Inter- and intra-individual variation in plasma and red blood cell vitamin E after supplementation.

To establish the range of individual blood responses to supplemental vitamin E, 30 healthy subjects ingested 75 mg of deuterium-labelled alpha-tocopherol with a standard breakfast. Blood was collected at 6, 9, 12, 27 and 51 h post ingestion and deuterated (d6) and non-deuterated (do) alpha-tocopherol concentrations were determined in plasma and red blood cells (RBC) by GC-MS. To examine intra-individual responses, 6 of these subjects were re-examined at 6-month intervals over a 30-month period. Post ingestion, the amount of d6-alpha-tocopherol in blood increased rapidly with time with maximal concentrations seen at 12 h (plasma) and 27 h (RBC) in most subjects. At these times, d6-alpha-tocopherol concentration ranged from 0.3-12.4 micromol/l in plasma and 0.6-4.09 micromol/l packed cell in RBC. Area under the curve calculations indicated inter-individual differences of alpha-tocopherol uptake to be 40-fold for plasma (12.9-493.3 micromol h/l) and 6-fold for RBC (24.4-146.1 micromol h/l packed RBC). Intra-individual variation in alpha-tocopherol uptake was small in comparison and remained relatively constant over the 30-month period. We conclude that vitamin E uptake varies widely in the normal population, although it is comparatively stable for an individual over time. These differences likely arise from variations in the regulation of vitamin E uptake and metabolism between subjects. Factors regulating this process must be better understood before the optimal intake of vitamin E can be ascertained.

The ferroxidase activity of caeruloplasmin is reduced in haemodialysis patients.

Increased free-radical production leading to oxidative stress may contribute to the development of cardiovascular complications in haemodialysis patients. The ferroxidase activity of caeruloplasmin forms an important component of antioxidant defences in body fluids. The aim of this study was to assess ferroxidase activity in haemodialysis patients. Venous blood was collected from 83 haemodialysis patients immediately prior to and after dialysis and from 52 healthy controls. Immunoreactive caeruloplasmin was measured by rate nephelometry, and ferroxidase activity determined by measuring loading of ferrous iron onto iron-free transferrin. A significant reduction in ferroxidase activity was observed in dialysis patients when compared with controls (37 +/- 1.20 and 46 +/- 1.14 mU/l, respectively; p < 0.001). Following dialysis, ferroxidase activity rose significantly to 41 +/- 1.16 mU/l, with a significant difference still remaining between control and patient ferroxidase activity (p < 0.005). Immunoreactive caeruloplasmin was found to be similar in all groups (before dialysis 0.40 +/- 0.07 g/l, after dialysis 0.39 +/- 0.07 g/l, control 0.42 +/- 0.09 g/l: p = NS). A significant difference in caeruloplasmin-specific activity was therefore observed between predialysis, postdialysis and control samples (97 +/- 2.31, 105 +/- 1.74 and 112 +/- 1.51 mU/g; p < 0.001, p < 0.01, respectively). Ferroxidase activity of caeruloplasmin is impaired in renal failure. Inhibition of caeruloplasmin ferroxidase activity in dialysis patients may contribute to increased oxidative stress in these patients.

Antioxidants, but not B-group vitamins increase the resistance of low-density lipoprotein to oxidation: a randomized, factorial design, placebo-controlled trial.

We have conducted an intervention trial to assess the effects of antioxidants and B-group vitamins on the susceptibility of low-density lipoprotein (LDL) to oxidation. A total of 509 men aged 30-49 from a local workforce were screened for total plasma homocysteine. The 132 selected (homocysteine concentration > or = 8.34 mumol/l) men were randomly assigned, using a factorial design, to one of four groups receiving supplementation with B group vitamins alone (1 mg folic acid, 7.2 mg pyridoxine, 0.02 mg cyanocobalamin), antioxidant vitamins (150 mg ascorbic acid, 67 mg alpha-tocopherol, 9 mg beta-carotene), B vitamins with antioxidant vitamins, or placebo. Intervention was double-blind. A total of 101 men completed the 8-week study. The lag time of LDL isolated ex vivo to oxidation (induced by 2 mumol/l cupric chloride) was increased in the two groups receiving antioxidants whether with (6.88 +/- 1.65 min) or without (8.51 +/- 1.77 min) B-vitamins, compared with placebo (-2.03 +/- 1.50) or B-vitamins alone (-3.34 +/- 1.08) (Mean +/- S.E., P < 0.001). Antibodies to malondialdehyde (MDA) modified LDL were also measured, but there were no significant changes in titers of these antibodies in any group of subjects whether receiving antioxidants or not. Contrast analysis showed that there was no interaction between antioxidants and B-group vitamins. This study indicates that while B-group vitamins lower plasma homocysteine they do not have an antioxidant effect. Thus B-group vitamins and antioxidants appear to have separate, independent effects in reducing cardiovascular risk.

Plasma glutathione peroxidase activity is reduced in haemodialysis patients.

Cardiovascular disease is the major cause of morbidity and mortality in patients with end-stage renal failure. Increased free radical production and antioxidant depletion may contribute to the greatly increased risk of atherosclerosis in these patients. Glutathione peroxidase (GPX) is an important antioxidant, the plasma form of which is synthesized mainly in the kidney (eGPX). The aim of this study was to assess the activity of eGPX in patients with end-stage renal failure on haemodialysis. Venous blood was collected from 87 haemodialysis patients immediately prior to and after dialysis and from 70 healthy controls. Serum eGPX activity was measured using hydrogen peroxide as substrate and immunoreactivity determined by ELISA. eGPX activity was significantly reduced in dialysis patients when compared to controls (106 +/- 2.7 and 281 +/- 3.6 U/l respectively, p < 0.001). Following haemodialysis, eGPX activity rose significantly to 146 +/- 3.8 U/l, p < 0.001, although remaining below control values (p < 0.005). Immunoreactive eGPX, however, was similar in all groups (pre-dialysis 14.10 +/- 1.26 microg/ml, post-dialysis 14.58 +/- 1.35 microg/ml, controls 15.20 +/- 1.62 microg/ml, p = NS). A decrease was observed in the specific activity of eGPX in patients when compared to controls (8.81 +/- 1.14, 10.71 +/- 1.54 and 21.97 +/- 1.68 U/mg respectively, p < 0.0001). eGPX activity is impaired in patients undergoing haemodialysis and so may contribute to atherogenesis in renal failure.

Carbamylation inhibits the ferroxidase activity of caeruloplasmin.

The ferroxidase activity of caeruloplasmin (EC 1.16.3.1) is an important antioxidant defence mechanism in man. In chronic renal failure proteins become carbamylated as a result of reactions with urea-derived cyanate. We have therefore investigated the effects of carbamylation on the ferroxidase activity of caeruloplasmin. Increasing degrees of carbamylation produce a progressive impairment of ferroxidase activity measured using o-dianisidine as substrate, and the ability of caeruloplasmin to load ferrous iron onto ovotransferrin is reduced. Carbamylation of caeruloplasmin may contribute to reduced antioxidant capacity in patients with renal failure.

Carbamylation of proteins and atherogenesis in renal failure.

The incidence of atherosclerosis is greatly increased in patients with chronic renal failure, and this increased risk is only partly explained by conventional risk factors. Carbamylation of proteins occurs in renal failure as a result of reactions between urea-derived cyanate and protein amino groups. A mechanism is proposed whereby oxidation of LDL cholesterol within the arterial wall may be enhanced as a result of carbamylation. This may be accentuated as a result of inhibition of antioxidant enzymes by carbamylation. The combination of these processes may lead to increased atherogenesis.