In vivo effect of N-ethylmaleimide (NEM) on the measurement of nitrate in plasma.

BACKGROUND: Bioavailability of nitric oxide in the body may be estimated by measuring the concentration of nitrate in plasma. However, it has not been reported whether sequestering of aminothiols in plasma affects the concentration of nitrate in the samples. OBJECTIVE: N-ethylmaleimide (NEM) sequesters aminothiols in plasma therefore we tested the in vivo effect of NEM on the concentration nitrate in plasma. SUBJECTS AND METHODS: Blood samples were collected from 56 healthy subjects in EDTA vials, EDTA vials containing PBS (pH7.4) and EDTA vials containing NEM dissolved in PBS. Nonparametric statistical tests were used to study the effect of NEM on the concentration of nitrate in plasma measured by the Griess reagent assay and by an HPLC method. RESULTS: The concentration of nitrate in plasma containing NEM dissolved in PBS was lower than plasma containing PBS and plasma without any reagent measured by the Griess reagent assay (p<0.001 and p<0.001). Similarly, the concentration of nitrate in plasma containing NEM measured by the Griess reagent assay was significantly lower than nitrate concentration in plasma measured by the HPLC method (p<0.001). CONCLUSION: Our findings suggest that plasma, which contains NEM, may not be suitable for the measurement of nitrate by the Griess reagent assay.

Effect of Omega-3 and Vitamins E + C Supplements on the Concentration of Serum B-Vitamins and Plasma Redox Aminothiol Antioxidant Status in Elderly Men after Strength Training for Three Months.

BACKGROUND: Data on redox plasma aminothiol status in individuals on strength training are very limited. Therefore, we studied the effect of omega-3 and vitamins E + C supplementation on the concentration of B-vitamins and redox aminothiol status in elderly men after strength training for 3 months. METHODS: Healthy men, age 60 ± 6 (mean ± SD) were randomly divided into 3 groups: group I received placebo (n = 17), group II consumed omega-3 (700 mg, n = 17), and group III consumed vitamins E + C (235 mg +1 g, n = 16) daily for 3 months. All participants completed a strength training program for the same period. RESULTS: The concentration of serum vitamin B12 decreased and the concentration of serum folate increased in group I after the intervention (p = 0.01, p = 0.009). The concentration of plasma 5-pyridoxal phosphate decreased in groups II and III (p = 0.03 and p = 0.01), whereas the concentration of serum uric acid decreased only in group II (p = 0.02). We detected an increase in the concentration of reduced form of aminothiols in all groups (p < 0.001). The red/ox plasma aminothiol status was significantly changed in all groups after the intervention (p < 0.05). CONCLUSION: Omega-3 and vitamins E + C supplementation affect the concentrations of serum B-vitamins and redox plasma aminothiol status in healthy elderly men on strength training.

Eating patterns are associated with biomarkers in a selected population of university students and employees.

The association between diet and CVD cannot be assigned to a single nutrient, but rather to a set of nutrients and non-nutrients, and eating pattern analyses have become an important tool in investigation of this relationship. Our objective was to investigate eating patterns in relation to nutrient intake and serum concentration of folate, vitamin B12 and TAG in ninety-five healthy adult participants. Dietary information was collected by an FFQ, and eating patterns were obtained by principal components analyses of thirty-three food groups. Three eating patterns were extracted, a sweet eating pattern identified by intakes of cakes, snacks, sugar-sweetened drinks and chocolates; a prudent eating pattern identified by vegetables, fruits and olive oil; and a traditional food pattern identified by red meat, lean fish and cheese. Blood samples were collected in the morning after an overnight fast. Linear regression analyses adjusted for age, BMI and smoking showed a negative association between the sweet eating pattern scores and the serum concentration of folate (ß = -2·31 (95 % CI -4·14, -0·45)) and a positive association with serum concentration of TAG (ß = 0·35 (95 % CI 0·12, 0·57)). The prudent eating pattern scores were positively associated with the serum concentration of folate (ß = 1·69 (95 % CI 0·44, 2·92)). In conclusion, a sweet eating pattern was associated with risk factors for CVD, whereas a prudent eating pattern was associated with protective factors.

Serum folate is significantly correlated with plasma cysteine concentrations in healthy industry workers.

BACKGROUND: A low concentration of serum folate is associated with an increased risk of cardiovascular disease. Extracellular cysteine is involved in aging, cancer and cardiovascular disease. The relationship between serum folate and plasma cysteine is poorly understood. Therefore, we investigated this relationship in industry workers, whose health has economic implications. METHODS: The concentration of serum folate was determined by the Access ImmunoAssay System Sanofi Pasteur. Plasma cysteine and homocysteine were measured by an ion-pair HPLC method. The concentrations of serum triglycerides were determined by an enzymatic colorimetric method. RESULTS: We detected a positive correlation between the concentration of serum folate and plasma cysteine, whereas the concentration of serum folate was negatively correlated with plasma homocysteine and serum triglycerides. In a multiple regression analysis with adjustment for age, BMI and smoking, serum folate as the dependent variable exhibited a strong relationship with plasma cysteine, and a negative relationship with plasma homocysteine and serum triglycerides. CONCLUSION: We observed significant correlations between serum folate, plasma cysteine and serum triglyceride concentrations in industry workers, implying that folate may modulate key aspects of the body's cysteine and lipid metabolism.

Association between folate intake from different food sources in Norway and homocysteine status in a dietary intervention among young male adults.

The aim of the present investigation was to study the effect of a dietary intervention which combined nutrition information with increased availability of vegetables, fruits and wholegrain bread. The effect of the intervention was determined by changes in the intake of vegetables, fruits, wholegrain bread and estimated nutrients. Furthermore, the study investigated whether changes in relative contribution from different food sources of folate were related to changes in the concentration of plasma total homocysteine (p-tHcy). The 5-month intervention study included 376 male recruits from the Norwegian National Guard, Vaernes (intervention group) and 105 male recruits from the Norwegian National Guard, Heggelia (control group). The study resulted in an increase in the total consumption of vegetables, fruits, berries and juice (P < 0.001) and of wholegrain bread (P < 0.001). The participants in the intervention group showed a higher increase in the intake of dietary fibre (P < 0.001) and folate (P < 0.001) compared with the control group. The relative contribution of folate intake from fruits, vegetables and wholegrain bread was higher in the intervention group compared with the control group (P < 0.001 for all). The increased intake of folate from wholegrain bread was inversely associated with a reduced concentration of p-tHcy (P = 0.017). In summary, the dietary intervention resulted in an increased intake of vegetables, fruits and wholegrain bread and a subsequent increase in folate intake from these food components. Reduction in the concentration of p-tHcy was significantly related to an increased folate intake due to an increased consumption of wholegrain bread.

Folate, related B vitamins, and homocysteine in childhood and adolescence: potential implications for disease risk in later life.

OBJECTIVES: Folate and the metabolically related B vitamins are an important priority throughout life, but few studies have examined their status through childhood and adolescence. The aims of the current study were to investigate age, gender, and lifestyle factors as determinants of folate, related B-vitamin status, and homocysteine concentrations among British children and adolescents and to propose age-specific reference ranges for these biomarkers, which, at present, are unavailable. PARTICIPANTS AND METHODS: Data from the National Dietary and Nutritional Survey of 2127 young people aged 4 to 18 years were accessed to provide a representative sample of British children. All of the subjects who provided a blood sample for homocysteine concentrations were included in the current study (n = 840). Of these, laboratory biomarkers of folate (serum and red cell folate: n = 832 and 774, respectively), vitamin B(12) (n = 828), vitamin B(6) (n = 770), and riboflavin (n = 839) were also examined. RESULTS: The biomarker status of all 4 of the relevant B vitamins decreased significantly with age. Correspondingly, homocysteine concentrations progressively increased, with median values of 5.6, 6.3, and 7.9 mumol/L for children aged 4 to 10 years, 11 to 14 years, and 15 to 18 years, respectively, and were higher in boys compared with girls (15-18 years only). Independent of age and gender, fortified breakfast cereal intake (consumed by 89% of the sample) was associated with significantly higher B-vitamin status and lower homocysteine concentrations. CONCLUSIONS: It is not generally appreciated that the well-established progressive increase in homocysteine from 4 to 18 years reflects decreases in the biomarker status of all 4 metabolically related B vitamins. We suggest age-specific laboratory reference ranges for homocysteine and related B-vitamin concentrations for potential use within a pediatric setting.

BMI, lipid profile, physical fitness and smoking habits of young male adults and the association with parental education.

BACKGROUND: Few studies have focused on the potential relationship between parental educational level and cardiovascular risk factors among young male adults. The aim of this study was to investigate cardiovascular disease risk factors among young men and whether body mass index (BMI), serum lipids, physical fitness and smoking habits were related to paternal and maternal education. METHODS: In this cross-sectional study 750 18- to 26-year-old male recruits participated. RESULTS: Linear regression analyses showed that the paternal education was inversely associated with BMI (P = 0.035) and the concentration of total cholesterol (P = 0.003) and low-density lipoprotein (LDL) (P = 0.014). Running performance was inversely related to cigarette smoking (P = 0.022) and the concentration of triacylglycerol (P = 0.001). BMI was positively related to the concentration of LDL (P = 0.002), total cholesterol/high-density lipoprotein (HDL) ratio (P < 0.001) and inversely related to the concentration of HDL (P < 0.001), running performance (P < 0.001) and muscular strength (P = 0.011). Recruits with low BMI, both high and low fitness, had a significantly better lipid profile than recruits with high BMI and low fitness (P

Comparison of high-pressure liquid chromatography (HPLC) and Griess reagent-spectroscopic methods for the measurement of nitrate in serum from healthy individuals in the Nordic countries.

OBJECTIVES: Bioavailability of NO can be estimated by measuring the concentration of nitrate (NO(3)) in serum. However, the methods used for the measurement NO(3) in plasma or serum show a great degree of variation. Therefore, we compared two analytical methods for the measurement of NO(3) in serum. DESIGN AND METHODS: The concentration of NO(3) in 600 serum samples collected from healthy individuals was determined by the HPLC and by the Griess reagent-spectroscopic method. RESULTS: The concentration of NO(3) in the samples was 29.4+/-16.1 micromol/L and 26.2+/-14.0 micromol/L (mean+/-SD) measured by HPLC and Griess reagent-spectroscopic method respectively (p<0.0001). We detected a significant correlation between the two methods (R=0.81, p<0.0001). CONCLUSIONS: A significant correlation between the two methods may suggest that either method can be used for the measurement of NO(3) in serum, however the Griess reagent-spectroscopic method measures lower concentrations of NO(3) than the HPLC method.

Changes in predictors and status of homocysteine in young male adults after a dietary intervention with vegetables, fruits and bread.

BACKGROUND: Elevated plasma total homocysteine (p-tHcy) is associated with increased risk of cardiovascular disease, and an inverse association has been shown between the dietary intake of B-vitamins, B-vitamin profile and the concentration of p-tHcy. AIM OF THE STUDY: The main objective of this investigation was to study the effect of a dietary intervention focusing on an increased intake of vegetables, fruits and bread. The effect of the dietary intervention was determined by the changes in plasma concentrations of tHcy, cysteine (cys), riboflavin, flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) and serum concentrations of folate and vitamin B(12). METHOD: An intervention study with duration of 5 months, including 541 male recruits from the Norwegian National Guard, Vaernes and a control group, including 209 male recruits from the Norwegian Army, Heggelia. RESULTS: The dietary intervention resulted in decreased concentration of p-tHcy (-10%, P = 0.002), p-cys (-6%, P < 0.001) and FMN (-11%, P = 0.310) and increased concentration of riboflavin (+23%, P < 0.001) and FAD (+10%, P = 0.008) in the intervention group compared with the control group. The change in p-tHcy concentration was positively related to the change in the concentration of p-cys (P < 0.001) and FMN (P = 0.035) and inversely related to the change in concentration of folate (P = 0.021). CONCLUSIONS: A dietary intervention program focusing on an increased intake of vegetables, fruits and bread showed a favourable effect on the concentration of p-tHcy and its metabolites. Our findings suggest that the changes in the concentration of p-cys, folate and FMN seem to be predictors of changes in the p-tHcy concentration.

A novel HPLC method for the measurement of thiobarbituric acid reactive substances (TBARS). A comparison with a commercially available kit.

OBJECTIVES: Malondialdehyde (MDA) as a part of thiobarbituric acid reacting substances (TBARS) is frequently used as an indicator of lipid peroxidation. Most methods for the measurement of TBARS require long derivatization time and addition of antioxidants in the samples. Furthermore, comparison of these methods with commercially available HPLC kits is lacking. DESIGN AND METHODS: We investigated column performance of five different columns, tested eight different acids for the hydrolysis of the samples, and estimated stability of derivatized plasma samples with different anticoagulants. The samples were derivatized with TBA. The peak for the TBA(2)-MDA adduct was separated and detected by HPLC. RESULTS: Performance of the Phenomenex Gemini column was best. PCA at the concentration of 0.1125 N was used in this method. Coefficient of variation (CV %) within the run and between the run was 4.1% and 6.7%, and analytical recovery was 90-94%. The retention time of the TBA(2)-MDA peak was 1.8 min. Reference intervals for TBARS in serum from 250 individuals were 0.53 and 2.1 micromol/l using our HPLC method and 0.07 and 0.24 micromol/l using the Chromsystems assay. Linear regression with log converted values revealed weak relationship between the two methods (r(2) = 0.064). CONCLUSIONS: Our HPLC method for the analysis of TBARS in serum and plasma is fast and accurate and therefore can be used in clinical studies.