Dietary protein supplementation increases peak bone mass acquisition in energy-restricted growing rats.

Peak bone mass is a major determinant of osteoporosis pathogenesis during aging. Respective influences of energy and protein supplies on skeletal growth remains unclear. We investigated the effect of a 5-mo dietary restriction on bone status in young rats randomized into six groups (n = 10 per group). Control animals were fed a diet containing a normal (13%) (C-NP) or a high-protein content (26%) (C-HP). The other groups received a 40% protein energy-restricted diet (PER-NP and PER-HP) or a 40% energy-restricted diet (ER-NP and ER-HP). High-protein intake did not modulate bone acquisition, although a metabolic acidosis was induced and calcium retention impaired. PER and ER diets were associated with a decrease in femoral bone mineral density. The compensation for protein intake in energy-restricted conditions induced a bone sparing effect. Plasma osteocalcin (OC) and urinary deoxypyridinoline (DPD) assays revealed a decreased OC/DPD ratio in restricted rats compared with C animals, which was far more reduced in PER than in ER groups. Circulating IGF-1 levels were lowered by dietary restrictions. In conclusion, both energy and protein deficiencies may contribute to impairment in peak bone mass acquisition, which may affect skeleton strength and potentially render individuals more susceptible to osteoporosis.

Influence of high and low protein intakes on age-related bone loss in rats submitted to adequate or restricted energy conditions.

Low energy and protein intake has been suggested to contribute to the increased incidence of osteoporosis in the elderly. The impact of dietary protein on bone health is still a matter of debate. Therefore, we examined the effect of the modulation of protein intake under adequate or deficient energy conditions on bone status in 16-month-old male rats. The animals were randomly allocated to six groups (n = 10/group). Control animals were fed a diet providing either a normal-protein content (13%, C-NP) or a high-protein content (26%) (C-HP). The other groups received a 40% protein/energy-restricted diet (PER-NP and PER-HP) or a normal protein/energy-restricted diet (ER-NP and ER-HP). After 5 months of the experiment, protein intake (13% or 26%) did not modulate calcium retention or bone status in those rats, although a low-grade metabolic acidosis was induced with the HP diet. Both restrictions (PER and ER) decreased femoral bone mineral density and fracture load. Plasma osteocalcin and urinary deoxypyridinoline levels were lowered, suggesting a decrease in bone turnover in the PER and ER groups. Circulating insulin-like growth factor-I levels were also lowered by dietary restrictions, together with calcium retention. Adequate protein intake in the ER condition did not elicit any bone-sparing effect compared to PER rats. In conclusion, both energy and protein deficiencies may contribute to age-related bone loss. This study highlights the importance of sustaining adequate energy and protein provision to preserve skeletal integrity in the elderly.

Long-term intake of a high-protein diet with or without potassium citrate modulates acid-base metabolism, but not bone status, in male rats.

High dietary protein intake generates endogenous acid production, which may adversely affect bone health. Alkaline potassium citrate (Kcit)(2) may contribute to the neutralization of the protein-induced metabolic acidosis. We investigated the impact of 2 levels of protein intake and Kcit supplementation on acid-base metabolism and bone status in rats. Two-month-old Wistar male rats were randomly assigned to 4 groups (n = 30 per group). Two groups received a normal-protein content (13%) (NP) or a high-protein (HP) content diet (26%) for 19 mo. The 2 other groups received identical diets supplemented with Kcit (3.60%) (NPKcit and HPKcit). Rats were pair-fed based on the ad libitum intake of the HP group. At 9, 16, and 21 mo of age, 10 rats of each group were killed. The HP diet induced a metabolic acidosis characterized by hypercalciuria, hypermagnesuria, and hypocitraturia at all ages. Kcit supplementation neutralized this effect, as evidenced by decreased urinary calcium and magnesium excretion by the HPKcit rats. Femoral bone mineral density, biomechanical properties, bone metabolism biomarkers (osteocalcin and deoxypyridinoline), and plasma insulin-like growth factor 1 levels were not affected by the different diets. Nevertheless, at 21 mo of age, calcium retention was reduced in the HP group. This study suggests that lifelong excess of dietary protein results in low-grade metabolic acidosis without affecting the skeleton, which may be protected by an adequate calcium supply.

Organic potassium salts or fibers effects on mineral balance and digestive fermentations in rats adapted to an acidogenic diet.

BACKGROUND: Fibers and potassium (K) organic salts in plant foods are liable to affect Ca and Mg balance at digestive and renal levels, respectively. K organic salts could counteract the acidifying effects of western diets and consequences of excess NaCl. AIM OF THE STUDY: To study this question, male rats were adapted to a basal acidifying low-K (LK) diet, or to diets supplemented with a fiber mix (LK/F), or K citrate (HK) or both (HK/F). RESULTS: HK and HK/F diets displayed a marked alkalinizing effect in urine and promoted citraturia, but this effect was not modulated by fibers. The effect of fibers on Ca digestive absorption was more potent than K citrate effect on Ca renal excretion. In contrast, K citrate effect on kidney Mg excretion was more effective than that of fibers on Mg digestive absorption, a maximal effect on Mg balance was observed in rats fed the HK/F diet. Digestive fermentations in rats fed the LK/F diet were characterized by high-propionic acid fermentations and succinate accumulation. In rats adapted to the HK/F diet, K citrate supplementation depressed succinate and increased butyrate concentrations. CONCLUSION: Organic anions arising from digestive fermentations seem to be not directly involved in the alkalinizing effects of plant foods. Fibers and organic K salts exert distinct effects on Ca and Mg metabolism, but with interesting interactions as to Mg balance, digestive fermentations and urine pH.

The effect of aging on intestinal absorption and status of calcium, magnesium, zinc, and copper in rats: a stable isotope study.

Many investigators have reported changes in mineral status with age but conflicting observations were done concerning mineral absorption. This study was conducted to clarify the effect of aging on intestinal absorption and status of minerals, using a stable isotope approach. To do so, 40 rats of different ages: 9, 22, 44, and 88 weeks were fed with a semi-purified diet for a total of 30 days. At the beginning of the 4th week, the rats received a stable isotope solution containing (44)Ca, (25)Mg, (67)Zn, and (65)Cu. Individual feces and urine were then collected during 4 consecutive days in order to measure stable isotopes by inductively coupled plasma/mass spectrometry (ICP/MS) and blood and tissues were sampled for mineral status determination. Intestinal absorption of (44)Ca and (67)Zn considerably decreased with age, whereas intestinal (25)Mg absorption decreased only moderately and intestinal (65)Cu absorption was unaffected. Plasma and bone calcium (Ca) were not modified with age whereas urinary Ca excretion considerably increased. Plasma and erythrocyte magnesium (Mg) levels were unaffected with age whereas urinary Mg excretion and Mg bone level decreased. Plasma zinc (Zn) level decreased and bone Zn level increased with age whereas red blood cell and liver Zn level and urinary Zn excretion remained unchanged. Plasma Cu level increased with age whereas liver and bone Cu levels and urinary Cu excretion remained unchanged. These results show that the effect of aging on the intestinal mineral absorption and status differ largely according to the mineral considered. Further studies are required under different nutritional conditions to explore the underlying mechanisms during aging and to adjust a better nutrition of the elderly.

Excess casein in the diet is not the unique cause of low-grade metabolic acidosis: role of a deficit in potassium citrate in a rat model.

This study examined the effects of a dietary model of protein excess and K anion salt deficit on the occurrence of metabolic acidosis in rat. Rats were adapted to diets containing either 13 or 26% casein, together with mineral imbalance, through lowering K/increasing sodium/omitting alkalinizing anions. For each protein level, a group of rats was supplemented with K citrate. Dietary K citrate resulted in neutral urinary pH, whatever the protein level. Urea excretion was higher in rats adapted to 26% casein than 13% casein diets, but K citrate enhanced this excretion and suppressed ammonium elimination. No citraturia could be observed in acidotic rats, whereas K citrate greatly stimulated citraturia and 2-ketoglutarate excretion. In conclusion, low-grade metabolic acidosis can occur with a moderate protein level in the diet. K citrate was apparently less effective in rats adapted to the 26% casein level than in those adapted to the 13% casein level with regard to magnesium, citrate and 2-ketoglutarate concentrations in urine.

Dietary inulin intake and age can significantly affect intestinal absorption of calcium and magnesium in rats: a stable isotope approach.

BACKGROUND: previous studies have shown that non-digestible inulin-type fructan intake can increase intestinal mineral absorption in both humans and animals. However, this stimulatory effect on intestinal absorption may depend on experimental conditions such as duration of fermentable fiber intake, mineral diet levels and animals' physiological status, in particular their age. OBJECTIVES: the aim of this study was to determine the effect of inulin intake on Ca and Mg absorption in rats at different age stages. METHODS: eighty male Wistar rats of four different ages (2, 5, 10 and 20 months) were randomized into either a control group or a group receiving 3.75% inulin in their diet for 4 days and then 7.5% inulin for three weeks. The animals were fed fresh food and water ad libitum for the duration of the experiment. Intestinal absorption of Ca and Mg was determined by fecal monitoring using stable isotopic tracers. Ca and Mg status was also assessed. RESULTS: absorption of Ca and Mg was significantly lower in the aged rats (10 and 20 mo) than in the young and adult rat groups. As expected, inulin intake increased Ca and Mg absorption in all four rat groups. However, inulin had a numerically greater effect on Ca absorption in aged rats than in younger rats whereas its effect on Mg absorption remained similar across all four rat age groups. CONCLUSION: the extent of the stimulatory effect of inulin on absorption of Ca may differ according to animal ages. Further studies are required to explore this effect over longer inulin intake periods, and to confirm these results in humans.

Effect of potassium salts in rats adapted to an acidogenic high-sulfur amino acid diet.

Low-grade metabolic acidosis, consecutive to excessive catabolism of sulfur amino acids and a high dietary Na:K ratio, is a common feature of Western food habits. This metabolic alteration may exert various adverse physiological effects, especially on bone, muscle and kidneys. To assess the actual effects of various K salts, a model of the Westernised diet has been developed in rats: slight protein excess (20 % casein); cations provided as non-alkalinising salts; high Na:K ratio. This diet resulted in acidic urine (pH 5.5) together with a high rate of divalent cation excretion in urine, especially Mg. Compared with controls, K supplementation as KCl accentuated Ca excretion, whereas potassium bicarbonate or malate reduced Mg and Ca excretion and alkalinised urine pH (up to 8). In parallel, citraturia was strongly increased, together with 2-ketoglutarate excretion, by potassium bicarbonate or malate in the diet. Basal sulfate excretion, in the range of 1 mmol/d, was slightly enhanced in rats fed the potassium malate diet. The present model of low-grade metabolic acidosis indicates that potassium malate may be as effective as KHCO3 to counteract urine acidification, to limit divalent cation excretion and to ensure high citrate concentration in urine.

Long-term moderate zinc supplementation increases exchangeable zinc pool masses in late-middle-aged men: the Zenith Study.

BACKGROUND: Zinc supplementation may be beneficial for health. Assessing exchangeable zinc pools may be a useful approach to evaluate zinc status. OBJECTIVE: We evaluated the effects of long-term supplementation with 2 moderate doses of zinc on the mass of exchangeable zinc pools. DESIGN: Three groups of healthy, late-middle-aged men (n = 16 per group) participated in a stable-isotope zinc kinetic study after 6 mo of daily supplementation with 0 (placebo), 15, or 30 mg Zn. At the end of the supplementation period, each subject received an intravenous injection of 0.89 mg (70)Zn, and the plasma zinc disappearance curve was monitored for the next 10 d. Two approaches were used to determine the characteristics of the exchangeable zinc pools: 1) formal 3-compartmental modeling and 2) a simplified determination of the total mass of the rapidly exchangeable zinc pool (EZP). RESULTS: In the placebo group, the exchangeable zinc pool masses for the 3 considered pools were as follows: 2.15, 12.7, and 100.5 mg Zn. The rapidly exchangeable zinc pool mass in the placebo group was 143 mg Zn. Zinc supplementation significantly increased the exchangeable zinc pool masses regardless of the approach used to determine these pools. In addition, these data confirm that exchangeable zinc pool masses correlate positively with total zinc intake and negatively with subject age and do not correlate with plasma zinc concentrations. CONCLUSION: Our data show that long-term supplementation with 2 moderate doses of zinc is an efficient way to increase exchangeable zinc pool masses in late-middle-aged men.

Effects of sulphate- and bicarbonate-rich mineral waters on net and fractional intestinal absorption and urinary excretion of magnesium in rats.

Magnesium (Mg) intake is below the recommended daily allowances in many developed countries. Mg-rich mineral waters can provide significant amounts of energy-free Mg and thus help to meet Mg requirements. We assessed the effects of different Mg-rich mineral waters on overall intestinal Mg absorption and urinary Mg excretion in 40 rats split into four groups: one received distilled water, another a solution of MgCl(2) and the others two different mineral waters, sulphated water (Hépar) and carbonated water (Badoit) mixed with the diet and as drinking water, for four weeks. The rats were given 3 mg of (26)Mg orally and 0.5 mg of (25)Mg intravenously. They were placed in metabolic cages, and diet consumption, and faeces and urine excretion were monitored during the last four days of the experiment. The rats were then sacrificed and blood was sampled. Mg levels in the diet, faeces, urine and biological samples were measured by atomic absorption spectrometry. Mg stable isotope measurements were performed by ICP/MS. Mg-rich mineral waters significantly increased net intestinal absorption of Mg by more than 30%, but the proportions of both apparent and true intestinal absorption of Mg were similar in all four groups. Thus, net and fractional retention of Mg were similar in the three Mg-supplemented groups. In conclusion, both types of Mg-rich mineral waters studied similarly increased both absorption and urinary excretion of Mg with no positive effect on the overall retention of Mg, probably because the Mg status of the rats was already satisfactory.

Effects of inulin-type fructans of different chain length and type of branching on intestinal absorption and balance of calcium and magnesium in rats.

BACKGROUND: Inulin-type fructans or chains with mainly beta[2-1] linked fructose molecules escape the ingestion procedure in the small intestine and are fermented by the microflora, and are known to increase colonic absorption of minerals in animals. The fermentation rate in the large bowel into short-chain fatty acids depends on the molecular mass and the structure of these food ingredients. It is thought that this colonic fermentation is the basis for the reported increase in mineral absorption. AIM OF THE STUDY: The purpose of the present study was twofold: a) to compare different types of fructans that differ in the sugar chain length and in chain branching; b) to determine the potential synergistic effect of a combination of inulin-type fructans with different chain lengths. METHODS: For this purpose, 50 adult male Wistar rats weighing 170 g each were used in this study. The rats were distributed into 5 groups and fed for 28 days a fiber-free basal purified diet or diet containing 10 % oligofructose (OF) (DP(av) 4), or 10 % HP-inulin (DP(av) 25), a blend of 50 % OF and 50 % HP-inulin, or a branched-chain inulin. RESULTS: During the first period, the rats went into a gradual adaptation, during which the rats received 2.5 % for 1 week and then 5 % for 1 week of the tested products. During the last 4 days of the experiment, feces and urine were monitored for mineral balance study. The animals were then sacrificed and blood, cecum and tibia were sampled for mineral status assessment. Our results showed that the ingestion of all the tested fructans led to a considerable cecal fermentation. All tested compounds increased the intestinal absorption and balance of Mg significantly. Interestingly, in the present experimental set-up, all tested compounds increased the intestinal absorption and balance of Ca numerically, but only the blend OF + HP-inulin increased apparent intestinal absorption and balance of Ca significantly. CONCLUSIONS: The different types of fructans studied in the present experiment seem to have similar activity on mineral absorption. However, the combination of OF and HP-inulin showed synergistic effects on intestinal Ca absorption and balance in rats. Further studies with other combinations of fructans need to be done to extend these findings.

Impact of whole wheat flour and its milling fractions on the cecal fermentations and the plasma and liver lipids in rats.

The aim of the present work was to evaluate the cholesterol-lowering potency of the different milling fractions of whole wheat flour, by investigating the effects of these wheat fractions (white flour, whole flour, and bran) on digestive fermentations and lipid metabolism in Wistar rats. Compared to the control, which was fiber-free, the different cereal fractions did not affect the daily food intake or weight gain. The white flour and whole flour diets markedly enlarged the cecum and elicited acidic fermentations (pH approximately 6.2), whereas bran was less effective. It appears that white flour rather promoted propionate-rich fermentations (+62%), whereas bran favored butyrate-rich fermentations (+178%). White flour or bran did not significantly affect total steroid excretion, but whole flour was effective (+41%). Both white flour and whole flour decreased cholesterol in the d < 1.040 fraction, but only whole flour significantly lowered cholesterolemia. However, all the cereal diets significantly decreased liver lipids, whole flour being the most potent (-54%). In conclusion, the totality of the wheat grain is important for cholesterol- and triglyceride-lowering effects, and the splitting up of the grain alters its health effects.

Fractional intestinal absorption of magnesium is directly proportional to dietary magnesium intake in rats.

The precise mechanisms of intestinal Mg absorption are still unclear and the possibility of an adaptative rise in the fraction of Mg absorbed as Mg intake is lowered is controversial. Mg deficiency has been studied extensively in rats where it is readily produced by dietary depletion. In this study, we investigated the effect of Mg intake on fractional absorption of Mg acutely and after adaptation to graded Mg intake in rats. For this purpose, male Wistar rats (n = 30) were fed a basal semipurified diet containing 600 mg Mg/kg (MgO) for 7 d. Three groups of 10 rats were then formed and fed the basal semipurified diet with 600, 300 or 150 mg Mg/kg, for 28 d. Apparent and true intestinal absorptions and fecal endogenous excretion of Mg were determined at the beginning and end of the experiment. As expected, plasma Mg levels were lower in the deficient groups than in the control by d 4 and differences were more marked at the end of the experiment. Erythrocyte Mg levels were significantly lower at the end of the experiment in the group fed the diet containing 150 mg Mg/kg. The amounts of Mg absorbed were directly proportional to the dietary Mg intakes in all three experimental groups at both testing periods. This indicated that the apparent and true intestinal absorption percentages of Mg were not different in rats fed the three different levels of dietary Mg. These results also show that fecal endogenous excretion of Mg was nearly directly proportional to the dietary Mg intakes. These results argue in favor of a passive diffusional process for intestinal Mg absorption.

Chronic oral administration of rhamnogalacturonan-II dimer, a pectic polysaccharide, failed to accelerate body lead detoxification after chronic lead exposure in rats.

Lead is a ubiquitous heavy metal and its toxicity remains an important public health issue. In previous work, we reported that ingestion of rhamnogalacturonan-II dimer (dRGII), a pectic polysaccharide, may decrease intestinal absorption and status of Pb in rats. Here, we evaluated the potential detoxifying effect of different doses of dRGII after chronic oral Pb exposure in rats. For this purpose, six groups of ten male Wistar rats weighing 150g were treated as follows: group A received a semi-purified control diet for 6 weeks; groups B, C, D, E and F received the same diet plus 3 mg Pb (as acetate) for 3 weeks. Group B was then killed. Groups C, D, E, and F continued to receive the semi-purified control diet containing 0, 2, 6 or 18g dRGII/kg diet for 3 additional weeks. During the last 5 d, a Pb conventional balance study was performed. Rats were then anaesthetized and tissues were sampled for Pb and essential minerals assay. The results showed that residual Pb in the added dRGII was not available for absorption. However, the added dRGII failed to induce any significant increase in faecal or urinary Pb excretion. Consequently, at the end of the study the intestinal Pb absorption and balance remained unchanged in the animals receiving the different doses of dRGII. In line with this, we showed that dRGII administration was not effective in decreasing tibia or kidney Pb levels in rats. In conclusion, Pb complexed by dRGII in fruits and vegetables and fruit juice is thus mostly unavailable for intestinal absorption. However, the addition of dRGII after chronic Pb exposure does not help Pb detoxification.

Exchangeable magnesium pool masses in healthy women: effects of magnesium supplementation.

BACKGROUND: Studying magnesium pools in the body with use of stable isotopes may be helpful for evaluating magnesium status. Data on the evaluation of magnesium pools in humans are scarce. OBJECTIVE: We undertook this study to evaluate the effects of a magnesium supplementation program on the size of the exchangeable body pools of magnesium and on classic indexes of magnesium status in healthy women with normal magnesium status. DESIGN: Ten healthy women participated in a kinetic study with magnesium stable isotopes before and after 8 wk of magnesium supplementation. Each woman received 3 supplements containing 5.08 mmol (122 mg) elemental Mg/d (366 mg/d). Before and at the end of the supplementation period, each woman received an intravenous injection of 1.67 mmol (40 mg) (25)Mg, and the plasma magnesium disappearance curve was followed for the next 7 d. Two methods were used to analyze the exchangeable pools of magnesium: 1) formal multicompartmental modeling and 2) a simplified estimation of the total mass of the rapidly exchangeable magnesium pool (EMgP). RESULTS: In these healthy women, exchangeable magnesium pools represented 11-12% of total body magnesium on the basis of multicompartmental analysis. The simplified estimation of EMgP overestimated the size of the exchangeable magnesium pools by approximately 45-50%. Eight weeks of magnesium supplementation did not significantly modify the size of the exchangeable magnesium pools, whereas urinary magnesium excretion was significantly higher after 8 wk of supplementation. CONCLUSION: Women with no clinical evidence of magnesium deficiency may not respond to short-term supplementation with increases in the mass of the exchangeable magnesium body pool or in magnesium turnover rates.