Erythrocyte Glutathione Reductase Activation Coefficient Values Research Articles

The Homozygous Hemoglobin EE Variant Is Associated with Poorer Riboflavin Status in Cambodian Women of Reproductive Age

Riboflavin is required for erythropoiesis, which is increased in people with hemoglobinopathies due to increased hemolysis and erythrocyte turnover. Dietary intake and status of riboflavin is poor in Cambodia, where hemoglobinopathies are common. We assessed the association between genetic hemoglobin disorders and riboflavin status in women of reproductive age in Cambodia. Venous blood samples from 515 Cambodian women of reproductive age, 18-45 y, were analyzed for biomarker status of riboflavin [erythrocyte glutathione reductase activation coefficient (EGRac)], genetic hemoglobin (Hb) disorders, and hematological indices. Linear regression analysis was used to estimate the association between EGRac with Hb, ferritin, and Hb genotypes. EGRac was log transformed in the analyses, and the regression coefficients represent the geometric mean differences. Genetic Hb disorders were present in 57% of the population, with the homozygous hemoglobin E variant (Hb EE) occurring in ∼10% of women (n=53). Deficient (EGRac ≥1.40) or marginal riboflavin status (EGRac ≥1.30 and <1.40) was observed in 92% (n=475) of women. The variant Hb EE genotype was associated with 18% (95% CI: 9%, 28%) higher geometric mean EGRac values than the normal Hb AA genotype (P<0.001). Although riboflavin biomarker deficiency or marginal status is widely prevalent in Cambodian women, lower riboflavin status was observed more frequently in women with the Hb EE genotype than in women with normal Hb AA. The relation between genetic Hb disorders and riboflavin warrants further investigation. This trial was registered at clinicaltrials.gov as NCT01593423 and NCT02481375.

Investigation of Blood Pressure during the First Trimester of Pregnancy in relation to the MTHFR C677T Polymorphism

AbstractThe common C677T polymorphism in the MTHFR gene encoding the folate-metabolising enzyme methylenetetrahydrofolate reductase is implicated in hypertension and hypertension in pregnancy. Hypertension affects up to 15% of all pregnancies and has been identified as a leading cause of maternal and neonatal morbidity and mortality worldwide. We previously reported higher systolic and diastolic blood pressure (BP) in non-pregnant women with the variant MTHFR 677TT genotype compared to CT/CC genotypes. In addition, randomised controlled trials (RCTs) in non-pregnant hypertensive adults from our Centre demonstrated that supplemental riboflavin (co-factor for MTHFR) lowers BP specifically in those with the TT genotype. However, the role of this common folate polymorphism and its interaction with riboflavin during pregnancy remains unclear. The aim of this study was to investigate the impact of MTHFR genotype and riboflavin status on BP in pregnancy. Data were generated from the ongoing Optimal Nutrition for the Prevention of Hypertension (OptiPREG) project. Pregnant women were recruited at the end of the first trimester from antenatal clinics in Northern Ireland and in the Republic of Ireland. Participants were screened for MTHFR genotype and BP was measured according to current clinical guidelines. Biomarker status of riboflavin was determined using the erythrocyte glutathione reductase activation coefficient (EGRac), a functional assay with higher EGRac values representing a lower status. Overall, 117 (11.6%) participants were identified with the variant MTHFR 677TT genotype. Both systolic and diastolic BP decreased from 8th to 16th gestational week (GW), however, this typical BP pattern was not observed in the TT genotype group. After adjusting for maternal age, GW and body mass index, women with the TT genotype at 12th GW had higher mean systolic (P 0.035) and diastolic (P 0.034) BP. When the results at the 12th GW were stratified by riboflavin status, the BP phenotype owing to this polymorphism was evident only among women with lower status (i.e. EGRac &gt; 1.30), with mean (SEM) systolic BP of 120.4 (3.1) mmHg compared to 112.6 (2.5) mmHg in those with higher status (EGRac ≤ 1.30) within the TT genotype group; in contrast, low versus high riboflavin status had no impact on BP in CT/CC genotype groups. These results suggest that MTHFR genotype influences BP during pregnancy and that riboflavin can exert an important modulating effect on BP in women with TT genotype. An RCT is required to fully investigate the role of MTHFR genotype and its interactive effect with riboflavin in BP during pregnancy.

Suboptimal Biochemical Riboflavin Status Is Associated with Lower Hemoglobin and Higher Rates of Anemia in a Sample of Canadian and Malaysian Women of Reproductive Age

Riboflavin is required for several redox reactions. Clinical riboflavin deficiency occurs mainly in low-income countries, where it is associated with anemia. The functional significance of suboptimal riboflavin status in different populations and its role in anemia is not well understood. We assessed the biomarker status of riboflavin and its association with hemoglobin concentration and anemia in women living in Vancouver, Canada, and Kuala Lumpur, Malaysia. Healthy nonpregnant, nonbreastfeeding women (19-45 y) were recruited from Canada ( n=206) and Malaysia (n=210) via convenience sampling. Fasting blood was collected to assess riboflavin status [erythrocyte glutathione reductase activity coefficient (EGRac)], hematological indicators, soluble transferrin receptor (sTfR), ferritin, vitamin A, folate, and vitamin B-12 concentrations. Linear and logistic regression models were used to assess the association of riboflavin status with hemoglobin concentration and anemia. EGRac (mean±SD) values were higher, indicating poorer riboflavin status, in Malaysian compared with Canadian women (1.49±0.17 compared with 1.38±0.11). Likewise, riboflavin biomarker deficiency (EGRac≥1.40) was significantly more prevalent among Malaysians than Canadians (71% compared with 40%). More Malaysian than Canadian women were anemic (hemoglobin <120 g/L; 18% compared with 7%). With use of linear regression (pooled sample; n=416), EGRac values were negatively associated with hemoglobin concentration (r=-0.18; P<0.001). This relation remained significant (P=0.029) after adjusting for age, parity, ethnicity, vitamin B-12, folate, sTfR, ferritin, and vitamin A. Women with riboflavin deficiency (EGRac≥1.40) were twice as likely to present with anemia (adjusted OR:2.38; 95% CI: 1.08, 5.27) compared with women with EGRac<1.40. Biochemical riboflavin deficiency was observed in Canadian and Malaysian women, with higher rates of deficiency among Malaysian women. Deficient biomarker status of riboflavin was a weak but significant predictor of hemoglobin and anemia, suggesting that the correction of riboflavin deficiency may potentially play a small protective role in anemia, but this requires further investigation.

P3566Ambulatory blood pressure response to riboflavin supplementation in adults with the C677T polymorphism in the folate metabolising enzyme methylenetetrahydrofolate reductase (MTHFR)

Abstract Background Epidemiological evidence suggests that the C677T polymorphism in the folate metabolising enzyme MTHFR is associated with a 24–87% increased risk of hypertension. Riboflavin (a cofactor for MTHFR) can modify this phenotype, as demonstrated in a series of RCTs, at this centre. ABPM measures BP over 24hrs and provides a more reliable assessment of an individual's BP compared to clinic BP. Aim To investigate the effect of riboflavin supplementation on BP (by ABPM) in adults with the MTHFR 677TT genotype. Methods Adults (n=3462) were recruited and screened for the MTHFRTT genotype and those eligible (n=81), were stratified by baseline SBP and randomised to receive riboflavin (10mg/day/16weeks) or placebo. Riboflavin status was measured using the erythrocyte glutathione reductase activation coefficient (EGRac) assay and clinic and ABPM were measured in accordance with clinical guidelines. Results Mean 24hr, daytime and night-time SBP was 6mmHg higher (p&lt;0.05) in adults with the TT v CC genotype. BP response to riboflavin was strongly dependent on baseline BP. In participants with a baseline SBP≥125mmHg, riboflavin resulted in a significant BP lowering in daytime SBP (Table). Day-time SBP response to riboflavin Treatment Placebo P-value Age 48.6 (11.3) 47.8 (12.0) 0.823 Male n (%) 17 (73.9) 12 (46.2) 0.093 BMI 28.8 (3.8) 27.2 (3.1) 0.131 Riboflavin status (EGRac)† Pre 1.33 (1.28–1.38) 1.29 (1.24–1.34) Post 1.20 (1.14–1.25) 1.31 (1.25–1.36) &lt;0.001 Change −0.12 0.02 Daytime BP (mmHg)‡ Pre 137.1 (132.7–141.3) 134.7 (130.3–139.1) Post 133.0 (128.9–138.1) 134.8 (130.6–139.1) 0.036 Change −3.8 −0.2 ANCOVA, adjusting for sex. †Higher EGRac values indicate lower riboflavin status; ‡mean of participant daytime/awake hrs personalised for each participant. Conclusion This is the first study to show, using ABPM, that riboflavin supplementation, targeted at adults with the MTHFR 677TT genotype, results in significant lowering of mean, day and night BP. Given the frequency of this genotype worldwide (approximately 10%, but as high as 30% in some populations) these findings could offer a personalised approach for BP management in at-risk sub-populations. Acknowledgement/Funding This work was supported by DSM Nutritional Products, Ltd and Wellcome Trust-Wolfston Northern Ireland Clinical Research Facility are acknowledged

Effect of riboflavin supply on student body's provision in north-western Poland with riboflavin measured by activity of glutathione reductase considering daily intake of other nutrients

The riboflavin nutritional status of 120 people, age 22–25, studying in Szczecin, Poland, together with contents of their daily food servings were studied. Body's provision with riboflavin was determined using the erythrocyte glutathione reductase activity coefficient (EGRAC) and was compared with a sample in which the enzyme activity was stimulated with flavin adenine dinucleotide. The information concerning diets was collected with the method of a 7-day food record prior to blood collection. Biochemical deficiency in riboflavin was observed in 33.7% of women and 25% of men. The resulting average EGRAC value was 1.02 for women and 0.88 for men. Assessment of significant differences in riboflavin provision between the sexes revealed better provision in the male group. The comparison of EGRAC values with riboflavin content in 7-day diets of the respondents showed that the average intake of this vitamin in the female group, in which biochemical deficiency was observed, amounted to 1.05 mg, whereas in the male group it was, on average, 1.39 mg. In the group of people in which the potential risk of riboflavin deficiency in the body was not observed, the level of this vitamin consumption was, on average, 1.43 mg and 1.8 mg in the female and male groups, respectively. Women with biochemical riboflavin deficiency consumed significantly less of all the analyzed nutrients in comparison with the people without riboflavin deficiency.

Riboflavin Status in Acutely Ill Patients and Response to Dietary Supplements

Although a number of studies have reported riboflavin deficiency in free-living older people, no data are available on riboflavin intake and status in older people during acute illness. To determine the riboflavin response to dietary supplements during acute illness, 297 hospitalized, acutely ill older patients are randomly assigned to receive a daily oral nutritional supplement containing 1.3 mg of riboflavin or a placebo for 6 weeks. Outcome measures are riboflavin intake and riboflavin biochemical status at baseline, 6 weeks, and 6 months using the erythrocyte glutathione reductase activation coefficient (EGRAC), a measure of riboflavin tissue saturation. EGRAC values are inversely proportional to riboflavin status. Fifty-six percent of patients (167/297) have suboptimal riboflavin status (EGRAC > 1.30). No significant correlation is found between EGRAC and either total energy or riboflavin intakes. Significant correlations are found between total energy intake and riboflavin intakes both in hospital and at home (r = 0.67, P < .0001 and r = 0.57, P < .0001, respectively). Smokers and patients with chronic obstructive pulmonary disease (COPD) have lower riboflavin status (high EGRAC values) compared with nonsmokers and those without COPD. Riboflavin status improves significantly in the supplement group at 6 weeks compared with the placebo group, but status declines between 6 weeks and 6 months, after patients stop taking the supplements. A high proportion of acutely ill patients have suboptimal riboflavin status. Supplementation with a physiological amount of riboflavin in a mixed-nutrient supplement significantly improves riboflavin status, but the effect is transient and status deteriorates again after patients stop taking the supplements.

Riboflavin status in acute ischaemic stroke

There is experimental evidence that riboflavin (vitamin B2) supplementation reduces oxidative damage and cerebral oedema following acute stroke. To measure riboflavin levels in acute stroke before and after supplementation with this vitamin. Ninety-six acute ischaemic stroke patients had their riboflavin status measured at baseline and then randomly assigned to receive 5 mg of oral riboflavin and other B-group vitamins within 12 h of the stroke onset and then daily or no B-vitamins for 14 days. Non-fasting venous blood was obtained at baseline, days 7 and 14 post-randomization for measurement of riboflavin status using erythrocyte glutathione reductase activity coefficient (EGRAC). EGRAC is a measure of riboflavin tissue saturation. This assay has the advantage of being extremely stable and sensitive. EGRAC values are inversely proportional to riboflavin status, so that values greater than 1.3 indicate biochemical deficiency. Fifty-one per cent of patients studied were riboflavin deficient at baseline. Fourteen days of riboflavin supplementation significantly improved the measure of B2 status compared with the control group. Seven out of 37 patients in the supplement group (19%) were riboflavin deficient compared with 22 out of 39 patients (56%) in the control group at the end of the treatment period (P=0.035 for the differences in cumulative changes between groups over 2 weeks). A high proportion of acute stroke patients were biochemically deficient of riboflavin immediately post-infarct. Supplementation with 5 mg of riboflavin for 2 weeks significantly improved riboflavin status; however, the clinical significance of these findings is not yet known.

Flavin adenine dinucleotide levels in erythrocytes of very low birthweight infants under vitamin supplementation.

Despite the potential dangers of over- and undersupplementation with riboflavin (vitamin B2), little is known about status, requirements and metabolism of this vitamin in very low birthweight (VLBW) infants. As reported here, it is possible to calculate the intraerythrocytic concentration of free flavin adenine dinucleotide (FAD), the predominant coenzyme form of riboflavin, on the basis of the erythrocyte glutathione reductase activation coefficient (EGRAC). This parameter was determined in 9 VLBW infants (median of birthweight 634 g, median of gestational age 24 weeks + 3 days) at intervals over a period of 6 weeks. All infants received riboflavin supplementation, either in milk (average intake: 175 micrograms/kg/day) and a multivitamin preparation (200 micrograms/day) or via total parenteral nutrition (TPN; 720 micrograms/kg/day). The average EGRAC was 1.07 (1.00-1.24) during enteral nutrition and 1.05 (1.00-1.18) during TPN. These values correspond to free FAD levels of 757 and 1,060 nM, which is 4 to 5 times higher than in cord blood or in breast-fed infants. EGRAC values between 1.15 and 1.35, representing free intracellular FAD levels of 350-150 nM, should be adequate in VLBW infants in order to minimize the hazards of flavin-induced radical reactions.

Riboflavin status of female athletes consuming normal diets

Riboflavin (B 2) status was determined in trained female athletes (n=13) and untrained female subjects (n=14). Groups were similar in age, height and weight. Subjects ingesting large quantities of supplemental or dietary riboflavin were excluded from the study. Dietary intake information was obtained and evaluated for B 2 content. Riboflavin status was evaluated using the erythrocyte glutathione reductase activity coefficient (EGRAC) and urinary riboflavin excretion. Results indicated that total riboflavin intake per 24 hr was significantly greater (p<0.05) in the athletic group. However, no significant difference in intake was noted when B 2 consumption was expressed as intake/ 1000 kcal consumed (1.0±0.4 mg and 0.9±0.2 mg, athletes and untrained, X±SD). While all subjects had normal EGRAC values, athletes tended to have slightly higher (p=0.09) values when compared to untrained females (1.06±0.06 vs 1.01±0.07). Total riboflavin excretion/24 hr was not different in athletes (98±51 mcg) versus untrained females (86±16 mcg) although riboflavin excretion/g creatinine was somewhat lower (p=0.1) in athletes (63±26 mcg/g) than in untrained females (82±28 mcg/g). Five athletes and five untrained subjects with the lowest riboflavin intakes were also compared. No significant differences were noted for EGRAC values or total urinary riboflavin excretion. However, athletes in the subgroup excreted significantly less (p<0.05) B 2/g creatinine than did the untrained subgroup (50.8±15.5 and 81.3±16.7 mcg/g, respectively). Findings from the present study suggest that female athletes were able to maintain adequate riboflavin status from dietary intake alone.

Bioavailability of riboflavin from fortified palm juice.

The bioavailability of riboflavin from fortified palm juice was assessed in young adult men, Riboflavin status was assessed from urinary riboflavin excretion and erythrocyte glutathione reductase activity coefficient (EGR-AC) while iron status was assessed from haemoglobin and serum ferritin concentrations. Although the consumption of unfortified palm juice made significant contribution to the meager riboflavin intake, it conferred no metabolic advantage. The consumption of fortified palm juice produced a marked reduction in EGR-AC values and a significant increase in urinary riboflavin excretion. Since iron release from storage sites may be flavin dependent, riboflavin deficiency may affect iron utilization. Fortification may prove effective in alleviating nutrient deficiencies, but the carrier vehicle must be acceptable to all age groups.

Riboflavin requirement of lactating swine.

Twenty-five crossbred gilts and 25 crossbred sows were used in an experiment to estimate the riboflavin requirement of lactating swine. During gestation the females were fed a 12% crude protein, corn-soybean meal diet that was marginal in riboflavin content, i.e., 2.3 mg/kg. After farrowing, litter size was equalized across lactation diets within each parity category. The basal, 14% crude-protein corn-soybean meal lactation diet was supplemented to provide five levels of dietary riboflavin: 1.25, 2.25, 3.25, 4.25 and 5.25 mg/kg. Five gilts and five sows were fed each dietary treatment. Each dam was provided her assigned diet ad libitum during the 24-d lactation; piglets were not provided supplemental feed. The erythrocyte glutathione reductase activity coefficient (EGRAC), an indicator of riboflavin status, was measured on blood samples obtained from the dams and their piglets on d 1 and d 24 postpartum. On d 1, the mean EGRAC of gilts was slightly higher than that of sows, while piglet EGRAC was similar regardless of maternal age. On d 24 gilts and their piglets had higher average EGRAC (P less than .01) than did sows and their piglets. Thus, there was a treatment x dam age interaction (P less than .05). Lactation performance criteria gave further evidence of similar treatment x age group interactions. Gilts fed the diet containing 1.25 ppm riboflavin had higher piglet mortality, consumed less feed and lost more weight (P less than .05) for each criterion than did sows fed the same diet. Despite these observations, the broken-line estimates of the riboflavin requirement, based on EGRAC, for gilts and sows were 16.3 and 16.2 mg/d, respectively. The EGRAC values for piglets closely paralleled those of their dams regardless of treatment, suggesting that there is no mechanism to protect the nursing piglets from a maternal dietary deficiency of riboflavin.

Growth, erythrocyte glutathione reductase and liver flavin as indicators of riboflavin status in turkey poults.

1. Turkey poults were fed on an isolated soya protein diet supplemented with 0, 2, 4 or 8 mg riboflavin/kg between 7 and 28 d of age. Maximum body weight was attained with the 4 and 8 mg diets, and poults fed on the supplemented diet did not survive until 21 d of age. The erythrocyte glutathione reductase activation coefficient (EGRAC) of the poults was inversely related to dietary riboflavin at all ages. At 28 d of age the values for EGRAC of poults fed on the 2, 4 and 8 mg diets were 2.25, 1.59 and 1.25 respectively; at 31 d, following a single oral dose of 10 mg riboflavin administered on day 28, the same poults had EGRAC values of 1.36, 1.17 and 1.26. 2. In a second experiment, poults were fed on diets supplemented with 0, 2, 4, 8 or 12 mg riboflavin/kg. Maximum body weight was attained with the 4, 8 and 12 mg diets. At 28 d of age the EGRAC values for the 0, 2, 4, 8 and 12 mg treatments were 2.36, 2.34, 1.59, 1.28 and 1.18 respectively. The concentration of flavin in the liver was not related to riboflavin intake, whereas total flavin in the liver was positively related, being on average 94, 206, 400, 440 and 413 micrograms flavin/liver for the 0, 2, 4, 8 and 12 mg treatments respectively. 3. It is concluded that EGRAC is a sensitive indicator of riboflavin status in the turkey poult and that an EGRAC of 1.6 indicates a marginal riboflavin status. It is also concluded that liver total flavin reflects merely the effect of riboflavin on growth and liver size.