Preferential Excretion Research Articles

Species-specific dechlorane plus isomer fractionation during bioaccumulation: phenomenon and potential mechanisms

The occurrence and behavior of dechlorane plus (DP), an additive chlorinated flame retardant, have been intensively studied since it was identified in 2006. The commercial products of DP are a mixture of two stereoisomers: syn-DP and anti-DP. Stereoselective bioaccumulation of DP isomers in biota was reported in field monitoring and laboratory experiments. This review summarizes stereoselective bioaccumulation of DP in biota samples and provides the potential mechanisms for this stereoselective bioaccumulation. Stereoselective enrichment of syn-DP was widely observed in fish, whereas selective enrichment of anti-DP was mainly found in some birds. This species-specific stereoselective enrichment of DP might reflect that two different types of DP isomer fractionation occurred in bioaccumulation between ectotherms and endotherms. Anti-DP is more readily metabolized through biotransformation in all animals. However, a preferential excretion of anti-DP in fish and syn-DP in birds was observed based on the available data. Both processes determine the DP isomer fractionation in bioaccumulation. A direct comparison in DP composition between biological samples and commercial products was conducted for most studies to determine the occurrence of stereoselective DP enrichment, which may lead to underestimating the potential stereoselective enrichment of DP in organisms. The factors which affected the DP isomer composition in organisms included the tissues or organs used, DP concentration, organisms’ trophic levels occupied, and sex. Inconsistent results were obtained considering the effects of these influence factors. The underlying cause of these inconsistent results is unclear based on present data. Further research on DP biotransformation and interactions between DP and biomacromolecule is needed.

Multixenobiotic resistance mechanism: Organ-specific characteristics in the fish Prochilodus lineatus and its role as a protection against genotoxic agents

The multixenobiotic resistance mechanism (MXR) can decrease intracellular genotoxic pressure through the efflux of compounds out of the cell. Thus, this work presents a temporal approach to evaluate the MXR activity and the occurrence of genotoxic damage in different organs of the fish Prochilodus lineatus after an intraperitoneal injection of benzo[a]pyrene (B[a]P). Although the liver and brain demonstrated rapid MXR induction (6 h), the occurrence of DNA damage was not prevented. However, these organs presented some return to DNA integrity after MXR activity. The kidney demonstrated the slowest response in the MXR induction (24 h), which may be related to the preferential excretion of B[a]P metabolites by this route. Moreover, the kidney MXR reduction at 96 h may be related to its role in the excretion of metabolites from all other metabolizing organs. The gills did not appear to play an essential role in xenobiotics efflux; however, their participation in biotransformation is exhibited through the occurrence of DNA damage. The integrated response of the organs in the dynamics for the maintenance of the organism integrity could be promoted by the circulation of the xenobiotic through the bloodstream, which corroborates the increase in the DNA damage in the erythrocytes at 6 h. Therefore, the ability to induce MXR was linked to the preservation of DNA integrity in the presence of B[a]P, since MXR acts to avoid the accumulation of xenobiotics inside the cell.

Mercury isotopes of key tissues document mercury metabolic processes in seabirds

Seabirds accumulate significant amounts of mercury (Hg) due to their long-life span together with their medium to high trophic position in marine food webs. Hg speciation and Hg isotopic analyses of total Hg in different tissues (pectoral muscles, liver, brain, kidneys, blood and feathers) were assessed to investigate their detoxification mechanisms. Three species with contrasted ecological characteristics were studied: the Antarctic prion (zooplankton feeder), the white-chinned petrel (pelagic generalist consumer) and the southern giant petrel (scavenger on seabirds and marine mammals). The difference of mass-dependent fractionation (MDF, δ202Hg) values between liver and muscles (up to 0.94 ‰) in all three seabirds strongly suggests hepatic demethylation of the isotopically lighter methylmercury (MeHg) and subsequent redistribution of the isotopically heavier fraction of MeHg towards the muscles. Similarly, higher δ202Hg values in feathers (up to 1.88 ‰) relative to muscles and higher proportion of MeHg in feathers (94–97%) than muscles (30–70%) likely indicate potential MeHg demethylation in muscle and preferential excretion of MeHg (isotopically heavier) in the growing feathers during moult. The extents of these key detoxification processes were strongly dependent on the species-specific detoxification strategies and levels of dietary MeHg exposure. We also found higher mass-independent fractionation (MIF, Δ199Hg) values in feathers relative to internal tissues, possibly due to different integration times of Hg exposure between permanently active organs and inert tissues as feathers. Hg isotope variations reported in this study show evidence of detoxification processes in seabirds and propose a powerful approach for deep investigation of the Hg metabolic processes in seabirds.

Urine metallomics signature as an indicator of pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest types of cancer. Its high mortality rate is attributed largely to the difficulty of early diagnosis. Analysis of urine is an excellent non-invasive approach to trace changes in biochemical reactions due to cancer development. Here we show remarkable differences in concentration of several essential metals: significantly lower levels of urinary calcium and magnesium and increased levels of copper and zinc in PDAC when compared to healthy controls, and demonstrate that a combined analysis of these essential metals are accurate indicators (sensitivity = 99.5%) for metal dyshomeostasis in PDAC. In addition, natural stable zinc isotope composition (δ66/64Zn) in urine reveals the preferential excretion of isotopically light zinc in PDAC (δ66/64Znmedian = -0.15‰) compared to healthy controls (δ66/64Znmedian = +0.02‰), likely supporting the dysregulation of metalloproteins. These findings demonstrate for the first time that metallomics is a promising approach for discovery of biomarkers for detection of patients with PDAC, completely non-invasively, using urine samples.

Enantiomer-specific accumulation and depuration of α-hexabromocyclododecane (α-HBCDD) in chicken (Gallus gallus) as a tool to identify contamination sources

A LC-ESI(−)-HRMS method dedicated to the analysis of 6 HBCDD enantiomers at trace levels in animal matrices was developed, using a cellulose based stationary phase with a particle size of 2.5 μm. This method was applied to a sample set derived from a kinetic study of α-HBCDD previously conducted in fast- and slow-growing chickens (Gallus gallus domesticus, n = 49, plus controls), in order to study the enantiomer specific accumulation and depuration of α-HBCDD in various tissues. Regarding abdominal adipose tissue, muscle and liver, the average enantiomeric fractions of α-HBCDD (EFα) for continuously exposed groups ranged between 0.434 and 0.467, with standard deviations below 0.014, showing a significant enrichment in (−)α enantiomer even accentuated for slow growing individuals during depuration with EFα reduced by about 0.020. Similar trends were observed for pooled plasma. Then, EFα of circulating plasma α-HBCDD appeared to closely reflect EFα in storage tissues and liver, suggesting some equilibrium. The racemic elimination of α enantiomer in excreta during the contamination phase indicated that no preferential gastrointestinal absorption took place. By contrast, preferential excretion of (−)α-HBCDD from the circulating compartment to the intestinal lumen occurred during the depuration. Finally, the method was applied to samples collected in three chicken farms, selected for total HBCDD levels in muscle in the ng/g range, as a tool to determine whether the contamination occurred ante- or post-mortem, according to the chiral signature. Ante-mortem contamination was hypothesised for 2 farms, with feed being excluded as contamination source.

Quantifying mercury isotope dynamics in captive Pacific bluefin tuna (Thunnus orientalis)

Abstract Analyses of mercury (Hg) isotope ratios in fish tissues are used increasingly to infer sources and biogeochemical processes of Hg in natural aquatic ecosystems. Controlled experiments that can couple internal Hg isotope behavior with traditional isotope tracers (δ13C, δ15N) can improve the applicability of Hg isotopes as natural ecological tracers. In this study, we investigated changes in Hg isotope ratios (δ202Hg, Δ199Hg) during bioaccumulation of natural diets in the pelagic Pacific bluefin tuna (Thunnus orientalis; PBFT). Juvenile PBFT were fed a mixture of natural prey and a dietary supplement (60% Loligo opalescens, 31% Sardinops sagax, 9% gel supplement) in captivity for 2914 days, and white muscle tissues were analyzed for Hg isotope ratios and compared to time in captivity and internal turnover of δ13C and δ15N. PBFT muscle tissues equilibrated to Hg isotope ratios of the dietary mixture within ∼700 days, after which we observed a cessation in further shifts in Δ199Hg, and small but significant negative δ202Hg shifts from the dietary mixture. The internal behavior of Δ199Hg is consistent with previous fish studies, which showed an absence of Δ199Hg fractionation during Hg bioaccumulation. The negative δ202Hg shifts can be attributed to either preferential excretion of Hg with higher δ202Hg values or individual variability in captive PBFT feeding preferences and/or consumption rates. The overall internal behavior of Hg isotopes is similar to that described for δ13C and δ15N, though observed Hg turnover was slower compared to carbon and nitrogen. This improved understanding of internal dynamics of Hg isotopes in relation to δ13C and δ15N enhances the applicability of Hg isotope ratios in fish tissues for tracing Hg sources in natural ecosystems.

Can polychlorinated biphenyl (PCB) signatures and enantiomer fractions be used for source identification and to age date occupational exposure?

Detailed polychlorinated biphenyl (PCB) signatures and chiral Enantiomer Fractions (EFs) of CB-95, CB-136 and CB-149 were measured for 30 workers at a transformer dismantling plant. This was undertaken to identify sources of exposure and investigate changes to the PCB signature and EFs over different exposure periods. Approximately 1.5g of serum was extracted and PCB signatures were created through analysis by comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometry (GC×GC–TOFMS) and EFs calculated following analysis by gas chromatography with high resolution mass spectrometry (GC–HRMS). A total of 84 PCBs were identified in the serum samples with concentrations of the 7 indicator PCBs ranging from 11–350ngg−1 of serum (1.2–39μgg−1 lipid). The PCB signatures were interpreted using principal component analysis (PCA) which was able to distinguish workers with background or recent minimal exposure from those with prolonged occupational exposure. Occupationally exposed individuals had a similar PCB profile to Aroclor A1260. However, individuals with prolonged exposure had depleted proportions of several PCB congeners that are susceptible to metabolism (CB-95, CB-101 and CB-151) and elevated proportions of PCBs that are resistant to metabolism (CB-74, CB-153, CB-138 and CB-180). The results also identified a third group of workers with elevated proportions of CB-28, CB-60, CB-66, CB-74, CB-105 and CB-118 who appeared to have been exposed to an additional source of PCBs. The results show near complete removal of the CB-95 E2 enantiomer in some samples, indicating that bioselective metabolism or preferential excretion of one enantiomer occurs in humans. By considering PCB concentrations along with detailed congener specific signatures it was possible to identify different exposure sources, and gain an insight into both the magnitude and duration of exposure.

Elevated levels of δ15N in riverine Painted Turtles (Chrysemys picta): trophic enrichment or anthropogenic input?

The natural abundance of stable isotopes of elements in animal tissue is influenced by both biotic and abiotic factors. Biotically, animals feeding at higher trophic levels are enriched in the ratio of 15N:14N (δ15N) relative to their food resources owing to the preferential excretion of 14N. Abiotically, increases in δ15N may also reflect different sources of biologically available nitrogen, including nitrogen resulting from denitrification of inorganic fertilizer. We studied variation in δ15N among freshwater turtle populations to assess spatial variation in δ15N and to determine whether this variation can be attributed to differences in nitrogen source or trophic enrichment. We examined nitrogen and carbon stable isotope ratios in duckweed (genus Lemna L.) and in Painted Turtles (Chrysemys picta (Schneider, 1783)) in aquatic ecosystems expected to be differentially affected by agricultural activity and denitrification of inorganic fertilizer. Across sites, C. picta δ15N was strongly correlated with Lemna δ15N and was elevated in sites influenced by agricultural activity. Furthermore, trophic position of turtles was not associated with δ15N but was consistent with expected values for primary consumers in freshwater systems, indicating that differences in tissue δ15N could be attributed to differences in initial sources of nitrogen in each ecosystem. Our results suggest that care must be taken when attributing differences in isotopic values of animal populations to trophic factors.

Tigecycline and cyclosporine interaction—an interesting case of biliary-excreted drug enhancing the oral bioavailability of cyclosporine

To the Editor: Stumpf et al. (2009) have reported an interesting observation of enhanced blood levels of cyclosporine in a renal transplant patient upon co-medication with tigecycline [1]. The interaction between the two agents was found to be acute in nature and necessitated a dose reduction of cyclosporine to avoid safety concerns. Upon the cessation of tigecycline treatment, the dosing of cyclosporine had to be restored to its original level to avoid graft rejection due to lower efficacy [1]. The observations made in this case study are important ones since there is a myth that drugs administered through two different routes are less likely to show a pharmacokinetic interaction. Additionally, it provides rational information in our understanding of the issues associated with an intravenously administered drug (tigecycline) versus an orally administered drug (cyclosporine). While the authors have proposed p-glycoprotein (pgp) effux transporters as possible culprits for this pharmacokinetic interaction [1], this communication attempts to bring in additional views for consideration to explain this intriguing phenomenon. Although Stumpf et al. do not report the route of administration of cyclosporine, it is presumed that the renal transplant patient was on a oral dose of cyclosporine [1]. As described by the authors [1], the oral bioavailability of cyclosporine is highly variable owing to the concomitant effects of cytochrome P450 (CYP) metabolism (CYP3A4/ 3A5) and pgp-related efflux transport activity [2]. Upon the addition of tigecyline treatment, there was an acute increase in the blood concentrations of cyclosporine suggesting an enhanced oral bioavailability of cyclosporine [1]. The altered bioavailability of cyclosporine could have been achieved by two different possibilities. Firstly, as described by Stumpf et al., since tigecycline is excreted in bile (almost 59% of the administered dose [3]), it is likely that tigecycline would have impaired pgp-mediated gut efflux of cyclosporine resulting in an increased oral bioavailability. Secondly, as an alternate possibility, since cyclosporine is also known to undergo biliary excretion, perhaps there was a preferential biliary excretion of tigecycline as opposed to cyclosporine (direct competition) and/or inhibition of the biliary excrection of cyclosporine by tigecycline leading to the increased blood concentrations of cyclosporine. However, paradoxically, cyclosporine has been shown to increase the plasma concentrations of mycophenolic glucuronide due to the direct inhibition of its biliary excretion pathway [4]. While the above speculative possibilities could account for the observed enhanced bioavailability of cyclosporine in the presence of tigecylcline, one has to closely evaluate the recently reported data from the interaction study of tigecycline and digoxin [5]. Interestingly, tigecycline failed to show an interaction with digoxin, a well known and widely used probe substrate for pgp-mediated efflux [5]. Therefore, a key question to ask should be why tigecylcine would preferentially affect the pgp efflux of cyclosporine but not digoxin. A recent work published by Igel et al. (2008) sheds some light on the specificity of the involvement of apically located pgp transporters for the absorption and transport of digoxin [6]. The use of a pgp inhibitor such as quinidine in the study effectively increased the absorption of digoxin [6]. However, the bioavailability and oral absorption of antipyrine, which undergoes paracellular absorption, was Eur J Clin Pharmacol (2009) 65:543–544 DOI 10.1007/s00228-009-0631-x

Repetitive Fragmentation Products of Albumin and α1-Antitrypsin in Glomerular Diseases Associated with Nephrotic Syndrome

Even if nephrotic syndrome is characterized by massive urinary loss of major plasma proteins, a clear structural characterization based on proteomics has never been reported. Urine and plasma of 23 patients with different idiopathic nephrotic syndromes (10 steroid-sensitive minimal-change nephropathy, seven steroid-resistant FSGS, and six membranous glomerulonephritis) were analyzed with two-dimensional electrophoresis in soft gel, Western blot, and matrix-assisted laser desorption/ionization time of flight mass spectrometry; 72 urinary components corresponded to fragments of albumin and/or of alpha1-antitrypsin. Several repetitive fragmentation motives and a few differences among different pathologies were found. Several (21 of 72) urinary albumin fragments also were detected in plasma, although in lower concentration, suggesting a preferential excretion. The bulk of components with low molecular weight were detected only in urine, suggesting an in situ formation; zymograms with albumin as substrate showed the presence in urine of specific proteases. A final but not secondary point was the characterization of albumin adducts that harbor both the COOH and NH2 terminal parts of the protein, suggesting the formation of new covalent chemical groups. Altogether, these new findings reveal unexpected structural and functional aspects of proteinuria that may play a key role in pathogenesis. Characterization of urinary fragmentation patterns should be extended to other renal diseases.

Design and evaluation of “3 + 1” mixed ligand oxorhenium and oxotechnetium complexes bearing a nitroaromatic group with potential application in nuclear medicine oncology

The synthesis and evaluation of a series of oxotechnetium and oxorhenium complexes containing a nitroaromatic moiety as potential radiopharmaceuticals for targeting tumour hypoxia is presented. 99mTc labelling was performed in high yield (> 85%) and radiochemical purity (> 90%). Their structure was corroborated by means of the rhenium complexes. Reduction potentials were in the range for bioreducible compounds. 99mTc complexes III–VI were selected for “in vivo” experiments in view of the results of cytotoxicity studies. Biodistribution in normal animals was characterized by high initial blood, lung and liver uptake, fast blood and soft tissue depuration and preferential excretion via the hepatobiliary system. Initial tumour uptake was moderate but tumour/muscle ratios for complexes III and IV, were favourable at all time points. Although the results are encouraging further development is still necessary in order to achieve higher tumour uptake and lower gastrointestinal activity.

An experimental study of nitrogen flux in llamas: is 14N preferentially excreted?

Nitrogen isotope analysis is now commonly used to investigate the diets, and to a lesser extent, the environments of ancient populations. These studies assume that mammals are predictably enriched in 15N over their food, and concomitantly, that 15N becomes increasingly concentrated as one moves up the food chain. The literature commonly states that this 15N-enrichment of mammalian tissues is due to preferential excretion of light nitrogen ( 14N), but there are few data to support this assertion. To address the gap, we conducted two nitrogen flux trials in which four llamas ( Lama glama) were fed high- and low-protein diets. The ratios of fecal nitrogen loss to urinary nitrogen loss were 0.30 and 0.88 on the high- and low-protein diets respectively. Feces were enriched in 15N by approximately +3‰ on both diets, whereas urinary nitrogen was depleted in 15N (−2.1‰) on the low-protein diet, but not significantly different from intake δ 15N on the high-protein diet. Most importantly, there was no statistically significant difference between dietary and total excreta δ 15N on either diet. Given these data and theoretical considerations, we argue that the nitrogen influx and efflux of adult mammals at steady state should be isotopically commensurate. However, during growth, diet change, thermal or nutritional stress, animals may not be at steady state and fractionation between intake and excreta δ 15N may occur.

Stereoselective urinary excretion of formoterol and its glucuronide conjugate in human.

Formoterol is an inhaled beta2-adrenoceptor agonist used as a racemic mixture of the active (R; R)- and inactive (S; S)-enantiomers (rac-formoterol). Glucuronidation is an important route of metabolism in humans which occurs faster for (S; S)-formoterol in human liver microsomes. The aim of this study was to investigate the stereoselectivity of urinary excretion of formoterol and its glucuronide conjugate after oral dosing with rac-formoterol. Seven nonsmoking volunteers (six males, one female) were included in the study. After an overnight fast, a single 60 micro g oral dose of rac-formoterol fumarate dihydrate was ingested. Urine samples were collected at 1 h intervals for the first 4 h, and at 6, 8, 12 and 24 h after dosing. Formoterol enantiomers were analysed by chiral h.p.l.c. assay and formoterol glucuronides were determined as formoterol enantiomers after enzymatic cleavage with beta-glucuronidase. The female subject displayed a different pattern of metabolism and statistical analysis was therefore limited to data for the six males. The median (range) of the total urinary excretion of formoterol was 37.8% (20.9-51.2%) of the dose. The medians (ranges) of the amounts of (R; R)- and (S; S)-formoterol and of (R; R)- and (S; S)-formoterol glucuronide excreted were 2.1 (1.0-2.9), 3.5 (2.6-3.8), 21.0 (13.1-31.0) and 10.3 (4.2-14.6)%, respectively, of the dose. Unchanged (S; S)-formoterol excretion was significantly greater than that of unchanged (R; R)-formoterol and (R; R)-formoterol glucuronide excretion was significantly greater than that of (S; S)-formoterol glucuronide. The total RR-formoterol (unchanged drug plus glucuronide) excreted was significantly greater than the total (S; S)-formoterol. Our study demonstrates that the urinary excretion of formoterol in male humans after oral administration of rac-formoterol is stereoselective with preferential excretion of the active (R; R)-formoterol as unchanged drug and glucuronide. The different pattern of metabolism in the female subject provides impetus for further studies of the effect of gender on the stereoselective metabolism and pharmacokinetics of formoterol.

Pharmacokinetics of organic anions in rats with arterial calcinosis.

1. Ageing induces calcium accumulation in the vascular system. The simplest experimental way of producing high degrees of arterial calcium overload is by administration of an overdose of vitamin D(3) to rats. The aim of the present study was to evaluate the pharmacokinetics of organic anions in rats with arterial calcinosis induced by an overdose of vitamin D(3). 2. We used bromosulfophthalein (BSP) and sulfanilamide (SA) as models of organic anions with preferential biliary and renal excretion, respectively. 3. Increases in the clearance and elimination rate constant of BSP were observed in treated rats. The clearance and the elimination rate constant for SA were also increased in rats with arterial calcinosis. 4. Variations in arterial hepatic blood flow, aspartate aminotransferase activity and liver calcium accumulation were not observed in treated rats. In contrast, treated rats had a lower renal blood flow and increased renal calcium levels. 5. In summary, rats with arterial calcinosis showed an increase in total body clearance of both BSP and SA, probably associated with modifications in their metabolism and/or in organ extraction. Alterations to hepatic and renal blood flow do not account for these phenomena.

Pharmacokinetic analysis of factors determining elimination pathways for sulfate and glucuronide metabolites of drugs. I: Studies by in vivo constant infusion

1. The hepatic and renal handling of glucuronides and sulphates of three phenolic compounds, 4-methylumbelliferone (4-MU), p-nitrophenol (pNP) and acetaminophen (APAP), were evaluated pharmacokinetically by in vivo constant infusion experiments in rat. It was shown that the urinary excretion rate at steady-state was larger than the biliary excretion rate for both glucuronides and sulfates, and sulfates, in particular, were extensively excreted into the urine. 2. For each glucuronide, however, biliary excretion clearances (CLb) calculated based on the total concentration and unbound concentration in the liver were much larger than the corresponding renal excretion clearances (CLr). Even in the case of sulfates, there was not any large difference between CLr and CLb based on the total and unbound concentration in tissues, which could not explain their extensive urinary excretion. From these results, these excretion clearances were recognized not to reflect necessarily the actual excretion rate obtained. 3. On the other hand, the tissue-to-plasma concentration ratio (Kp) of both glucuronides and sulfates for every phenolic compound was much higher in the kidney than that in the liver. The results suggested that one of the most important determinants for the preferential excretion of these conjugates into the bile or urine is the extent of disposition of each compound to the liver or kidney. 4. In addition, Kp of both glucuronides and sulfates in the liver, where these conjugates are mainly formed, was small. The Kp of sulfates was quite low, suggesting that sulfates generated in the liver were subject to extensive sinusoidal efflux.

Quantification of urinary excretion of ethosuximide enantiomers and their major metabolites in the rat.

A chiral gas chromatographic assay has been developed for quantitative analysis of ethosuximide and its major metabolites in rat urine. The extraction procedure was found to be precise and reproducible. Recovery was in the range of 94-98%, intraday CV(%) was 0.92% for (S)-ethosuximide (50 microg/ml) and 0.51% for (R)-ethosuximide (50 microg/ml). Interday CV(%) was 1.12% for (S)-ethosuximide and 0.72% for (R)-ethosuximide. The limit of detection was determined to be around 0.01 microg/ml for each enantiomer. Following administration of rac-ethosuximide by i.v., i.p. and oral routes, unchanged ethosuximide was detected in urine up to 72 h after drug administration. The appearance of all detected metabolites occurred within 24 h of drug administration. Significantly more (S)-ethosuximide was excreted unchanged than (R)-ethosuximide with all three routes studied. A substantial amount of the drug was eliminated as the 2-(1-hydroxyethyl)-2-methylsuccinimide (2 pairs of diastereoisomers). Much less drug was eliminated as the 2-ethyl-3-hydroxy-2-methylsuccinimide with only one diastereoisomer observed. Examination of the one pair of diastereoisomers of 2-(1-hydroxyethyl)-2-methylsuccinimide that was resolved showed preferential excretion of one isomer. Comparison of both pairs of diastereoisomers showed that one pair was formed in preference to the other with a ratio of approximately 0.8:1. It is concluded that stereoselective metabolism of ethosuximide occurs.

Different activity of ATP dependent transport across the canalicular membrane for tributylmethylammonium and triethylmethylammonium as a potential mechanism of the preferential biliary excretion for tributylmethylammonium in the rat.

The mechanism(s) responsible for the significantly higher biliary excretion of tributyl methyl ammonium (TBuMA) than of triethyl methyl ammonium (TEMA) was investigated in canalicular liver plasma membrane vesicles (cLPM). The uptake of [3H]TBuMA and [3H]TEMA into cLPM in the presence of a pH gradient or ATP was measured by a rapid filtration technique. The uptake of substrates into the vesicle was significantly increased by an outwardly directed pH gradient. The pH dependent uptake was saturable and cross-inhibited by the other organic cation, indicating that TEMA and TBuMA share a common transport mechanism. Kinetic analysis revealed the two compounds show similar characteristics for the pH-gradient dependent uptake. Thus, the organic cation/H+ exchange mechanism does not appear to explain the significant difference in biliary excretion of the organic cations. In the presence of ATP, however, uptake into cLPM was readily observed for TBuMA while TEMA uptake was negligible. Inhibition studies with typical P-glycoprotein substrates indicated the uptake may be mediated by the P-glycoprotein. Differences between TBuMA and TEMA in reactivity for an ATP dependent transport process, rather than for an organic cation/H+ exchanger, may be responsible for the markedly different biliary excretion of TBuMA and TEMA.

Separatory determination of diastereomeric ibuprofen glucuronides in human urine by liquid chromatography/electrospray ionization-mass spectrometry.

A method for the separatory determination of diastereomeric isomers of glucuronic acid conjugates of ibuprofen having a carboxyl group at the chiral center by liquid chromatography (LC)/electrospray ionization (ESI)-mass spectrometry (MS) has been developed. The authentic specimens of acyl glucuronides of R(-)- and S(+)-ibuprofen were chemically synthesized by the Mitsunobu reaction. In the ESI mode, the glucuronides were characterized by an abundant quasi-molecular ion [M-H]-, and the formation of the negative ion was markedly influenced by a drift voltage. The resolution of diastereomeric isomers was achieved on a Develosil ODS-HG-5 column with 20 mM ammonium acetate (pH 5.0):acetonitrile (5:2, v/v) as a mobile phase where diastereomers were monitored with a corresponding quasi-molecular ion. After oral administration of racemic ibuprofen, a preferential excretion of (S)-ibuprofen glucuronide into the urine was observed.

Calcium and Strontium Metabolic Studies in Patients on Capd

Strontium is known to affect calcium metabolism both experimentally and in clinical studies on conditions other than end-stage renal failure (ESRF) and continuous ambulatory peritoneal dialysis (CAPD). To investigate Sr metabolism in relation to that of Ca in ESRF patients undergoing CAPD, and the possible influence of the duration of treatment. Cross-sectional observational study. University medical center and Institute of Nuclear Physics. Twenty-four patients on CAPD; 14 chronic renal failure (CRF) patients not on dialysis, and 52 healthy controls. Calcium and Sr content of serum, urine or dialysate effluent, and selected dietary products. Calcium and Sr are absorbed by the intestinal tract of healthy subjects with equal efficiency. Serum Ca levels were considerably lower in CRF patients than in healthy subjects and patients on CAPD (p < 0.001). Serum Sr was significantly higher in both CAPD and CRF patients than in healthy controls (p < 0.001). The Sr/Ca ratio in the sera of the healthy subjects was defined by the preferential excretion of Sr over Ca by the kidney. This preferential excretion was lost during renal failure. During treatment there was a tendency for the uptake of both Ca and Sr to increase. Strontium is accumulated in the body during renal failure and CAPD cannot restore normal levels. Considering the varying effects of different doses of Sr on bone metabolism experimentally, it would be interesting to determine by further studies the possible significance of the observed Sr accumulation for renal bone disease.

Accumulation of glycated albumin in end-stage renal failure: Evidence for the principle of “physiological microalbuminuria”

In light of the growing understanding of the toxic effects of glycated albumin and of the preferential excretion of this substance, the excretion of glycated albumin could be considered a physiologic function of the kidney. Furthermore, if the increased load of glycated albumin in diabetic patients results in glycated albumin excretion rates in the range of 20 to 200 μg/min, might this not be considered “physiologic microalbuminuria”? The hypothesis is presented that microalbuminuria composed of glycated albumin is a homeostatic renal function. Although some proteins are glycosylated for their normal physiologic function, many proteins are glycated nonenzymatically according to ambient blood glucose. Albumin is subject to nonenzymatic glycation in all humans, but at increased rates in diabetic patients. Glycated albumin induces changes in the microvasculature and glomerulus that may lead to endothelial dysfunction and diabetic nephropathy, respectively. Renal excretion of glycated albumin is enhanced compared with native albumin. To explore this potential homeostatic function of the kidney, patients with impaired renal function were studied to determine whether glycated albumin accumulates. Plasma levels of glycated albumin were determined in diabetic and nondiabetic patients on hemodialysis. Hemoglobin A 1c was used as an index of the rate of nonenzymatic glycation of proteins. Hemoglobin A 1c was increased in the diabetic subjects but was normal in the nondiabetic group (7.9% ± 0.5% v 6.2% ± 0.2%, respectively; P < 0.01). On the other hand, the glycated albumin was elevated in both groups and was not significantly different between them (1.95% ± 0.15% in the diabetic patients v 1.75% ± 0.14% in the nondiabetic patients; P = NS). The results of this study provide the first clinical evidence supporting the hypothesis that the excretion of glycated albumin is a homeostatic renal function.