Convergent validity, ceiling, and floor effects of the English-ISYQOL against established quality of life questionnaires (SRS-22r and SAQ) and curve angles in adolescents with idiopathic scoliosis.

Scoliosis significantly impacts Quality of Life (QOL). Current quality of life questionnaires for adolescents with idiopathic scoliosis (AIS) have limitations. A new questionnaire for measuring QOL in AIS called the Italian Spine Youth Quality of Life (ISYQOL) has been developed to address these limitations but the English translation has not yet been validated. To determine the ceiling and floor effects, and the convergent validity of the ISYQOL questionnaire against established QOL questionnaires and Cobb angle in AIS. One hundred consecutive females with AIS, (10-18 years old), treated non-operatively. The English translation of the ISYQOL was compared to the following established questionnaires: Scoliosis Research Society-22r and the Spinal Appearance Questionnaire. The participants were 100 females (13.89+/-1.8 years) with 28.75+/-13.9° curve angles. The convergent validity of the ISYQOL score (60.3+/-12.44) was supported by significant correlation with the SRS-22r total score, function, pain, self-image, and mental health scores (r = 0.70, 0.54, 0.57, 0.52 and 0.50, respectively), and with the SAQ general, waist, and expectations domains (r = -0.6. -.52, and -0.56, respectively). Correlation with the Cobb angle was (r = -.37)(see Table 1). No ceiling effect was observed in the ISYQOL. Ceiling effects were observed for the SRS-22r and the SAQ. The ISYQOL demonstrated evidence of convergent validity. This study supports its suitability for QOL research in AIS. ISYQOL appears more likely to detect changes in evaluative studies than the SRS- 22r and the SAQ.

Single oral doses of 13C forms of pteroylmonoglutamic acid and 5-formyltetrahydrofolic acid elicit differences in short-term kinetics of labelled and unlabelled folates in plasma: potential problems in interpretation of folate bioavailability studies.

Single (13)C6-labelled doses of pteroylmonoglutamic acid (PteGlu; 634 nmol) or 5-formyltetrahydrofolic acid (431-569 nmol) were given to fasted adult volunteers, and the rise in total and (13)C-labelled plasma 5-methyltetrahydrofolic acid metabolite monitored over 8 h by HPLC and liquid chromatography-MS. The dose-adjusted area under the curve (AUC) for total (labelled plus unlabelled) plasma 5-methyltetrahydrofolic acid following a 5-formyltetrahydrofolic acid test dose was 155 % that obtained following a PteGlu test dose. Surprisingly, an average 60 and 40 % of the total plasma 5-methyltetrahydrofolic acid response to [(13)C6]PteGlu and [(13)C6]5-formyltetrahydrofolic acid, respectively, was unlabelled; an observation never before reported. Short-term kinetics of plasma [(13)C6]5-methyltetrahydrofolic acid showed a slower initial rate of increase in plasma concentration and longer time to peak following an oral dose of [(13)C6]PteGlu compared with that for an oral dose of [(13)C6]5-formyltetrahydrofolic acid, while the [(13)C6]5-methyltetrahydrofolic acid AUC for [(13)C6]5-formyltetrahydrofolic acid was 221 % that for [(13)C6]PteGlu. These data indicate that PteGlu and 5-formyltetrahydrofolic acid, which are thought to be well absorbed (about 90 %) at physiological doses, exhibit dramatically different rates and patterns of plasma response. A limitation in the rate of reduction of PteGlu before methylation could result in slower mucosal transfer of [(13)C6]5-methyltetrahydrofolic acid derived from [(13)C6]PteGlu into the plasma. This, when coupled with an observed similar plasma clearance rate for [(13)C6]5-methyltetrahydrofolic acid metabolite derived from either folate test dose, would yield a comparatively smaller AUC. These findings suggest potential problems in interpretation of absorption studies using unlabelled or labelled folates where the rate of increase, the maximum increase, or the AUC, of plasma folate is employed for test foods (mainly reduced folates) v. a 'reference dose' of PteGlu.

Increased DNA oxidative susceptibility without increased plasma LDL oxidizability in Type II diabetes: effects of alpha-tocopherol supplementation.

In vivo supplementation studies of the antioxidant alpha-tocopherol in human Type II diabetes have used surrogate, rather than direct, markers of oxidative damage/antioxidant protection and have used higher doses of alpha-tocopherol than used in coronary secondary prevention trials. We tested the hypothesis that oral alpha-tocopherol in a dosage regimen used in secondary prevention trials would reduce directly observed oxidatively induced single-strand breaks in lymphocyte DNA in Type II diabetes. We studied 40 people with Type II diabetes and 30 controls in a randomized, double-blind, placebo-controlled trial of 400 i.u. of oral alpha-tocopherol daily for 8 weeks. Lymphocyte DNA single-strand breaks and low-density lipoprotein (LDL) particle size and oxidizability were measured at baseline, after 8 weeks, and after 4 weeks washout. Polymorphisms in the gene for the antioxidant enzyme paraoxonase-1 gene (position 192) were measured. The diabetics had increased DNA oxidative susceptibility (P=0.008), without increased LDL oxidative susceptibility. There was a direct relationship between DNA oxidative susceptibility and baseline plasma alpha-tocopherol in the diabetes group alone (r=0.421, r(2)=0.177 and P=0.023), but DNA and LDL oxidative susceptibility were not influenced by alpha-tocopherol supplementation in either group in this regimen. Paraoxonase-1 gene polymorphisms did not contribute to LDL or DNA oxidative susceptibility or response to alpha-tocopherol. Increased DNA oxidative susceptibility, therefore, can occur in Type II diabetes without increased LDL oxidative susceptibility, but alpha-tocopherol supplementation in this regimen has no influence on DNA or LDL oxidative susceptibility in Type II diabetes or controls. Polymorphisms in the paraoxonase gene (position 192) are not associated with differences in oxidative susceptibility or responses to alpha-tocopherol.

No significant effects of lutein, lycopene or beta-carotene supplementation on biological markers of oxidative stress and LDL oxidizability in healthy adult subjects.

OBJECTIVE: The objective of this study was to determine the effect of individual carotenoid supplementation on biochemical indices of oxidative status in apparently healthy adult males. METHODS: The study was a placebo controlled single blind study. Healthy male volunteers (n= 175) were assigned to four groups. They received daily supplements of beta-carotene (15 mg), lutein (15 mg), lycopene (15 mg) and placebo for three months. The effects of the supplementation on antioxidant status were monitored by plasma carotenoid, vitamin C and A levels, glutathione (GSH and GSSG) concentrations, protein SH groups. erythrocyte antioxidant enzyme activities (Cu-Zn SOD, Se-GSH-Px) and susceptibility of LDL to copper-induced oxidation. RESULTS: beta-carotene, lycopene and lutein supplementation led to significant plasma and LDL increases in each of these carotenoids, without modifications of other carotenoid levels in plasma or in LDL. The supplementation failed to enhance the resistance of LDL to oxidation or to modify the LDL polyunsaturated/ saturated fatty acid ratio. Vitamin C, GSH, protein SH groups and antioxidant metalloenzyme activities were also unchanged. CONCLUSION: We did not observe beneficial or adverse effects of lutein, lycopene or beta-carotene supplementation on biomarkers of oxidative stress. In apparently healthy subjects, carotenoid supplementation does not lead to significantly measurable improvement in antioxidant defenses.

A European carotenoid database to assess carotenoid intakes and its use in a five-country comparative study.

A food frequency questionnaire (FFQ) and carotenoid database with information on alpha- and beta-carotene, lutein, lycopene and beta-cryptoxanthin was prepared and used to compare the carotenoid intakes in five European countries: UK, Republic of Ireland, Spain, France and The Netherlands. Eighty, age- (25-45 years) and sex-matched volunteers were recruited in each of the five countries. A FFQ and carotenoid database was prepared of the most commonly consumed carotenoid rich foods in the participating countries and the information was used to calculate frequency and intake of carotenoid-rich foods. The median total carotenoid intake based on the sum of the five carotenoids, was significantly higher (P < 0.05) in France (16.1 mg/day) and lower in Spain (9.5 mg/day,) than the other countries, where the average intake was approximately 14 mg/day. Comparison of dietary source of carotenoids showed that carrots were the major source of beta-carotene in all countries except Spain where spinach was most important. Likewise, carrots were also the main source of alpha-carotene. Tomato or tomato products, were the major source of lycopene. Lutein was mainly obtained from peas in Republic of Ireland and the UK, however, spinach was found to be the major source in other countries. In all countries, beta-cryptoxanthin was primarily obtained from citrus fruit. Comparing the data with that from specific European country studies suggests that the FFQ and carotenoid database described in the present paper can be used for comparative dietary intake studies within Europe. The results show that within Europe there are differences in the specific intake of some carotenoids which are related to different foods consumed by people in different countries.

Serum concentrations of carotenoids and vitamins A, E, and C in control subjects from five European countries.

High intakes of fruits and vegetables, or high circulating levels of their biomarkers (carotenoids, vitamins C and E), have been associated with a relatively low incidence of cardiovascular disease, cataract and cancer. Exposure to a high fruit and vegetable diet increases antioxidant concentrations in blood and body tissues, and potentially protects against oxidative damage to cells and tissues. This paper describes blood concentrations of carotenoids, tocopherols, ascorbic acid and retinol in well-defined groups of healthy, non-smokers, aged 25-45 years, 175 men and 174 women from five European countries (France, UK (Northern Ireland), Republic of Ireland, The Netherlands and Spain). Analysis was centralised and performed within 18 months. Within-gender, vitamin C showed no significant differences between centres. Females in France, Republic of Ireland and Spain had significantly higher plasma vitamin C concentrations than their male counterparts. Serum retinol and alpha-tocopherol levels were similar between centres, but gamma-tocopherol showed a great variability being the lowest in Spain and France, and the highest in The Netherlands. The provitamin A: non-provitamin A carotenoid ratio was similar among countries, whereas the xanthophylls (lutein, zeaxanthin, beta-cryptoxanthin) to carotenes (alpha-carotene, beta-carotene, lycopene) ratio was double in southern (Spain) compared to the northern areas (Northern Ireland and Republic of Ireland). Serum concentrations of lutein and zeaxanthin were highest in France and Spain; beta-cryptoxanthin was highest in Spain and The Netherlands; trans-lycopene tended to be highest in Irish males and lowest in Spanish males; alpha-carotene and beta-carotene were higher in the French volunteers. Due to the study design, the concentrations of carotenoids and vitamins A, C and E represent physiological ranges achievable by dietary means and may be considered as 'reference values' in serum of healthy, non-smoking middle-aged subjects from five European countries. The results suggest that lutein (and zeaxanthin), beta-cryptoxanthin, total xanthophylls and gamma-tocopherol (and alpha- : gamma-tocopherol) may be important markers related to the healthy or protective effects of the Mediterranean-like diet.

Increased fruit and vegetable consumption: potential health benefits.

In this European Union project, a Core human study was conducted in Ireland, Northern Ireland, Spain, France and The Netherlands. Oxidative and antioxidant status, vegetable and fruit consumption, and carotenoid intake of volunteers from different countries were compared. Response to increased carotenoid intake was determined. Attention was paid to whether the antioxidant capability of beta-carotene, lutein and lycopene demonstrated in vitro was apparent in relation to increased oxidation resistance of low-density-lipoprotein (LDL). Other (complementary) studies were undertaken and included determination of: protective effects of carotenoid-rich foods against LDL and DNA oxidative damage; carotenoid absorbability; barriers to increased vegetable consumption; and carotenoid content of fruits and vegetables frequently consumed in Europe. Our results demonstrated that carotenoid supplementation did not increase LDL oxidation resistance. However, increased consumption of carotenoid-rich fruits and vegetables did increase LDL oxidation resistance, and higher plasma concentration of total and specific carotenoids (pre-supplementation) was associated with lower DNA damage.

Erythrocyte folate analysis: saponin added during lysis of whole blood can increase apparent folate concentrations, depending on hemolysate pH.

BACKGROUND: The analysis of red cell folate (RCF) depends on complete hemolysis of erythrocytes, and it is assumed that complete hemolysis is achieved by 10-fold dilution of whole blood with hypotonic solutions of 10 g/L ascorbic acid/ascorbate. This report challenges this assumption. METHODS: The conventional method of erythrocyte lysis was modified to include saponin, a known effective hemolyzing agent. The influence of saponin was determined at various lysate pHs, using the microbiological (Lactobacillus rhamnosus) folate assay. The effect of saponin during lysate preparation was subsequently compared with either the effect of 30 s of sonication or a single 1-h freeze-thaw cycle. RESULTS: Saponin addition was found to increase assayable RCF up to ninefold, depending on lysate pH. Sonication of lysates had no effect, and freezing-thawing lysates once did not always guarantee complete hemolysis. Lysates created with 10 g/L ascorbic acid (a historically widely used diluent) without pH adjustment produced assayable folate concentrations significantly lower than optimal. CONCLUSIONS: A lysing agent should be incorporated into RCF assays to guarantee complete hemolysis. Ten-fold dilution of blood with 10 g/L ascorbic acid, without pH adjustment, produces lysates with pHs (pH 4.0) below the point (pH 4.7) at which hemoglobin can denature irreversibly. The optimum pH for hemolysates is approximately 5.0.

Influence of increased fruit and vegetable intake on plasma and lipoprotein carotenoids and LDL oxidation in smokers and nonsmokers.

BACKGROUND: Epidemiological studies suggest a cardioprotective role for carotenoid-rich foods. Smokers have a high risk of cardiovascular disease and low dietary intake and plasma concentrations of carotenoids. The aim of this study was to determine the carotenoid response of smokers and nonsmokers to increased intake of 300-400 g of vegetables and its effect on LDL oxidation. METHODS: After a depletion period of 8 days, 34 healthy females (18 nonsmokers, 16 smokers) were supplemented with beta-carotene- and lutein-rich (green) and lycopene-rich (red) vegetable foods, each for 7 days. RESULTS: Baseline concentrations (mean +/- SD) of plasma beta-carotene (0.203+/-0.28 micromol/L vs. 0.412+/-0.34 micromol/L; P <0.005) and lutein (0.180 +/-0.10 vs. 0.242+/-0.11 micromol/L; P<0.05) but not lycopene (0.296+/-0.10 vs. 0.319+/-0.33 micromol/L) were significantly lower in smokers compared with nonsmokers. After supplementation, the change (supplementation minus depletion) in plasma beta-carotene (0.152+/- 0.43 vs. 0.363+/-0.29 micromol/L in smokers vs. nonsmokers; P = 0.002) and LDL lutein (0.015+/-0.03 vs. 0.029+/-0.03 micromol/mmol cholesterol; P = 0.01) was significantly lower in smokers than nonsmokers. Green-vegetable supplementation had no effect on the resistance of LDL to oxidation (lag-phase) in either group. After red-vegetable supplementation, plasma and LDL lycopene concentrations were increased in both groups, but only nonsmokers showed a significant increase in the lag-phase (44.9+/-9.5 min at baseline, 41.4+/-6.5 min after depletion, and 49.0+/-8.9 min after supplementation; P<0.01) compared with depletion. CONCLUSIONS: In this short-term intervention study, a dietary intake of >40 mg/day of lycopene by a group of nonsmoking individuals significantly reduced the susceptibility of LDL to oxidation, whereas an equivalent increase in lycopene by a group of smokers showed no such effect.

Measurement of cellular repair activities for oxidative DNA damage.

Two related assays capable of determining cell extract repair activities for different oxidative lesions in DNA are described. Both assays measure the incorporation of radiolabeled nucleotides during repair of an oxidatively damaged template in a cell-free system. The assays differ in the type of oxidative damage present in the DNA. In one, singlet oxygen is used to generate predominantly 8-oxo-2'-deoxyguanosine lesions. In the other, hydroxyl radicals are used to generate a broad spectrum of damage including oxidized bases and strand breaks. Assay conditions were adjusted to ensure that radiolabel incorporation was directly proportional to cell extract repair activity. These assays represent sensitive tools for investigating the regulation of repair systems for oxidative DNA damage.

Effects of lycopene and lutein supplementation on the expression of functionally associated surface molecules on blood monocytes from healthy male nonsmokers.

It has been suggested that dietary carotenoids can enhance immune function. Supplementation with beta-carotene (15 mg daily) was previously shown to enhance human monocyte function. To examine the effect of other dietary carotenoids, two similar independent studies were done. Healthy adult male nonsmokers were randomly assigned to receive lycopene (study 1), lutein (study 2), or placebo for 26 days, followed by the alternative treatment for another 26 days. The expression of functionally related monocyte surface molecules was quantified by laser flow cytometry before and after each treatment period. There was a significant increase in plasma levels of each carotenoid following dietary supplementation, but the effects on monocyte surface molecule expression were not as striking as those observed after beta-carotene supplementation. These findings emphasize that it cannot be assumed that the effect of one carotenoid will be the same as another, even at the same level of intake.

Beta-carotene and lycopene, but not lutein, supplementation changes the plasma fatty acid profile of healthy male non-smokers.

Polyunsaturated fatty acids (PUFAs) are highly susceptible to free radical attack. In vitro studies of carotenoids--including beta-carotene, lycopene, and lutein--have shown them to be effective quenchers of singlet oxygen, to have good radical-trapping properties, or to be effective peroxyl radical scavengers (or to have a combination of these qualities). If carotenoids act as antioxidants in vivo, then arguably, plasma PUFA should be conserved. The objective of the current study was to answer the question "Does supplementation with beta-carotene, lycopene, or lutein, at dietarily achievable levels, over a time period known to significantly increase circulating carote concentrations, lead to an observable increase in fasting plasma PUFA?" The normal diets of human volunteers were supplemented with either 15 mg/day beta-carotene (n = 25), lycopene (n = 23), or lutein (n = 21) for 26 days in three independent double-blind, placebo-controlled supplementation studies. Supplementation with beta-carotene increased plasma linoleic acid but left the polyunsaturated:saturated (P:S) fatty acid ratio unaltered. In contrast, supplementation with lycopene reduced linoleic acid, which resulted in a large decrease in the P:S ratio. Lutein supplementation had no effect. It was concluded that neither beta-carotene, lycopene, nor lutein supplementation engender antioxidant effects that lead to the widespread general conservation of plasma PUFAs. Beta-carotene and lycopene supplementation appear to interact with the metabolism of linoleic acid, the "essential" fatty acid, resulting in either an increase (beta-carotene) or decrease (lycopene) in its plasma concentration. Alterations in plasma 18:2 or P:S ratios could ultimately lead to changes in tissue cellular membrane composition and hence to alterations in membrane fluidity and cell-surface protein expression.

Vitamin E supplementation and oxidative damage to DNA and plasma LDL in type 1 diabetes.

OBJECTIVE: To determine the effect of 400 IU/day of the antioxidant vitamin E on the susceptibility of plasma LDL and lymphocyte DNA to oxidative damage in type 1 diabetes. RESEARCH DESIGN AND METHODS: We studied 42 patients with type 1 diabetes and 31 age- and sex-matched control subjects in a randomized prospective double-blind placebo-controlled trial by using 400 IU/day of oral vitamin E for 8 weeks. Measurements were made of single-strand breaks in lymphocyte DNA at baseline and after hydrogen peroxide-induced stress (comet assay) and of copper-induced LDL oxidization and plasma antioxidant profiles. RESULTS: Plasma LDL and lymphocyte DNA were more resistant to induced oxidative change in the type 1 diabetes group than in control subjects. Vitamin E supplementation reduced LDL oxidizability in the control subjects but not in the type 1 diabetes group and had no effect on oxidative DNA damage in either group. The type 1 diabetes group had a significantly poorer plasma antioxidant profile with lower mean serum concentrations of alpha-tocopherol and most carotenoids than control subjects. CONCLUSIONS: Plasma LDL and lymphocyte DNA appear to be more resistant to oxidative change in type 1 diabetic subjects than in control subjects, and there was no evidence of oxidatively induced DNA or LDL change in type 1 diabetes. This study does not support the hypothesis of oxidative damage in these patients, and a dose of vitamin E (400 IU/day) that reduced LDL oxidative susceptibility in control subjects did not do so in patients with type 1 diabetes.

High performance liquid chromatography method for the determination of pyridoxal-5-phosphate in human plasma: how appropriate are cut-off values for vitamin B6 deficiency?

OBJECTIVES: Application of a HPLC (high performance liquid chromatography) method, using cyanide derivatisation, to the determination of plasma pyridoxal-5-phosphate (PLP) concentrations as an indicator of vitamin B6 adequacy. SETTING: The study was performed at the Institute of Food Research, Norwich, UK. Blood samples were taken at the Institute, at Health Centres, or in the volunteer's home. SUBJECTS: 51 adolescent, 131 adult, 68 non-institutionalized elderly and 44 aged (>73 y) volunteers were recruited from local authority schools, local Health Centres and General Practitioners. RESULTS: The mean PLP recovery was 92.8%. The intra- and inter-assay coefficients of variation were 2.8% and 5.2% respectively. Mean PLP concentrations for males and females, respectively, were: adolescents (13-14 y), 36.4 and 43.5 nM; adults (20-64 y), 39.2 and 40.0 nM; elderly (68-73 y), 34.8 and 35.3 nM; aged (>73 y), 57.8 and 49.0 nM. Percentages of subjects with PLP concentrations <34.4 nM were over 26% in all population groups. Mean vitamin B6 intakes (microg/g protein intake), as assessed by weighed dietary records, were all above reference nutrient intakes (15 microg/g protein). CONCLUSIONS: An HPLC method, using cyanide derivitisation, has been applied to the determination of plasma PLP. Comparisons of results for local population groups with current cut-off values for plasma PLP, show large numbers of volunteers at risk of vitamin B6 deficiency although this is not reflected by vitamin B6 intakes calculated from food tables. The 34.4 nM cut-off value for value for plasma PLP, indicating deficiency, is questioned.

Oxidative insult specifically decreases levels of a mitochondrial transcript.

The effects of oxidative insult, applied with hydrogen peroxide, on gene transcript levels in a human lymphocyte cell line (Molt-17) were investigated using mRNA differential display. Several cDNA fragments corresponding to putatively up- or down-regulated transcripts were isolated. One of these was found to hybridize to two discrete transcripts on Northern blots of Molt-17 cell RNA. The more abundant transcript, that has previously been demonstrated to correspond to the mRNA for mitochondrial ATPase subunits 8 and 6, was unaffected by the hydrogen peroxide treatment. In contrast, levels of the rarer, larger transcript were consistently reduced in a rapid, sustained, and dose-dependent manner following hydrogen peroxide treatment. Prior supplementation of the cells with beta carotene provided some protection against the reduction in levels of this transcript following hydrogen peroxide treatment. In contrast, vitamins C and E had no effect at the concentrations tested. We have now cloned the cDNA corresponding to this stress-responsive transcript and demonstrated that it is an incompletely processed product of the mitochondrial genome encompassing ATPase subunits 8 and 6 plus the adjacent gene for cytochrome c oxidase subunit 3. This decrease in one specific mitochondrial transcript may represent a novel mechanism for differential expression of mitochondrially-encoded genes.

Changes in plasma carotenoid and vitamin E profile during supplementation with oil palm fruit carotenoids.

It has been hypothesized that supplementation with a single source of carotenoids causes perturbations in the plasma level of diet-derived carotenoids and that this may explain the lack of association between disease rates and the intake of carotenoid supplements. This article describes the effect of supplementation with an oil palm fruit extract, rich in beta-and alpha-carotene, on the plasma carotenoid profile of 15 healthy women volunteers. Volunteers were supplemented for 35 days with 15 mg/d of total carotenoids. Blood samples were taken at regular intervals during the supplementation period and analyzed for a range of carotenoids. Results indicate that the hydrocarbon carotenoid components of the supplement are absorbed and appear in the plasma disproportionately to the ratios in the supplement and that the plasma concentration of diet-derived lutein, a dihydroxy carotenoid ((3R,3S,6R)-beta,epsilon-carotene-3,3-diol), is depressed, whereas that of lycopene is unaffected. Plasma alpha-tocopherol concentrations were unaffected by supplementation. It is concluded that supplementation with carotenoids from a single source results in plasma carotenoid profile changes that are not predictable from a knowledge of supplement composition and that such changes should be monitored and considered when drawing conclusions as to the effect of carotenoid supplementation on health outcomes.

Oxidative DNA damage measured in human lymphocytes: large differences between sexes and between countries, and correlations with heart disease mortality rates.

The 'antioxidant hypothesis' proposes that vitamin C, vitamin E, carotenoids, and other antioxidants occurring in fruit and vegetables afford protection against heart disease and cancer by preventing oxidative damage to lipids and to DNA, respectively. To test elements of this hypothesis, we have measured blood levels of dietary antioxidants, and 8-oxodeoxyguanosine (8-oxo-dG) concentrations in lymphocyte DNA, in healthy men and women from five European countries: France, Ireland, The Netherlands, Spain, and the U.K. Volunteers, aged 25 45, all nonsmokers, gave blood samples before and after a 12-wk carotenoid supplementation regime. Vitamin C was measured in plasma and vitamin E and carotenoids were measured in serum by high-performance liquid chromatography (HPLC). 8-oxo-dG was assayed by HPLC (with coulometric detection) in DNA isolated from lymphocytes from the same blood samples. Mean values were calculated for groups of volunteers at each sampling time according to country, sex, and supplementation (between 9 and 24 individual samples contributing to each mean). We found that 8-oxo-dG levels in lymphocyte DNA vary significantly according to sex and country. A low mean 8-oxo-dG concentration is seen in DNA of women from all five countries, and of men from France and Spain. 8-oxo-dG is significantly higher (up to about threefold) in lymphocyte DNA from men in Ireland and the U.K. Oxidative DNA damage is not significantly affected by carotenoid supplementation; nor is there any association with mean baseline levels of antioxidants, which are generally similar in the five countries. The five countries sampled lie on an axis from northern to southern Europe with a steep gradient in terms of premature heart disease. There is a strong association between premature coronary heart disease mortality in men and the mean levels of 8-oxo-dG for the five countries (r = 0.95, P < 0.01). Women have low coronary heart disease mortality rates, which do not correlate with 8-oxo-dG. In terms of cancer deaths, only colorectal cancer in men shows a significant positive correlation (r = 0.91, P < 0.05), and stomach cancer in women is negatively correlated with DNA oxidation (r = -0.92, P = 0.01).

Erythrocyte folate analysis: a cause for concern?

Neural tube defects can be prevented by adequate intake of periconceptional folate, and inverse associations between folate status and cardiovascular disease and various cancers have been noted. Thus, there is renewed interest in the analysis of red cell folate (RCF) as an indicator of folate deficiency risk. Assessment of the assumptions that underpin RCF assays indicates that many are false. Published literature suggests that increased deoxy-hemoglobin (which can bind RCF electrostatically) yields more assayable folate, and increased oxy-hemoglobin (which cannot bind RCF) yields less assayable folate. It is argued that as deoxy-hemoglobin picks up oxygen and switches quaternary structure, any bound folate must, on purely theoretical grounds, become physically "trapped". Venous blood taken for analysis is 65% to 75% saturated with oxygen, and pro-rata "trapping" will lead to serious underestimation of RCF. Hence, doubt is cast over the validity of all previous RCF values. Some strategies for accurately assessing RCF are suggested.

Beta-carotene enhances the recovery of lymphocytes from oxidative DNA damage.

Epidemiological studies have revealed a strong correlation between high intake of fruit and vegetables and low incidence of certain cancers. Micronutrients present in these foods are thought to decrease free radical attack on DNA and hence protect against mutations that cause cancer, but the fine details of the causal mechanism have still to be elucidated. Whether dietary factors can modulate DNA repair--a crucial element in the avoidance of carcinogenesis--is an intriguing question that has not yet been satisfactorily answered. In order to investigate the effects of beta-carotene on oxidative damage and its repair, volunteers were given a single 45 mg dose and lymphocytes taken before and after the supplement were treated in vitro with H2O2. DNA strand breaks and oxidised pyrimidines were measured at intervals, to monitor the removal of oxidative DNA damage. We found inter-individual variations in response. In cases where the baseline plasma beta-carotene concentration was high, or where supplementation increased the plasma concentration, recovery from oxidative damage (i.e. removal of both oxidised pyrimidines and strand breaks) was relatively rapid. However, what seems to be an enhancement of repair might in fact represent an amelioration of the continuing oxidative stress encountered by the lymphocytes under in vitro culture conditions. We found that culture in a 5% oxygen atmosphere enhanced recovery of lymphocytes from H2O2 damage.