Circulating Ionized Magnesium as a Measure of Supplement Bioavailability: Results from a Pilot Study for Randomized Clinical Trial.

Oral supplementation may improve the dietary intake of magnesium, which has been identified as a shortfall nutrient. We conducted a pilot study to evaluate appropriate methods for assessing responses to the ingestion of oral magnesium supplements, including ionized magnesium in whole blood (iMg2+) concentration, serum total magnesium concentration, and total urinary magnesium content. In a single-blinded crossover study, 17 healthy adults were randomly assigned to consume 300 mg of magnesium from MgCl2 (ReMag®, a picosized magnesium formulation) or placebo, while having a low-magnesium breakfast. Blood and urine samples were obtained for the measurement of iMg2+, serum total magnesium, and total urine magnesium, during 24 h following the magnesium supplement or placebo dosing. Bioavailability was assessed using area-under-the-curve (AUC) as well as maximum (Cmax) and time-to-maximum (Tmax) concentration. Depending on normality, data were expressed as the mean ± standard deviation or median (range), and differences between responses to MgCl2 or placebo were measured using the paired t-test or Wilcoxon signed-rank test. Following MgCl2 administration versus placebo administration, we observed significantly greater increases in iMg2+ concentrations (AUC = 1.51 ± 0.96 vs. 0.84 ± 0.82 mg/dL·24h; Cmax = 1.38 ± 0.13 vs. 1.32 ± 0.07 mg/dL, respectively; both p < 0.05) but not in serum total magnesium (AUC = 27.00 [0, 172.93] vs. 14.55 [0, 91.18] mg/dL·24h; Cmax = 2.38 [1.97, 4.01] vs. 2.24 [1.98, 4.31] mg/dL) or in urinary magnesium (AUC = 201.74 ± 161.63 vs. 139.30 ± 92.84 mg·24h; Cmax = 26.12 [12.91, 88.63] vs. 24.38 [13.51, 81.51] mg/dL; p > 0.05). Whole blood iMg2+ may be a more sensitive measure of acute oral intake of magnesium compared to serum and urinary magnesium and may be preferred for assessing supplement bioavailability.

Manganese accumulation in bone following chronic exposure in rats: steady-state concentration and half-life in bone.

Literature data indicate that bone is a major storage organ for manganese (Mn), accounting for 43% of total body Mn. However, the kinetic nature of Mn in bone, especially the half-life (t(1/2)), remained unknown. This study was designed to understand the time-dependence of Mn distribution in rat bone after chronic oral exposure. Adult male rats received 50 mg Mn/kg (as MnCl2) by oral gavage, 5 days per week, for up to 10 weeks. Animals were sacrificed every 2 weeks during Mn administration for the uptake study, and on day 1, week 2, 4, 8, or 12 after the cessation at 6-week Mn exposure for the t(1/2) study. Mn concentrations in bone (MnBn) were determined by AAS analysis. By the end of 6-week's treatment, MnBn appeared to reach the steady state (T(ss)) level, about 2-3.2 fold higher than MnBn at day 0. Kinetic calculation revealed t(1/2)s of Mn in femur, tibia, and humerus bone of 77 (r=0.978), 263 (r=0.988), and 429 (r=0.994) days, respectively; the average t(1/2) in rat skeleton was about 143 days, equivalent to 8.5 years in human bone. Moreover, MnBn were correlated with Mn levels in striatum, hippocampus, and CSF. These data support MnBn to be a useful biomarker of Mn exposure.

[Comparative study of magnesium salts bioavailability in rats fed a magnesium-deficient diet].

The purpose of this study was to compare efficiency of compensation of alimentary Mg deficiency after administration of 12 organic and 8 inorganic magnesium salts and to evaluate the ability of vitamin B6 to accelerate their effect. Two hundred eighty rats were placed on a Mg-deficient diet (Mg content (15 mg/kg) and demineralized water for 7 weeks. Twelve control rats were fed a basal diet (Mg content 500 mg/kg). Starting from day 49 of the Mg-deficient diet, the rats were given magnesium salts (50 mg magnesium and 5 mg pyridoxine per kg): Mg chloride, Mg sulphate, Mg oxide, M nitrate, Mg thiosulphate, Mg hydrophosphate, Mg carbonate, Mg trisilicate, Mg (L-, D- and DL-) aspartate, Mg (L- and DL-) pyroglutamate, Mg succinate, Mg glycinate, Mg orotate, Mg taurate, Mg lactate or their combination with vitamin B6 (5 mg/kg b.w.). Erythrocyte and plasma Mg levels were measured by spectrophotometry following the colour reaction between Mg and titanium yellow. Mg L-aspartate compensated for magnesium deficit more effectively and faster than all other salts. Mg chloride showed the highest efficiency among inorganic magnesium salts. Both Mg chloride and Mg L-aspartate in combination with vitamin B6 caused statistically significant compensation of magnesium deficit.

[Effects of mangesium salts and their combinations with vitamin B6 on oxalates crystalluria in rats fed with pyridoxine-deficient diet].

We studied the effects of oral magnesium (Mg) salts either alone or in combination with pyridoxine hydrochloride in rats on pyridoxine-deficient diet. Fifty-four male rats were randomized into two groups and were fed either a standard diet or a pyridoxine-deficient diet for 3 weeks. A significant rise of the EGOT index ( > 1.5), oxaluria (from 74.8 +/- 5.2 to 117.9 +/- 12.3 mcM/l, p = 0.035), and crystalluria in rats fed with pyridoxine deficient diet were revealed. Oral Mg chloride, Mg L-aspartate either alone or in combination with pyridoxine in comparison with magnesium sulfate, magne B6 (Mg lactate with pyridoxine) and pyridoxine alone were administered (50 mg of magnesium and/or 5 mg of pyridoxine per kg body weight). Magnesium salts in combination with pyridoxine lowered an oxalate level and crystalluria whereas magnesium salts alone reduced only crystalluria. Antilithis effects of Mg L-aspartate and Mg chloride in combination with pyridoxine were comparable with those observed in magne B6 or pyridoxine treatment and were significantly higher than in magnesium sulfate treatment.

A streamlined method to predict hepatic clearance using human liver microsomes in the presence of human plasma.

INTRODUCTION: Human liver microsomal incubations are often used to predict the metabolic lability of new chemical entities. The clearance values are scaled-up from in vitro data and mathematically corrected for plasma protein binding, or in some cases the free fraction ratio of plasma to microsomes, using well-established scaling methods such as the well-stirred model. This can be time consuming for multiple compounds since it requires separate experiments to determine in vitro lability, and free fraction. METHODS: We attempted to streamline clearance predictions by combining experiments into one. Firstly, we combined the free fraction experiments into one free fraction ratio by measuring the partitioning of compound between plasma and microsomes, and by applying this experimental ratio to clearance predictions found that it performed at least as well as free fractions determined separately. We also incubated compounds with plasma added to the incubation mixture and compared the predicted clearances to values determined using traditional mathematical protein binding corrections. RESULTS: Consistently, incubations with added plasma resulted in CL predictions closer to literature values than incubations only mathematically corrected for protein binding. For example, incorporating plasma into a ketamine incubation resulted in a CL value of 15.1 mL/min/kg, compared with a value of 10.2 using mathematical binding corrections. The literature value is 16.4 mL/min/kg. DISCUSSION: This work characterizes this new method and compares it to the traditional microsomal incubation method using several literature compounds, and suggests that streamlining the methods may generate quality data faster and with less resource investment.

Functional mapping of neural pathways in rodent brain in vivo using manganese-enhanced three-dimensional magnetic resonance imaging.

This work presents three-dimensional MRI studies of rodent brain in vivo after focal and systemic administration of MnCl2. Particular emphasis is paid to the morphology and dynamics of Mn2+-induced MRI signal enhancements, and the physiological mechanisms underlying cerebral Mn2+ uptake and distribution. It turns out that intravitreal and intrahippocampal injections of MnCl2 emerge as useful tools for a delineation of major axonal connections in the intact central nervous system. Subcutaneous administrations may be exploited to highlight regions involved in fundamental brain functions such as the olfactory bulb, inferior colliculus, cerebellum and hippocampal formation. Specific insights into the processes supporting cerebral Mn2+ accumulation may be obtained by intraventricular MnCl2 injection as well as by pharmacologic modulation of, for example, hippocampal function. Taken together, Mn2+-enhanced MRI opens new ways for mapping functioning pathways in animal brain in vivo with applications ranging from assessments of transgenic animals to follow-up studies of animal models of human brain disorders.

Effect of Mg2+ on guanine nucleotide sensitivity of ligand binding to serotonin1A receptors from bovine hippocampus.

The serotonin1A (5-HT1A) receptor is an important member of the superfamily of seven transmembrane domain G-protein coupled receptors (GPCRs). We report here that guanine nucleotide sensitivity of agonist binding to hippocampal 5-HT1A receptors is dependent on the concentration of Mg2+. Our results show that agonist binding to 5-HT1A receptors is relatively insensitive to guanine nucleotides in the absence of Mg2+. In contrast to this, the specific antagonist binding is insensitive to guanine nucleotides, even in the presence of Mg2+. These results point out the requirement of an optimal concentration of Mg2+ which could be used in assays toward determining guanine nucleotide sensitivity of ligand binding to GPCRs such as the 5-HT1A receptor. Our results provide novel insight into the requirement and concentration dependence of Mg2+ in relation to guanine nucleotide sensitivity for the 5-HT1A receptor in particular, and GPCRs in general.

Uptake of MnCl2 and mangafodipir trisodium in the myocardium: a magnetic resonance imaging study in pigs.

PURPOSE: To examine the changes in the longitudinal relaxation times (DeltaR1) induced in pig myocardium and blood following injections of 5, 10, and 15 micromol mangafodipir trisodium (Mn-DPDP) or MnCl2/kg of body weight (b.w.). MATERIALS AND METHODS: Twelve pigs were divided into two groups, one group receiving MnCl2 and the other receiving Mn-DPDP. Three consecutive doses of contrast agent (5, 10, and 15 micromol/kg of b.w.) were injected in each animal with a 40-minute time interval between each dose. Measurements of T1 in blood and myocardium were made 5, 15, 25, and 35 minutes after each injection. Additionally, relaxivity measurements in blood samples were performed. RESULTS: An increase in myocardial R1 was observed for both contrast agents at all concentration levels tested. This increase peaked 5 minutes after injection and then declined. An increase could still be detected 35 minutes after injection. The effect was larger when using MnCl2 than when using Mn-DPDP. CONCLUSION: The dissociation kinetics of Mn2+ from the DPDP ligand limits the relaxation increase of Mn-DPDP relative to that of MnCl2. On the other hand, the toxicity of MnCl2 may exclude it from clinical use.

Three-dimensional MRI of cerebral projections in rat brain in vivo after intracortical injection of MnCl2.

In this study we investigated the potential of in vivo MRI detection of axonal Mn2+ transport for tracing neuronal projections originating in the sensorimotor cortex in healthy and lesioned rat brains. Special attention was given to the potential of visualizing neuronal sprouting of central nervous system across the midline. After injecting unchelated MnCl2 into the forelimb area of sensorimotor cortex of 18 healthy and 10 lesioned rats corticofugal projections could be traced through the internal capsule to the cerebral peduncle and the pyramidal decussation. Although the neuronal tract was visible as early as 6 h after MnCl2 injection, best contrast was achieved after 24-48 h. Beside the cortico-spinal tract, the cortico-thalamic fibres were also visualized by anterograde Mn2+ transport. Cortico-striatal fibres were partially masked by the very high signal near the MnCl2 injection site but could be discerned as well. Slight, diffuse signal enhancement of cortical tissue contralateral to the MnCl2 injection site in healthy rat brains suggests interhemispheric connections or passive diffusion of Mn2+. However, enhanced fibre tract contrast connecting both hemispheres was visible 16 weeks after onset of focal photothrombotic cortical injury. In conclusion our study has shown that we were able to visualize reproducibly the main descending corticofugal projections and interhemispheric connections by non-invasive MRI after localized injection of MnCl2. The appearance of interhemispheric Mn2+-enhanced fibres after photothrombotic focal injury indicates that the method may bear potential to follow non-invasively gross plastic changes of connectivity in the brain after injury.

Adsorption of cationized bovine serum albumin onto epithelial crypt fractions of the rat colon.

The purpose of the study was to characterize mucosal attachment of a cationized model protein, bovine serum albumin (BSA), onto the various fractions of colonic crypts epithelium in the rat. BSA was labeled with fluorescein isothiocyanate (FITC) and its surface net electric charge was modified from negative to positive. Attachment of the cationized protein (CF-BSA) onto rat colonic epithelium was performed by incubation of colonic everted sacs in medium containing cationized or non-cationized FITC-labeled BSA. Using a nonenzymatic isolation procedure, colonocytes were harvested from five horizontal fractions of the colonic crypts. BSA adhesion to the isolated colonocytes was quantified spectrofluorometrically. In addition, the effect of increasing concentrations of Mg(2+) on the adsorption of the cationized BSA onto the surface of colonic epithelium was evaluated by measuring its ability to displace the adhered BSA from its binding sites. BSA cationization facilitated protein adherence to the colon epithelium in a crypt depth-dependent manner. The largest extent of adherence was observed in the outer layer (first fraction) of the colon. Binding persisted to approximately half the depth of the crypts. The relation between CF-BSA concentration in the incubation medium and the amount of CF-BSA adsorbed onto the colonic epithelium was exponential in nature. The addition of electrolyte (Mg(2+)) caused a detachment of the CF-BSA. The adsorption process was characterized by Langmuir's adsorption isotherm. It is concluded that cationized BSA could be useful as a targetable drug platform in cases where the target site is the gastrointestinal epithelium.

Paracellin-1 is critical for magnesium and calcium reabsorption in the human thick ascending limb of Henle.

BACKGROUND: A new protein, named paracellin 1 (PCLN-1), expressed in human thick ascending limb (TAL) tight junctions, possibly plays a critical role in the control of magnesium and calcium reabsorption, since mutations of PCLN-1 are present in the hypomagnesemia hypercalciuria syndrome (HHS). However, no functional experiments have demonstrated that TAL magnesium and calcium reabsorption were actually impaired in patients with HHS. METHODS: Genetic studies were performed in the kindred of two unrelated patients with HHS. Renal magnesium and calcium reabsorption in TAL were analyzed in one homozygous affected patient of each family, one patient with extrarenal hypomagnesemia (ERH), and two control subjects (CSs). RESULTS: We found two yet undescribed mutations of PCLN-1 (Gly 162 Val, Ala 139 Val). In patients with HHS, renal magnesium and calcium reabsorptions were impaired as expected; NaCl renal conservation during NaCl deprivation and NaCl tubular reabsorption in diluting segment were intact. Furosemide infusion in CS markedly increased NaCl, Mg, and Ca urinary excretion rates. In HHS patients, furosemide similarly increased NaCl excretion, but failed to increase Mg and Ca excretion. Acute MgCl(2) infusion in CS and ERH patient provoked a dramatic increase in urinary calcium excretion without change in NaCl excretion. When combined with MgCl(2) infusion, furosemide infusion remained able to induce normal natriuretic response, but was unable to increase urinary magnesium and calcium excretion further. In HHS patients, calciuric response to MgCl(2) infusion was blunted. CONCLUSION: This study is the first to our knowledge to demonstrate that homozygous mutations of PCLN-1 result in a selective defect in paracellular Mg and Ca reabsorption in the TAL, with intact NaCl reabsorption ability at this site. In addition, the study supports a selective physiological effect of basolateral Mg(2+) and Ca(2+) concentration on TAL divalent cation paracellular permeability, that is, PCLN-1 activity.

Bioavailability of US commercial magnesium preparations.

Magnesium deficiency is seen with some frequency in the outpatient setting and requires oral repletion or maintenance therapy. The purpose of this study was to measure the bioavailability of four commercially-available preparations of magnesium, and to test the claim that organic salts are more easily absorbed. Bioavailability was measured as the increment of urinary maginesium excretion in normal volunteers given approximately 21 mEq/day of the test preparations. Results indicated relatively poor bioavailability of magnesium oxide (fractional absorption 4 per cent) but significantly higher and equivalent bioavailability of magnesium chloride, magnesium lactate and magnesium aspartate. We conclude that there is relatively poor bioavailability of magnesium oxide, but greater and equivalent bioavailability of magnesium chloride, lactate, and aspartate. Inorganic magnesium salts, depending on the preparation, may have bioavailability equivalent to organic magnesium salts.

cAMP-activated anion conductance is associated with expression of CFTR in neonatal mouse cardiac myocytes.

In this study, patch-clamp techniques were applied to cultured neonatal mouse cardiac myocytes (NMCM) to assess the contribution of cAMP stimulation to the anion permeability in this cell model. Addition of either isoproterenol or a cocktail to raise intracellular cAMP increased the whole cell currents of NMCM. The cAMP-dependent conductance was largely anionic, as determined under asymmetrical (low intracellular) Cl(-) conditions and symmetrical Cl(-) in the presence of various counterions, including Na(+), Mg(2+), Cs(+), and N-methyl-D-glucamine. Furthermore, the cAMP-stimulated conductance was also permeable to ATP. The cAMP-activated currents were inhibited by diphenylamine-2-carboxylate, glibenclamide, and an anti-cystic fibrosis transmembrane conductance regulator (CFTR) monoclonal antibody. The anti-CFTR monoclonal antibody failed, however, to inhibit an osmotically activated anion conductance, indicating that CFTR is not linked to osmotically stimulated currents in this cell model. Immunodetection studies of both neonatal mouse heart tissue and cultured NMCM revealed that CFTR is expressed in these preparations. The implication of CFTR in the cAMP-stimulated Cl(-)- and ATP-permeable conductance was further verified with NMCM of CFTR knockout mice [cftr(-/-)] in which cAMP stimulation was without effect on the whole cell currents. In addition, stimulation with protein kinase A and ATP induced Cl(-)-permeable single-channel activity in excised, inside-out patches from control, but not cftr(-/-) NMCM. The data in this report indicate that cAMP stimulation of NMCM activates an anion-permeable conductance with functional properties similar to those expected for CFTR, thus suggesting that CFTR may be responsible for the cAMP-activated conductance. CFTR may thus contribute to the permeation and/or regulation of Cl(-)- and ATP-permeable pathways in the developing heart.

Blood and urinary magnesium kinetics after oral magnesium supplements.

A study was conducted to compare the pharmacokinetic profile of three oral magnesium supplements--magnesium chloride solution, slow-release magnesium chloride tablets, and magnesium gluconate tablets--at 16 mmol/dose. Twelve healthy normomagnesemic subjects were evaluated during an initial baseline study, followed by three magnesium supplementation studies. Supplements were administered in a randomized, crossover fashion at weekly intervals. During each of the four trials, subjects followed the same routines and consumed identical diets. Magnesium concentrations were measured in urine samples collected from 0 to 4, 4 to 8, 8 to 12, and 12 to 24 hours. Intraleukocyte, total serum, and ultrafiltrable magnesium were measured in blood samples drawn at 0, 1, 2, 3, 4, 8, 12, and 24 hours. Compared with baseline, 24-hour urinary magnesium excretion significantly increased (P < 0.05) after the administration of the magnesium chloride solution and also increased after the administration of the other supplements, but the difference was not significant. The 24-hour areas under the curve (AUCs) for total serum, ultrafiltrable, and leukocyte magnesium were greater after the administration of each of the supplements when compared with baseline, although the differences were not statistically significant. Differences in delta AUCs (supplement AUC minus baseline AUC) for total magnesium, ultrafiltrable magnesium, and 24-hour urinary magnesium excretion were statistically different from zero or between supplements. Statistically significant differences (P < 0.05) in total serum, ultrafiltrable, and leukocyte magnesium concentrations were observed at various time points. These results suggest that there were no major differences in the overall effect of these supplements on total serum, ultrafiltrable, and leukocyte magnesium concentrations but do reveal differences in the time-concentration profiles in magnesium levels in blood and urine among the three supplement forms.

Mode of action of GR69153, a novel catechol-substituted cephalosporin, and its interaction with the tonB-dependent iron transport system.

GR69153 is a novel cephalosporin incorporating a catechol-substituted 7-aminothiazolyl-oxime. The antibiotic is actively transported into gram-negative cells via iron-regulated outer membrane proteins regulated by the tonB product. This transport enhances bactericidal activity most significantly at low concentrations, essentially removing the permeability barrier for antibiotic uptake.

Magnesio plasmático y umbral de fibrilación ventricular en perros anestesiados / Plasmatic magnesium and ventricular fibrillation threshold in anesthetized dogs

En perros anestesiados con pentobarbital sódico se estudió el efecto de la administración parenteral de soluciones de sulfato y cloruro de magnesio sobre algunas variables electrofisiológicas vinculadas a su potencial efecto de la administración parenteral de soluciones de sulfato y cloruro de magnesio sobre algunas variables electrofisiológicas vinculadas a su potencial efecto antiarrítmico. De los resultados obtenidos se puede concluir que tanto el sulfato como el cloruro de magnesio prolongan el período refractario efectivo ventricular (PREV), efecto probablemente relacionado con la prolongación del intervalo QTc. La administración de magnesio prolongó el intervalo AH probablemente por bloquear el canal de Ca++. El cloruro de magnesio no modificó el umbral de fibrilación ventricular (UFV), pero el sultato de magnesio lo hizo descender en forma significativa. Este efecto pudo estar relacionado con la disminución del potasio plasmático inducida por la administración de sulfato de magnesio y a la elevación del magnesio plasmático, porque la administración de sulfato de sodio, que también disminuyó el K+ plasmático, no modificó el UFV. Los efectos antiarrítmicos observados con la aministraciRon de magnesio en pacientes normomagnesémicos podrían ser explicados por una prolongación del PREV. Sin embargo, la disminución del UFV observada en los experimentos con sulfato de magnesio constituye un efecto potencialmente peligroso que merece ser investigado (AU)

Magnesio plasmático y umbral de fibrilación ventricular en perros anestesiados / Plasmatic magnesium and ventricular fibrillation threshold in anesthetized dogs

En perros anestesiados con pentobarbital sódico se estudió el efecto de la administración parenteral de soluciones de sulfato y cloruro de magnesio sobre algunas variables electrofisiológicas vinculadas a su potencial efecto de la administración parenteral de soluciones de sulfato y cloruro de magnesio sobre algunas variables electrofisiológicas vinculadas a su potencial efecto antiarrítmico. De los resultados obtenidos se puede concluir que tanto el sulfato como el cloruro de magnesio prolongan el período refractario efectivo ventricular (PREV), efecto probablemente relacionado con la prolongación del intervalo QTc. La administración de magnesio prolongó el intervalo AH probablemente por bloquear el canal de Ca++. El cloruro de magnesio no modificó el umbral de fibrilación ventricular (UFV), pero el sultato de magnesio lo hizo descender en forma significativa. Este efecto pudo estar relacionado con la disminución del potasio plasmático inducida por la administración de sulfato de magnesio y a la elevación del magnesio plasmático, porque la administración de sulfato de sodio, que también disminuyó el K+ plasmático, no modificó el UFV. Los efectos antiarrítmicos observados con la aministraciRon de magnesio en pacientes normomagnesémicos podrían ser explicados por una prolongación del PREV. Sin embargo, la disminución del UFV observada en los experimentos con sulfato de magnesio constituye un efecto potencialmente peligroso que merece ser investigado