Liquid chromatography/tandem mass spectrometric analysis of penicillamine for its pharmacokinetic evaluation in dogs.

Copper storage disease occurs in multiple dog breeds and is one of the most common causes of chronic hepatitis in this species. The disease is caused by hereditary defects in copper metabolism in conjunction with high dietary copper levels. The progressive copper accumulation leads to hepatitis, cirrhosis, and eventually death if left untreated. Copper chelators are critical in modulating the effects of this disease. It is therefore of significant practicality to understand the pharmacokinetic (PK) parameters of chelating agents, particularly since they are oftentimes quite expensive. A liquid chromatography-tandem mass spectrometric (LC/MS/MS) method was developed to measure plasma levels of one of the most common chelators, d-penicillamine. The compound was discovered to exist in two forms, monomeric and dimeric, and various chemical derivatizations were tried to force the compound into one form or the other. Eventually, the simplest approach was individual determination of penicillamine and its dimer, with summation of the two quantities. This enabled determination of canine PK parameters for penicillamine based on comparison of oral and intravenous administration of the drug, including time to maximum drug level (Tmax), concentration at maximum (Cmax), clearance (Cls) and volume of distribution (Vdss). The drug was found to exist predominantly in the dimeric form in plasma, which is incapable of chelating copper owing to lack of free sulfhydryl groups and must therefore provide a storage form of the drug in equilibrium with its monomeric form in vivo. Mechanisms are discussed for the electrospray-induced fragmentation of penicillamine as well as of its dimer.

Coordination-Mediated Synthesis of Purification-Free Bivalent 99mTc-Labeled Probes for in Vivo Imaging of Saturable System.

In the synthesis of technetium-99m (99mTc) labeled target-specific ligands, the presence of a large excess of unlabeled ligands over 99mTc in the injectate hinders target accumulation of 99mTc-labeled ligands by competing for target molecules. To circumvent the problem, we recently developed a concept of the metal coordination-mediated multivalency, and proved the concept with a 99mTc-labeled trivalent compound [99mTc(CO)3(CN-RGD)3]+. In this study, D-penicillamine (Pen) was selected as a chelating molecule and a cyclic RGDfK peptide was conjugated to Pen via a hexanoic linkage (Pen-Ahx-c(RGDfK)). 99mTc complexation reaction, and the stability, integrin αvß3 binding affinity, and biodistribution of the 99mTc-labeled probe were investigated to evaluate the applicability of the concept to bivalent probes. 99mTc-[Pen-Ahx-c(RGDfK)]2 was obtained over 95% radiochemical yields under low Pen-Ahx-c(RGDfK) concentration (50 µM). 99mTc-[Pen-Ahx-c(RGDfK)]2 showed approximately 10-times higher integrin αvß3 binding affinity than the monovalent compounds, Pen-Ahx-c(RGDfK) and c(RGDyV). In biodistribution studies, the tumor accumulation of 99mTc-[Pen-Ahx-c(RGDfK)]2 was decreased to 77% and 43% of HPLC-purified (Pen-Ahx-c(RGDfK)-free) 99mTc-[Pen-Ahx-c(RGDfK)]2 by the presence of 5 nmol of unlabeled Pen-Ahx-c(RGDfK) and Re-[Pen-Ahx-c(RGDfK)]2, respectively. 99mTc-[Pen-Ahx-c(RGDfK)]2 provided tumor image without removing unlabeled ligand, while a 99mTc-labeled monovalent probe prepared from a monovalent ligand could not. These findings indicate the availability of the design concept to prepare 99mTc-labeled bivalent probes with a variety of 99mTc core and other metallic radionuclides of clinical relevance.

Disposition of d-penicillamine, a promising drug for preventing alcohol-relapse. Influence of dose, chronic alcohol consumption and age: studies in rats.

Pharmacokinetic studies concerning d-penicillamine (an acetaldehyde sequestering agent) are scarce and have not evaluated the influence of chronic ethanol consumption and age on its disposition. Since recent preclinical studies propose d-penicillamine as a promising treatment for alcohol relapse, the main aim of the present work was to evaluate the influence of these two factors on d-penicillamine disposition in order to guide future clinical studies on the anti-relapse efficacy of this drug in alcoholism. Additionally, the effect of the administered dose was also evaluated. To this end, three studies were carried out. Study 1 assessed the influence of dose on d-penicillamine disposition, whereas studies 2 and 3 evaluated, respectively, the influence of chronic alcohol consumption and age. Rapid intravenous administrations of 2, 10 and 30 mg/kg of d-penicillamine were performed using young or adult ethanol-naïve rats or adult ethanol-experienced (subjected to a long-term ethanol self-administration protocol) rats. Pharmacokinetic parameters were derived from the biexponential model. Statistical analysis of CL, normalized AUC0 (∞) , V1 and k10 revealed that disposition, in the range plasma concentrations assayed, is non-linear both in young ethanol-naïve and in adult ethanol-experienced rats. Notably, no significant changes in t1/2 were detected. Chronic ethanol consumption significantly reduced CL values by 35% without affecting t1/2 . d-Penicillamine disposition was equivalent in young and adult animals. In conclusion, although DP pharmacokinetics is non-linear, the lack of significant alterations of the t1/2 would potentially simplify the clinical use of this drug. Chronic consumption of ethanol also alters d-penicillamine disposition but, again, does not modify t1/2.

Novel D-penicillamine carrying nanoparticles for metal chelation therapy in Alzheimer's and other CNS diseases.

Metal ions accumulate in the brain with aging and in several neurodegenerative diseases. Aside from the copper storage disease, Wilson's disease, recent attention has focused on the accumulation of zinc, copper and iron in the Alzheimer's disease (AD) brain and the accumulation of iron in Parkinson's disease. In particular, the parenchymal deposition of beta-amyloid (Abeta) and its interaction with metal ions has been postulated to play a role in the progression of AD. Thus, the strategy of lowering brain metal ions and targeting the interaction of Abeta peptide and metal ions through the administration of chelators has merit. Our recent finding that nanoparticle delivery systems can cross the blood-brain barrier has led us to investigate whether chelators delivered conjugated to nanoparticles could act to reverse metal ion induced protein precipitation. In the present studies, the Cu (I) chelator D-penicillamine was covalently conjugated to nanoparticles via a disulfide bond or a thioether bond. Nanoparticle-chelator conjugates were stable between pH 6-8 in aqueous suspension if stored at 4 degrees C, and did not aggregate when challenged with salts and serum. Release of D-penicillamine from the nanoparticles was achieved using reducing agents such as dithiothreitol (as a model for glutathione). Nanoparticles treated only under reducing conditions that released the conjugated D-penicillamine were able to effectively resolubilize copper-Abeta (1-42) aggregates. These results indicate that nanoparticles have potential to deliver D-penicillamine to the brain for the prevention of Abeta (1-42) accumulation, as well as to reduce metal ion accumulation in other CNS diseases.

Clinical pharmacokinetics of D-penicillamine.

Penicillamine exists as 2 stereoisomers, but only the D-isomer is used therapeutically. Its chemical reactivity derives from its functional groups, of which the thiol group seems the most important. It is difficult to determine penicillamine in biological fluids because of its instability, the presence of endogenous compounds with a thiol function, and the various chemical forms in which it occurs, namely reduced free penicillamine, penicillamine bound to proteins, and internal (P-S-S-P) and mixed (P-S-S-C) disulphides. The earliest assay methods (colourimetry, isotopic methods, gas-phase chromatography) were neither sensitive nor specific. High performance liquid chromatography with electrochemical detection has led to a more specific assay for D-penicillamine, with detection based on either derivatisation reactions or on electro-oxidisation of the thiol function. With dual-electrode detectors (Au/Hg) disulphides can be assayed directly. D-penicillamine is absorbed rapidly but incompletely (40 to 70%) in the intestine, with wide interindividual variations. Food, antacids and, in particular, iron reduce absorption of the drug. Its bioavailability is also dramatically decreased in patients with malabsorption states. The peak plasma concentration occurs at 1 to 3 hours after ingestion, regardless of dose, and is of the order of 1 to 2 mg/L after an oral dose of 250 mg; some investigators have reported a double peak in plasma, which is probably not due to an enterohepatic cycle. The concentration in plasma then decreases rapidly, generally following a biphasic curve. When long term treatment is discontinued, there is a slow elimination phase lasting 4 to 6 days, which suggests that there is a 'deep compartment' or 'slow pool of the drug reversibly bound to tissues', particularly the skin. This may explain the persistence of its therapeutic effect and the occurrence of undesirable side effects after treatment has been stopped. During long term treatment plasma concentrations are highly variable between individuals. They do not seem to be correlated with the activity or the toxicity of D-penicillamine in patients with rheumatoid arthritis. More than 80% of plasma D-penicillamine is bound to proteins, particularly albumin. The rest is mainly in the free reduced form or as disulphides. Only a small portion of the dose is metabolised in the liver to S-methyl-D-penicillamine. The route of elimination is mainly renal; disulphides represent the main compounds found in the urine. Faecal excretion corresponds mainly to the non-absorbed fraction of the drug.

Clinical pharmacokinetics of slow-acting antirheumatic drugs.

The pharmacokinetics of the slow acting antirheumatic drugs (SAARDs), hydroxychloroquine, chloroquine, penicillamine, the gold complexes and sulphasalazine, in humans have been studied. For all these drugs, both in controlled clinical trials and in empirical observations from rheumatological practice, delays of several months are reported before full clinical effects are achieved. Variability in response is also characteristic of these agents. Pharmacokinetic factors may partially explain these clinical observations. Delays in the achievement of steady-state concentrations or of concentrations likely to have a therapeutic benefit may occur because of slow drug accumulation. Variable concentrations may arise after standard administered doses because of interindividual pharmacokinetic variability. These factors are likely to contribute to the delay in response and the variable response, respectively. Pharmacokinetics of the antimalarials, hydroxychloroquine and chloroquine, are characterised by extensive tissue sequestration with reported volumes of distribution in the thousands of litres. Both drugs have reported elimination half-lives of greater than 1 month. A 2- to 3-fold range occurs in the fraction of an oral dose absorbed from a tablet formulation. Variable interindividual clearance is also reported. Hydroxychloroquine and chloroquine are administered as racemates. Enantioselective disposition of both compounds occurs, again with notable interindividual variability. Sulphasalazine is split in the large intestine into sulphapyridine, proposed to be the active compound in rheumatoid arthritis, and mesalazine (5-aminosalicylic acid). Sulphapyridine is metabolised partly by acetylation, the rate of which is under genetic control. A wide range of sulphapyridine steady-state concentrations are reported after standard doses of sulphasalazine. The gold complexes are administered either intramuscularly or in an oral form (auranofin). Gold is widely distributed in the body. Very long terminal elimination half-lives and slow accumulation rates are reported. Penicillamine is administered orally. Its bioavailability is variable and may decrease by as much as 70% in the presence of food, antacids and iron salts. Penicillamine forms disulphide bonds with many proteins in the blood and tissues, creating potential slow release reservoirs of the drug. Like the other SAARDs, gold complexes and penicillamine are found in a wide range of blood concentrations after administration in standard doses to different individuals. More research must be conducted into the concentration-effect relationships of the SAARDs before the pharmacokinetic characteristics of these drugs can be used effectively to optimise patient therapy.

Penicillamine in rheumatoid arthritis. A problem of toxicity.

Penicillamine has proved an effective second line agent in rheumatoid arthritis. Its use, however, is limited by its toxicity. Long term studies show that only between 30 and 40% of patients started on penicillamine are still taking the drug at 2 years. Toxicity is the chief reason for stopping treatment, the commonest adverse effects requiring cessation of therapy being proteinuria (10 to 13%), skin rashes (5 to 9%), gastrointestinal events (5%) and thrombocytopenia or leucopenia (2 to 5%). A number of autoimmune syndromes may rarely be induced by penicillamine. HLA-B8, Dr3 positive individuals and poor sulfoxidisers are at increased risk of developing toxicity. Meticulous supervision of penicillamine therapy is required to minimise toxicity.

Biotransformation and cytotoxic properties of NO-donors on MCF7 and U251 cell lines.

Previous studies have shown a role for nitric oxide (NO) as a cytotoxic effector. In the present work, two chemically different NO-donors such as glyceryl trinitrate (GTN) and S-nitroso-N-acetylpenicillamine (SNAP) were evaluated for both NO release and cytostatic/cytotoxic properties. Nitrite accumulation in the supernatant of MCF-7 and U251 cell lines indicated a greater and quickly release of NO derived from SNAP. A time-course of hemoglobin absorption spectral changes showed a greater release of NO derived from GTN in presence of cells compared to the values observed in the media, confirming that the release of NO by GTN can be enzymatic and non-enzymatic. On the contrary, SNAP generated NO without contribution of cellular components and saturated oxyhemoglobin quickly, within 2 hours. Both NO-donors inhibited thymidine incorporation in a similar manner and dose-dependently in U251 cells, but not in MCF-7 cells, where SNAP at the highest tested dose of 1000 microM induced only a 33% cytostatic effect. About trypan blue exclusion test, after 24 h GTN and SNAP, releasing similar amounts of NO, showed comparable cytotoxic effects on U251 cells (50% dead cells), but not on MCF-7 cells, where GTN resulted more cytotoxic. From our data, the "in vitro" antitumoral activity of NO-donors seems to be related to the type of tumor cell lines, to the amount and duration of NO release.

Absorption enhancement of a protein drug by nitric oxide donor: effect on nasal absorption of human granulocyte colony-stimulating factor.

The objective of this study was to evaluate the effect of a nitric oxide (NO) donor, S-nitroso-N-acetyl-DL-penicillamine (SNAP), on the nasal absorption of recombinant human granulocyte colony-stimulating factor (rhG-CSF) in rabbits and to evaluate the irritation (cytotoxicity) potential of the NO donor on the mucosal membrane using a cultured cell system (strain KB, human epidermoid carcinoma of the floor of the mouth). Significantly higher serum G-CSF concentration and increased total leukocyte count in the peripheral blood were observed after coadministration of rhG-CSF (100 microg/kg) with SNAP at various doses (0.3-3.3 mg/kg). The serum G-CSF concentration and the increased total leukocyte count were markedly decreased by the presence of the NO scavenger, 2-(4-carboxyphenyl)-4,4,5,5-tetramethyl-imidazole-1-oxyl 3-oxide sodium salt (carboxy-PTIO), in combination with rhG-CSF and SNAP. However, no significant inhibitory effect of glutathione (peroxynitrite scavenger) on the absorption-enhancing effect of SNAP was observed. These results suggest that carboxy-PTIO inhibits the absorption-enhancing effect of NO released from SNAP. We found that SNAP has a very low potential for cytotoxicity, as evaluated by the cell detachment assay, release of lactate dehydrogenase (LDH) from cultured cells and morphological observations of nasal tissue of rabbits. It is concluded that a NO donor such as SNAP is a promising absorption enhancer for nasal protein-drug delivery.

Improvement of intestinal absorption of macromolecules by nitric oxide donor.

Nitric oxide (NO) is one of the most versatile mediators in mammalian biology. In the present study, we investigated the absorption-enhancing effects of an NO donor, 3-(2-hydroxy-1-methylethyl-2-nitrosohydrazino)-N-methyl-1-propa namine (NOC7), on drugs that are poorly absorbed from the gastrointestinal tract. NOC7 significantly increased the jejunal absorption of fluorescein isothiocyanate dextrans (FDs) of different average molecular weights (4000-20,000). This enhancing effect decreased as the FD molecular weight increased. Another NO donor, S-nitroso-N-acetyl-DL-penicillamine (SNAP), also increased the absorption of FD-4 from the jejunum. The absorption enhancement effect of NOC7 significantly decreased after coadministration with an NO scavenger, 2-(4-carboxyphenyl)-4,4,5, 5-tetramethylimidazole-1-oxyl-3-oxide, sodium salt. Furthermore, the enhancement effect of NOC7 was reversed shortly after cessation of the enhancer treatment. Little damage by NOC7 to the intestinal mucosa was observed in terms of release of lactose dehydrogenase and protein from the intestinal mucosa. NOC7 also increased the absorption of FD-4 by the colon and rectum. The findings suggest that an NO donor can improve the absorption of macromolecules from all regions of the rat intestine with very little mucosal damage and that an NO donor can act as a potent absorption enhancer.

Application of an electrochemical detector with a graphite electrode to liquid chromatographic determination of penicillamine and captopril in biological samples.

A high-performance liquid chromatographic determination of penicillamine and captopril in rat serum, liver and kidney samples is described. An electrochemical detector with a graphite working electrode at a potential of +0.9 V vs the Ag/AgCl reference electrode is used for the detection system. Linear responses of the peak height to the amount of samples injected were obtained in a range of 0.1-500 ng on-column and 0.5-500 ng on-column for penicillamine and captopril with correlation coefficients of 0.997 and 0.995, respectively. Detection limits at a signal-to-noise ratio of 3 were 20 and 300 pg for penicillamine and captopril, respectively. The graphite electrode has a long lifetime of about 4 months with continuous use, even with the high voltage supplied. The analytical application of this method to the determination of penicillamine and captopril in biological samples was successful.

The amelioration of hepatocyte oxidative stress injury by nitric oxide released from S-nitroso-N-acetyl penicillamine: a study in immobilized perfused hepatocytes.

S-nitroso-N-acetyl penicillamine (SNAP, 0.1-0.5 mM) caused release of nitric oxide (NO) into the perfusion medium of immobilized hepatocytes. Oxidative injury of hepatocytes was evoked by tert-butyl hydroperoxide (TBH, 1 mM) and the functional and morphological ultrastructural integrity of the cells was monitored. At the end of a 270-min perfusion period, SNAP-induced NO reduced lactate dehydrogenase leakage in TBH-injured hepatocytes as compared to untreated TBH-injured cells (122% +/- 5 vs. 146% +/- 6 of control levels), lipid peroxides production (2.7 +/- 0.2 vs. 3.7 +/- 0.3 nmol/10(6) cells), increased O2 consumption (26 +/- 2 vs. 12 +/- 1 nmol/10(6) cells) although urea synthesis was reduced. SNAP improved the formation of granules in the Golgi complex as compared to untreated TBH-injured hepatocytes and preserved the ultrastructural architecture of mitochondria and the smooth endoplasmic reticulum. The present data support a possible protective role of NO in oxidative liver injury.

Labeling of penicillamine di sulfide with technetium-99m.

Complex forming conditions of Penicillamine di sulfide with 99mTc have been specified. Labeling of penicillamine di sulfide with 99mTc by direct reduction with SnCl2 did not give favorable good results while the 99mTc complex of penicillamine can be easily obtained. Ligand exchange reaction with 99mTc-gluconate was attempted and a 95% labeling efficiency was obtained. Radiopharmaceutical potential of 99mTc-PADS (99mTc-Penicillamine di sulfide) has been investigated with a gamma camera in rabbits and the complex was found to be uptaken mostly by the liver and kidneys.

The metabolism and disposition of D-penicillamine in the DA-strain rat.

Radiolabelled [35S]-D-penicillamine was administered orally to DA-strain rats. After 72 hours approximately 65% of the dose was excreted with no significant differences between male and female animals. The major urinary product was inorganic sulphate with small amounts of D-penicillamine. Penicillamine disulphide, penicillamine-cysteine, S-methyl penicillamine and N-acetyl penicillamine were also found as metabolites. The female rat excreted significantly less sulphate (P less than 0.1) and more penicillamine disulphide (P less than 0.01) than the male. The residual radioactivity was found in the carcass, with slight concentration in the gut, skin, kidney, bladder and liver.

Pharmacokinetics and relative bioavailability of D-penicillamine in fasted and nonfasted dogs.

BACKGROUND: D-Penicillamine is the most commonly used copper-chelating agent in the treatment of copper-associated hepatitis in dogs. Response to therapy can be variable, and there is a lack of pharmacokinetic information available for dogs. Coadministering the drug with food to alleviate vomiting has been recommended for dogs, which contradicts recommendations for drug administration to humans. HYPOTHESIS: Coadministration of d-penicillamine with food decreases relative bioavailability and maximum plasma drug concentrations (C(max)) in dogs. ANIMALS: Nine purpose-bred dogs with a median body weight of 17.0 kg. METHODS: Dogs received D-penicillamine (12.5 mg/kg PO) fasted and with food in a randomized, crossover design. Blood samples were collected before and 0.25, 0.5, 1, 2, 3, 4, 8, 12, and 24 hours after dosing. Total d-penicillamine concentrations were measured using liquid chromatography coupled with tandem quadrupole mass spectrometry. Pharmacokinetic parameters were calculated for each dog. RESULTS: Two fasted dogs (22%) vomited after receiving d-penicillamine. Mean C(max) ± standard deviation (SD) was 8.7 ± 3.1 µg/mL (fasted) and 1.9 ± 1.6 µg/mL (fed). Mean area under the plasma concentration curve ± SD was 16.9 ± 5.9 µg/mL·h (fasted) and 4.9 ± 3.4 µg/mL·h (fed). There were significant reductions in relative bioavailability and C(max) in fed dogs (P < .001). CONCLUSIONS AND CLINICAL IMPORTANCE: Coadministration of d-penicillamine with food significantly decreases plasma drug concentrations in dogs. Decreased drug exposure could result in decreased copper chelation efficacy, prolonged therapy, additional cost, and greater disease morbidity. Administration of d-penicillamine with food cannot be categorically recommended without additional studies.

Efficacy of D-penicillamine, a sequestering acetaldehyde agent, in the prevention of alcohol relapse-like drinking in rats.

RATIONALE: Nowadays, very few approved anti-relapse treatments for alcoholism exist, and their overall efficacy can be considered moderate. An exciting rationale drug development opportunity for the treatment of chronic alcoholism is the use of acetaldehyde sequestering agents. Although these compounds are able to attenuate or prevent most of the behavioral and neurochemical effects of ethanol, the efficacy of acetaldehyde sequestration, by using agents such as D-penicillamine (DP), in relapse prevention has not been studied yet. OBJECTIVES: The aim of this study was to analyze the effects of DP treatment on the alcohol deprivation effect (ADE) in long-term ethanol-experienced rats as a model of relapse behavior and measure drug plasma and brain levels during treatment. METHODS: Rats were subcutaneously implanted with mini-osmotic pumps delivering 0, 0.25, or 1 mg/h of DP during 1 week. The efficacy to prevent ADE was determined. DP plasma and brain levels achieved during its subcutaneous administration were measured. In a second experiment, animals received bilateral infusions of 0 or 1.5 µg/h of DP directly into pVTA, and the appearance of ADE was evaluated. RESULTS: One milligram per hour, but not 0.25 mg/h, DP infusion prevented ADE and reduced the total ethanol preference in animals. DP plasma concentrations associated with ADE suppression were around 3-4 µg/ml, and brain DP levels in these conditions were about 2-3 % of those found in plasma. Intra-pVTA DP administration also suppressed ADE. CONCLUSION: DP is able to prevent alcohol-relapse-like drinking in rats suggesting that this drug may be a useful new tool in the management of relapse in alcohol-dependent patients.

Synthesis, characterization and biological evaluation of novel neutral fac-M(CO)3(SNO) complexes (M=Re, 99mTc) bearing the o-methoxyphenylpiperazine moiety.

The synthesis, characterization and biological evaluation of two new neutral tricarbonyl fac-M(CO)(3)(SNO) (M=Re, (99m)Tc) bearing o-methoxyphenylpiperazine as pharmacophore and S-functionalized cysteine or penicillamine as chelators are reported. Competition binding tests showed good affinity for the 5-HT(1A) receptor (8 and 54 nM for 4a and 4b, respectively). Biodistribution studies in healthy animals showed high initial blood and liver uptake, fast blood and tissue depuration and negligible brain uptake.

Polypeptide conjugates of D-penicillamine and idarubicin for anticancer therapy.

We investigated anticancer therapy with a novel combination of D-penicillamine (D-pen) and Idarubicin (Ida) in a synthetic dual drug conjugate (DDC). D-pen and Ida were covalently linked to poly(α)-L-glutamic acid (PGA) via reducible disulfide and acid-sensitive hydrazone bonds, respectively. The DDCs showed cell uptake and sustained release of the bound drugs in conditions mimicking the intracellular release media (10mM glutathione and pH 5.2). The in-vitro cytotoxicity of DDCs was comparable to unconjugated Ida in several sensitive and resistant cancer cell lines and correlated with the rate of cell uptake. In a single equivalent-dose pharmacokinetic study, DDCs enhanced the drug exposure by 7-fold and prolonged the plasma circulation half-life (t(1/2)) by 5-fold over unconjugated Ida. The therapeutic index of DDCs was 2-3-fold higher than unconjugated drugs. DDCs caused 89% tumor growth inhibition compared to 60% by unconjugated Ida alone and led to significant enhancement in the median survival (17%) of athymic nu/nu mice bearing NCI-H460 tumor xenografts.

99mTc-d-penicillamine-glucuronide: synthesis, radiolabeling, in vitro and in vivo evaluation.

The current study was aimed at synthesizing a glucuronide derivative of D-penicillamine (D-PA) to be used for imaging purposes. First of all, D-PA-glucuronide (D-PA-Glu) was synthesized by experimental treatments starting with uridine 5'-diphospho-glucuronosyltransferase enzyme rich microsome preparate. Then, the synthesized compound was labeled with technetium ((99m)Tc) by using a reduction method with stannous chloride. Quality controls were performed by using high-performance liquid chromatography and thin-layer radio chromatography (TLRC). Radiolabeling yield of (99m)Tc-D-PA-Glu was more than 98% according to TLRC results. In vitro evaluations of radiolabeled complexes were investigated on PC-3 human prostate cancer cells. (99m)Tc-D-PA-Glu exhibited more accumulation on PC-3 cells versus (99m)Tc-D-PA at 240 minutes. In order to determine its radiopharmaceutical potential, biodistribution studies were carried out in male Albino Wistar rats. The biodistribution results of (99m)Tc-D-PA-Glu, showed the highest uptake in prostate at 120 minutes postinjection with the main excretion route being through kidneys and bladder. (99m)Tc-D-PA-Glu and (99m)Tc-D-PA have exhibited different biodistribution results.