A perpendicular field electromagnet with a 250 mm access bore.

We present a laboratory electromagnet capable of generating magnetic fields up to ±0.48 T, specifically designed as a perpendicular flux source for thin film samples in an ambient environment. The magnet features a 250 mm diameter clear access bore above the sample plane, thus offering compatibility with a wide variety of experimental apparatus. Despite its generous size, the magnet thermally dissipates less than 1 kW at maximum field. A shaped ferromagnetic core is used to amplify and homogenize the field B, leading to an estimated uniformity of ±1.5 mT (≲0.3%) in B within a 28 mm2 zone at maximum field. The sample stage is thermally regulated and isolated from the magnet, enabling temperature control with ±5 mK precision even at elevated magnetic fields.

Erythropoiesis equivalence, pharmacokinetics and immune response following repeat hematide administration in cynomolgus monkeys.

Hematide is a synthetic PEGylated peptidic erythropoiesis stimulating agent (ESA) that is presently being developed for the correction of anemia in patients with chronic renal failure. Unlike currently marketed ESAs, Hematide does not possess any sequence homology to erythropoietin (EPO) and has not elicited moribund immune responses in animal safety studies thereby allowing the generation of a robust safety package. Animals administered marketed ESAs develop anti-EPO antibodies that null the effect of the administered ESA and neutralize endogenous EPO, resulting in severe anemia that precludes the interpretation of chronic safety studies. The primary objective of this study is to determine whether Hematide-specific antibodies are generated when male monkeys are exposed to high Hematide doses (10 mg/kg, intravenous [IV] and subcutaneous [SC]) administered at frequent dosing intervals (every two weeks) for a total of 9 doses; secondary objectives are to evaluate whether developed antibodies impact pharmacokinetics (PK) and pharmacology. In this study, no Hematide-specific antibodies were detected. Hematide exhibits a prolonged plasma half-life and slow clearance by either IV or SC administration. Hematide induced significant erythropoiesis with reticulocytosis and subsequent increases in red blood cells, hematocrit and hemoglobin (Hgb) levels. No erythropoietic differences were noted between the IV and the SC dosed groups with mean +/- SD Hgb levels of 20.9 +/- 2.5 and 20.3 +/- 2.1 g/dL, respectively, occurring on Day 48, corresponding to Hgb increases of 6.5 and 6.7 g/dL, respectively, over pre-dose levels. In conclusion, Hematide is a potent erythropoiesis stimulating agent that exhibits plasma persistence in monkeys. Similar erythropoietic responses were produced following IV and SC administration. The absence of antibody development suggests that Hematide, at the doses and regimen described, has a low immunogenic potential in cynomolgus monkeys.

Effect of glutathione peroxidase activity on lipid peroxidation in biological membranes.

Results are presented indicating that, although glutathione peroxidase activity inhibits lipid peroxidation in membranes, it does not appear to do so by reducing membrane lipid peroxides to lipid alcohols, as has been shown by others to be the case for free fatty acid peroxides in solution. Lipid peroxidation was studied in an enzymic system (microsomal NADPH oxidase) and in a non-enzymic system (mitochondria plus ascorbate). A study of the fatty acids in the phospholipids of microsomes and mitochondria demonstrated that detectable amounts of hydroxy fatty acids were not formed in the membranes when the latter were incubated in the presence of the glutathione peroxidase system even under conditions known to have generated significant levels of lipid peroxides in the membrane. Fatty acid analyses of the microsomal and mitochondrial particles indicated that glutathione peroxidase activity inhibited loss of polyunsaturated fatty acids when these organelles were exposed to peroxidizing conditions. If glutathione peroxidase activity were inhibiting the formation of malondialdehyde (a product of lipid peroxidation) by converting peroxide groups to alcohols, the loss of the constitutive polyunsaturated fatty acids in the membrane should not have been appreciably affected by addition of the peroxidase system. The protective effect cannot be due to quenching of an autocatalytic type of lipid peroxidation (at least in the microsomal system) since it has been established that the microsomal enzyme system (NADPH oxidase) catalyzes a continuous attack on microsomal polyunsaturated fatty acyl groups during the reaction and that the peroxidative process is not autocatalytic in nature. It appears, therefore, that glutathione peroxidase activity must exert its effect on this system by preventing free radical attack on the polyunsaturated membrane lipids in the first place. A possible mechanism for the interruption of a free radical attack on the lipids is proposed.

Studies on the properties of the singlet oxygen-like factor produced during lipid peroxidation.

The singlet oxygen reaction product of various trapping agents is observed during enzymic and nonenzymic peroxidation of microsomes as well as during the peroxidation of pure lipids extracted from microsomes. We now wish to report that purified fatty acid hydroperoxide alone, as well as peroxidized microsomal lipid and cumene hydroperoxide also form the singlet oxygen reaction product with 2,5-diphenylfuran. The reaction product (cis-1,2-dibenzoylethylene) was observed to be formed in an anaerobic system, with or without EDTA. The data indicate that a reaction of hydroxyl radicals with 2,5-diphenylfuran cannot account for the formation of dibenzoylethylene in these systems. These results are consistent with a hypothesis that the singlet oxygen-like factor was formed from the lipid peroxides per se and, in addition, supports the possibility that either the peroxides can react directly with diphenylfuran to produce dibenzoylethylene or that the self-reaction of organic peroxides may form an intermediate product which can react directly with singlet oxygen-trapping agents to produce substances which are identical to a reaction of the trapping agents with singlets oxygen.

Peptidyl transferase substrate specificity with nonaromatic aminoacyl analogues of puromycin.

A series of puromycin analogues, 3'-N-(S-substituted L-cysteinyl) puromycin aminonuleosides, has been prepared and examined as substrates for ribosomal peptidyl transferase. S-Substituted N-tert-butyloxycarbonyl-L-cysteines were coupled with puromycin aminonucleoside using dicyclohexylcarbodiimide and N-hydroxysuccinimide. Removal of the t-Boc blocking group with anhydrous trifluoroacetic acid gave the desired puromycin analogues. Kinetic studies indicate that the nonaromatic aminoacyl analogues of puromycin are effective substrates for the peptidyl transferase reaction. In addition, the discovery of the existence of hydrophilic character beyond the region normally occupied by hydrophobic amino acid R groups of the aminoacyladenyl termini of tRNA molecules, and the proper exploitation of this information, has provided the first active purmoycin analogue possessing a hydrophilic amino acid.

Puromycin analogues. Effect of aryl-substituted puromycin analogues on the ribosomal peptidyltransferase reaction.

A series of ortho- and para-substituted L-phenylalanylpuromycin analogues were synthesized and evaluated as substrates for the peptidyltransferase reaction of Escherichia coli ribosomes. Kinetic results reveal that substitution of the p-methoxy group of the puromycin molecule alters the peptidyltransferase activity of the molecule with the following decreasing order of substrate efficiencies: p-NH2 greater than p-NHCOCH3 greater than p-NO2 = p-NHCO(CH2)2CH3 greater than p-NHCOCH2Br. However, the inability of the ribosome to tolerate a nitro group at the ortho position of the phenylalanine ring precluded the use of the photosensitive puromycin analogue, 2-nitro-4-azidophenylalanylpuromycin aminonucleoside (7a), as a photoaffinity label for the peptidyltransferase site.

Internalization of recombinant tissue-type plasminogen activator by isolated rat hepatocytes is via coated pits.

The uptake and internalization of tissue-type plasminogen activator (t-PA) by freshly isolated rat hepatocytes was investigated. Electron microscopic examination of the uptake of t-PA-colloidal gold conjugates (t-PA-gold) by isolated rat hepatocytes showed that t-PA-gold was internalized via coated pits. This was inhibited with excess t-PA. Uptake of 125I-t-PA by isolated rat hepatocytes was a rapid, saturable, and specific process. The initial rate of specific uptake was 0.1 fmol/10(6) cells per min. The specific uptake plateaued at 1.4 fmol/10(6) cells by 30 min and declined to 0.8 fmol/10(6) cells at 2 h. Depletion of cellular ATP by 85-90% did not affect the initial rate of specific uptake. However, specific uptake by ATP-depleted hepatocytes at 30 min was reduced by 37%. By 2 h specific uptake by ATP-depleted hepatocytes was only 5% lower than by untreated hepatocytes, suggesting that processing of t-PA and/or its receptor is ATP-dependent. Uptake of 125I-t-PA was temperature dependent. Specific uptake was reduced by approximately 20% at 22 degrees C and by 70% at temperatures below 16 degrees C. Finally, inhibition of coated pit formation by K(+)-depletion with nigericin decreased the uptake of 125I-t-PA. This inhibition was shown to be K(+)-specific since treatment with nigericin in the presence of K+ did not inhibit coated pit formation or 125I-t-PA uptake. A threshold K(+)-depletion level for inhibition of coated pit formation was also demonstrated since treatment under conditions that reduced cellular K+ by only 54% had no effect on coated pit formation or 125I-t-PA uptake. These data support our hypothesis that internalization of t-PA by isolated rat hepatocytes is via coated pits and suggest that uptake of t-PA is a receptor-mediated process.

Dog liver N-methyltransferase. A drug-metabolizing enzyme.

A non-specific N-methyltransferase was demonstrated in dog liver. This enzyme is different from other N-methylating systems, especially in terms of substrate and species specificity. The enzyme catalyzes the methylation of a variety of endogenous and exogenous amines; of the compounds studied, SK&F 64139 (7,8-dichloro-1,2,3,4-tetrahydroisoquinoline) was found to be the best substrate. The enzyme utilized S-adenosylmethionine but not 5-methyltetrahydrofolate as a methyl donor, and it had a pH optimum at 8.0. Study of SK&F 64139 with the partially purified enzyme indicated that this dog liver N-methyltransferase had very low Km and high Vmax values for SK&F 64139. Methylation of SK&F 64139 was not observed with the monkey or rat liver enzyme preparation. This finding is in accordance with the fact that SK&F 64139 is methylated extensively in the dog, but not in other species. The ability of this enzyme to methylate a number of arylalkylamines suggests its possible importance in drug biotransformation.

Selective early loss of polypeptides in liver microsomes of CCl4-treated rats. Relationship to cytochrome P-450 content.

Treatment of rats with carbon tetrachloride (CCl4) resulted in early reproducible losses of either one or two specific polypeptides (depending on the inducing agent with which the animals had been treated) in the molecular weight range of the multiple forms of cytochrome P-450. The loss was correlated with a decrease in total cytochrome P-450 content in the microsome. The results of this study and those in the accompanying report indicate that CCl4 was metabolized by a specific form of cytochrome P-450 (52,000 daltons), which was rapidly destroyed in the process. The early loss of this peptide occurred simultaneously with the previously demonstrated production of highly reactive trichloromethyl radicals (CCl3). This polypeptide, which was shown to disappear from liver microsomes following treatment of rats with CCl4 was demonstrated in the accompanying report to be the form of cytochrome P-450 specifically required for production of the highly reactive trichloromethyl radical in a reconstituted monooxygenase system.

Specificity of a phenobarbital-induced cytochrome P-450 for metabolism of carbon tetrachloride to the trichloromethyl radical.

Evidence is presented which demonstrates that the first polypeptide to disappear in liver microsomes of phenobarbital-induced rats treated with CC14 was the 52,000 dalton p-450 cytochrome. Data are also presented which show that this form of cytochrome P-450 was capable of generating the trichloromethyl radical from CCl4 in a reconstituted system containing the purified cytochrome, NADPH-cytochrome P-450 reductase, NADPH, CCl4, and the spin-trapping agent, phenyl-t-butyl nitrone. Other cytochrome P-450 fractions not containing the 52,000 dalton form did not produce this radical. The formation of this highly reactive radical may have resulted in localized damage to the cytochrome, causing the cytochrome either to be released from the microsomal membrane or to form large aggregates which did not migrate in the gel electrophoretic procedures employed.

Clinical pharmacology of bruceantin by radioimmunoassay.

During the phase I clinical trial of a new antitumor agent, bruceantin, the pharmacology was studied in 18 cancer patients. The drug was infused intravenously (IV) for 3 h at doses ranging from 1 to 3.6 mg/m2 per day for 5 days. The plasma drug disappearance curves were biphasic, with a fast initial half-life of less than 15 min. The second half-life (t1/2 beta) varied from 0.7 to 38 h among different patients and was not dose-related. The difference between the t1/2 beta on day 1 and that on day 5 was not significant. In patients with normal liver function, the mean plasma concentration at the end of infusion was 22 ng/ml, and the value of the area under the concentration X time curve (AUC) was 111 (ng/ml)h. In contrast, in patients with abnormal liver function the corresponding values were 115 ng/ml and 830 (ng/ml)h, respectively. In addition, these patients had a slower elimination half-life of 10.9 h and a decreased total clearance of 157 ml/min/m2, as compared with 2.6 h and 671 ml/min/m2, respectively, for the normal group. All these differences were statistically significant. Patients with abnormal liver function developed more severe toxicity, including fever, severe nausea, vomiting, and hypotension. Two patients with severe hepatic dysfunction received a reduced dose and developed no toxicity. These results demonstrated the importance of the effects of liver dysfunction on drug disposition and showed that the dosage should be reduced in patients with hepatic dysfunction.

Uptake of human recombinant tissue-type plasminogen activator by rat hepatocytes in vivo: an electron microscope autoradiographic study.

The hepatic uptake of recombinant human tissue-type plasminogen activator (tPA) has been studied by electron microscope autoradiography (EMARG) of serial hepatic biopsies taken from anaesthetized, laparotomized rats following intravenous injection of 125I labeled tPA. Serial blood samples showed both radiolabel and biologic activity to be eliminated from circulation with an initial half-life of approximately two minutes. Grain half-distance distribution profiles and grain density analysis showed that the para-sinusoidal region of the hepatic parenchymal cell is the only site in the liver to concentrate radiolabeled tPA after intravenous injection. These data support the hypothesis that the parenchymal cell is the principal cell responsible for hepatic clearance of tPA from circulation and suggest that receptor mediated endocytosis may be the mechanism of cellular uptake.

Evidence for superoxide-dependent reduction of Fe3+ and its role in enzyme-generated hydroxyl radical formation.

This report describes studies yielding additional evidence that superoxide anion (O2) production by some biological oxidoreductase systems is a potential source of hydroxyl radical production. The phenomenon appears to be an intrinsic property of certain enzyme systems which produce superoxide and H2O2, and can result in extensive oxidative degradation of membrane lipids. Earlier studies had suggested that iron (chelated to maintain solubility) augmented production of the hydroxyl radical in such systems according to the following reaction sequence: O2 + Fe3+ leads to O2 + Fe2+ Fe2+ + H2O2 leads to Fe3+ + HO-+OH-. The data reported below provide additional support for the occurrence of these reactions, especially the reduction of Fe3+ by superoxide. Because the conditions for such reactions appear to exist in animal tissues, the results indicate a mechanism for the initiation and promotion of peroxidative attacks on membrane lipids and also suggest that the role of antioxidants in intracellular metabolism may be to inhibit initiation of degradative reactions by the highly reactive radicals formed extraneously during metabolic activity. This report presents the following new information: (1) Fe3+ is reduced to Fe2+ during xanthine oxidase activity and a significant part of the reduction was oxygen dependent. (2) Mn2+ appears to function as an efficient superoxide anion scavenger, and this function can be inhibited by EDTA. (3) The O2-dependent reduction of Fe3+ to Fe2+ by xanthine oxidase activity is inhibited by Mn2+, which, in view of statement 2 above, is a further indication that the reduction of the iron involves superoxide anion. (4) Free radical scavengers prevent or reverse the Fe3+ inhibiton of cytochrome c3+ reduction by xanthine oxidase. (5) The inhibition of xanthine oxidase-catalyzed reduction of cyt c3+ by Fe3+ does not affect uric acid production by the xanthine oxidase system. (6) The reoxidation of reduced cyt c in the xanthine oxidase system is markedly enhanced by Fe3+ and is apparently due to enhanced HO-RADICAL formation since the Fe3+-stimulated reoxidation is inhibited by free radical scavengers, including those with specificity for the hydroxyl radical.

Tolerability, pharmacokinetics and concentration-dependent hemodynamic effects of oral CF101, an A3 adenosine receptor agonist, in healthy young men.

OBJECTIVES: To assess safety, tolerability, pharmacokinetics and hemodynamic effects of oral CF 101, an A3 adenosine receptor (A3AR) agonist, in healthy men. METHODS: One single and 1 repeated dose, parallel-group, ascending dose, double-blind and placebo-controlled study in normal volunteers. In the single dose study, n = 15 subjects received 1, 5 or 10 mg oral CF101; in each group 1 subject received placebo, the remainder active CF101. In the repeat-dose study, n = 28 subjects received repeated 12-hourly oral doses of CF 101 (2, 3, 4 or 5 mg) for 7 days, in each group 2 subjects received placebo, the remainder active CF101. TEST MATERIALS: Single-dose study: CF101 in 30% Cremophor RH40. Multiple-dose sudy: CF101 in 0.5% methylcellulose suspension. Both studies: the corresponding vehicles were used as placebos. Galenicals were prepared remotely from the clinical study site to ensure double-blind nature of the study. RESULTS TOLERABILITY: Single doses up to 5 mg CF101 were safe and well-tolerated. However, the single dose of 10 mg CF101 was associated with flushing, tachycardia, nausea and vomiting, which were viewed as dose-limiting in normal volunteers. Single doses of CF101 (as well as the first of the multiple doses) were associated with increases in heart rate (8 - 24 beats/min after 5 mg and 18 - 55 beats/min after 10 mg). Multiple doses up to 4 mg 12-hourly for 7 days were safe and well-tolerated. However, the 5 mg multiple-dose group reported headache, drowsiness, hot flushes and dizziness on standing; this declined with dosing duration and was not dose-limiting in this study. Adverse events were commonest near t(max). RESULTS PHARMACOKINETICS: For oral CF101, the t(max) was always 1 - 2 h post-dose and t 1/2 about 9 h, in both the single- and multiple-dose studies. For a single 5 mg dose (mean +/- SD) C(max) = 81.6 +/- 23.6 ng/ml in the single dose study, and 63.6 +/- 22.0 ng/ml after the first of the multiple doses; AUC if was 904.0 +/- 221.9 ng.h/ml and 596.1 +/- 196.6 ng.h/ml for the 2 studies, respectively. After 7 days of multiple dosing there was little change, and AUC(0-24h) = 601.0 +/- 163.6 ng.h/ml. These pharmacokinetic parameters were linearly proportional to dose in the other treatment groups. RESULTS PHARMACODYNAMICS: Increases in heart rate were related to plasma concentration and evident only in the upper range of concentrations observed. There were no changes on ECG monitoring beyond sinus tachycardia, and, in particular, no evidence of PR prolongation in any subject (n = 43). In comparison with single doses, this response was almost absent after 7 days of dosing. Leucocytosis (increases up to about 1.5 x 10(9)/l after 5 and 10 mg) was similarly transient and reversible after multiple dosing. CONCLUSIONS: Single oral doses up to 5 mg CF101 and repeated doses up to 4 mg 12-hourly for 7 days were safe and well-tolerated. Multiple-dose CF101 pharmacokinetics were unchanged and predictable from single-dose estimates, and were linearly proportional to dose. Increases in heart rate and neutrophil count were reversible during multiple dosing and were not dose-limiting in the repeat dose study. CF101 warrants further study for its efficacy in treating human disease.

A fast algorithm for designing stack filters.

Stack filters are a class of nonlinear filters with excellent properties for signal restoration. Unfortunately, present algorithms for designing stack filters can only be used for small window sizes because of either their computational overhead or their serial nature. This paper presents a new adaptive algorithm for determining a stack filter that minimizes the mean absolute error criterion. The new algorithm retains the iterative nature of many current adaptive stack filtering algorithms, but significantly reduces the number of iterations required to converge to an optimal filter. This algorithm is faster than all currently available stack filter design algorithms, is simple to implement, and is shown in this paper to always converge to an optimal stack filter. Extensive comparisons between this new algorithm and all existing algorithms are provided. The comparisons are based both on the performance of the resulting filters and upon the time and space complexity of the algorithms. They demonstrate that the new algorithm has three advantages: it is faster than all other available algorithms; it can be used on standard workstations (SPARC 5 with 48 MB) to design filters with windows containing 20 or more points; and, its highly parallel structure allows very fast implementations on parallel machines. This new algorithm allows cascades of stack filters to be designed; stack filters with windows containing 72 points have been designed in a matter of minutes under this new approach.

An overview of the physiological effects of sustained high +Gz forces on human being.

In the search for the advantage in aerial combat, increasingly higher performance aircraft are being developed that impose ever greater acceleration force loads on the human pilot. The symptomatology associated with increasing +Gz acceleration forces, culminating in G-induced loss of consciousness caused by the cessation of cerebral blood circulation are described. The inability of the normal cardiovascular system mechanisms to provide adequate circulation to the brain and the eyes, and the resulting associated symptomatology and physiological changes are discussed. The normal weight, hydrostatic pressure and physiological ventilation/perfusion gradients in the lungs are exaggerated under high +Gz forces resulting in increased pulmonary arterio-venous shunting. This causes impairment of circulatory oxygenation and may also result in acceleration induced atelectasis. The effects of +Gz acceleration forces on the renal system and the limitations of the cervical musculature are also discussed. This paper serves to describe the human physiological responses to increased +Gz acceleration forces in an attempt to provide a better understanding of the body's reaction to such demanding physical stressors.

Aromatization of 7,8-dichloro-1,2,3,4-tetrahydroisoquinoline by rat liver microsomes.

The in vitro aromatization of 7,8-dichloro-1,2,3,4-tetrahydroisoquinoline (DCTQ) has been studied. Incubation of DCTQ with various rat liver subcellular fractions in the presence and absence of cofactors suggested that oxidative reactions catalyzed by microsomal enzymes were involved in this aromatization pathway. In addition to the aromatization product, 7,8-dichloroisoquinoline, three other metabolites were detected in the 9000g supernatant and microsomal incubations. By comparing the chromatographic and spectral data of the metabolites with those obtained for synthetic compounds, these three metabolites were identified as the hydroxylamine, nitrone, and the partially oxidized product (3,4-dihydro) of DCTQ. When added to microsomes, the hydroxylamine and the 3,4-dihydro derivatives were also metabolized to the 7,8-dichloroisoquinoline, and the conversions were NADPH and oxygen dependent. These findings, together with kinetic data, suggested that the aromatization of DCTQ catalyzed by rat liver microsomes was a stepwise oxidative reaction, with N-hydroxylation of DCTQ as the initial step.

Photoaffinity labeling of the ribosomal peptidyl transferase site with synthetic puromycin analogues.

A photoaffinity labeling puromycin analogue, Nepsilon-(2-nitro-4-azidophenyl)-L-lysinyl puromycin aminonucleoside (NAP-Lys-Pan), was synthesized and used for investigation of the peptidyl transferase center of 70S riobsomes. Visible light irradiation of NAP-Lys-Pan led to covalent linkage of the analogue with Escherichia coli ribosomes. In a subsequent step, poly(uridylic acid) was employed to direct Ac[14C]Phe-tRNA to the P sites of the photolabeled ribosomes. Transpeptidation of Ac[14C]phenylalanine to the bound NAP-Lys-Pan resulted in selective incorporation of radioactive label into the peptidyl transferase A site. Dissociation of the ribosomes into subunits, and digestion of the RNA components, indicated that the radioactive label was incorporated into a protein fraction of the 50S subunit.

Use of a post-column immobilized beta-glucuronidase enzyme reactor for the determination of diastereomeric glucuronides of fenoldopam in plasma and urine by high-performance liquid chromatography with electrochemical detection.

A post-column enzyme reactor, containing beta-glucuronidase immobilized on controlled-pore glass beads, was developed for use in the high-performance liquid chromatographic (HPLC) analysis of glucuronide metabolites using electrochemical detection. The reactor performance was evaluated with glucuronide conjugates of the new antihypertensive agent, fenoldopam [6-chloro-2,3,4,5-tetrahydro-1-(4-hydroxyphenyl)-1H-3-benzazepine-7,8-di ol]. These conjugates, which are electrochemically inactive at 0.6 V vs. Ag/AgCl, were separated by HPLC and passed directly into the post-column beta-glucuronidase reactor, which converted the glucuronides to their electrochemically active aglycone, fenoldopam. The enzyme reactor converted greater than 80% of the entering glucuronide to fenoldopam and produced a linear response for fenoldopam glucuronide in the range 0.4-200 ng injected on-column. The reactor performance was optimal when the mobile phase (methanol-acetate buffer) contained 0-25% methanol, but the efficiency gradually declined thereafter until, at 50% methanol, the reactor was inactive. The working pH range for the mobile phase was 5.5-8.0, with a performance optimum at pH 6.0. The reactor displayed marked stability during usage (greater than 4 months) and during storage (greater than 6 months). The reactor did not hydrolyze the 8-O-sulfate conjugate of fenoldopam but did convert the 1(R) and 1(S) diastereomers of fenoldopam-7-O-beta-glucuronide and 1(S)-fenoldopam-8-O-beta-glucuronide to fenoldopam. An assay was developed for 1(R)-fenoldopam-7-O-beta-glucuronide in plasma and urine by using the deschloro, des-4'-hydroxy analogue of fenoldopam glucuronide as the internal standard. The assay was linear in the range 4-1600 ng/ml. The within-day and between-day coefficients of variation for the method were less than 7% at three plasma fenoldopam glucuronide concentrations.