In vitro studies of fatty acid metabolism in vitamin B6 deficient rats.

The oxidation of [1-14C]palmitate, [1-14C]linoleate, and [1-14C]arachidonate as well as their incorporation into lipids was investigated in vitamin B6 deficient rats using liver slices and isolated mitochondria. In experiments on fatty acid oxidation in liver slices, we observed a decrease in the production of 14CO2 from radioactive palmitate and linoleate, but not from arachidonate. In the case of mitochondria, we detected no difference between the deficient rats and the controls with linoleats; however, there was an increase in the oxidation of palmitate in the deficient rats as compared to the controls. Experiments on the incorporation of radioactivity into lipids showed an increase in radioactivity from [1-14C]palmitate and [1-14C]linoleate in the triglyceride, phospholipid and cholesterol fractions of deficient rats. In the case of [1-14C]arachidonate, no difference was observed between the two groups of rats. These experiments showed that major changes occurred in metabolism of palmitate and linoleate and strongly suggested that the decrease in the arachidonate content previously observed in rat tissues could not be explained by alteration in the metabolism of this fatty acid.     

Oxidant/antioxidant status in liver tissue of vitamin B6 deficient rats

BACKGROUND AND AIMS: The aim of this study was to evaluate the lipid peroxidation and antioxidant function in liver tissue of vitamin B6 deficient rats and investigate relationship among these parameters in either group. METHODS: Twenty-four male rats with a weight of 48-59 g were used for the experiment. The rats were divided into control (n=12) and vitamin B6 deficient groups. After 4 weeks of feeding, the rats were killed by cervical dislocation and liver tissues were removed. Biochemical measurements in liver tissue were carried out using a spectrophotometer. RESULTS: Liver tissue antioxidant potential, superoxide dismutase, glutathione peroxidase, glutathione-S-transferase, total (enzymatic plus non-enzymatic) superoxide scavenger activity, non-enzymatic superoxide scavenger activity, glutathione levels were significantly lower in vitamin B6 deficient rats than in control group. However, liver tissue glutathione reductase activity, and MDA values were significantly higher in vitamin B6 deficient rats than in control group. CONCLUSIONS: These results explicitly indicate that vitamin B6 deficiency causes a decrease in antioxidant defense system and an increase in oxidant stress in liver tissue in rats.     

Levels of neurotransmitters in brain of vitamin B12 deficient rats.

The levels of norepinephrine, dopamine and 5-hydroxytryptamine in brain homogenates of vitamin B12-deficient rats have been investigated. The norepinephrine levels were significantly decreased in the deficient animals compared to controls. The two major catabolic pathways of norepinephrine e.g. monoamine oxidase and catechol-O-methyl transferase did not show significant variations. Both acetyl cholinesterase and butiryl-cholinesterase markedly decreased in the plasma of the vitamin B12-deficient rats.     

Induction of pyridoxal phosphate-dependent enzymes in vitamin B-6 deficient rats.

The effectiveness of dietary and hormonal treatments in inducing several pyridoxal phosphate-(PLP)-dependent enzymes has been examined in vitamin B-6 deficient rats. Holo- and apoenzymes of serine dehydratase and ornithine aminotransferase were inducible in both control and deficient rats by feeding them 80% casein diets or by injecting them with glucagon. Holo- and apotyrosine aminotransferase were induced in both control and deficient rats by injecting them with glucagon or with dexamethasone phosphate. Phosphoenolpyruvate carboxykinase, a non-PLP-dependent enzyme, was inducible in both control and deficient rats by glucagon treatment if the rats were fed, but not if they had been starved. The degree of induction of certain enzymes depended upon whether rats were fed ad libitum, starved overnight, or fed a protein-free diet prior to the induction period. Phosphoenolpyruvate carboxykinase activities were about the same in both control and deficient rats. In vitamin B-6 deficient rats, both uninduced and induced activities of serine dehydratase, ornithine aminotransferase, and tyrosine aminotransferase assayed in the prsence of PLP, but not in its absence, either equaled or exceeded control values under most experimental conditions. Synthesis of excess of apoenzyme of PLP-dependent enzymes generally accounted for the high total enzyme activity in deficient rats. Differences between values for control and deficient rats could not be accounted for by differences in liver cyclic AMP concentrations nor were they apparently related to reduced food intake of the deficient rats. High apoenzyme concentration during depletion of coenzyme would tend to prevent depletion of active enzyme.     

Cystathionine excretion in relation to vitamin B-6 nutriture of rats: strain differences and effects of DL- or L-methionine supplementation.

Wistar and Long-Evans rats excreted significantly more cystathionine during days 27 to 31 of a vitamin B-6 deficiency than did Spague-Dawley rats. Wistar rats were therefore chosen for experiments designed to test the effects of dietary DL- and L-methionine supplementation upon cystathionine excretion of vitamin B-6-deficient and control rats. Methionine supplementation did not significantly affect mean daily food consumption or weight gain of the control animals. L-Methionine-supplemented, deficient rats gained significantly more during the first 15 days of the experiment and less during the last 11 days than did the other deficient groups. No cystathionine was detected in the urine of methionine-supplemented or unsupplemented control rats between days 8 and 26 of the experiment; but measurable quantities of cystathionine were present in the urine of all deficient groups by days 8 to 12. Deficient animals fed DL-methionine excreted up to 2.33 mg more cystahionine per 4-day period than did the unsupplemented deficient rats, whereas rats fed L-methionine excreted up to 7.44 mg more per 4-day period than did the unsupplemented ones. This difference in cystathionine response was highly significant and indicates that the L supplement stressed the vitamin B-6-deficient rats more than did the DL supplement. An explanation for this may be that the D isomer in the DL supplement was not as efficiently absorbed and/or metabolized as the L form.     

Stability of some pyridoxal phosphate-dependent enzymes in vitamin B-6 deficient rats.

The effects of lowering the liver pyridoxal phosphate (PLP) concentration by vitamin B-6 deficiency on the stability of several rat liver enzymes were examined. Three PLP-dependent enzymes (serine dehydratase, ornithine-delta-aminotransferase, and tyrosine aminotransferase) and two non-PLP-dependent enzymes (glucose-6-phosphate dehydrogenase and phosphoenolpyruvate carboxykinase) were induced in vitamin B-6 deficient and control rats by feeding them high-protein diets or by injecting them with glucagon or dexamethasone. The decline of each activity was followed after withdrawal of the inducer. Serine dehydratase activity declined more rapidly in vitamin B-6 deficient than in control liver; however, ornithine aminotransferase and tyrosine aminotransferase activities were equally stable in deficient and control liver. Ornithine aminotransferase was predominantly in holoenzyme form in both control and deficient rats, whereas tyrosine aminotransferase was predominantly in apoenzyme form in both groups. The proportion of serine dehydratase in apoenzyme was less stable than the holoenzyme. Activity changes of glucose-6-phosphate dehydrogenase and phosphoenolpyruvate carboxykinase in control and vitamin B-6 deficient rats were similar. The results suggest that differences in the stability of PLP-dependent enzymes in vitamin B-6 deficient rats depend upon differences in the proportions of these enzymes existing as holo- and apoenzyme.     

Acute ethanol ingestion by humans and subacute treatment of rats increase urinary folate excretion

Previous studies with rats showed that acute treatment with ethanol (4 g/kg) produce a marked increase in urinary folate levels, followed by a decrease in plasma folate levels. Analogous studies with human volunteer subjects using a lower dose of ethanol showed that there were small, but statistically significant increases in urinary folate levels after four hours. The initial ethanol dose was 1.0 g/kg with a single supplement of 0.1-0.2 g/kg to maintain ethanol blood levels at about 100 mg/dl for six hours. Further studies with rats were designed to test the cumulative effects of repeated daily doses of ethanol. Male Sprague-Dawley rats were treated for 1, 2, 3, or 4 days either with ethanol orally in 4 doses of 1 g/kg each at 0, 1, 2, and 3 hours or with glucose orally in 4 isocaloric doses. Urine was collected at timed intervals up to 12 hours after each daily dose. The pattern of the increase in urinary folate levels was similar in all groups, whether treated for 1, 2, 3 or 4 days. These results suggest that repeated ethanol treatment can lead to a marked cumulative folate loss via increased urinary excretion and that increased urinary folate excretion may contribute to the development of folate deficiency in humans.     

B vitamin supplementation reduces excretion of urinary dicarboxylic acids in autistic children

Urinary dicarboxylic acids are an important source of information about metabolism and potential problems especially connected with energy production, intestinal dysbiosis, and nutritional individuality in autistic children. A diet rich in vitamins and macroelements is a new idea of intervention in autism. The objective of the present study was to test the hypothesis that vitamin B2, vitamin B6, and magnesium supplementation is effective in reducing the level of dicarboxylic acids in the urine of autistic children. We examined the levels of succinic, adipic, and suberic acids in the urine of autistic children before and after vitamin supplementation. Thirty children with autism received magnesium (daily dose, 200 mg), vitamin B6 (pyridoxine; daily dose, 500 mg), and vitamin B2 (riboflavin; daily dose, 20 mg). The treatment was provided for a period of 3 months. Organic acids were determined using gas chromatography/mass spectrometry. Before supplementation, the levels of succinic, adipic, and suberic acids in the urine of autistic children were 41.47 ± 50.40 μmol/mmol creatinine, 15.61 ± 15.31 μmol/mmol creatinine, 8.02 ± 6.08 μmol/mmol creatinine; and after supplementation, the levels were 9.90 ± 8.26 μmol/mmol creatinine, 2.92 ± 2.41 μmol/mmol creatinine, and 2.57 ± 3.53 μmol/mmol creatinine, respectively. The results suggest that the supplementation reduces the level of dicarboxylic acid in the urine of autistic children.     

The differences of the metabolism related to vitamin B6-dependent enzymes among vitamin B6-deficient germ-free and conventional rats.

The differences of the metabolism related to vitamin B6-dependent enzymes were investigated using germ-free and conventional rats. There was a significant difference in the boyd weight gain between vitamin B6-deficient germ-free and conventional rats after about 30 days of the experiment, and the body weight gain was much less in the deficient germ-free rats than in the deficient conventional ones. Urinary excretion of xanthurenic acid was higher in the deficient germ-free rats than in the deficient conventional ones after 18 days. There was a significant difference in the activities of kynurenine aminotransferase in mitochondrial fractions of germ-free rats, but not in mitochondrial fractions of conventional ones. The activities of aspartate and alanine aminotransferases, with or without pyridoxal phosphate, significantly decreased in the deficient germ-free rats, but not in the deficient conventional ones. These findings indicate that the degree of vitamin B6 deficiency was more severe in the deficient germ-free rats than in the deficient conventional ones, and also suggest that intestinal microflora may have some effects on vitamin B6-deficient conventional rats.     

Evaluation of urinary excretion of l-hydroxypyrene and thioethers in workers exposed to bitumen fumes

Summary Biological monitoring of exposure to bitumen fumes during road-paving operations was carried out. In order to evaluate the biological uptake of the workers, the nonselective urinary thioether assay and a selective method for the determination of urinary l-hydroxypyrene were used. Urinary thioether data of exposed workers were higher than those of nonexposed subjects. The effect of smoking, however, was stronger than the effect of occupational exposure. Levels of l-hydroxypyrene in road-paving workers were significantly higher than those in control subjects. The 1-hydroxypyrene level was also influenced by smoking habits, but the effect of occupational exposure was stronger. Our present data suggest that enhanced urine levels of both thioethers and 1-hydroxypyrene in bitumen workers may indicate an increased genotoxic risk. Furthermore, our results demonstrate the applicability of the 1-hydroxypyrene assay after occupational exposure to petroleum-based products.     

Quantitative assessment of motor and sensory function in vitamin B-6 deficient rats

Ataxia, weakness and changes in peripheral sensitivity are consequences of vitamin B-6 deficiency. This experiment was conducted to quantify these effects, using tests commonly employed in toxicology studies. Female Long-Evans rats (mean weight=232g) were blocked by weight into 3 treatment groups (n=12): AL-CON, fed AIN-76A diet ad libitum; DEF, fed the same diet devoid of pyridoxine; and PF-CON, pair-fed to DEF. Motor function was evaluated with hindleg gait analysis, grip-strength, and whole-body pull-up. Sensory function was evaluated using tail-flick and landing foot-spread. Deficiency was detected with a tryptophan-load test on day 9 of a simultaneous experiment with an identical feeding protocol. On day 9, abnormalities were detected and quantified by gait analysis—angle and width of step were affected by diet (P<0.05) although there were no differences in body weights and food intakes at that time. The overall effect of diet on angle and width was highly significant (P<0.001). Food intake of DEF was depressed by week 2; at week 10, body weight and feed efficiencies were significantly different among groups. Except for a transient effect on tail-flick, no other test was affected by diet. Gait analysis may have potential as an index of vitamin B-6 status under these experimental conditions.