Carbohydrate metabolism in erythrocytes of copper deficient rats.

Dietary copper deficiency is known to adversely affect the circulatory system of fructose-fed rats. Part of the problem may lie in the effect of copper deficiency on intermediary metabolism. To test this, weanling male Long-Evans rats were fed for 4 or 8 weeks on sucrose-based diets containing low or adequate copper content. Copper deficient rats had significantly lower plasma and tissue copper as well as lower plasma copper, zinc-superoxide dismutase activity. Copper deficient rats also had a significantly higher heart:body weight ratio when compared to pair-fed controls. Direct measurement of glycolysis and pentose phosphate pathway flux in erythrocytes using (13)C NMR showed no differences in carbon flux from glucose or fructose to pyruvate but a significantly higher flux through the lactate dehydrogenase locus in copper deficient rats (approximately 1.3 times, average of glucose and glucose + fructose measurements). Copper-deficient animals had significantly higher erythrocyte concentrations of glucose, fructose, glyceraldehyde 3-phosphate and NAD(+). Liver metabolite levels were also affected by copper deficiency being elevated in glycogen and fructose 1-phosphate content. The results show small changes in carbohydrate metabolism of copper deficient rats.

Proposed phytic acid standard including a method for its analysis.

A method is described that accurately and rapidly quantifies the free and total phosphorous content of a commercially available, purified, phytic acid preparation. This allows its use as a standard for phytic acid determinations in foods. The method involves a wet ashing step followed by phosphorous measurement with a 1-amino-2-naphthol-4-sulfonic acid-molybdate reagent in a microplate reader at 660 nm. The procedure can be performed in 3 h with as little as 50 mg sample.

Elemental composition of anatomically distinct regions of rat liver.

Experiments were conducted to test the commonly held assumption that analysis of a portion of rat liver is representative of the elemental concentration of the whole organ. Male Sprague-Dawley rats (initial body weight approximately 250 g) fed a chow diet or weanling male Long-Evans rats (initial body weight approximately 50 g) fed a semipurified diet with or without copper in the mineral premix were sacrificed after 4 wk on their respective diets and livers were dissected into seven portions representing major anatomical divisions of this organ. Elemental analyses by atomic absorption spectroscopy (calcium, magnesium, iron, zinc, copper, manganese), atomic emission spectroscopy (sodium, potassium), or colorimetric assay (phosphorus) demonstrated no statistically significant differences in composition of these nine elements among anatomical regions of liver. Dietary copper deficiency led to equivalently reduced copper concentration in all portions of rat liver and did not cause any other significant alterations in liver composition of these nine elements within the 4 wk of these studies. These results confirm the validity of the common assumption that analysis of a portion of rat liver can be representative of the elemental composition of the whole organ. This conclusion will allow more analyses to be performed on fewer animals, thereby reducing animal use and reagent costs without sacrificing analytical accuracy.

A method for the determination of type I iodothyronine deiodinase activity in liver and kidney using 125I-labelled reverse triiodothyronine as a substrate.

OBJECTIVES: A detailed method for the determination of iodothyronine deiodinase type 1 (DI-1) activity is described. The objective of the present method development was to consolidate the effective procedures of previous methods and produce an efficient assay that can be easily reproduced. DESIGN AND METHODS: This method uses a 5',-125I labelled rT3 as substrate and ion-exchange chromatography to separate released ionic iodine. Released 125I- collected in the eluate is counted, and the results used to calculate DI-1 activity. RESULTS: Results were found to be linear for tissue homogenates containing 3-11 mg protein.mL-1. Day-to-day coefficient of variation of liver homogenate was determined to be 13%. CONCLUSIONS: This method was found to be reliable, reproducible, and sample sizes as small as 10 microL could be readily assayed. The use of centrifuge filter units to contain the ion-exchange medium decreased handling of the material, and potential sources of error.

Magnesium status and excretion in age-matched subjects with normal and elevated blood pressures.

Plasma, erythrocyte, and leucocyte magnesium (Mg) concentration, as well as urinary Mg, calcium (Ca), and phosphorus (P) excretion were determined in male subjects with elevated diastolic blood pressure (DBP) > 90 mmHg. These parameters were compared to those in normotensive age-matched (DBP < 85 mmHg) and drug-treated hypertensive subjects. Mg and Ca were determined by atomic absorption spectroscopy and P was determined by a colorimetric method. Urinary excretion was expressed in terms of creatinine (Cr) excretion. There were no significant differences in plasma, erythrocyte, or leucocyte Mg concentrations. Both urinary Ca and Mg were significantly decreased in the group with elevated blood pressure (BP) and the drug-treated group compared to the normotensive group. Urinary P was elevated in the drug-treated hypertensive group. Of all the variables studied, urinary Mg was the only one that was significantly inversely correlated with both systolic (SBP) and DBP. These data suggest that decreased urinary Mg, which may be indicative of a decreased Mg intake, is associated with hypertension.

Effects of dietary calcium on the metabolism of trace elements in male and female rats.

The effect of dietary calcium on the metabolism of iron, zinc, copper, and manganese in male and female rats was investigated. For 3 or 6 weeks the rats were fed three diets containing: (1) 0.26, (2) 0.52, or (3) 2.08% Ca. The apparent absorption of iron was depressed by the high calcium diet, and manganese absorption was highest in the low calcium groups. Generally there was a decrease in the absorption of minerals from 3 to 6 weeks. With an increase in the dietary calcium the absorption of Ca and P decreased. The liver iron concentration in the females fed diet 3 decreased from about 600 to 200 microg/g dry weight. The high calcium intake also caused a slight increase in the heart calcium levels in both sexes. However, diet 3 prevented kidney calcification in the female rats at 6 weeks and this was attributed to a dramatic decrease in the urinary phosphorus, although the calcium had increased about 40 times. In males, on the other hand, the high calcium diet caused some kidney calcification.

The effect of dietary copper and zinc on cholesterol metabolism.

The effects of different amounts of dietary zinc and copper on cholesterol metabolism in the rat were investigated. The levels of the minerals used were comparable to those likely to occur in a normal mixed North American diet. Although the different levels affected the serum, liver, and femur concentrations of the minerals, they had no effect on the levels of serum and liver, total, esterified or free cholesterol, or on high density lipoprotein cholesterol. It was concluded that dietary copper and zinc, at levels likely to occur in a normal mixed diet, are not significant factors in cholesterol metabolism.

Beneficial effect of zinc supplementation on reproduction in rats fed rapeseed protein concentrate.

Three groups of 33 90-day-old female Sprague-Dawley rats were fed, ad libitum, the following diets for 2 weeks before breeding. Diet 1 (D1) contained 20% protein from casein, diet 2 (D2) had the same level of protein from Tower rapeseed (Brassica napus) protein concontrate (RPC) and diet 3 (D3) was the same as D2 with a zinc supplement (70 mg/l) in the drinking water. From each group 6 animals were killed before breeding and 5-9 animals were killed at 1 and 2 weeks of gestation and post-partum. From each rat, blood, thyroids, liver and femur were collected for the determination of zinc, copper, iron, manganese, calcium and magnesium. As a measure of the reproductive performance, body weight, number of pups in the uterus or delivered live or dead, and gestations days before parturition were recorded. The pups were examined for obvious deformities and also analysed for the above mineral elements by atomic absorption spectroscopy. In group D2, levels of zinc in maternal serum, liver, femur and in the pups were significantly lower than the comparable levels in the other two groups. The zinc supplemented RPC-fed group did not show the anorexia experienced by the unsupplemented group and there was neither a significant difference between reproductive performances of groups D1 and D3 nor was there any significant difference between the zinc levels determined. It was concluded that the toxic symptoms caused by RPC feeding was attributable to zinc deficiency probably caused by the high phytate level in the RPC.

Beneficial effect of zinc supplementation on reproduction in rats red rapeseed protein concentrate.

Three groups of 33 90-day-old female Sprague-Dawley rats were fed, ad libitum, the following diets for 2 weeks before breeding. Diet 1 (D1) contained 20% protein from casein, diet 2 (D2) had the same level of protein from Tower rapeseed (Brassica napus) protein concontrate (RPC) and diet 3 (D3) was the same as D2 with a zinc supplement (70 mg/l) in the drinking water. From each group 6 animals were killed before breeding and 5--9 animals were killed at 1 and 2 weeks of gestation and post-partum. From each rat, blood, thyroids, liver and femur were collected for the determination of zinc, copper, iron, manganese, calcium and magnesium. As a measure of the reproductive performance, body weight, number of pups in the uterus or delivered live or dead, and gestation days before parturition were recorded. The pups were examined for obvious deformities and also analyzed for the above mineral elements by atomic absorption spectroscopy. In group D2, levels of zinc in maternal serum, liver, femur and in the pups were significantly lower than the comparable levels in the other two groups. The zinc supplemented RPC-fed group did not show the anorexia experienced by the unsupplemented group and there was neither a significant difference between reproductive performances of groups D1 and D3 nor was there any significant difference between the zinc levels determined. It was concluded that the toxic symptoms caused by RPC feeding were attributable to zinc deficiency probably caused by the high phytate level in the RPC.

Bioavailability of zinc in infant cereals.

The bioabailability of zinc in three cereals was determined by a rat assay, using total femur zinc as the indicator. The basal diet contained 25% egg white solids and 0.6 microgram zinc per gram. The infant cereals were included in the diets at three graded levels to supply 3--12 microgram zinc per gram. Zinc sulphate was used as a standard source, with and without supplemental iron (185, 370, 555 microgram/g, respectively) at 3, 6 and 9 microgram zinc per gram with a view to determine the effect of excess iron on zinc availability. Added iron from sodium iron pyrophosphate was not found to have any effect on the bioavailability of zinc from zinc sulphate. The responses for the three cereals were linear but the regression lines representing them and the standard source did not have a common intercept. However, the ratio of the slope of the regression line for the test source to that of the standard indicated that the cereal containing soy protein was a better source of zinc (ratio = 0.49) than the barley cereal (9.10) or the rice cereal (0.32). These ratios did not show any correlation with the actual iron-precipitable phytate phosphorus content or the reported crude fibre content.

Lead concentration in the livers of Canadians.

Liver specimens were collected at 114 (83 males and 31 females) autopsies on accident victims and on 4 stillborn infants in seven Canadian Cities. The age of the individuals ranged from newborn to 89 years with about 70% being 19--65 years. The level of lead in the samples was determined by atomic absorption spectroscopy using a carbon rod atomizer. The concentration of lead in liver was low (microgram/g: Mean, 0.37, Median 0.28) at birth and during infancy. In the stillborn infants it ranged from 0.30 to 0.77 (Mean 0.51). 7ale adults had an average level of 1.78 (Median 1.41) whereas in adult females the corresponding values were 0.65 and 0.57. The liver lead concentration peaked at about the end of the fourth decard in adult males. There was no indication of any regional differences.

Bioavailability of zinc in milk and soy protein-based infant formulas.

Total femur zinc of young rats was used to evaluate the biological availability of zinc in milk and soy protein-based infant formulas. A zinc deficient diet (0.8 mug Zn/g) containing egg white protein was supplemented with graded levels of zinc from zinc sulfate, milk and soy protein-based infant formulas. A plot of total femur zinc (log) after feeding the diet for 3 weeks versus the zinc added to the diet gave a linear relationship over the range of 0, 3, 6, 9 and 12 mug/g added zinc. By using a slope-ratio bioassay model, the relative biological availability of endogenous and added zinc in milk-based formula was estimated to be 0.86 and that of soy-based formula 0.67 (zinc sulphate = 1.00) with corresponding 95% fiducial limits being 0.82 to 0.91 and 0.62 to 0.71. Thus, to provide equivalent amounts of available zinc, the total zinc content of the soy protein-based formula would need to be at least 20% higher than that of the formula containing milk protein.

Liver storage iron in Canadians.

Liver samples were collected at 105 (78 males and 27 females) autopsies carried out on accident victims in seven cities of Canada. The age of individuals ranged from newborn to 89 years but about 75% of them were 19 to 65 years old. The liver iron stores of males were high at birth (150 to 300 mug/g), decreasing to around 100 mug/g in the second year. This level was maintained up to about 15 years and then increased to approximately 250 mug/g in 2 to 3 years. After 55 years, the hepatic iron stores tended to decrease. Although the number of samples from females was small, there was an indication of high stores (above 200 mug/g) at birth followed by a decline to values from 50 to 150 mug/g. This level was generally maintained in females throughout life. Storage iron in the livers of stillborn infants was high, ranging from 200 to 1600 mug/g. Analysis of liver samples from 64 persons (50 males and 14 females) who died of cardiovascular diseased showed that males under 45 years had lower iron stores than the corresponding accident victims.

Effect of the matrix of the standard on results of atomic absorption spectrophotometry of zinc in serum.

Zinc in pooled human serum, diluted 10-fold with demineralized water, was determined by flame atomic absorption spectrophotometry. Recoveries of zinc added to the serum were calculated by use of standards prepared in water, physiological saline, synthetic serum, or dilute HNO3 (10 ml/liter). The flow rates were essentially the same (9.1 to 9.3 ml/min) for water, diluted synthetic serum or actual human serum, and diluted nitric acid. The mean apparent percentage recoveries (plus or minus SD) were the best (102.6 plus or minus 2.5) when the standards were prepared in nitric acid; the values obtained when standards in water were used were too high (111.5 plus or minus 3.9); those with standards prepared in physiological saline (92.0 plus or minus 3.2) or synthetic serum (92.3 plus or minus 2.5) were too low.