Uremic Middle Molecules Research Articles

Nonzero Finite Wall Resistance Solution With Constant Dialyzate for Various Mass Transfer Quantities

This article deals with the theoretical investigations into dialytic mass transfer in parallel-plate hemodialyzers. The theoretical solutions presented are closed-form analytical solutions involving only exponential and algebraic polynomial functions, and are simple, economical, and convenient to use. With the help of Laplace transforms and the Galerkin method, the expressions for mixed mean concentration, local fluid-wall Sherwood number, logarithmic fluid-wall Sherwood number, local fluid Sherwood number, logarithmic fluid Sherwood number, clearance, and local concentration have been obtained. Data obtained in the present contribution are compared with earlier results and are found to be in the excellent agreement. Two solutes, urea and vitamin B12, were considered. It is observed from the above-mentioned comparison that the effect of variable dialyzate concentration on various mass transfer quantities is significant in the case of uremic small molecules. In the case of uremic middle molecules, the nonuniform concentration distribution in the dialyzate channel is only slightly affected.

Separation, identification of uremic middle molecules, and preliminary study on their toxicity

Background Compounds accumulating in uremic serum with molecular mass from 300 to 5000 Da are called uremic middle molecules (UMMs). In our previous work, two UMM fractions A and B were obtained from uremic sera, urine, and normal urine by gel permeation chromatography (GPC), and six UMMs from subfraction A3 of uremic plasma and normal urine were purified and characterized. Methods Urine and serum samples from uremic patients and healthy subjects were isolated by GPC, ion exchange chromatography (IEC), and reversed-phase high-performance liquid chromatography (RP-HPLC). Moreover, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) were used to characterize the compounds. The effects of subfraction A3 on renal function were studied in rabbit models with chronic renal failure (CRF). Results A compound with molecular weight 1007.94 in subfraction A3 was determined to be an octapeptide by mass spectrometry, with an amino acid sequence of Val–Val–Arg–Gly–Cys–Thr–Trp–Trp. Two CRF rabbits injected with A3 died in 5 days, while the other two CRF rabbits (no injection) survived a few days. By multistep chromatography and MALDI-TOF MS, another 11 endogenous compounds were found not only in the subfraction B9 of uremic sera but also in that of normal urine. Conclusion Seventeen endogenous middle molecular compounds were found in fractions A and B of uremic plasma and normal urine, among them an octapeptide with M W 1007.94 in subfraction A3. Preliminary experimental results on rabbits indicate that subfraction A3 could accelerate the death of rabbits with CRF.

Isolation and purification of uremic middle molecules by multi-step liquid chromatography

Isolation and comparison of uremic sera and urine and normal sera and urine were performed by gel permeation chromatography, anion exchange chromatography and reversed-phase high performance liquid chromatography. Two uremic middle molecular fractions (A and B) were obtained from uremic sera and urine and normal urine by gel permeation chromatography, but not from normal sera. The anion exchange chromatographic results of fraction A from different origins demonstrate that subfraction A-3 could be excreted in urine by healthy subject, but accumulated in uremic serum for renal failure of patient with uremia. After desalinization subfraction A-3 was analyzed by MALDI-TOF-MS. The results show that subfraction A-3 consists of six compounds with molecular weight 839, 873, 1007.94, 1106, 1680 and 2015 respectively. Finally, by reversed-phase high performance liquid chromatography, subfraction A-3 was further resolved into six independent fractions. Thus, the isolation and purification of six middle molecular compounds in subfraction A-3 came true by our method.

Influence of Uremic Middle Molecules on In Vitro Stimulated Lymphocytes and lnterleukin-2 Production

The influence of uremic middle molecules on in vitro lymphocyte blast transformation and interleukin-2 production was studied in 12 patients on long-term hemodialysis and 12 healthy control subjects with normal kidney function. Middle molecules inhibit phytohemoagglutinin induced lymphocyte proliferation in a concentration dependent fashion, achieving more than 50% inhibition of tritiated thymidine incorporation after 72 hrs in culture. In addition, there was less interleukin-2 production in the long-term hemodialysis patients studied compared with healthy control subjects.

Influence of Uremic Middle Molecules on In Vitro Stimulated Lymphocytes and Interleukin-2 Production

Influence of Uremic Middle Molecules on In Vitro Stimulated Lymphocytes and Interleukin-2 Production

Effect of uremic middle molecules removed during hemofiltration on the enzyme activity in rat liver cytosol

It is well known that the products of protein catabolism and some of the intermediates including middle molecules (MM) are a biochemical feature of uremic serum and also the activity of serum alanine aminotransferase decreases relatively. To examine the toxicity of MM, we investigate the effect of MM removed by HF on aminotransferases and peptidases in rat liver cytosol. The following enzymes were selected: aminotransferases (GOT, GPT), leucine aminopeptidase (LAP), gamma-glutamyltranspeptidase (gamma GTP). MM samples were as follows: preparation of MM fraction from each 10 l HF fluid at half intervals of former (F-) and latter (L-) for 5 h. HF of two patients (5 weeks) by 1KD ultrafiltration, and group separation into hydrophobic (Mo) and hydrophilic (Mi) by XAD-4 resin (F-Mo, L-Mo, F-Mi, L-Mi). Except for GOT, the effect of MM were found at the activities decreased on LAP (26-30%), on GPT (5-6%), and increased on gamma GTP (4-23%), as compared to control. We found a little difference in the results by the character of MM (Mo, Mi) and by the intervals of HF (F-, L-). These results suggest that MM might play a role in the formation of uremic peptides.

Identification of an ascorbic acid metabolite among "uremic middle molecules"

Among uremic toxins in the middle molecular mass range, 1H, 13C-nuclear magnetic resonance, ultraviolet spectrometry, and chromatographic analyses allow identification of the main component of the so-called "2-5-3 fraction" as ascorbic acid 2-sulfate, a conjugated metabolite of ascorbic acid. We previously (Clin Nephrol 1986;25:212-8) showed an inhibitory effect of the 2-5-3 fraction on microtubule formation. Therefore, we tested the action of ascorbic acid 2-sulfate and its synthetized enantiomers on tubulin polymerization. Because these molecules did not exert any inhibitory effect, we hypothesize that the 2-5-3 fraction is a mixture of compounds in which only a very low quantity of the inhibitory factor is present.

Fluorescent component found in uremic middle-molecules in the hemofiltrate of a dialysis patient

Fluorescent component found in uremic middle-molecules in the hemofiltrate of a dialysis patient

Hydrophobic uremic middle molecules in hemofiltrate of a dialysis patient

Hydrophobic uremic middle molecules in hemofiltrate of a dialysis patient

Inhibitory effect of peak 2-4 of uremic middle molecules on platelet aggregation.

It is well-known that uremic patients present prolonged bleeding times as a common complication. Factors responsible for this disorder have been extensively investigated. In order to elucidate the possible role of uremic middle molecules as responsible for the bleeding tendency observed in uremia, we have studied the effect on platelet aggregation of middle molecules obtained from uremic plasma by Sephadex and ion-exchange chromatography. Our results show that some purified middle molecular fractions have a specific inhibitory activity on platelet aggregation and suggest an important role for these compounds in the pathogenesis of uremic bleeding.

Identification of two uremic toxins by nuclear magnetic resonance and mass spectrometry.

A few uremic toxins (UT) previously called uremic middle molecules were isolated in pure form and identified In a previous study, by using a three step chromatographic separation we had isolated ten uremic toxins and had identified two of them The first is a double conjugate of glucuronidate-o-hydroxyhippuric acid identical to Furst’s compound The second is a glucuronic acid conjugate of o-hydroxybenzoic acid. In the present paper, we report the identification by 1H nuclear magnetic resonance and mass spectrometry of two new uremic toxins represented by peaks 2-5-2 and 2-5-5.

Identification by nuclear magnetic resonance and mass spectrometry of a glucuronic acid conjugate of o-hydroxybenzoic acid in normal urine and uremic plasma.

An endogenous compound (included in the fraction of uremic toxins often called the "uremic middle molecules") was separated from plasma of uremic patients and urine from normal persons. As elucidated by mass spectrometry, enzymatic hydrolysis, and 1H, 13C nuclear magnetic resonance, it appears to be a conjugate of o-hydroxybenzoic acid with glucuronic acid. Its presence in urine of healthy subjects indicates its physiological character.

Action on mitochondrial calcium metabolism of an ionophorous compound isolated from uremic plasma or normal urine.

An ionophorous compound that is one of the uremic middle molecules is able to inhibit the mitochondrial storage of calcium. Its active concentration is equivalent to that found in uremic plasma. This result can explain the diminution of phosphate calcium granules observed in mitochondria from uremic children. Moreover, this phenomenon may be involved in the calcium pool decrease observed in chronic renal insufficiency.

A compound from uremic plasma and from normal urine isolated by liquid chromatography and identified by nuclear magnetic resonance.

A compound present in normal urine and in ultrafiltrates of uremic plasma in the fraction of so-called "uremic middle molecules" was isolated by liquid chromatography. Preliminary studies, including amino acid analysis, characterization of uronic acids, and ultraviolet spectroscopy, show that the molecule contains glycine, a uronic acid, and an aromatic ring. Characterization by 1H and 13C nuclear magnetic resonance spectrometry shows conclusively that this compound is a double conjugate of glucuronidate--o-hydroxyhippuric acid, which has been previously described by Zimmerman et al., using quite different techniques of isolation and identification (Clin Nephrol 14: 107, 1980; FEBS Lett 129: 237, 1981).

Inhibition of platelet function by uremic middle molecules.

To further define the platelet abnormality responsible for uremic bleeding, we studied platelet aggregation with adenosine diphosphate, ristocetin, and collagen in serum fractions obtained by Sephadex G-15 chromatography. We found that uremic patients had considerable inhibition in several peaks of middle molecular range, but the findings were inconsistent and not clearly related to the degree of uremia.

Anion exchange chromatography and double-diffusion cells for the study of middle molecules

We report herein a study of uremic middle molecules with techniques which allow their analysis without separation from protein. We have avoided the use of size exclusion techniques and filtration for size discrimination. In their place, we have used direct anion exchange chromatography for partial isolation of the compounds. Most of the middle-sized molecules which have been defined have been anionic, and most investigators have used DEAE-Sephadex in their isolation. For size discrimination, we have used diffusive properties through cellulosic membranes and the well-known inverse relationship of molecular weight to diffusibility [1]. Diffusive passage across membranes is dependent on molecular weight and radius. Problems such as variable protein layering, seen in convective transfer (filtration) [2], are avoided in diffusion studies.

Separation of one uremic middle molecules fraction by high performance liquid chromatography

Since the hypothesis of Babb et al [1, 2] concerning the existence of uremic middle molecules (UMM), it is now known these substances in the molecular weight range of 300 to 2000 daltons accumulate in body fluids of patients with chronic renal disease and in normal urine.

Uremic middle moleculesの糖脂質代謝に及ぼす影響について

Uremic middle moleculesの糖脂質代謝に及ぼす影響について

An automatic analytical system for quantitative determination of uremic middle molecules

A system for automatic analyses of medium-sized compounds (middle molecules-MM) is described, applying gel filtration combined with gradient ion-exchange chromatography and using an automatic sample injector, motoric selector valves, and a minicomputer, which controls the system. Quantitation is performed by integrating the peak areas obtained. Using the automatic analyzer 7 MM solutes could be separated, detected, and quantified. The method permits a sampling rate of 28 samples per week. The precision of the method was 9.5%. The error in a single determination based on duplicate samples was 10% and in duplicate determinations was 7%. The automatic, analytical system is a prerequisite for performing sereening studies within a reasonable period of time. The present automatic, analytical system has been tested during a period of 3–5 years, and used in more than 4000 determinations.

A study of the cardiotoxicity of uremic middle molecules on embryonic chick hearts.

Ultrafiltrates from uremic patients can induce, in vitro, strong modifications of beating frequency of auricular fragments of the embryonic chick heart. According to concentration, effects increase from tachycardia to cessation of cardiac beats. Complete reversion is always observed after washing with normal medium. Only fraction 7c of middle molecules exhibits this cardiotoxicity, other fractions are without effect, even in high concentration. Uremic middle molecules strongly modify cardiac beats in vitro.