Hydroxytyrosol, tyrosol and caffeic acid effects on hydrogen peroxide-induced DNA damage, hydroperoxide generation and redox enzyme gene expression were studied in oxidative-stress-sensitive human prostate cells (PC3). Hydroxytyrosol led to lower levels of hydroperoxides, DNA damage, and mRNA levels of classic glutathione peroxidase (GPx) for all the studied concentrations. Only hydroxytyrosol was effective at low concentrations (10 μM). TYROSOL REDUCED DNA OXIDATION ONLY AT HIGH (>50 Μm) concentrations and increased hydroperoxides, GPx and phospholipid hydroperoxide GPx mRNA levels. Caffeic acid elicited effects between those of the other two phenolics. Results indicate that hydroxytyrosol is the only significant antioxidant phenolic in olive oil and may be the major component accounting for its beneficial properties. Tyrosol appeared to exhibit pro-oxidant effects (only at high concentrations) and caffeic acid was neutral. Both number and position of hydroxyl groups appear to play a role in the cellular effects of hydroxytyrosol.

Our knowledge on the absorption of folate is incomplete. The deconjugation process as a possible limiting factor in the absorption of folates was investigated. The study also attempted to validate the use of the area under the serum response curve (AUC) from food compared with folic acid as a proxy variable for food folate bioavailability. Folate absorption was determined in healthy ileostomy volunteers (n 11) using a single-dose short-term protocol. In a randomised crossover design, volunteers received spinach meals and a supplement. Based on analysis of test meals and ileostomy effluents, there was no difference in folate absorption between spinach with a mono-:polyglutamate ratio 40:60 and the same spinach with a 100:0 ratio. The absolute absorption of spinach folate (79 %) calculated from the difference between folate intake and folate content of ileostomy effluents was approximately equal to the relative absorption (81 %) calculated from the AUC after consumption of spinach meals in relation to the AUC after consumption of the folic acid supplement. We conclude that the deconjugation process is not a limiting factor in the absorption of spinach folates. Comparison of AUC of food folate v. folic acid in a short-term protocol may be suitable for assessing food folate bioavailability.

Conjugated linoleic acid (CLA) refers to a group of positional and geometric isomers of linoleic acid. Studies using animal models have shown that CLA reduces adiposity, improves plasma lipoprotein metabolism and insulin sensitivity and reduces arteriosclerosis. Whilst CLA may have therapeutic potential with regard to coronary artery disease risk factors in human subjects, there has been little investigation into its effects in human subjects. This current study investigated the effects of dietary supplementation using two isomeric blends of CLA on triacylglycerol (TAG)-rich lipoprotein metabolism and reverse cholesterol transport in human subjects and evaluates whether CLA modulated cardiovascular disease risk factors. Fifty-one normolipidaemic subjects participated in this randomised double-blind placebo-controlled intervention trial. Subjects were randomly assigned to receive 3 g cis-9,trans-11–trans-10,cis-12 isomeric blend (50: 50) or a cis-9,trans-11–trans-10,cis-12 isomeric blend (80: 20) CLA or linoleic acid (control)/d for 8 weeks. The 50: 50 CLA isomer blend significantly reduced (P≤0·005) fasting plasma TAG concentrations. The 80: 20 CLA isomer blend significantly reduced (P≤0·05) VLDL-cholesterol concentrations. CLA supplementation had no significant effect on LDL-cholesterol, HDL-lipid-protein composition or reverse cholesterol transport. CLA supplementation had no effect on body weight, plasma glucose and insulin concentrations. Fatty acid analysis revealed that the cis-9,trans-11 CLA isomer was incorporated into total plasma lipids following supplementation with both isomeric blends of CLA. The present study demonstrates that CLA supplementation significantly improves plasma TAG and VLDL metabolism in human subjects. The study confirms that some of the cardio-protective effects of CLA that were shown in animal studies are relevant to man.

The present experiment aimed to determine the effects of supplements of folic acid (FA) alone or in combination with vitamin B12 on folate and homocysteine metabolism in gestating nulliparous Yorkshire–Landrace (YL) and multiparous Landrace (LD) occidental sows and multiparous Chinese Meishan–Landrace (ML) sows. LD sows were randomly assigned to two treatments: 0 or 15 mg FA/kg diet while YL and ML sows were assigned to three treatments: 0 mg FA/kg diet, 15 mg FA/kg or 15 mg FA+160 μg vitamin B12/kg diet. Supplements were given from the oestrus preceding insemination up to slaughter on day 15 of gestation. At slaughter, a uterine flush was collected to determine uterine contents of homocysteine, methionine, tetrahydrofolate (THF), 5-methyl-THF, pyridoxal 5-phosphate (P5P) and vitamin B12. Blood samples were taken at first oestrus, at insemination and on days 5, 10 and 15 of gestation to determine plasma concentrations of homocysteine, methionine, THF, 5-methyl-THF, P5P, vitamin B12 and relative total folate-binding capacity. In occidental sows (YL and LD), the FA supplement tended to decrease uterine flush content of homocysteine (P=0·06) and concentrations of plasma homocysteine (P=0·09). Nulliparous YL sows had lower concentrations of plasma homocysteine, methionine, THF and 5-methyl-THF (P<0·05) than multiparous LD sows. Multiparous ML and LD sows had similar concentrations of plasma THF, 5-methyl-THF, methionine and vitamin B12, but ML sows had lower concentrations of plasma homocysteine (P<0·05). The vitamin B12 supplement increased concentrations of plasma vitamin B12 (P<0·05) both in multiparous ML and nulliparous YL sows, but had no effect on the composition of either uterine flush or plasma. The present results showed also that sows had a low vitamin B12 status (<200 pg/ml) and high circulating homocysteine levels (>15 μM) during the first 15 d of gestation. Furthermore, the vitamin B12 content in uterine secretions represented between 180 and 300 % of the total content in plasma. The low plasma concentrations of homocysteine in multiparous ML sows suggest a more efficient remethylation pathway which may not be dependent upon dietary supply of FA or vitamin B12. In nulliparous YL sows, low concentrations of both homocysteine and methionine suggest that the methionine requirement for protein deposition might have reduced the amount of methionine available for the methylation pathway. The results of the present experiment suggest that the reduction of uterine homocysteine may be an important aspect of the role of FA supplement on the uterine environment in occidental sows. The presence of high levels of vitamin B12 in uterine secretions merits further investigation in relation to embryonic development.

Free radicals and reactive species produced in vivo can trigger cell damage and DNA modifications resulting in carcinogenesis. Dietary antioxidants trap these species limiting their damage. The present study evaluated the role of vitamins C and E in the prevention of potentially premalignant modifications to DNA in the human stomach by supplementing patients who, because of hypochlorhydria and possible depletion of gastric antioxidants, could be at increased risk of gastric cancer. Patients undergoing surveillance for Barrett's oesophagus (n 100), on long-term proton pump inhibitors were randomized into two groups: vitamin C (500 mg twice/d) and vitamin E (100 mg twice/d) for 12 weeks (the supplemented group) or placebo. Those attending for subsequent endoscopy had gastric juice, plasma and mucosal measurements of vitamin levels and markers of DNA damage. Seventy-two patients completed the study. Plasma ascorbic acid, total vitamin C and vitamin E were elevated in the supplemented group consistent with compliance. Gastric juice ascorbic acid and total vitamin C levels were raised significantly in the supplemented group (P=0·01) but supplementation had no effect on the mucosal level of this vitamin. However, gastric juice ascorbic acid and total vitamin C were within normal ranges in the unsupplemented group. Mucosal malondialdehyde, chemiluminescence and DNA damage levels in the comet assay were unaffected by vitamin supplementation. In conclusion, supplementation does not affect DNA damage in this group of patients. This is probably because long-term inhibition of the gastric proton pump alone does not affect gastric juice ascorbate and therefore does not increase the theoretical risk of gastric cancer because of antioxidant depletion.

Tobacco and alcohol consumption are strongly related to other cardiovascular and cancer risk factors. The aim of the present study was to analyse the association of nutrient intake, blood lipid variables and leisure-time physical activity with tobacco and alcohol consumption status. Participants were recruited in a cross-sectional population-based survey, including cardiovascular risk factor measurements and evaluation of physical activity and diet intake in a Mediterranean population (n 1748). Multiple linear regression analysis, adjusted for several confounders, showed a direct association of saturated fatty acids (g and % total energy intake), dietary cholesterol intakes and serum triacylglycerol with smoking. An inverse association was observed for smoking and unsaturated fatty acids (% energy intake), vitamin C, α-tocopherol and β-carotene intakes, leisure-time physical activity and HDL-cholesterol. These associations were not observed for alcohol drinking. After adjusting for the confounders earlier mentioned, low dietary intakes of vitamin C and dietary fibre were more likely in heavy-smokers as compared with non-smokers (odds ratio 1·74 (95 % CI 1·07, 2·73) and 1·94 (95 % CI 1·29, 2·92) of low vitamin C (<60 mg/d) and dietary fibre intakes (<10 g/d) respectively). Alcohol consumption was directly associated with HDL-cholesterol and triacylglycerol, and attenuated the effects of smoking on HDL-cholesterol. These results suggest that the dietary intake of fibre and several antioxidant components of the Mediterranean diet is reduced in smokers, who also show an adverse lipid profile. However, the worst triacylglycerol levels are associated with the combination of heavy smoking and heavy alcohol drinking. Moderate alcohol consumption was not associated with an unhealthy diet pattern or adverse lipid profile. The health benefits of the Mediterranean diet appear to be strongly counteracted by smoking.

We have reported that blood pressure was elevated in 3-month-old rats whose mothers were Fe-restricted during pregnancy. These animals also had improved glucose tolerance and decreased serum triacylglycerol. The aim of the present study was to determine whether these effects of maternal nutritional restriction, present in these animals at 3 months of age, can be observed in the same animals in later life. Pulmonary and serum angiotensin converting enzyme (ACE) concentrations were also measured to investigate whether the renin–angiotensin system was involved in the elevation of blood pressure observed in the offspring of Fe-restricted dams. Systolic blood pressure was higher in the offspring of Fe-restricted dams at 16 months of age. Heart and kidney weight were increased as a proportion of body weight in the offspring of Fe-restricted dams. The pulmonary ACE concentration was not significantly different between the groups. The serum ACE concentration was significantly elevated in the offspring of Fe-restricted dams at 3 but not 14 months of age. There was a strong correlation between serum ACE levels at 3 and 14 months of age. Glucose tolerance and serum insulin were not different between the maternal diet groups. Serum triacylglycerol tended to be lower in the offspring of Fe-restricted dams. There were no differences in serum non-esterified fatty acids or serum cholesterol between the maternal diet groups. This study provides further evidence that maternal nutrition has effects on the offspring that persist throughout life. At 16 months of age, the elevation of blood pressure in Fe-restricted offspring does not appear to be mediated via changes in ACE levels. Both cardiac hypertrophy and decreased serum triacylglycerol have also been observed in Fe-restricted fetuses, suggesting that these changes may be initiated in utero.

We have tested the effect of dietary fatty acids on aspects of innate and specific adaptive T helper (Th) 1- and Th2-driven immune responses in a murine sensitisation model using dinitrochlorobenzene as sensitiser. Six groups of fifteen BALB/c mice were fed diets containing 30 % fat (by energy) for 8 weeks. Diets were rich in saturated fatty acids, n-6 polyunsaturated fatty acid (PUFA), or n-3 PUFA, each at a sufficient (11, 35 and 68 mg/kg) and a supplemented vitamin E level (1028, 1031 and 1030 mg/kg respectively). Feeding n-6 PUFA marginally decreased % phagocytosing cells at the low vitamin E level, but had no other effects on immune function. The n-3 PUFA diets decreased production of prostaglandin E2 while increasing oxidative burst and tumour necrosis factor α production. In addition adaptive Th1-driven responses (immunoglobulin, Ig)G2a, IgG2b, interferon-γ:interleukin 4) were decreased, whereas Th2-driven and mucosal immune responses were increased (IgE) or unaffected (IgG1, IgA). Combination with high levels of α-tocopherol did not affect the reduced prostaglandin E2 production, augmented the increase of tumour necrosis factor α production and tended to ameliorate the selective suppressive effects of n-3 PUFA on certain Th1-driven effects (interferon-γ:interleukin 4 ratio and IgG2a levels). We conclude that the sensitisation model appears useful for application in nutrition research. It allows a broad assessment of the effects of dietary intervention on various aspects of immune responsiveness, and as such provides a valuable model to assess, characterise and rank effects of foods and/or nutrients on a range of immune functions, including Th1–Th2 polarisation.

Early postnatal overnutrition (PNO) induced by restricting litter size in rats leads to increased body-weight (BW) and body-fat gain in later life. PNO rats are used as an animal model of moderate obesity and early hyperinsulinism. We investigated whether the increased adiposity could be due to a decreased energy expenditure. Male newborn Wistar rats were raised in litters of either two (SL) or twelve pups (NL), weaned at 4 weeks of age and subsequently fed ad libitum. BW was recorded continuously until 12 weeks of age. Daily energy intake, total daily energy expenditure (EE, measured by indirect calorimetry) and body composition were measured in weaned pups at 5, 8 and 12 weeks of age. SL rats displayed increased BW compared with NL rats from week 2 to 5 and again from week 10 to 12. Lean body mass, body fat and protein content and total EE were increased in SL rats at week 5. The same linear correlation described the relationship between BW and total EE in NL and SL rats. At week 8 to 12 no differences in energy metabolism could be found, but the total fat content was increased in SL rats at week 12. Energy balance, i.e. assimilated energy minus EE, was no different between SL and NL at any time that it was measured. We conclude that although PNO rats display increased adiposity in early life, there seem to be no long-lasting effects on energy metabolism in later life, even if a tendency to increased adiposity can still be detected.

The fasting concentration of non-esterified fatty acids (NEFA) and the degree to which it declines during an oral glucose tolerance test are closely associated with insulin resistance and glucose intolerance. However, relatively few studies have described possible environmental determinants of NEFA concentrations. Physical activity is likely to be related to NEFA levels, but habitual activity level is difficult to quantify in epidemiological studies. In particular, it is unclear whether NEFA is more closely related to cardio-respiratory fitness or to habitual energy expenditure. In order to quantify these relationships, we analysed data from the Ely prospective population-based study in which 931 subjects underwent a glucose tolerance test with measurements of cardio-respiratory fitness and 4 d energy expenditure by heart-rate monitoring, a technique previously validated against whole-body calorimetry and doubly-labelled water. In order to estimate the latent variables of usual fitness and energy expenditure, a subset of 190 subjects underwent repeat testing on three further occasions over 1 year. In analyses adjusting only for age and sex, energy expenditure and cardio-respiratory fitness were both negatively correlated with the total area under the NEFA curve following the oral glucose load (standardised β coefficients -0·030 and -0·039 respectively; both P<0·001) However, further adjustment for degree of obesity and bivariate measurement error suggested that the effect of energy expenditure was significantly greater than that for fitness (-0·047 and -0·005 respectively). These results suggest that the area under the NEFA curve in the oral glucose tolerance test, a measure of insulin sensitivity, is strongly associated with the habitual level of physical activity.

The effectiveness of a health and nutrition education programme, in changing certain chronic disease risk factors, was assessed after the 6 years intervention period was completed. The school-based intervention programme was applied to all children registered in the first grade (age 5·5–6·5 years) in 1992 in two counties of Crete, while the children from a third county served as a control group. In order to assess the effectiveness of the intervention, a variety of biological and behavioural parameters were measured before and following completion of the intervention in a randomly selected school-based sample of 602 intervention group (IG) and 444 control group (CG) pupils. At the end of the 6-year period, it was found that biochemical indices generally improved significantly more in the IG compared with the CG (mean change for IG v. CG was -0·27 v. -0·12 mmol/l for total cholesterol (TC); -0·07 v. +0·24 for TC:HDL and -0·13 v. +0·14 for LDL:HDL). Similarly, the changes observed in the anthropometric variables in the two groups were in favour of the IG (+3·68 v. +4·28 kg/m2 for BMI; +2·97 v. +4·47 mm for biceps skinfold). Total energy intake and consumption of total fat and saturated fat increased significantly less in the IG compared with the CG (+747·7 v. 1534·7 kJ (+178·7 v. +366·8 kcal); +5·9 v. +18·8 g and +0·8 v. +5·1 g respectively), while time devoted to leisure time physical activity and cardiovascular run test performance increased significantly more in the IG (+281 v. +174 min/week and +2·5 v. +1·2 stages respectively). The findings of the present study underline the importance of such programmes in health promotion and disease prevention. Although the long-term effects of these programmes can only be assessed by tracking this population through to adolescence and adulthood, these programmes seem to have the potential to lead to a healthier lifestyle and thus a reduction in risk factor levels.

This study estimated total body water (TBW) in four groups (twelve per group; sedentary and highly trained men and women) at the time of 2H dosing (T0) and after a 3·5 h equilibration period (Teq). Standard TBW calculations were employed at T0 (no correction for disproportionate urinary tracer loss) and Teq (correction for urinary tracer loss only), plus those calculations that corrected for a disproportionate urinary tracer loss and insensible tracer loss respectively. The measurement of body density enabled the four TBW estimates to be compared for the determination of three-compartment % body fat (BF). The very small difference between the standard and corrected T0 TBW data was not significant (P=0·914) and no Group×TBW interaction was identified (P=0·125). These results reflect the closeness of the 2H concentration in the urine produced during the equilibration period and the Teq saliva samples. The associated mean % BF values were essentially identical. Although correcting for insensible 2H losses in addition to urinary losses at Teq produced a statistically significant (P<0·001) lower mean TBW (about 200 g) than the standard calculation, this translated to a small difference in % BF (0·3). The larger difference (about 500 g, P<0·001) between the two (T0, Teq) corrected TBW calculations was also associated with a small body composition difference (0·1 % BF), which was less than the propagated error (0·3 % BF) for the three-compartment body composition model. Corrections to the standard calculations of TBW at T0 and Teq for a protocol employing a brief equilibration period (3·5 h) were therefore of marginal use for improving the accuracy of % BF estimates. The TBW difference over time (T0v. Teq) also had little impact on % BF values.