Erythrocyte Glutathione Reductase Activity Research Articles

II. Multicentre clinical trial of diets for low birthweight infants: interim analysis of short term clinical and biochemical effects of diet

In 3 of 5 collaborating centres, two clinical trials comparisons are being made between infants randomly assigned to a preterm formula (PTF) or banked milk (BBM) fed as sole diets or in conjunction with maternal milk. (n=200 at second interim analysis). BBM fed infants showed: 1. increased incidence of: severe hyponatraemia (Na <126 mmol/1, p <0.05), radiological and biochemical evidence of rickets of prematurity (hypophosphataemia and higher alkaline phosphatase p <0.01) and riboflavin deficiency (erythrocyte glutathione reductase activation coefficient >1.3, p<0.01); 2. higher peak plasma bilirubin and more prolonged hyperbilirubinaemia (p<0.01), 3. higher plasma prolactin and growth hormone concentration (p<0.01). PTF fed infants showed 1. increased: growth rates (reported elsewhere), gastric pooling (p<0.01) and latent systemic anaphylactic sensitisation to cows milk (basophil histamine release to cowsmilk and anti IgE challenge, p<0.01); 2. faster postnatal fall in plasma IgG and rise in IgM (p<0.05); 3. lower plasma calcium (week 1) and potassium (p<0.02) 4. higher platelet count (p<0.05) and plasma concentration of some amino acids, notably threonine (p<0.0001). We conclude that patterns of short term response to diet are complex and cannot be used to predict an overall clinical benefit of a feeding regime - their clinical significance can be assessed only by follow up studies, the principle concern of these trials.

Erythrocyte glutathione reductase activity and acquired trisomy #8 in various hematologic disorders

The gene for erythrocyte glutathione reductase (E-GR) activity has been assigned to chromosome #8. In the present series, we examined the E-GR activity in 14 cases with chronic myelodyplastic syndrome (CMS, preleukemia), atypical acute myelogenous leukemia (AML), or chronic myelogenous leukemia (CML), with and without acquired trisomy #8. No difference in the incidence of high levels of this enzyme was found between two groups, i.e., those with and without trisomy #8 suggesting the existence of a complex regulatory system in addition to chromosome #8.

Effect of riboflavin supplementation on riboflavin nutriture of a secondary school population in Nigeria

Riboflavin status was assessed in 42 secondary school students before and after supplementing the food intake with 5 mg riboflavin daily for 7 days. Energy, protein, and riboflavin intakes were determined on foods actually consumed by each student. Riboflavin nutriture was based on urinary riboflavin excretion and erythrocyte glutathione reductase activity coefficient. The energy and riboflavin intakes of the students were 68 to 82% and 80 to 88%, respectively, of the recommended allowance. The basal urinary riboflavin excretion was 0.335 mg/g creatinine and increased significantly to 3.51 mg/g creatinine after supplementation. The basal erythrocyte glutathione reductase activity coefficient values indicated an overall prevalence of 38% biochemical ariboflavinosis (erythrocyte glutathione reductase activity coefficient >1.30) and dropped significantly (p < 0.001) from 1.26 to 1.08. The results confirm that urinary riboflavin is of limited value in the assessment of riboflavin status while erythrocyte glutathione reductase activity coefficient more precisely assesses metabolic availability of riboflavin and more accurately detects biochemical ariboflavinosis.

Relation of riboflavin nutriture in healthy elderly to intake of calcium and vitamin supplements: evidence against riboflavin supplementation

The status of riboflavin nutriture was evaluated in 24 healthy elderly female residents of a private, nonprofit facility for the care of ambulatory elderly. Riboflavin intake by history was greater than or equal to the recommended dietary allowances (RDA) for this nutrient in all but three subjects, and the average intake in the group as a whole was 50% greater than the RDA. Confirmatory of the findings by history, the status of riboflavin nutriture was excellent in nearly all subjects as evaluated by urinary riboflavin excretion and erythrocyte glutathione reductase activity coefficient. By contrast, calcium intake was greater than or equal to the RDA in ony four of the 24 subjects. The adequacy of calcium intake was found to depend upon a sufficiently high percentage of the total dietary intake of riboflavin being derived from milk and dairy products. It was observed that individual calcium intakes were less than 80% of the RDA unless 40% or more of the total intake of riboflavin was derived from milk and dairy products rather than from other food sources. In those subjects taking daily supplementation with a single multivitamin tablet containing low levels of riboflavin, the total intake of riboflavin and its urinary excretion were increased similarly, suggesting that even small amounts of riboflavin are not retained by elderly subjects consuming a diet adequate in riboflavin. This study suggests that careful selection of B vitamin sources can be made for older individuals to assure that the intakes of both calcium and riboflavin are satisfactory, and furthermore that the use of riboflavin in vitamin supplements, even at very low doses, is largely wasted in healthy elderly individuals consuming adequate amounts of this vitamin from food sources.

Changes in glutathione and glutathione metabolizing enzymes in erythrocytes and lymphocytes of mice as a function of age

1. 1. Changes in reduced glutathione levels in liver, lung and whole blood of female Swiss-Webster mice with age were determined. In addition, glutathione content, and glutathione S-transferase and glutathione reductase activities of erythrocytes and lymphocytes of mice as a function of age were examined. 2. 2. Reduced glutathione content increased in liver, lung, whole blood, erythrocytes and lymphocytes with age from 3 to 9 months, reached a maximum level at 9 months of age and decreased thereafter with advanced age in all tissues. 3. 3. Glutathione S-transferase and glutathione reductase activities in erythrocytes and lymphocytes increased with age from 3 to 9 months, reached maximum activities at 9 months and decreased thereafter with advanced age. 4. 4. The glutathione content of erythrocytes from animals 18 months of age decreased by 56% as compared to 9 month old mice, while the activities of glutathione S-transferase and glutathione reductase decreased by 56 and 48%, respectively, over the same age span.

The distribution of supposed indicators of nutritional status in elderly patients.

The results of a survey of indicators of nutritional status in patients on admission to wards in the Department of Health Care of the Elderly in Nottingham are reported. Results outside conventional limits of normality are common. Little or none of the observed variance in the biochemical indicators in this population is explained by age or sex. Weight, mid-arm circumference, triceps skinfold thickness, albumin, retinol binding protein and retinol tend to be lower in patients who subsequently die, in patients who are clinically classified as thin or wasted, in patients with poor appetites and in patients with a diagnosis of malignancy. Plasma vitamin C, erythrocyte glutathione reductase activation coefficient and erythrocyte transketolase activation coefficient do not show these trends and are likely to be determined by different factors.

Depressed erythrocyte glutathione reductase activity in sickle cell disease

In a group of 27 sickle cell disease patients ranging in age from 2 yr 3 months to 43 yr, 13 (48%) were found to have depressed erythrocyte glutathione reductase activity suggesting riboflavin deficiency. Glutathione reductase activity coefficients did not correlate with riboflavin intakes which were calculated from 3-day diet records returned by 16 patients. Other causes of riboflavin deficiency including decreased absorption, altered metabolism, or increased excretion of the vitamin must be considered. The potential effect of depressed erythrocyte glutathione reductase activity in the sickle cell disease process is discussed.

EFFICACY OF VITAMIN SUPPLEMENTATION IN CHRONIC ALCOHOLICS UNDERGOING DETOXIFICATION

Ninety-seven admissions to an alcoholic unit were allocated to one of three groups receiving either oral vitamin supplements, intravenous vitamin therapy or placebo during a fiveday detoxification regime. Thiamine, riboflavin and pyridoxine status was assessed by measurement of erythrocyte transketolase (ETK), glutathione reductase (EGR) and glutamateoxaloacetate transaminasc (EGOT) activities on Days 0, 2 and 5. Vitamin status as assessed by these parameters improved in most patients by Day 5 irrespective of the treatment regime. However, two-thirds of those patients originally thiamine-deficient, as assessed by basal ETK, were still deficient after fivedays, irrespective of vitamin therapy received. Assessment of thiamine status by basal ETK yielded more patients rated as deficient than did assessment by the activation coefficient for transketolase ( a -ETK or ‘TPP effect’). EGOT was elevated in nearly all patients studied, whilst erythrocyte pyridoxal-5-phosphate concentrations were low, indicating that in alcoholics, EGOT is a poor indicator of pyridoxine status.

Effects of exercise on riboflavin requirements of young women

The riboflavin requirement of young women during periods of sedentary living and exercise was determined during a 12-wk metabolic study. The study was divided into a 6-wk no exercise period followed by a 6-wk exercise period in which subjects jogged around a track for 20 to 50 min/day. Twelve subjects, aged 19 to 27 yr, were fed a basic diet containing 0.6 mg riboflavin/1000 kcal of intake. Riboflavin intake was increased by 0.2 mg/1000 kcal increments by provision of riboflavin in a glucose polymer mixture. Linear regression analysis was used to estimate the riboflavin intake required for an erythrocyte glutathione reductase activity coefficient of 1.25 during both the no exercise and exercise periods. Individual riboflavin requirements ranged from 0.62 to 1.21 mg/1000 kcal before exercise and 0.63 to 1.4 mg/1000 kcal during the exercise periods. Riboflavin requirement could not be related to the kilocalorie intake or lean body mass of the subjects. It is concluded that healthy young women require more riboflavin to achieve biochemical normality than the 1980 Recommended Dietary Allowances and that exercise increases riboflavin requirements.

Riboflavin Status of Rats Fed Two Levels of Protein during Energy Deprivation and Subsequent Repletion

Adult male rats were fed either a low protein (LP) or moderate protein (MP) diet (0.6 and 1.8 g casein per rat per day, respectively) and 30 µg riboflavin (RF) while their energy intakes were progressively restricted to about 30% (26 kcal/day) of the predepletion ad libitum consumption. Total weight losses in 4 weeks averaged 17.3 ± 0.6 and 12.5 ± 0.7% from the initial body weights for the LP and MP groups, respectively, despite equal energy deprivation. Although liver riboflavin concentrations (micrograms/gram) were significantly higher in the energy-restricted groups than in either the predepletion or the ad libitum-fed control groups, total liver riboflavin remained at the predepletion level in both restricted groups. During energy deprivation, muscle riboflavin decreased significantly from the predepletion level and was not affected by protein intake. Subsequent repletion with 100 µg riboflavin/day and energy at the level voluntarily consumed by the LP group increased body and liver weights with both levels of protein, but liver riboflavin increased only in the MP group. Muscle riboflavin did not return to the predepletion level in either restricted group. Erythrocyte glutathione reductase activity coefficients remained normal for all groups during the depletion-repletion cycle.

Phosphoribosyl pyrophosphate synthetase and glutathione reductase in erythrocytes from hyperuricaemic and gout patients

The enzymic activities of erythrocyte phosphoribosyl pyrophosphate synthetase (EC 2.7.6.1) and glutathione reductase (EC 1.6.4.2) have been measured in 54 primary gout patients, 35 individuals having hyperuricaemia and 51 healthy controls. Statistical analyses have shown a significant increase ( p < 0.01) in the enzymic activity of erythrocyte PRPP synthetase in both the hyperuricaemic and gout groups compared with the controls. No correlation between activity and age was found in any of the three clinical groups. A significant decrease ( p ∼- 0.01) was found in the enzymic activity of red cell glutathione reductase in the gout group compared with the other two groups. The biochemical significance of the changes in enzymic activities of the two enzymes in primary gout is discussed.

Nutritional status in a healthy elderly population: riboflavin

Riboflavin status in 270 free-living and healthy elderly was determined from dietary intake (3-day food records) and erythrocyte glutathione reductase activity coefficients (EGR-AC). High EGR-ACs (greater than 1.35) indicate poor riboflavin nutriture. Mean dietary intakes of riboflavin were 1.86 +/- 0.64 (SD) for males and 1.58 +/- 0.69 mg/day for females. Approximately 45% of the population were taking some supplemental riboflavin and total riboflavin intakes ranged from 0.65 to 165 mg/day. The mean EGR-AC for those taking supplemental riboflavin was significantly lower than that of the nonsupplemented group (1.06 and 1.16, respectively). Only three subjects had EGR-ACs greater than 1.35. A significant correlation was found between total riboflavin intake and EGR-AC (r = 0.53). In a separate population of 667 volunteers between the ages of 20 and 87 yr, a significant decrease in mean EGR-AC with age was found. The mean EGR-AC for those over 60 yr and not taking a supplement was 1.16 +/ 0.10 compared to 1.23 +/- 0.11 for those from 20 to 29 yr old. Inadequate riboflavin nutriture appears to be more of a problem for younger than older adults.

Growth and Riboflavin Status of Rats Fed Different Levels of Protein and Riboflavin

The relationship between riboflavin and protein utilization was studied in 5-week-old male Sprague-Dawley rats, by using a factorial design with three levels of riboflavin (8, 16 and 24 µg per rat per day) and protein (1.0, 1.6 and 2.2 g casein per rat per day) in a 9-week experiment. With the lowest level of casein, protein intake was growth limiting, and the level of riboflavin intake had no effect on either weight gain or liver nitrogen retention. With the two higher levels of casein, both weight gain and liver nitrogen retention increased with riboflavin intake, but 24 µg riboflavin per day was inadequate for maximal utilization of nitrogen from 2.2 g casein. Neither protein nor riboflavin intake affected the concentration of liver nitrogen per gram of fresh tissue. Increasing the protein intake from 1.0 to 1.6 g increased riboflavin retention in the liver, but additional protein had no further effect. Liver and muscle (gastrocnemius) riboflavin concentrations, as micrograms per gram wet tissue, increased with riboflavin intake. At the two higher intakes of riboflavin, tissue riboflavin levels decreased and the erythrocyte glutathione reductase activity coefficients (EGR-AC) increased with protein intake. These findings are consistent with the view that the effect of protein on riboflavin requirement is related to the rate of growth and not to protein intake, per se.riboflavin-protein interdependence riboflavin status erythrocyte glutathione reductase activity coefficient

Impact of long term, low dose B-complex vitamin supplements on vitamin status and psychomotor performance of rural school boys

Rural school boys having a very high incidence of biochemical vitamin deficiencies, were supplemented with recommended daily requirement of B-complex vitamins or placebo, for 1 year. The objective was to use the data for deriving the interpretive guidelines for biochemical tests of vitamin nutrition status. The supplemented children had significantly superior biochemical status of riboflavin, pyridoxine and folic acid as judged by erythrocyte glutathione reductase and erythrocyte aspartate aminotransferase activation coefficients, and RBC folate levels respectively. However, even in the supplemented group there was a wide scatter in biochemical values, precluding the use of the data for deriving interpretive guidelines. More than 50 percent of the boys in this group continued to have biochemical evidence of deficiency of one or more vitamins using reported criteria. However, selective psychomotor testing revealed beneficial effects of supplements on arm-hand steadiness test. Thus, functional impact of vitamin supplements may be seen even in the absence of clear-cut clinical or biochemical change.

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.

Diet and vitamin nutrition of the high esophageal cancer risk population in Linxian, China.

In order to examine the nutritional status of the population of Linxian (in Henan, China) known to be at high risk for esophageal cancer (EC), we analyzed blood samples and conducted 3-day dietary surveys on 3 groups of normal adults, age 40-50, from Henan province. Two groups were from Linxian, where the EC mortality rate is about 138/100,000 (Group C, consisting of volunteers with a family history of EC, and Group NC, consisting of those who did not have a family history of EC). The third group, Group F, was from Fanxian, where the EC mortality rate is about 24/100,000. The average plasma retinol level was found to be about 0.27 micrograms/ml in all 3 groups; about 13%-20% of the individuals had a level less than 0.2 micrograms/ml. The average plasma carotene level of all 3 groups appeared to be adequate, ranging from 0.5-1.0 micrograms/ml. About 23% of the subjects in both Groups C and NC had low (less than 2 micrograms/ml) plasma ascorbate levels; in Group F, 16% had low ascorbate levels. Less than 12% of the individuals in each of the 3 groups had erythrocyte glutathione reductase activation coefficients less than 1.2, indicating a widely occurring riboflavin deficiency. Consumption of alcoholic beverages was not found to be a contributing factor to EC. The results of the present study reveal a deficiency of vitamins in the study populations, but do not indicate that the average nutritional status of the volunteers in Group C was worse than that of Groups NC and F with regard to vitamin A, ascorbate, and riboflavin.

Riboflavin status during pregnancy

The riboflavin status of 20 nonpregnant and 60 pregnant women (in the 2nd trimester, 3rd trimester, and early postpartum) was determined by the erythrocyte glutathione reductase activation test. None of the nonpregnant subjects but 26 pregnant subjects (21.7% in the 2nd trimester, 20.8% in the 3rd trimester, and 29.6% in early postpartum) had an activity coefficient greater than 1.20, indicative of biochemical deficiency of riboflavin. The deficiency developed at any of the three stages of pregnancy under study. Follow-up of individual cases revealed no progressive deterioration in riboflavin nutriture with advancement of pregnancy. The mean intake of riboflavin was higher than the recommended intake and revealed a significant negative correlation with activity coefficient values at the 3rd trimester. No significant correlation of riboflavin status with the outcome of pregnancy was noted. The effect of numbers of previous pregnancies, history of oral contraceptive usage, smoking, and alcohol showed no consistent effect on the percentage incidence of deficiency at all the three periods.

Effect of riboflavin deficiency on estrous cyclicity in pigs.

Previous studies have shown that riboflavin is needed for reproduction. A deficiency of this vitamin causes a lack of cyclicity and interferes with pregnancy. Because the exact means by which riboflavin affects reproduction is unknown, we conducted two experiments to determine the effect of a riboflavin deficiency on plasma and urinary sex steroids. Riboflavin status was measured by the determination of erythrocyte glutathione reductase activity coefficients (EGR-AC). In experiment 1, 13 male and 12 female pigs were bled by venipuncture for 3 consecutive days to determine if the EGR-AC was affected by the sex of the animal and if the EGR-AC fluctuated from day to day. Neither sex nor day of sampling significantly affected EGR-AC. The mean EGR-AC was 1.43± 0.11. In experiment 2, 20 crossbred cycling gilts were randomly allotted to two treatments: riboflavin adequate (control) and riboflavin deficient cornstarch-soybean meal diets. Gilts were checked daily for estrus. Blood was obtained by venipuncture at 1000 h every 2 weeks after the establishment of the deficiency. Plasma was frozen at −20°C and assayed for progesterone (P4) and estradiol-17β(E2). After 28 days, the first urine sample from two consecutive days was collected and assayed for estrone sulfate (E1 S) after enzymatic hydrolysis. Gilts on the riboflavin deficient diet became anestrous ∿63 days after the beginning of the experiment. The average time elapsed between consecutive estrous periods increased with length of time on the deficient diet. The control gilts had elevated (P<0.01) plasma P4 levels between Days 4–18 with peak concentrations of 28.3 ± 1.0 ng/ml on Day 12 of the cycle. In contrast, P4 levels in riboflavin deficient gilts were significantly elevated (P<0.001) over control gilts (64.2 ± 1.0 ng/ml). The E2 plasma concentrations in control gilts were low (19.6 ± 1.9 pg/ml) from Days 2–18, but were elevated (64.7 ± 2.3 pg/ml; P<0.01) 2 days before the onset of estrus. The riboflavin deficient gilts displayed significantly elevated E2 levels (82.6 ± 1.8 pg/ml; P<0.001). The E1 S urine concentrations were elevated in the control gilts with peak levels of 22.1 ± 2.3 ng/mg creatinine on Day 20. The riboflavin deficient guts had low E1 S levels throughout the test period (4.1 ± 0.8 ng E1 S/mg creatinine). These data indicate a loss of ovarian cyclicity and the presence of abnormal plasma concentrations and urinary excretion of sex steroids for riboflavin deficient gilts.

Riboflavin status in Gambian pregnant and lactating women and its implications for Recommended Dietary Allowances

Riboflavin status by the erythrocyte glutathione reductase (NAD(P)H2:glutathione oxidoreductase EC1.6.4.2) test, and dietary riboflavin intake, were measured in 156 pregnant and lactating women in Keneba, a rural Gambian village, and in 59 pregnant and lactating women in Cambridge, England. The Keneba women were studied longitudinally, for up to 19 months. In Keneba, where the mean daily riboflavin intake was 0.5 mg, the mean erythrocyte glutathione reductase activation coefficient was 1.78 and there was a marked deterioration of biochemical status near parturition. In Cambridge, where the mean intake was 2.3 mg/day, the mean activation coefficient was 1.19. A vitamin fortified diet supplement, given to lactating women in Keneba for 8 months, raised their mean riboflavin intake for 0.5 to 1.5 mg/day and reduced the mean activation coefficient to 1.42. It is therefore likely that an intake even greater than the current United Kingdom or United States recommended daily amounts would be needed to achieve biochemical 'normality' in these women.

1253 PHOTOTHERAPY AND RIBOFLAVIN STATUS

Erythrocyte glutathione reductase(EGR)activity has been used to detect riboflavin(B2) deficiency in premature infants under phototherapy(PHOTO). We investigated variables that might influence PHOTO induced B2 deficiency. 27 infants(BW 1.55 kg and GA 32.5 wks) were studied from 1 to 7 days of age. 14 Infants received PHOTO and 13 served as controls. In the PHOTO group, mean age of onset of PHOTO was 38.5±13.5 hours (range 15-52 hrs) and the mean duration of PHOTO was 39.7±23 hrs (range 17-95 hrs). The mean daily irradiance administered via continuous blue or white PHOTO was 7.8±5.9 nm/cm2 (range 2.8-17 nm/cm2) in the blue spectral range. The PHOTO group had a mean pre-treatment bilirubin of 7.7±1.3 mg/dl and post-treatment bilirubin of 7.6±1.9 mg/dl. Food samples were analyzed for B2 content and the calculated B2 intake for the PHOTO group was 0.09±.09 mg/day and for the control group was 0.14±.12 mg/day. EGR activity as expressed by the activity coefficient(AC) was measured daily. There were no significant differences in the ACs of the control or PHOTO groups. No infants of either group had evidence of B2 deficiency, i.e., AC>1.20. Although our mean duration of PHOTO was less than in previous reports of PHOTO-induced B2 deficiency, 5 of our infants who had normal ACs received >40 hrs of PHOTO with moderate to high irradience and had B2 intakes of ≥0.1 mg/day. B2 deficiency in the premature under PHOTO does not necessarily occur and is not strictly a function of duration of PHOTO or B2 intake.