol/L, 844 (±291) nmol/L and 11.6 (±3.9) μmol/L, respectively. The mean BMI was 22.8 (±3.0; 15.6–33.3) kg/m2, and only 72 subjects (17%) had BMI > 26.0 kg/m2. Mean plasma folate decreased as BMI increased (p-trend < 0.01), whereas mean erythrocyte folate and plasma tHcy were similar regardless of BMI (p-trends = 0.49 and 0.28, respectively).Our data indicate that the interpretation of plasma folate concentrations to assess folate nutritional status is complicated by BMI, although the impact of BMI on plasma folate was relatively small. It is important to take this association into account for the selection of subjects for future large-scale studies. The mechanism of this inverse association between BMI and plasma folate concentrations should be investigated.
Keywords: Folate; Blood; Homocysteine; Body mass index; Human

Impact of spinach consumption on DNA stability in peripheral lymphocytes and on biochemical blood parameters: results of a human intervention trial by Beate Moser; Thomas Szekeres; Christian Bieglmayer; Karl-Heinz Wagner; Miroslav Mišík; Michael Kundi; Oliwia Zakerska; Armen Nersesyan; Nina Kager; Johann Zahrl; Christine Hoelzl; Veronika Ehrlich; Siegfried Knasmueller (587-594).
A controlled intervention trial was conducted to assess the impact of spinach consumption on DNA stability in lymphocytes and on health-related biochemical parameters.The participants (n = 8) consumed homogenised spinach (225 g/day/person) over a period of 16 days. DNA migration was monitored in single cell gel electrophoresis—comet assays under standard conditions, which reflect single- and double-strand breaks, after treatment of nuclei with lesion-specific enzymes (formamidopyrimidine glycosylase, FPG and endonuclease III, ENDO III) and after treatment of intact cells with H2O2 before, during and after intervention.While no reduction in DNA damage was observed under standard conditions after different time intervals of spinach intake, other endpoints, namely ROS sensitivity and DNA migration attributable to the formation of oxidatively damaged DNA bases (i.e. pyrimidines-ENDO III-sensitive sites and purines-FPG sensitive sites) were reduced 6 h after consumption of the first portion and after 11 days of continuous consumption. In the case of ENDO III-sensitive sites, also after 16 days, a decrease in comet formation was observed. At the end of a 40 days washout period, the DNA stability parameters were not significantly different from the background values. Other biochemical parameters which were significantly altered by spinach intake were the folate (+27%) and homocysteine (−16%) concentrations in blood, and it was found in an earlier human study that folate may prevent oxidative damage to DNA bases.Taken together, our results show that moderate consumption of spinach causes protection against oxidative DNA damage in humans and that this phenomenon is paralleled by alterations of health-related biochemical parameters.
Keywords: Intervention trial; Spinach; DNA damage; Antioxidant

Direct evidence that (−)-epicatechin increases nitric oxide levels in human endothelial cells by Tatjana Brossette; Claas Hundsdörfer; Klaus-Dietrich Kröncke; Helmut Sies; Wilhelm Stahl (595-599).
The dietary flavanol (−)-epicatechin has been suggested to mediate its vasodilatory effect by increasing nitric oxide levels in endothelial cells.To directly prove the formation of nitric oxide (NO) in human endothelial cells (HUVEC) in vitro by trapping NO to yield a fluorescent nitrosamine.HUVEC were treated with (−)-epicatechin; nitrite and NO formation were determined by reductive chemiluminescence detection and the NO-sensitive fluorophore 5-methoxy-2-(1H-naphthol[2,3-d]imidazol-2-yl)-phenol copper complex (MNIP-Cu), respectively. MNIP was synthesized in a rapid and convenient one-step microwave reaction. Endothelial nitric oxide synthase (eNOS) mRNA levels and mRNA stability were measured.Incubation with (−)-epicatechin (0.3–10 μM) led to elevated NO levels in HUVEC measured via reductive chemiluminescence detection and visualized as the fluorescent NO derivative of MNIP. Expression of eNOS mRNA and mRNA stability were not affected by (−)-epicatechin treatment within the time frame studied.(−)-Epicatechin augments the level of NO in endothelial cells, a process suggested to be responsible for the vasodilatory properties of the compound.
Keywords: (−)-Epicatechin; eNOS; HUVEC; Chemiluminescence; Fluorescence microscopy; NO