Relationship between vitamin B-6 status and elevated pyridoxal kinase levels induced by theophylline therapy in humans

Theophylline administration to seven healthy male volunteers resulted in a rapid and significant decline in both plasma and erythrocyte pyridoxal-5'-phosphate levels. Total erythrocyte pyridoxal kinase levels increased during 15 wk of theophylline treatment from a mean initial activity of 19.23 +/- 5.03 (mean +/- SD) to 62.64 +/- 11.59 nmol pyridoxal-5'-phosphate formed/(g hemoglobin.h). Although plasma pyridoxal levels remained normal, the threefold increase in total erythrocyte pyridoxal kinase activity levels did not normalize plasma and erythrocyte pyridoxal-5'-phosphate levels. Pyridoxal-5'-phosphate hydrolysis was not affected by theophylline therapy. Increased pyridoxal oxidation was confirmed by elevated urinary 4-pyridoxic acid excretion after 15 wk of theophylline treatment. Mean erythrocyte alanine aminotransferase activity declined by 70%, and aspartate aminotransferase activity declined by 50%, indicating that decreased availability of pyridoxal-5'-phosphate can have widespread metabolic consequences. We conclude that the effect of theophylline on vitamin B-6 metabolism is not transitory and cannot be overcome by elevated intracellular levels of pyridoxal kinase. However, pyridoxine supplementation (10 mg/d for 1 wk) normalized indices of vitamin B-6 status and reversed the downward trend in both alanine aminotransferase and aspartate aminotransferase activity levels.     

Vitamin B6 nutritional status in asthma: the effect of theophylline therapy on plasma pyridoxal-5'-phosphate and pyridoxal levels

Plasma pyridoxal-5'-phosphate concentrations were significantly lower (p less than 0.001) in a group of 28 asthmatic women when compared to 33 controls. Plasma pyridoxal levels in the two groups were not different. Theophylline was administered to a group of 17 volunteers and resulted in large reductions in plasma pyridoxal-5'-phosphate levels, while plasma pyridoxal levels and urinary 4-pyridoxic acid excretion were unaffected by theophylline therapy. An in vitro study showed that theophylline did not interfere with the high performance liquid chromatography assay for pyridoxal-5'-phosphate, indicating that theophylline could affect liver metabolism of vitamin B6.     

Vitamin B-6 intakes and plasma B-6 vitamer concentrations of men and women, 19-50 years of age

The vitamin B-6 intakes and plasma B-6 vitamer levels of healthy nonsupplemented men and women, 19-24 and 25-50 years, were compared. The subjects did not take nutrient supplements or medications or use tobacco products. Subjects were grouped as follows: eight, 19-24 y men; nine, 25-50 y men; 11, 19-24 y women; and 13, 25-50 y women. The estimated vitamin B-6 intakes, obtained via 24-h recalls followed by 2-d food records, of the two groups of men were significantly higher (P < 0.05) than those of the two groups of women. Thirty-five percent of the women reported consuming less than the Estimated Average Requirement for vitamin B-6. The four gender: age groups had similar B-6 vitamer concentrations of plasma pyridoxal-5'-phosphate, 4-pyridoxic acid, pyridoxine, pyridoxamine, and pyridoxamine-5'-phosphate. Males 25-50 y had significantly higher (P < 0.05) plasma pyridoxal concentrations than the two groups of females. All subjects had pyridoxal-5'-phosphate concentrations indicative of vitamin B-6 adequacy. Generally the plasma B-6 vitamer concentrations of these men and women, 19-24 and 25-50 years of age, all having adequate vitamin B-6 status, were similar.     

Relationship between maternal and neonatal vitamin B6 metabolism: perspectives from enzyme studies.

Enzymes involved in vitamin B6 metabolism, i.e., pyridoxal kinase, pyridoxamine (pyridoxine) 5'-phosphate oxidase, and pyridoxal 5'-phosphate phosphatase, were assayed in hemolysates prepared from cord, maternal, and control blood samples. Mean cord and control pyridoxamine (pyridoxine) 5'-phosphate oxidase activities were significantly higher than maternal activities (p less than 0.001 and p less than 0.05, respectively). A significant correlation (p less than 0.001) was observed between maternal and cord vitamin B6-metabolizing enzymes. Cord pyridoxal 5'-phosphate levels correlated significantly with maternal pyridoxal 5'-phosphate levels (p less than 0.001) and with cord pyridoxal kinase activity (p less than 0.05). Maternal pyridoxal 5'-phosphate levels appear to be the most important factor determining fetal vitamin B6 status.     

Activities of the hepatic enzymes of vitamin B6 metabolism for patients with cirrhosis

Patients with cirrhosis and other hepatic diseases frequently exhibit lower concentrations of plasma pyridoxal 5'-phosphate (PLP), which is derived primarily from liver. To determine the biochemical basis for this abnormality, the enzymes of vitamin B6 metabolism--pyridoxal kinase, pyridoxine (pyridoxamine) 5'-phosphate oxidase, PLP phosphatase(s), and pyridoxal oxidase(s)--were analyzed in liver. The activities of the two biosynthetic enzymes, pyridoxal kinase and pyridoxine (pyridoxamine) 5'-phosphate oxidase were similar for both. The phosphatase activities were significantly higher (mean +/- SD of 9.55 +/- 8.03 versus 3.97 +/- 2.36 nmol X min X mg protein, p less than 0.05) for cirrhotics. Pyridoxal oxidase activities appeared slightly lower for cirrhotics. There was considerable variation in many indices of liver function, which suggests that the defects contributing to altered vitamin B6 metabolism may be complex and individualistic. These analyses have shown that cirrhotics are capable of apparently normal PLP synthesis and that increased hepatic dephosphorylation may be responsible for low levels of plasma PLP.     

Pyridoxine supplementation: effect on lymphocyte responses in elderly persons

The effect of pyridoxine supplementation on lymphocyte responsiveness was investigated in 15 persons aged 65-81 y. Eleven subjects received 50 mg/d pyridoxine HCl (PN). Four subjects received a placebo. Lymphocyte proliferation to T and B cell mitogens, lymphocyte subpopulations with monoclonal antibodies, and plasma pyridoxal 5'-phosphate (PLP) were measured before and after 1 and 2 mo of supplementation. After 1 and 2 mo plasma PLP levels increased by 195 +/- 88 nM and 201 +/- 84 nM, respectively, in subjects receiving PN. With PN supplementation, lymphocyte proliferation increased significantly in response to phytohemagglutinin (p less than 0.01), pokeweed mitogen (p less than 0.01), and Staphylococcus aureus (Cowain I) (p less than 0.05). For PN-treated subjects with low presupplement plasma PLP levels, lymphocyte blastogenesis also increased significantly (p less than 0.01) in response to concanavalin A. Percentages of T3+ and T4+ but not T8+ cells increased significantly (p less than 0.05) in PN-treated subjects. These results suggest that improving vitamin B-6 status is important in stimulating immunocompetence in the elderly.     

Effect of oral vitamin B6 supplementation on in vitro platelet aggregation

A randomized, double-blind study was conducted with 12 healthy adult males to determine the effects of oral pyridoxine supplementation on in vitro platelet aggregation. Following a 4-wk baseline period, half the subjects received 100 mg/day of pyridoxine X HCl while the remaining subjects received a placebo for 6 wk. In vitro platelet responses to ADP and collagen and the plasma pyridoxal 5'-phosphate (PLP) concentrations were measured at biweekly intervals. Plasma PLP concentrations increased significantly (p less than 0.001) for those receiving the vitamin B6 compared to baseline values or compared to those receiving the placebo. However, there was no significant effect of increased levels of plasma PLP on collagen-stimulated platelet aggregation and only a slight effect on ADP-stimulated aggregation. Acute administration of 100 mg pyridoxine X HCl failed to alter the in vitro response of platelets to either ADP or collagen. Reevaluation of conclusions based solely on in vitro studies suggesting the use of pyridoxine as an effective in vivo antithrombotic agent may be warranted.     

Effect of maternal pyridoxine X HCl supplementation on the vitamin B-6 status of mother and infant and on pregnancy outcome

The effect of maternal pyridoxine X HCl (PN-HCl) supplementation on the vitamin B-6 status of pregnant women and their infants at birth and on pregnancy outcome was investigated. Volunteer subjects were randomly assigned a daily vitamin B-6 supplement containing 0, 2.6, 5, 7.5, 10, 15 or 20 mg of PN-HCl in a double-blind study. The mean dietary vitamin B-6 intake of the group was 1.43 +/- 1.28 mg/day as estimated from 24-hour dietary recalls. Maternal plasma pyridoxal 5'-phosphate (PLP) levels were positively correlated with vitamin B-6 supplementation at 30 weeks gestation (r = 0.55, P less than 0.0005) and at delivery (r = 0.54, P less than 0.01). Cord plasma PLP levels reached a maximum when maternal PN-HCl supplementation was 7.5 mg and greater. Supplemental PN-HCl at the 7.5-mg level was required to prevent a decrease in maternal plasma PLP at delivery. Apgar scores at 1 minute after birth were higher (P less than 0.05) for infants whose mothers took 7.5 mg or more supplemental PN-HCl than for infants of mothers who took 5 mg or less. These findings indicate that a vitamin B-6 intake between 5.5 and 7.6 mg/day (diet plus supplement as pyridoxine equivalents) was required to maintain maternal plasma PLP levels at term at a level comparable to initial values.     

Pyridoxal-5'-phosphate and pyridoxal biokinetics in male Wistar rats fed graded levels of vitamin B-6

Biokinetic parameters of plasma pyridoxal-5'-phosphate (PLP) and pyridoxal (PL) disposition were studied in male Wistar rats (age 8 mo) fed a purified diet containing less than 0.5, approximately 3 or approximately 6 mg pyridoxine.HCl/kg diet from weaning, with animals fed the 6 mg/kg diet serving as the control group. Basal plasma PLP concentration was lower in both the less than 0.5 and 3 mg/kg diet groups than in control animals (98 +/- 12, 314 +/- 40 and 514 +/- 56 nmol/L, respectively). Basal plasma PL concentration was lower in the less than 0.5 mg/kg diet group only [60 nmol/L (measured in pooled samples), 190 +/- 73 and 235 +/- 63 nmol/L for less than 0.5, 3 and 6 mg/kg diet groups, respectively]. In both the less than 0.5 and 3 mg/kg diet groups, PLP clearance was lower than in control rats (0.158 +/- 0.025, 0.131 +/- 0.040 and 0.240 +/- 0.051 L.h-1.kg body weight-1, respectively). In the less than 0.5 mg/kg diet group, PLP synthesis was more efficient than in control animals (34.7 +/- 9.3, 12.1 +/- 2.5 and 16.7 +/- 11.4% for less than 0.5, 3 and 6 mg/kg diet groups, respectively). In both the less than 0.5 and 3 mg/kg diet groups, volume of distribution of PLP as well as of PL was larger than in controls. It is concluded that B-6 vitamer metabolism is influenced by vitamin B-6 status. The metabolic pathway involved (PLP synthesis and/or PLP degradation) was observed to depend on degree of vitamin B-6 deficiency.     

Transport and metabolism of pyridoxine and pyridoxal in mice

[3H]Pyridoxine or [3H]pyridoxal in physiological amounts was orally administered to mice and the distribution of isotope between the six recognized forms of vitamin B6 and pyridoxic acid was determined at different times after the administration in the intestine, liver, blood, and brain. After 7 min about 50% of the radioactivity in pyridoxine and pyridoxal had been absorbed by the intestine and transported to the blood and other organs. When [3H]pyridoxine was administered, labeled pyridoxal, pyridoxal- and pyridoxine-phosphate were found in the intestine and liver, and labeled pyridoxine could not be detected in the peripheral blood but substantial amounts of labeled pyridoxal and pyridoxal-phosphate were found in the blood. The time course of the blood [3H]pyridoxal levels following the administration of [3H]pyridoxine was similar to that following the administration of [3H]pyridoxal. These results suggest that the intestine and/or liver play a major role in converting dietary pyridoxine to circulating pyridoxal which is taken up and phosphorylated by other organs. Moreover, most of the blood [3H]pyridoxal was shown to be located in the plasma. This localization may facilitate utilization by the organs.     

Retention of 14C label is lower in old than in young Wistar rats after oral dosing with [14C]pyridoxine.

Ten young (6-mo-old) and ten old (31-mo-old) male Wistar rats fed a purified diet (250 g casein and 6 mg pyridoxine.HCl per kg) from weaning were given either a single oral dose or five repeated oral doses of 14C-labeled pyridoxine. At various times after dosing animals of each age group were killed. Absorption of orally dosed [14C]pyridoxine.HCl was not found to be different between young and old rats. Total body retention of 14C label administered was modestly but significantly lower in old than in young rats. However, distribution of 14C label over various tissues and among the various B-6 vitamers was similar. No significant age-related differences were observed in the biokinetic parameters derived from urinary excretion data. Contrary to the findings for 14C label distribution, age-related differences were observed for absolute level of tissue 14C-labeled vitamers. The lower [14C]pyridoxal-5'-phosphate content in liver and muscle, and [14C]pyridoxamine-5'-phosphate content in liver, of old animals indicated an age-related difference in liver and muscle vitamin B-6 disposition. In both young and old rats, and in both liver and muscle tissue, pyridoxamine-5'-phosphate was observed to be a faster-exchanging tissue vitamin B-6 pool than pyridoxal-5'-phosphate.     

Effect of oral contraceptives and pyridoxine on the metabolism of vitamin B6 and on plasma tryptophan and alpha-amino nitrogen.

The effect of supplementary pyridoxine on the metabolism of vitamin B6 as well as plasma tryptophan and alpha-amino nitrogen was determined in women using oral contraceptive agents. Ten women who were taking oral contraceptive agents and 11 who had never taken them served as subjects. Blood from the various biochemical measurements was drawn from fasting subjects before and after they had received an oral dose of 50 mg of pyridoxine-HCl daily fo 2 days. The use of oral contraceptive agents had no effect on the levels of blood vitamin B6, plasma pyridoxal phosphate, and plasma tryptophan. The activity of erythrocyte glutamic oxaloacetic transaminase was higher (P less than 0.05) in the oral contraceptive agent users than in the nonusers but the stimulation in vitro by pyridoxal phosphate was similar for the two groups. Plasma alpha-amino nitrogen was slightly lower in the oral contraceptive agent users than in the nonusers, but the difference was not statistically significant. The rise in blood vitamin B6 in response to pyridoxine was similar in the two groups, but the rise in plasma pyridoxal phosphate tended to be lower in the oral contraceptive agent treated subjects. Following pyridoxine supplementation, the basal activity of erythrocyte glutamic oxalocetic transaminase increased (P less than 0.01) in both groups of subjects and the stimulation in vitro by pyridoxal phosphate decreased correspondingly. Plasma tryptophan and alpha-amino nitrogen were unaffected by the supplementary pyridoxine.     

Vitamin B-6 deficiency prolongs the time course of evoked dopamine release from rat striatum

Vitamin B-6-deficient animals exhibit motor abnormalities. To investigate the possible physiologic alterations in the dopaminergic nervous system in vitamin B-6 deficiency, dopamine release in the striatum of vitamin B-6-deficient rats was determined using in vivo electrochemistry. Male Sprague-Dawley rats, 3 wk old, weighing 50-60 g, were randomly assigned to a control (7 mg pyridoxine HCl/kg diet), vitamin B-6-deficient (0 mg pyridoxine HCl/kg diet), or pair-fed (7 mg pyridoxine HCl/kg diet) group. After 8 wk of dietary treatment, plasma concentrations of pyridoxal 5'-phosphate as well as the striatal pyridoxal 5'-phosphate and pyridoxamine 5'-phosphate were significantly lower in the vitamin B-6-deficient group than in the control and pair-fed groups. The dopamine concentrations of the striatum and the magnitude of the dopamine release after local application of KCl did not differ among the groups. However, the time required for KCl-evoked dopamine release to reach its peak level was significantly longer for the vitamin B-6-deficient rats than for controls. In addition, the decay time from the peak to one-half of the KCl-evoked dopamine release was also significantly prolonged in vitamin B-6-deficient rats compared with the control group. The results indicate that the cellular content of dopamine does not reflect the functional state of dopaminergic neurons in vitamin B-6 deficiency. The time course for release of dopamine and decay of the released dopamine is prolonged by vitamin B-6 deficiency, which might contribute to the motor abnormalities of the deficient rats.     

Vitamin B6 levels in rheumatoid arthritis: effect of treatment.

Patients with rheumatoid arthritis were found to have low plasma pyridoxal phosphate. Treatment with 50-150 mg/day of pyridoxine hydrochloride for 3 months caused a rise in pyridoxal phosphate in most cases to more than 3 times pretreatment levels. There was, however, no improvement in the clinical aspects of rheumatoid arthritis.     

Differences in vitamin B6 status indicator responses between young and middle-aged women fed constant diets with two levels of vitamin B6

The responses of adult women to two levels of vitamin B6, was conducted with five young and eight middle-aged subjects. A constant diet (2.3-2.4 mg vitamin B6 per day) was fed for four weeks followed by three weeks of the same diet supplemented with 8.0 mg pyridoxine. Plasma pyridoxal 5'-phosphate (PLP), plasma and urinary vitamin B6, and urinary 4-pyridoxic acid (4-PA) were determined. The older women had significantly lower plasma PLP, plasma and urinary vitamin B6 and slightly higher urinary 4-PA values on normal vitamin B6 intakes. With supplementation, only the difference in urinary total vitamin B6 remained significant. Tryptophan load tests revealed no significant between-group differences in xanthurenic acid or kynurenic acid excretion. These results demonstrate an age-related difference in vitamin B6 status indicators in women under controlled dietary intake of vitamin B6.     

Vitamin B-6 nutrition status and cigarette smoking

We investigated the vitamin B-6 status in smokers, nonsmokers, and exsmokers by measuring both B-6 aldehyde vitamers, pyridoxal-5'-phosphate (PLP) and pyridoxal (PL), in the plasma as well as in the erythrocyte compartment. Two hundred eighty-six healthy, sedentary male workers from a middle-income group were investigated. There were 159 smokers, 59 exsmokers, and 68 nonsmokers. Plasma PLP and PL concentrations were significantly lower in smokers than in the nonsmokers and exsmokers whereas erythrocyte PLP and PL did not differ significantly between groups. Because PLP mainly functions as an intracellular coenzyme, the clinical significance of a depressed plasma PLP concentration alone is uncertain. It is concluded that circulating plasma PLP is labile and not necessarily indicative of intracellular PLP concentrations. The measurement of erythrocyte PLP and/or PL may be more informative about vitamin B-6 status than is plasma PLP alone.     

Genetic polymorphism of glutamate-pyruvate transaminase (alanine aminotransaminase): influence on erythrocyte activity as a marker of vitamin B-6 nutritional status

A new, sensitive, two-step method free from interference by hemoglobin that measures erythrocyte glutamate-pyruvate transaminase (E-GPT) activity is described. Several aspects of E-GPT activity as an index of vitamin B-6 nutritional status were investigated with this method. 1) GPT shows a structural genetic polymorphism with two common alleles resulting in three phenotypes. In a population study (n = 92) E-GPT activity differed significantly (p less than 0.001) among the three phenotypic groups. Plasma pyridoxal-5'-phosphate concentrations in the three groups did not differ significantly. Therefore, E-GPT activity can only be used to assess vitamin B-6 nutritional status if GPT phenotype is accounted for. 2) Pyridoxine supplementation (10 mg/d) significantly (p less than 0.0001) increased E-GPT activity and decreased (p less than 0.0001) the percentage stimulation by pyridoxal-5'-phosphate in vitro although the absolute amount of in vitro stimulation by pyridoxal-5'-phosphate changed only marginally. 3) Inorganic phosphate inhibits in vitro activation of E-GPT by pyridoxal-5'-phosphate.     

Vitamin B-6 metabolism in premature infants

The response of premature infants to intravenous pyridoxine or pyridoxal was studied by measuring serum and erythrocyte pyridoxal 5'-phosphate (PLP). In the first study serum PLP was measured in 28 infants periodically through day 28. Infants less than 30 wk gestational age (GA) had no serum PLP response to the administration of pyridoxine. Infants greater than or equal to 30 wk GA had significantly greater concentrations of PLP by day 3. In the second study there was a negligible response of serum PLP in nine infants less than or equal to 28 wk GA to supplementation of pyridoxine or pyridoxal. However, erythrocyte PLP and whole-blood total vitamin B-6 concentrations increased in both groups, indicating the presence of a substantial amount of the vitamin in the circulation of the infants. Whereas the functional significance of these observations is not known, it appears that in premature infants, serum PLP may not be an appropriate indicator of vitamin B-6 status.     

Effect of pyridoxal 5'-phosphate on the 1,25-dihydroxyvitamin D3 receptor system

The effect of pyridoxal 5'-phosphate on the 1,25-dihydroxyvitamin D3 receptor system has been studied by using pig intestinal chromatin. Pyridoxal 5'-phosphate did not affect the binding of 1,25-dihydroxyvitamin D3 to its receptor extracted from chromatin with hypertonic KCl, although in the presence of pyridoxal 5'-phosphate 1,25-dihydroxyvitamin D3-receptor complexes were not readily precipitated with polyethylene glycol. In contrast, pyridoxal 5'-phosphate showed a potency to dissociate the 1,25-dihydroxyvitamin D3 receptor from chromatin in a dose-dependent manner. A low concentration of pyridoxal 5'-phosphate was as effective as hypertonic KCl in dissociating the receptor from chromatin, while pyridoxine, p-nitrophenyl phosphate, or inorganic phosphate was much less effective. These observations suggest the inhibitory effect of pyridoxal 5'-phosphate on the recognition of 1,25-dihydroxyvitamin D3 by its receptor system.     

Elevated plasma vitamers of vitamin B6 in patients with chronic renal failure on regular haemodialysis.

Plasma pyridoxal-5'-phosphate (PL-5'-P), pyridoxal (PL), pyridoxine (PN), and 4-pyridoxic acid (4-PA) were measured by high-performance liquid chromatography (HPLC) in 39 patients (15 male, 24 female) with chronic fenal failure undergoing regular haemodialysis and 46 healthy controls (28 male, 18 female). All three vitamers of vitamin B6 and the metabolite were significantly elevated in the haemodialysis patients. Mean PL-5'-P and PN concentrations were 20 times the mean in controls. Only one patient took a vitamin B6 supplement. In view of the neurotoxicity of supranutritional intakes of PN in normal humans we suggest that supplements of PN be carefully monitored when administered to patients with chronic renal failure.