The global profiling of the whole protein complement of the genome expressed in a particular cell or organ, or in plasma or serum, makes it possible to identify biomarkers that respond to alterations in diet or to treatment, and that may have predictive value for the modelling of biological processes. Proteomics has not yet been applied on a large scale in nutritional studies, yet it has advantages over transcriptome profiling techniques in that it directly assesses the entities that carry out the biological functions. The present review summarizes the different approaches in proteomics research, with special emphasis on the current technical ‘workhorses’: two-dimensional (2D)-PAGE with immobilized pH gradients and protein identification by MS. Using a work-flow approach, we provide information and advice on sample handling and preparation, protein solubilization and pre-fractionation, protein separation by 2D-PAGE, detection and quantification via computer-assisted analysis of gels, and protein identification and characterization techniques by means of MS. Examples from nutritional studies employing proteomics are provided to demonstrate not only the advantages but also the limitations of current proteome analysis platforms.

The aim of the present study was to investigate the effect of Chinese soft-shelled turtle whole egg powder (TE) on cholesterol metabolism in Sprague–Dawley rats to determine whether it has a cholesterol-lowering effect. Forty male Sprague–Dawley rats were fed a high-fat diet supplemented with TE (0, 0·75, 1·50 or 3·00 g/kg body weight) administrated by gavage for 24 weeks. Serum total cholesterol (TC), HDL-cholesterol (HDL-C), LDL-cholesterol (LDL-C) and faecal total bile acids levels were determined by enzymatic methods. Faecal steroid concentrations were measured by GC. Means and standard deviations were calculated where appropriate for values, and the data were tested by one-way ANOVA. After 24 weeks of feeding a high-fat diet with TE supplementation, serum TC and LDL-C levels, liver cholesterol and liver lipid levels were reduced in rats. TE supplementation did not affect the faecal output, but significantly increased steroid concentrations in faeces, indicating increased steroids excretion. The faecal bile acid excretion was also increased as evidence by elevated mRNA level of liver cytochrome P450, family 7, subfamily A, polypeptide 1. Our results demonstrated that the TE does have a cholesterol-lowering effect by increasing the excretion of total bile acids and neutral steroids.

It has previously been demonstrated that soya protein, which contains isoflavones and low methionine–glycine and lysine–arginine ratios, has a hypocholesterolaemic effect. In the present study, the hypocholesterolaemic effects of an isoflavone-enriched casein diet (HDI) and a single-cell protein-based diet (SCP) devoid of isoflavones but with low methionine–glycine and lysine–arginine ratios were investigated in obese Zucker rats after 6 weeks of feeding. The control diet contained casein, which has high ratios of methionine–glycine and lysine–arginine. HDI and SCP feeding reduced the concentrations of total cholesterol and cholesteryl esters in plasma and liver, and changed the fatty acid composition of the hepatic cholesteryl esters. Faecal cholesterol and bile acid levels were markedly higher in SCP-fed rats than in controls, whereas HDI feeding had only minor effects. However, both HDI and SCP feeding increased the hepatic gene expression of cholesterol 7α hydroxylase. In contrast, the hepatic acyl-CoA synthetase and acyl-CoA:cholesterol acyltransferase activities and the gene expression of the LDL receptor were increased by HDI, but not by SCP feeding. The present results suggested that the cholesterol-lowering effect of SCP was related to the enterohepatic circulation, whereas HDI seemed to lower the plasma cholesterol via the circulation. Plasma homocysteine level was reduced in rats fed HDI and SCP compared to rats fed casein. In summary, diets enriched in isoflavones or containing proteins with low methionine–glycine and lysine–arginine ratios lowered the plasma cholesterol and homocysteine levels, changing the plasma profile from atherogenic to cardioprotective.

Fibres with a range of abilities to perturb cholesterol homeostasis were used to investigate how the serum cholesterol-lowering effects of insoluble dietary fibres are related to parameters of intestinal cholesterol absorption and hepatic cholesterol homeostasis in mice. Cholestyramine, chitosan and cellulose were used as examples of fibres with high, intermediate and low bile acid-binding capacities, respectively. The serum cholesterol levels in a control group of mice fed a high fat/high cholesterol (HFHC) diet for 3 weeks increased about 2-fold to 4·3 mm and inclusion of any of these fibres at 7·5 % of the diet prevented this increase from occurring. In addition, the amount of cholesterol accumulated in hepatic stores due to the HFHC diet was reduced by treatment with these fibres. The three kinds of fibres showed similar hypocholesterolaemic activity; however, cholesterol depletion of liver tissue was greatest with cholestyramine. The mechanisms underlying the cholesterol-lowering effect of cholestyramine were (1) decreased cholesterol (food) intake, (2) decreased cholesterol absorption efficiency, and (3) increased faecal bile acid and cholesterol excretion. The latter effects can be attributed to the high bile acid-binding capacity of cholestyramine. In contrast, incorporation of chitosan or cellulose in the diet reduced cholesterol (food) intake, but did not affect either intestinal cholesterol absorption or faecal sterol output. The present study provides strong evidence that above all satiation and satiety effects underlie the cholesterol-lowering properties of insoluble dietary fibres with moderate or low bile acid-binding capabilities.

The plasma membrane oxidoreductase (PMOR) activity, which mainly utilises ascorbate as intracellular electron donor, represents a major mechanism for cell-dependent reduction of extracellular oxidants and might be an important process used by the erythrocytes to keep a reduced plasma environment. We previously reported that in human erythrocytes, myricetin and quercetin act as intracellular substrates of a PMOR showing a novel mechanism whereby these flavonoids could exert beneficial effects under oxidative stress conditions. Here, we evaluated the ability of different flavonoids (quercetin, myricetin, morin, kaempferol, fisetin, catechin, luteolin, apigenin, acacetin, rutin, taxifolin, naringenin, genistein) and of two in vivoO-methylated metabolites of quercetin (isorhamnetin and tamarixetin) to be substrates of PMOR, by comparing their antioxidant capacity (i.e. direct interaction with the oxidant ferricyanide or with the free radical 1,1-diphenyl-2-picryl-hydrazil) with their ability to penetrate the erythrocytes and donate electrons to the PMOR. The results obtained indicate that, although most of the flavonoids display significant antioxidant activities, only those (quercetin, myricetin, fisetin) that combine the cathecol structure of the B ring (responsible for the reducing activity) with the 2,3 double bond and 4-oxo function of the C ring (responsible for the uptake by erythrocytes) can act as intracellular substrates for PMOR. It is of note that the metabolites of quercetin enter erythrocytes and donate electrons to the PMOR as the parent compound. The present data show a relationship between the flavonoid structures and their ability to provide electrons to the PMOR, suggesting an additional mechanism whereby dietary flavonoids may exert beneficial effects in man.

Trans fatty acids are found mainly in processed foods. It has been shown that when their intake is high, total cholesterol, LDL-cholesterol and triacylglycerols are elevated, while HDL-cholesterol decreases. To evaluate a possible effect of these compounds, even in low amounts, C57Bl/6J mice were fed for 7 weeks a diet containing 13·6 % energy as partially hydrogenated rapeseed oil-enriched diet (Trans diet). The Trans diet contained 3 % energy as trans 18 : 1 fatty acid (elaidic acid). Control mice were on an isologous diet containing native rapeseed oil (Rapeseed diet) in which trans fatty acids were undetectable. Total, free and HDL-cholesterol as well as reverse cholesterol transport did not change. However, plasma triacylglycerol and VLDL levels increased. Hepatic gene expression in the Trans v. Rapeseed diet were compared using quantitative RT–PCR. The Trans diet produced a 2–3-fold elevation in mRNA of fatty acid synthase and microsomal transfer protein mRNA, explaining (at least in part) the observed increase in triacylglycerols and VLDL. In addition, mice on the Trans diet developed a deficiency in plasma vitamin E accompanied by a higher concentration of F2-isoprostanes, indicative of increased oxidative stress. The 78 kDa glucose-related protein (GRP78) mRNA expression increased 3–4-fold in liver, suggesting that a response against apoptosis was provoked by lipid peroxidation.

The activity of the somatotropic axis was analysed in juvenile rainbow trout (Oncorhynchus mykiss) fed either a fishmeal-based diet (FM) or graded levels of plant proteins to replace 50 % (PP50 diet), 75 % (PP75 diet) or 100 % (PP100 diet) of the fishmeal protein. For this purpose, partial cloning and sequencing of the gene encoding rainbow trout growth hormone receptor (GHR) was first accomplished by RT-PCR, using degenerate primers based on the sequences of non-salmonid fish GHR. Growth rates and energy retention were lowered by the PP75 and PP100 diets and a concurrent and progressive increase in plasma levels of growth hormone (GH) was found. However, no changes in hepatic GH binding and total plasma insulin-like growth factor (IGF)-I levels were observed among the four experimental groups. This fact agrees with the lack of changes in hepatic measurements of GHR and IGF-I transcripts. No consistent changes in IGF transcripts were found in peri-visceral adipose tissue and skeletal muscle, but GHR mRNA was up-regulated in the peri-visceral adipose tissue of fish fed the PP75 and PP100 diets, which would favour the lipolytic action of GH. Two specific bands (47 and 33 kDa) of IGF-binding proteins were found in the plasma of all analysed fish, but the sum of the two integrated areas increased progressively with plant protein supply, which might reflect a reduced free IGF availability. Therefore, in our experimental model, the growth impairment could be due, at least in part, to a lowered availability of biologically active IGF (free IGF fraction) rather than to liver GH desensitization or defect in IGF synthesis and release at the systemic and/or paracrine–autocrine level.

Soya foods may protect against the development of breast cancer. Insulin-like growth factor (IGF)-1 is under investigation as a possible link between nutrition and cancer. We examined the effect of soya foods on circulating IGF-1 and IGF binding protein (BP)-3 levels among 196 healthy premenopausal women in a 2-year randomised nutritional trial. The intervention group consumed two daily servings of soya foods including tofu, soya milk, soya nuts and soya protein powder (equivalent to 50 mg isoflavones and 5–22 g soya protein per serving); the controls maintained their regular diet. Five serum samples at baseline, 3, 6, 12, and 24 months were collected in the morning during the luteal phase and analysed for IGF-1 and IGFBP-3 by double-antibody ELISA. We applied mixed models to investigate the intervention effect and predictors of serum levels while considering the repeated measurement design. Adherence with the study regimen was high and dropout rates were acceptable. Randomisation resulted in similar mean IGF-1 and IGFBP-3 levels by group. We did not observe a significant intervention effect on IGF-1, IGFBP-3, and their molar ratio during the entire study period. However, urinary isoflavone excretion during the study period was positively associated with IGF-1 (P=0·04) and the IGF-1:IGFBP-3 ratio (P=0·06). The effect was consistent over time. Adding soya foods to the diet of premenopausal women does not appear to lower serum levels of IGF-1 and IGFBP-3; if anything, the greater protein intake from soya may lead to a small increase in IGF-1 serum levels.

An investigation was carried out to determine whether there were significant changes in nutrient intake over 17 years of adult life. The Medical Research Council National Survey of Health and Development is a longitudinal study of a nationally representative cohort of singleton births in the UK in 1946. Of this cohort, 1253 survey members provided information on diet recorded in a 5 d diary at age 36 years in 1982, 43 years in 1989 and 53 years in 1999. The outcome measures were mean intakes of energy, macronutrients, minerals and vitamins. There were significant changes in the intake of most nutrients in 1999 compared with previous years. Intakes of fat, Na, Fe and Cu have fallen, but there was a rising trend in the intakes of Ca, P, carotene, thiamin, pyridoxine, folic acid and vitamins C, D and E in both men and women. Additionally, intakes of K, Mg and vitamin K1 have risen in women. There were significant gender differences, women showing a higher percentage rise in the intakes of carotene, riboflavin, folic acid, vitamin C and vitamin E. These changes were related to changes in the consumption of certain key foods, such as the increased consumption of fruit and vegetables and a shift away from whole milk, butter and red meat. Most of these trends are in line with accepted nutritional guidelines. How far these changes are due to consumer choice and real changes in food composition or are due to artefacts inherent in the methodology is discussed.

The aim of this report is to describe breastfeeding duration and introduction of foods in Swedish infants born 1997–9, in relation to current recommendations. A secondary aim is to examine breastfeeding duration and introduction of certain allergenic foods in allergy-risk families (for whom allergy-preventive advice has been issued). Out of 21 700 invited infants, screening questionnaires were completed for 16 070 infants after delivery. Parents to 11 081 infants completed a follow-up questionnaire regarding breastfeeding and introduction of foods and 9849 handed in detailed food diaries at 1 year of age. The percentages of infants who were exclusively breast-fed at 3, 6 and ≥9 months of age were 78·4, 10·1 and 3·9, respectively. The corresponding percentages for partial breastfeeding were 87·8, 68·9 and 43·6. Gluten-containing foods were introduced to 66 % of infants between 4 and 6 months, as recommended at the time of the study, and one-quarter had stopped breastfeeding when gluten was introduced. More than 90 % of parents introduced the first sample of solid food during months 4–6, as recommended. Fish and eggs had been introduced during the first year in 43 % and 29 %, respectively, of infants with atopic heredity. Exclusive breastfeeding duration and time of introduction of solid foods, including gluten, seemed to have been in line with Swedish recommendations at the time, although gluten was often introduced late, and not during ongoing breastfeeding as recommended. The adherence to allergy-preventive advice was less than optimal in infants with atopic heredity.

The relationship between malnutrition and social support was first suggested in the mid-1990s. Despite its plausibility, no empirical studies aimed at obtaining evidence of this association could be located. The goal of the present study was to investigate such evidence. A case–control study was carried out including 101 malnourished children (weight-for-age National Center for Health Statistics/WHO 5th percentile) aged 12–23 months, who were compared with 200 well-nourished children with regard to exposure to a series of factors related to their social support system. Univariate and multiple logistic regressions were carried out, odds ratios being adjusted for per capita family income, mother's schooling, and number of children. The presence of an interaction between income and social support variables was also tested. Absence of a partner living with the mother increased risk of malnutrition (odds ratio 2·4 (95 % CI 1·19, 4·89)), even after adjustment for per capita family income, mother's schooling, and number of children. The lack of economic support during adverse situations accounted for a very high risk of malnutrition (odds ratio 10·1 (95 % CI 3·48, 29·13)) among low-income children, but had no effect on children of higher-income families. Results indicate that receiving economic support is an efficient risk modulator for malnutrition among low-income children. In addition, it was shown that the absence of a partner living with the mother is an important risk factor for malnutrition, with an effect independent from per capita family income, mother's schooling, and number of children.

The present study compared the nutritional status of schoolchildren from recently settled, ethnic minority tribespeople with those from a Persian village in southern Iran. Height and weight were measured and blood was collected from school children at three time points over 1·5 years. Supplemental Fe was provided to children with low Hb after the first screening. Twenty-one per cent of the children were wasted, 57 % were stunted and 23 % were anaemic. No statistically significant difference in the prevalence of wasting, stunting and anaemia was found between gender or ethnic groups. Children over the age of 12 years had a higher prevalence of wasting than children aged below 12 years. In a sub-sample of forty-one children the average BMI-for-age decreased. Fe supplementation increased Hb levels to normal in most children, but did not increase Fe level in a few children. Dietary deficiency of micronutrients, especially Zn and Fe, probably accounts for the high prevalence of stunting and anaemia in these children. Infection with Helicobacter pylori is another possible explanation for the Fe-deficiency anaemia. Further investigation is in progress to determine the cause(s) of the observed deficiencies.

The role of dietary glycaemic index (GI) and glycaemic load (GL) in disease aetiology is of increasing interest. However, nutritional factors related to dietary GI and GL are not well understood from a population perspective. We aimed to investigate the relation ship between GI and GL and dietary intake at the food and nutrient level. Study subjects were 1071 non-diabetic adults from the Insulin Resistance Atherosclerosis Study, Exam I, 1992–4. Usual dietary intake was assessed with a 114-item modified Block food frequency questionnaire. Published GI values were assigned to food line items. Correlation and regression analyses were conducted. Intake of white bread, beer, meats and fries/fried potatoes was positively associated with average GI, as was fat, starch and alcohol intake (before and after energy adjustment). Intake of fruits and low-fat milk was inversely associated with GI, as were intakes of mono- and disaccharides, and fibre. GL was positively correlated with carbohydrate foods and inversely with non-carbohydrate foods. Gender-specific regression models identified eight food groups explaining 63 % (men) and 55 % (women) total GI variation after adjusting for demographics; 70 % of variation in GL was explained by eleven (men) and nine (women) food groups, respectively. Although the GI of a food is an indicator of the ability of carbohydrates to raise blood glucose, dietary GI, unlike GL, appears to reflect more dimensions of diet than just carbohydrates, such as the combination of foods consumed. This may have implications for the interpretation of dietary GI in epidemiologic studies.

In order to examine the consumption of dietary supplements among urban adults and the impact of psychological stress on supplement use in relation to lifestyle, 375 interviews of a population-based sample of urban Japanese in 2002 were analysed. The usage of various supplements, stress process (daily stressors, psychological moderators, stress outcomes), personal health practices (smoking, alcohol drinking, physical exercise, fruit and vegetable juice consumption, health-conscious eating habits) and other background factors were measured. We examined the impacts of stress on the use of vitamin tablets and capsules, vitamin-enriched health drinks and health drinks for intestinal adjustment. The percentages of these three categories of supplement user were 26·9, 18·7 and 35·7 %, respectively. After adjusting for potential confounders, subjects with ‘two or more’ daily stressors out of the eight stressors investigated consistently showed 2-fold higher levels of consumption of either vitamin tablets and capsules or vitamin-enriched drinks compared with their counterparts with ‘one or less’ daily stressors. Stress-outcome indicators also related, to a greater or less extent, to the elevated consumption of various supplements. Further lifestyle-stratified analyses revealed that the stress–supplementation relationships were weaker in subjects fulfilling more than three of the five investigated health practices (i.e. the healthy lifestyle group), but stronger in subjects with fewer than two healthy practices (i.e. the less healthy lifestyle group). In conclusion, dietary supplement consumption is independently associated with stress in urban adults. The uncontrolled use of supplements for the self-medication of stress or to compensate for unhealthy behaviour represents a health concern for the general population.

α-Lactalbumin is a tryptophan-rich protein fraction. A diet enriched with α-lactalbumin increases the ratio of tryptophan to the other large neutral amino acids, which may in turn increase brain serotonin content. In stress-vulnerable individuals, α-lactalbumin improved mood and attenuated the cortisol response after experimental stress. The aim of the present study was to investigate the effects of an α-lactalbumin-enriched diet on mood and stress response in recovered depressed subjects and healthy controls. Forty-three subjects (twenty-three recovered depressed and twenty healthy subjects) received α-lactalbumin and casein (placebo) on separate days, in a double-blind randomised crossover design. On both occasions, subjects underwent a stress test (an unsolvable mental arithmetic task with loud noise). The stress test affected mood in both conditions. Although the α-lactalbumin diet led to the expected rises in tryptophan and tryptophan:large neutral amino acids ratio, only minimal effects were found on mood and cortisol response to experimental stress. The results were the same for recovered depressed patients and controls. A 1 d diet enriched with α-lactalbumin is not sufficient to prevent a stress-induced mood deterioration or a cortisol response in unmedicated, recovered depressed subjects. Future studies may investigate the effects of longer-term diets or may investigate different samples (e.g. medicated patients).

The present study describes food habits in adolescents in thirty-five countries and regions (European countries/regions, Israel, Canada and the USA), based on the food-frequency questions from the cross-sectional Health Behaviour in School-aged Children survey of the year 2001–2. A total of 162 305 pupils of 11, 13 or 15 years of age completed an anonymous standardised questionnaire on health and lifestyle factors during one school hour. Large differences in food habits between countries were found: the consumption frequency of fruit varied from on average 2·8 to 5 d/week, the consumption of vegetables varied from on average 2·4 to 5·5 d/week, the consumption of soft drinks varied from 2·1 to 5 d/week and sweet consumption from 2·6 to 5 d/week. Methodological problems in comparing between countries are discussed. Nonetheless, the results indicate a need for national and international health promotion programmes to improve adolescents' food habits.

It has been reported that green tea has a thermogenic effect, due to its caffeine content and probably also to the catechin, epigallocatechin-3-gallate (EGCG). The main aim of the present study was to compare the effect of a mixture of green tea and Guarana extracts containing a fixed dose of caffeine and variable doses of EGCG on 24 h energy expenditure and fat oxidation. Fourteen subjects took part to this randomized, placebo-controlled, double-blind, cross-over study. Each subject was tested five times in a metabolic chamber to measure 24 h energy expenditure, substrate oxidation and blood pressure. During each stay, the subjects ingested a capsule of placebo or capsules containing 200 mg caffeine and a variable dose of EGCG (90, 200, 300 or 400 mg) three times daily, 30 min before standardized meals. Twenty-four hour energy expenditure increased significantly by about 750 kJ with all EGCG–caffeine mixtures compared with placebo. No effect of the EGCG–caffeine mixture was observed for lipid oxidation. Systolic and diastolic blood pressure increased by about 7 and 5 mmHg, respectively, with the EGCG–caffeine mixtures compared with placebo. This increase was significant only for 24 h diastolic blood pressure. The main finding of the study was the increase in 24 h energy expenditure with the EGCG–caffeine mixtures. However, this increase was similar with all doses of EGCG in the mixtures.

Anthropometric parameters and catch-up growth were prospectively evaluated in fifty late-diagnosed children with coeliac disease aged 2·25–10 years after 1–4 years of adhering to a strict gluten-free diet (GFD). The anthropometric parameters were expressed as Z scores relative to National Centre for Health Statistics standards using Epi Info 2000 (weight-for-height Z score (WHZ) and height-for-age Z score (HAZ)). Catch-up growth was evaluated by repeated measures. ANOVA, overall significance by an F test and pair-wise comparisons for estimated marginal means using the least significant difference. At the time of enrolment, no significant difference was observed in WHZ and HAZ between children diagnosed before (group 1) or after (group 2) 4 years of age. On follow-up, HAZ was significantly higher in group 1 after the first and third years of the GFD (P=0·04 and 0·02, respectively), with a non-significant increase after completing 4 years of the GFD (P=0·22). Feeding the GFD resulted in an overall significant (F=3·99, P=0·011) increase in HAZ up to 4 years of follow-up. However, the catch-up in height was incomplete, with stunting in sixteen (55·4 %) of twenty-nine children after 3 years and in seven (46·6 %) of fifteen children after 4 years on the GFD. Pair-wise comparisons demonstrated a linear catch-up growth during the initial follow-up on GFD. Treatment with the GFD did not result in an overall significant increase in WHZ up to 4 years of follow-up (F=1·01, P=0·42). Our results suggest that, in children with late-diagnosed coeliac disease, treatment with a GFD leads to a normalisation of body mass and a significant but incomplete recovery in HAZ during 4 years of follow-up.

Phytosterols contain an unsaturated ring structure and therefore are susceptible to oxidation under certain conditions. Whilst the cytotoxicity of the analogous cholesterol oxidation products (COP) has been well documented, the biological effects of phytosterol oxidation products (POP)have not yet been fully ascertained. The objective of the present study was to examine the cytotoxicity of β-sitosterol oxides and their corresponding COP in a human monocytic cell line (U937), a colonic adenocarcinoma cell line (CaCo-2) and a hepatoma liver cell line (HepG2). 7β-Hydroxysitosterol, 7-ketositosterol, sitosterol-3β,5α,6β-triol and a sitosterol-5α,6α-epoxide–sitosterol-5β,6β-epoxide (6:1) mixture were found to be cytotoxic to all three cell lines employed; the mode of cell death was by apoptosis in the U937 cell line and necrosis in the CaCo-2 and HepG2 cells. 7β-Hydroxysitosterol was the only β-sitosterol oxide to cause depletion in glutathione, indicating that POP-induced apoptosis may not be dependent on the generation of an oxidative stress. A further objective of this study was to assess the ability of the antioxidants α-tocopherol, γ-tocopherol and β-carotene to modulate POP-induced cytotoxicity in U937 cells. Whilst α/γ-tocopherol protected against 7β-hydroxycholesterol-induced apoptosis, they did not confer protection against 7β-hydroxysitosterol-or 7-ketositosterol-induced toxicity, indicating that perhaps COP provoke different apoptotic pathways than POP. β-Carotene did not protect against COP- or POP-induced toxicity. In general, results indicate that POP have qualitatively similar toxic effects to COP. However, higher concentrations of POP are required to elicit comparable levels of toxicity.