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Fluorescence microscopy has emerged as an attractive technique to probe the intracellular processing of Pt-based anticancer compounds. Herein, we reported the first through-bond energy transfer (TBET) fluorescent probe NPR1 designed for sensitive detection and quantitation of Pt II complexes. The novel TBET probe was successfully applied for ratiometric fluorescence imaging of anticancer Pt II complexes such as cisplatin and JM118 in cells. Capitalizing on the ability of the probe to discriminate between Pt II complexes and their Pt IV derivatives, the probe was further applied to study the activation of Pt IV prodrug complexes that are known to release active Pt II species after intracellular reduction., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Several mobile phase additives (i.e., organic acids and their ammonium salts) were used to modulate the chromatographic retention of cyanocobalamin and its cis-diaminemonochloroplatinum(II) conjugate, depending on the specific nature of the stationary phase. Regardless of the mobile phase additive, the positively charged cyanocobalamin-cis-diaminemonochloroplatinum(II) conjugate was systematically less retained than cyanocobalamin on a conventional octadecyl-silica column. In contrast, the amide-embedded C18 column exhibited a progressive increase in the conjugate retention time upon changing the mobile phase additive from organic (acetic, formic and trifluoroacetic) acids to ammonium salts, ultimately leading to an inversion of the elution order. This change of retention was interpreted by invoking the interplay between hydrophobic interactions, hydrogen bonding between the conjugate and the polar amide groups and the ion-pairing ability of the lyophilic counterions, whereby the acetate anion was found to be the most suitable to control the solute retention., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

This study reports the transport characteristics of the pharmaceutical compounds carboplatin and cisplatin, and their respective derivatives, in saturated sand and soil columns. Pharmaceuticals are recognized as emerging pollutants of soil and water resources, but studies of the transport characteristics of organometallic pharmaceuticals in soil-water environments are rare. A recent study of oxaliplatin transport in natural soil raises the question of whether or not its behavior is representative of all Pt-based pharmaceuticals behavior in soil-water systems. To address this question, transport behaviors of carboplatin and cisplatin species were studied individually in packed sand columns under unamended conditions, and in packed soil columns under unamended and acetate-amended conditions. In contrast to oxaliplatin, carboplatin species exhibited very low affinity to both sand and soil surfaces: the retention of injected carboplatin was 3% and <6% for sand and soil, respectively. The affinity to soil was practically the same under the different redox conditions. The affinity of carboplatin to sand and soil surfaces was much smaller than the reported oxaliplatin affinity and the values reported in the literature. Cisplatin exhibited transport behavior similar to that of oxaliplatin in soil, including mild sensitivity to redox conditions (e.g., higher retention under acetate-amended conditions), overall exhibiting retention of 64-70% of the injected species. However, cisplatin also exhibited a similar retention in sand (retention of 45-53%), unlike the cases of carboplatin and oxaliplatin. The results indicate that similarly structured pharmaceuticals can exhibit very different transport characteristic in natural soil-water environments, and should therefore be studied and assessed individually., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Highly efficient, operationally stable, and pure-color organic light-emitting diodes (OLEDs) are of considerable significance for developing practical wide-color-gamut displays. Further, we have demonstrated the feasibility of an efficient pure green phosphorescent OLED (PHOLED) by employing a narrow-band platinum complex and a top-emitting structure. The utilization of the thermally activated delayed fluorescence (TADF) material as the phosphorescent host is expected to serve as a promising solution for obtaining operationally stable PHOLEDs with high color purity. However, the emission spectrum of the platinum complex in the TADF host exhibits a considerably broad emission spectrum. This study investigates the cause of the spectral change by evaluating the photoluminescence spectra of the platinum complex in various hosts exhibiting different molecular structures. The triazine unit in the host material was observed to result in exciplex formation between the lowest unoccupied molecular orbital (LUMO) of the host and the highest occupied molecular orbital (HOMO) of the platinum complex. Therefore, the TADF material that sterically hinders the triazine unit is considered to be suitable to prevent both exciplex formation and spectral broadening.

Quantitative screening for potential drug-protein binding is an essential step in developing novel metal-based anticancer drugs. ICP-MS approaches are at the core of this task; however, many applications lack in the capability of large-scale high-throughput screenings and proper validation. In this work, we critically discuss the analytical figures of merit and the potential method-based quantitative differences applying four different ICP-MS strategies to ex vivo drug-serum incubations. Two candidate drugs, more specifically, two Pt(IV) complexes with known differences of binding affinity towards serum proteins were selected. The study integrated centrifugal ultrafiltration followed by flow injection analysis, turbulent flow chromatography (TFC), and size exclusion chromatography (SEC), all combined with inductively coupled plasma-mass spectrometry (ICP-MS). As a novelty, for the first time, UHPLC SEC-ICP-MS was implemented to enable rapid protein separation to be performed within a few minutes at > 90% column recovery for protein adducts and small molecules. Graphical abstract Quantitative screening for potential drug-protein binding is an essential step in developingnovel metal-based anticancer drugs.

This study reports the transport characteristics of the organometallic anticancer compound oxaliplatin and its derivatives in natural soil-water environments. Although pharmaceuticals and their derivatives have for many years been detected in water resources, and linked to toxicological impacts on ecological systems, their transport in soil and groundwater is not fully understood. Specifically, studies that describe transport of organometallic pharmaceuticals in porous media are rare, and the transport characteristics of platinum complexes have received little attention. Oxaliplatin transport was studied in sand, as a function of two added natural chelators (citrate and humic acid), and in soil, under four continuously monitored, environmentally-relevant redox conditions: oxic, nitrate reducing, iron reducing and methanogenic. In sand, oxaliplatin species retention was about 7%, and affected only mildly by added citrate, and by humic acid under buffered pH. Transport with unbuffered humic acid was affected significantly by pH variations, and exhibited strong retention at pH < 8. In soil, unexpectedly similar breakthrough patterns of oxaliplatin species were found for all redox conditions, exhibiting linear, reversible retention of 79-87%. The strongest retention was observed under iron reducing conditions, whereas the weakest retention was under oxic conditions. Increased cation activity appears to promote weaker sorption. The results indicate that soil composition is the leading factor affecting oxaliplatin species mobility and fate in the soil-water environment, followed by the weaker factors of redox conditions and cation activities., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Purpose: The aim of this pilot study was to evaluate surface contamination by platinum drugs in the environment of patients in ICUs and wards treated by hyperthermic intraperitoneal chemotherapy (HIPEC)., Methods: The monitoring included 12 HIPEC treatments from four hospitals during the following 3 days after perfusion. A total of 33 urine and 33 drainage fluids from HIPEC patients and 160 wipe samples from several surfaces (urine/drainage bags, floors, gloves) were taken during the study period., Results: In urine, the highest platinum concentrations were measured on the first day after perfusion. Median platinum concentrations were 1260 ng/ml for patients after cisplatin perfusion and 11,000 ng/ml for oxaliplatin treatment. Concentrations decreased until day three to 413 ng/ml cisplatin and 529 ng/ml oxaliplatin, respectively. In drainage liquids, platinum concentrations were generally lower. Platinum concentrations from surfaces of bags and floors ranged from 0.01 to 439 pg/cm(2) (median: urine bag 2.77 pg/cm(2), drainage bag 0.22 pg/cm(2), floor left 0.14 pg/cm(2), floor right 0.24 pg/cm(2)), with the highest contamination found on the outer surface of the urine bags. Samples from nurses' protective gloves ranged between 0.03 and 12 pg/cm(2) (median: 0.2 pg/cm(2))., Conclusions: High platinum-drug concentrations in urine and drainage liquids are the main source of contamination. Therefore, safe handling of these liquids is the best way to avoid cross-contamination on surfaces in wards and ICUs. Our results show that it is possible to take care of HIPEC patients without high contaminations during the first 3 days.

To enhance the functionality of Pt-based reagents, several strategies have been developed that utilize Pt compounds modified with small, reactive handles. This Account encapsulates work done by us and other groups regarding the use of Pt(II) compounds with reactive handles for subsequent elaboration with fluorophores or other functional moieties. Described strategies include the incorporation of substituents for well-known condensation or nucleophilic displacement-type reactions and their use, for example, to tether spectroscopic handles to Pt reagents for in vivo investigation. Other chief uses of displacement-type reactions have included tethering various small molecules exhibiting pharmacological activity directly to Pt, thus adding synergistic effects. Click chemistry-based ligation techniques have also been applied, primarily with azide- and alkyne-appended Pt complexes. Orthogonally reactive click chemistry reactions have proven invaluable when more traditional nucleophilic displacement reactions induce side-reactivity with the Pt center or when systematic functionalization of a larger number of Pt complexes is desired. Additionally, a diverse assortment of Pt-fluorophore conjugates have been tethered via click chemistry conjugation. In addition to providing a convenient synthetic path for diversifying Pt compounds, the use of click-capable Pt complexes has proved a powerful strategy for postbinding covalent modification and detection with fluorescent probes. This strategy bypasses undesirable influences of the fluorophore camouflaged as reactivity due to Pt that may be present when detecting preattached Pt-fluorophore conjugates. Using postbinding strategies, Pt reagent distributions in HeLa and lung carcinoma (NCI-H460) cell cultures were observed with two different azide-modified Pt compounds, a monofunctional Pt(II)-acridine type and a difunctional Pt(II)-neutral complex. In addition, cellular distribution was observed with an alkyne-appended difunctional Pt(II)-neutral complex analogous in structure to the aforementioned difunctional azide-Pt(II) reagent. In all cases, significant accumulation of Pt in the nucleolus of cells was observed, in addition to broader localization in the nucleus and cytoplasm of the cell. Using the same strategy of postbinding click modification with fluorescent probes, Pt adducts were detected and roughly quantified on rRNA and tRNA from Pt-treated Saccharomyces cerevisiae; rRNA adducts were found to be relatively long-lived and not targeted for immediate degradation. Finally, the utility and feasibility of the alkyne-appended Pt(II) compound has been further demonstrated with a turn-on fluorophore, dansyl azide, in fluorescent detection of DNA in vitro. In all, these modifications utilizing reactive handles have allowed for the diversification of new Pt reagents, as well as providing cellular localization information on the modified Pt compounds.

We have developed six dihydroxidoplatinum(IV) compounds with cytotoxic potential. Each derived from active platinum(II) species, these complexes consist of a heterocyclic ligand (HL) and ancillary ligand (AL) in the form [Pt(HL)(AL)(OH)2](2+), where HL is a methyl-functionalised variant of 1,10-phenanthroline and AL is the S,S or R,R isomer of 1,2-diaminocyclohexane. NMR characterisation and X-ray diffraction studies clearly confirmed the coordination geometry of the octahedral platinum(IV) complexes. The self-stacking of these complexes was determined using pulsed gradient stimulated echo nuclear magnetic resonance. The self-association behaviour of square planar platinum(II) complexes is largely dependent on concentration, whereas platinum(IV) complexes do not aggregate under the same conditions, possibly due to the presence of axial ligands. The cytotoxicity of the most active complex, exhibited in several cell lines, has been retained in the platinum(IV) form., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Wide use of platinum-based chemotherapeutic regimens for the treatment for carcinoma calls for a simple and selective detection of platinum compound in biological samples. On the basis of the platinum(II)-base pair coordination, a novel type of aptameric platform for platinum detection has been introduced. This chemiluminescence (CL) aptasensor consists of a designed streptavidin (SA) aptamer sequence in which several base pairs were replaced by G-G mismatches. Only in the presence of platinum, coordination occurs between the platinum and G-G base pairs as opposed to the hydrogen-bonded G-C base pairs, which leads to SA aptamer sequence activation, resulting in their binding to SA coated magnetic beads. These Pt-DNA coordination events were monitored by a simple and direct luminol-peroxide CL reaction through horseradish peroxidase (HRP) catalysis with a strong chemiluminescence emission. The validated ranges of quantification were 0.12-240 μM with a limit of detection of 60 nM and selectivity over other metal ions. This assay was also successfully used in urine sample determination. It will be a promising candidate for the detection of platinum in biomedical and environmental samples.

The binding affinity of a series of square planar platinum(II) compounds of the type [Pt(A(L))(I(L))](2+), where A(L) is 1,2-diaminoethane and I(L) are 1,10-phenanthroline (phen), 4-methyl-1,10-phenanthroline (4Mephen), 5-methyl-1,10-phenanthroline (5Mephen), 4,7-dimethyl-1,10-phenanthroline (47Me2phen), 5,6-dimethyl-1,10-phenanthroline (56Me2phen) or 3,4,7,8-tetramethyl-1,10-phenanthroline (3478Me4phen) has been reinvestigated using Synchrotron Radiation Circular Dichroism (SRCD) spectroscopy. The additional peaks exhibited considerably greater intensity than those observed between 200 and 400 nm affording additional binding affinity determinations. In addition, the authors have reviewed the various mathematical approaches used to estimate equilibrium binding constants and thereby demonstrate that their mathematical approach, implemented with Wolfram Mathematica, has merit over other methods.

Dicycloplatin is a new generation supramolecular platinum-containing anti-cancer drug. Due to its structure, it is difficult to differentiate dicycloplatin from physical mixtures of carboplatin and cyclobutane dicarboxylate, and confounding results may arise during drug characterization. To solve this problem, this study aims to provide a reliable and reproducible standard for the determination of dicycloplatin. A simple method for dicycloplatin quality control has been developed using X-ray powder diffraction (XRPD) and high performance liquid chromatography (HPLC). XRPD allowed the control of impurities and dissociation of the dicycloplatin active ingredient to less than 1%, and HPLC allowed the monitoring and control of the relative molar ratio of carboplatin and cyclobutane dicarboxylate within the purity range. The study proved for the first time that the dicycloplatin supramolecule is substantially different from a physical mixture of carboplatin and cyclobutane dicarboxylate.

We report here FDCPt1, a novel selective fluorescent sensor for monofunctional platinum species. In the presence of such species, FDCPt1 exhibits a 70-fold increase in fluorescence emission, and can be used to monitor the metabolism of Pt(II)-based complexes in colorectal cancer cells. This probe is therefore expected to be valuable in studying changes in Pt coordination and distribution during chemotherapy.

The aim of this study was to evaluate air and surface contaminations, and internal contamination of healthcare workers during open-abdomen HIPEC using oxaliplatin. Platinum (Pt) was measured in urine of exposed workers and in multiple air and surface samples. Three successive HIPEC procedures were investigated in each of the two hospitals participating in the study. Analysis of air samples did not detect any oxaliplatin contamination. Heavy contamination of the operating table, the floor at the surgeon's feet, and the surgeon's overshoes were observed. Hand contamination was observed in surgeons using double gloves for intra-abdominal chemotherapy administration, but not in those using three sets of gloves. Pt was not detected in urine samples obtained after HIPEC (<5 ng/L). The main risk of HIPEC is related to direct or indirect skin exposure and can be prevented by correct use of adapted protective equipment.

The cytotoxic, platinum-based anticancer drugs, cisplatin, carboplatin and oxaliplatin, enter the aquatic environment largely in municipal wastes via excretion from outpatients undergoing chemotherapy. The environmental behaviour, effects and fate of these drugs are, however, unknown. In this study, the adsorption of the drugs to untreated and chemically modified (oxide-free and organic-free) sediment was examined in both river water and low salinity (S=3.2) estuarine water in order to determine the nature and extent of their interactions with suspended particles. In all cases, adsorption isotherms were linear, and the slopes of the relationships, or distribution coefficients (KDs), ranged from about 10(2) to 10(3) ml g(-1). Overall, adsorption decreased in the order: cisplatin>carboplatin>oxaliplatin; in river water and: cisplatin>carboplatin, oxaliplatin; in estuarine water. There was no clear dependence of adsorption on sediment treatment but, for all sediment types, both cisplatin and carboplatin adsorption was greater in river water than in estuarine water. Qualitatively, these observations are consistent with the rates of formation of reactive, aquated degradation products and the dependencies of these rates on aqueous chloride concentration. We predict that during transport through an estuarine turbidity maximum (of suspended sediment concentration=1 g L(-1)), up to about 45% of cisplatin and 35% of carboplatin are filtered out from the aqueous phase but that no more than 7% of oxaliplatin is retained., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Cisplatin, carboplatin, and oxaliplatin represent three generations of platinum based drugs applied successfully for cancer treatment. As a consequence of the employment of platinum based cytostatics in the cancer treatment, it became necessary to study the mechanism of their action. Current accepted opinion is the formation of Pt-DNA adducts, but the mechanism of their formation is still unclear. Nanomaterials, as a progressively developing branch, can offer a tool for studying the interactions of these drugs with DNA. In this study, fluorescent CdTe quantum dots (QDs, λem = 525 nm) were employed to investigate the interactions of platinum cytostatics (cisplatin, carboplatin, and oxaliplatin) with DNA fragment (500 bp, c = 25 μg/mL). Primarily, the fluorescent behavior of QDs in the presence of platinum cytostatics was monitored and major differences in the interaction of QDs with tested drugs were observed. It was found that the presence of carboplatin (c = 0.25 mg/mL) had no significant influence on QDs fluorescence; however cisplatin and oxaliplatin quenched the fluorescence significantly (average decrease of 20%) at the same concentration. Subsequently, the amount of platinum incorporated in DNA was determined by QDs fluorescence quenching. Best results were reached using oxaliplatin (9.4% quenching). Linear trend (R(2) = 0.9811) was observed for DNA platinated by three different concentrations of oxaliplatin (0.250, 0.125, and 0.063 mg/mL). Correlation with differential pulse voltammetric measurements provided linear trend (R(2) = 0.9511). As a conclusion, especially in the case of oxaliplatin-DNA adducts, the quenching was the most significant compared to cisplatin and nonquenching carboplatin., (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The interaction between oxaliplatin and the model protein ubiquitin (Ub) was investigated in a top-down approach by means of high-resolution electrospray ionization mass spectrometry (ESI-MS) using diverse tandem mass spectrometric (MS/MS) techniques, including collision-induced dissociation (CID), higher-energy C-trap dissociation (HCD), and electron transfer dissociation (ETD). To the best of our knowledge, this is the first time that metallodrug-protein adducts were analyzed for the metal-binding site by ETD-MS/MS, which outperformed both CID and HCD in terms of number of identified metallated peptide fragments in the mass spectra and the localization of the binding sites. Only ETD allowed the simultaneous and exact determination of Met1 and His68 residues as binding partners for oxaliplatin. CID-MS/MS experiments were carried out on orbitrap and ion cyclotron resonance (ICR)-FT mass spectrometers and both instruments yielded similar results with respect to number of metallated fragments and the localization of the binding sites. A comparison of the protein secondary structure with the intensities of peptide fragments generated by collisional activation of the [Ub+Pt-(chxn)] adduct [chxn = (1R,2R)-cyclohexanediamine] revealed a correlation with cleavages in solution phase random coil areas, indicating that the N-terminal β-hairpin and α-helix structures are retained in the gas phase.

A micellar electrokinetic chromatography (MEKC) method was developed for the determination of cisplatin, carboplatin, and oxaliplatin in pharmaceutical formulations. The background electrolyte consisted of a phosphate buffer (pH 7.0; 25 mM) with sodium dodecyl sulfate (80 mM). The applied voltage was 30 kV and the sample injection was performed in the hydrodynamic mode. All analyses were carried out in a fused silica capillary with an internal diameter of 50 μm and a total length of 64.5 cm. The detection of target compounds was performed at 200 nm. Under these conditions, a complete separation of cisplatin, carboplatin and oxaliplatin was achieved in less than 10 min. The MEKC-UV method was validated and trueness values between 99.7% and 100.8% were obtained with repeatability and intermediate precision values of 0.7-1.4% and 1.1-1.7%, respectively for the three drugs. This method was found appropriate for controlling pharmaceutical formulations containing platinum complexes and successfully applied in quality control at the Geneva University Hospitals., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A capillary electrophoresis-based method to characterize a PEGylated liposomal drug formulation of the anti-cancer agent oxaliplatin was developed. Pharmaceutical characterization in terms of determination of the free and total oxaliplatin concentrations in the liposomal formulation was successfully performed allowing calculation of the percentage of encapsulated drug and encapsulation efficiency. The trapping efficiency was likewise calculated. The capillary electrophoresis method allowed liposome characterization in the intended formulation media (sucrose solution with low electrolyte concentration), and the attained results were consistent with inductively coupled plasma mass spectrometry measurements. Accelerated drug leakage studies were initiated by the sonication of the PEGylated formulation, using an ultrasound probe, subsequently the drug leakage was determined by capillary electrophoresis. The results obtained with the PEGylated liposomes demonstrate that capillary electrophoresis may be a useful tool for the characterization of liposomal drug formulations., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Cholinesterase (ChE) and ethoxyresorufin-O-deethylase (EROD) were of special interest to this study as these biochemical tools have been widely used for the determination of exposure to pollutants. In this study, the freshwater oligochaete Limnodrilus profundicola was tested for its potential as a bioindicator of freshwater pollution. For this purpose, the ChE and EROD activities of L. profundicola and the level of polycyclic aromatic hydrocarbons (PAH) of water samples collected from different sites along the Curuksu stream on the Menderes River (the ancient Meander) running through south-western Turkey were studied. First, these activities were characterized using, as model substrates, acetylthiocholine (ATC), propionylthiocholine (PTC), and butyrylthiocholine (BTC). Then, the in vivo effects of insecticides and pollutants on these activities were investigated. L. profundicola were exposed to various doses of methyl-parathion, methomyl, and deltamethrin. Although significant inhibition of ChE was detected with each of the insecticides, the highest level of inhibition was observed with methyl-parathion. In addition to the inhibition of ChE, the activity of EROD was induced by exposure to oil-contaminated sediments. Thus, although L. profundicola has a reputation for being very resistant to pollution (although it is not insensitive to it), we demonstrated that it may potentially be used as a bioindicator species for contaminant exposure when ChE and EROD are used as biomarkers., (Copyright © 2010 Wiley Periodicals, Inc.)

A simple and rapid method using GC-MS and GC-FPD for the determination of pesticide contamination in processed food has been developed. Pesticides were extracted from a sample with ethyl acetate in the presence of anhydrous sodium sulfate, then cleaned up with a combination of mini-columns, such as macroporous diatomaceous earth, C18, GCB (graphite carbon black) and PSA. Recovery tests of 57 pesticides (known to be toxic or harmful) from ten kinds of processed foods (butter, cheese, corned beef, dried shrimp, frozen Chinese dumplings, grilled eels, instant noodles, kimchi, retort-packed curry and wine) were performed, and the recovery rates were mostly between 70% and 120%. This method can be used to judge whether or not processed foods are contaminated with pesticides at potentially harmful levels.

A simple and sensitive method is described for the determination of platinum surface contamination originating from cisplatin, carboplatin and oxaliplatin. Following extraction from swabs and preconcentration with the cloud point extraction (CPE) method, detection was by graphite furnace atomic absorption spectrometry (GFAAS). After desorption of platinum compounds from the swab, CPE involved on preconcentration of platinum in aqueous solution with diethyldithiocarbamate (DDTC) as chelating agent and Triton X-114 as extraction medium. DDTC is not only a chelating agent, but may also be a good candidate for the inactivation of platinum compounds. DDTC is recommended by the Word Health Organization (WHO) for the destruction of platinum-based anticancer drugs. The main factors affecting CPE efficiency, pH of the sample solution, concentrations of DDTC and Triton X-114, equilibration temperature and incubation time, were evaluated in order to enhance sensitivity of the method. The desorption of platinum compounds from the swab was investigated in parallel. Since platinum is bound to DDTC, it must exchange with copper in order to enhance platinum atomizing by GFAAS. A preconcentration factor of 29 was obtained for 10 mL of a platinum solution at 10 microg mL(-1). In optimal conditions, the limit of detection was 0.2 ng mL(-1), corresponding to 2.0 ng of platinum metal on the swab. Absorbance was linear between 0.7 and 15 ng mL(-1). The proposed method was applied for the determination of surface contamination by platinum compounds with correct results., (2009 Elsevier B.V. All rights reserved.)

Oxaliplatin is an important anti-cancer drug that has been approved for the treatment of colorectal cancer. It is known that oxaliplatin, like other Pt-based drugs, interacts with DNA to form cytotoxic Pt-DNA adducts that disrupt important biological processes such as DNA replication and protein synthesis. Linear ion trap electrospray ionisation mass spectrometry (ESI-MS) was employed to study the interaction of oxaliplatin with DNA nucleobases. It was shown that oxaliplatin formed adducts with all four DNA nucleobases when present individually and in combination in solution. Multiple-stage tandem mass spectrometry (MS(n)) enabled the fragmentation pathways of each adduct to be established. In addition, proposed structures for each product ion were obtained from the MS data. When all four bases were present together with the drug at near-equal molar concentrations, adducts containing predominantly adenine and guanine were formed, confirming that the drug preferentially binds to these nucleobases. A large molar excess of drug was required to ensure the formation of cytosine and thymine adducts in the presence of adenine and guanine. Even with a large excess of oxaliplatin, only mono-adducts of these nucleobases were observed when all four nucleobases were present.

CIEF with whole column imaging detection (WCID) was used to investigate the interaction of platinum-based anticancer drugs, cis-platinum(II) diamine dichloride (cisplatin) and [SP-4-2-{1R-trans)]-(1,2-cyclohexanediamine-N,N')[ethanedioata(2-)-O,O']platinum (oxaliplatin), with human hemoglobin A(0) (Hb). This technique facilitates the investigation and characterization of the formation of adducts between drugs and proteins. Cisplatin and oxaliplatin were mixed with the target protein at different concentrations (0:1, 1:1, 1:10, 1:50, and 1:100), and the reaction mixtures were incubated for 0, 0.5, 1, 12, 24, 48, and 72 h at 37 degrees C in a water-bath. The focused Hb-drug adduct profiles were imaged by WCID. At higher drug to protein molar ratios (for both oxaliplatin and cisplatin), the results exhibit significant changes in the peak shapes and heights, which may indicate the destabilization of the protein. However, the conformational change was less evident at lower molar ratios. In addition, a major pI shift was observed for the oxaliplatin reaction mixtures (for 1:10, 1:50, and 1:100 ratios). In comparison with previously reported findings obtained by other analytical methods, conclusions were drawn about the validity of CIEF as a simple and convenient method for the investigation of protein-drug interactions. These results may provide useful information for further understanding the activity and toxicity of these chemotherapeutic drugs and improving their clinical performance.

Metal-based anticancer agents are frequently used in the treatment of a wide variety of cancer types. The monitoring of these anticancer agents in biological samples is important to understand their pharmacokinetics, pharmacodynamics, and metabolism. In addition, determination of metals originating from anticancer agents is relevant to assess occupational exposure of health care personnel working with these drugs. The high sensitivity of inductively coupled plasma mass spectrometry (ICP-MS) has resulted in an increased popularity of this technique for the analysis of metal-based anticancer drugs. In addition to the quantitative analysis of the metal of interest in a sample, ICP-MS can be used as an ultrasensitive metal selective detector in combination with speciation techniques such as liquid chromatography. In the current review we provide a systematic survey of publications describing the analysis of platinum- and ruthenium-containing anticancer agents using ICP-MS, focused on the determination of total metal concentrations and on the speciation of metal compounds in biological fluids, DNA- and protein-adducts, and environmental samples. We conclude that ICP-MS is a powerful tool for the quantitative analysis of metal-based anticancer agents from multiple sample sources., ((c) 2008 Wiley Periodicals, Inc.)

Antitumoral Pt-containing drugs present side effects like nephrotoxicity and ototoxicity. Several systematic experiments have been carried out with Wistar rats treated with cisplatin, carboplatin, and oxaliplatin to study Pt-drugs accumulation and elimination, and Pt-biomolecule distribution in the cells and cytosols of ear, kidney, and liver. Inductively coupled plasma-mass spectrometry (ICP-MS) analysis shows a cisplatin accumulation capability between oxaliplatin (the highest) and carboplatin (the lowest). The maximum concentration of Pt in all the organs studied was achieved around the first week after cisplatin treatment. During the first 30 days, the elimination was very fast, decreasing in the subsequent 60 days in all the organs. Analysis of cytosols by liquid chromatography (LC)-ICP-MS showed an analogous behavior. In most samples, the distribution of the three drugs in the cellular and cytosolic fractions was similar for all the tissues. For kidney and ear, approximately 60% and 30%, respectively, of the metal accumulated was present in the cytosol, the cytosolic fractions smaller than 50 KDa being especially important. Cisplatin-biomolecule interaction strength under denaturing conditions was evaluated by LC-ICP-MS and showed a quite strong bond.

The present work focuses on the fate of two cancerostatic platinum compounds (CPC), cisplatin and carboplatin, as well as of two inorganic platinum compounds, [PtCl(4)](2-) and [PtCl(6)](2-) in biological wastewater treatment. Laboratory experiments modelling adsorption of these compounds onto activated sludge showed promising specific adsorption coefficients K(D) and K(OC) and Freundlich adsorption isotherms. However, the adsorption properties of the investigated substances were differing significantly. Adsorption decreased following the order cisplatin>[PtCl(6)](2-)>[PtCl(4)](2-)>carboplatin. LogK(D)-values were ranging from 2.5 to 4.3 , logK(OC) from 3.0 to 4.7. A pilot membrane bioreactor system (MBR) was installed in a hospital in Vienna and fed with wastewater from the oncologic in-patient treatment ward to investigate CPC-adsorption in a sewage treatment plant. During three monitoring periods Pt-concentrations were measured in the influent (3-250 microg l(-1) Pt) and the effluent (2-150 microgl(-1) Pt) of the treatment plant using ICP-MS. The monitoring periods (duration 30d) revealed elimination efficiencies between 51% and 63% based on averaged weekly input-output budgets. The derived logK(D)-values and logK(OC)-values ranged from 2.4 to 4.8 and from 2.8 to 5.3, respectively. Species analysis using HPLC-ICP-MS proofed that mainly carboplatin was present as intact drug in the influent and--due to low logK(D)--in the effluent of the MBR.

Hygienic standardization of soil sarin, soman, and Vx levels established the threshold and subthreshold concentrations of the agents from the general sanitary, translocational, and migratory water and migratory air safety indices. Substantiation of their maximum allowable concentrations (MAC), the limiting safety index is a migratory air index for soman and sarin and a migratory water index for Vx. Soil MACs of sarin, soman, and Vx are 2 x 10(-4), 1 x 10(-4), and 5 x 10(-5) mg/kg, respectively.

The title compound, [PtCl2(C3H9N)(NH3)], was obtained from potassium tetrachloroplatinate(II) by a two-step route. Ab initio crystal structure determination was carried out using X-ray powder diffraction techniques. Patterson and Fourier syntheses were used for the atom locations and the Rietveld technique for the final structure refinement. The Pt coordination is close to planar, with Cl atoms in a cis orientation. Molecules are combined into groups of two molecules, with antiparallel PtN2Cl2 planes and a shortest Pt...Pt distance of 3.42 angstroms. The molecule groups are packed in a parquet motif into corrugated layers parallel to ab. The molecules in the layers are linked by H-N...Cl hydrogen bonds.

Platinum originating from the excreted cancerostatic platinum compounds (CPC) cisplatin, carboplatin and oxaliplatin was monitored over a period of 28 days in the wastewater of the oncologic ward of the Vienna University Hospital. Concentration levels ranging from 4.7 to 145 microg L(-1) were measured by inductively coupled plasma mass spectrometry (ICP-MS). An average ratio of weekly drug emission/drug consumption of 0.27+/-0.12 was assessed. Model studies were carried out for fundamental understanding of CPC interaction with the solid phases present at different stages of the water cycle. Wastewater and activated sludge were spiked with CPC at concentration levels as found in the sewer of the oncologic ward. The platinum concentration remaining in the tested solution was measured after 24 h of incubation. Depending on pH, the three substances exhibited considerably different adsorption rates in wastewater. At pH 7, cisplatin was adsorbed by 88%, whereas only 26% of carboplatin and 54% of oxaliplatin were removed from the aqueous phase. Adsorption by activated sludge was higher, less affected by pH variation and comparable for all investigated CPC (96% for cisplatin, 70% for carboplatin and 74% for oxaliplatin at pH 6.8). In a next step, the dependence of CPC adsorption was tested for wastewater and activated sludge of different sampling sites. Strong variations were found only for wastewater, whereas activated sludge showed more consistent elimination rates (average values: cisplatin 92%, carboplatin 72%, and oxaliplatin 78%). These findings indicate that the major part of the excreted CPC is adsorbed by the solid phase in the water cycle and is thus expected to be removed from the wastewater by sewage treatment plants.

We report the interaction of intact human holo-transferrin (holo-Tf) with oxaliplatin (an anticancer drug), and the characterization of a complex composed of (1:1) intact holo-Tf and the parent oxaliplatin molecule using nanoelectrospray ionization quadrupole time-of-flight mass spectrometry (nanoESI-QTOF-MS). The molecular weight of this complex was determined to be 80,077 Da, which was an increase of 397 mass units compared to the protein alone (79,680 Da), suggesting that a parent drug molecule was bound to the intact protein. We further examined the interaction between the intact protein and oxaliplatin using size-exclusion high-performance liquid chromatography/inductively coupled plasma mass spectrometry (HPLC/ICPMS). The protein complex and free oxaliplatin were separated by HPLC and quantitatively determined by simultaneous monitoring of both 195Pt and 56Fe using ICPMS. The HPLC/ICPMS detected both Pt and Fe signals at retention time of 2.6 min, identifying the protein-drug complex. The Fe signal at 2.6 min did not change with the increase in incubation time of the reaction mixture containing holo-Tf and oxaliplatin, while the Pt signal at the same retention time increased over time, further demonstrating that the formation of this complex does not affect the protein-bound Fe. The binding constant of the (1:1) intact human holo-Tf-oxaliplatin complex was determined to be 7.7x10(5) M-1. Both nanoESI-MS and HPLC/ICPMS results support that the holo-Tf and parent oxaliplatin molecules form complexes through non-covalent binding, suggesting that holo-Tf may be a useful carrier for oxaliplatin delivery., (Copyright (c) 2005 John Wiley & Sons, Ltd.)

1H-detected indirect NMR techniques were used to determine 15N and 195Pt NMR parameters for a series of organoplatinum(IV) complexes and one platinum(II) complex containing nitrogen-based azobispyridine, bispyridyltetrazine, and bipyrimidine ligands. The inverse technique permitted the detection of small 4J(Pt,H) and 5J(Pt,H) long-range couplings and the acquisition of 15N NMR data in natural isotopic abundance via nJ(N,H) intra- and inter-ligand couplings, but failed in cases where coherence transfer is quenched by rapid relaxation of the metal atom. In one case, analysis of satellite patterns in a set of 1H,15N, 1H,195Pt and 1H,13C correlation spectra allowed a positive sign to be determined for 1J(Pt,15N). Qualitative arguments are presented to explain the observed 15N coordination shifts in complexes with different azine ligands in terms of azine-M dative bond formation and LnM-azine back-donation., (2004 John Wiley & Sons, Ltd.)

Four trans-planaramineplatinum(II) complexes code named YH9, YH10, YH11 and YH12, each of the form trans-PtL(NH(3))Cl(2) where L = 2-hydroxypyridine and 3-hydroxypyridine, imidazole, and imidazo(1,2-alpha)pyridine for YH9, YH10, YH11 and YH12, respectively. All of the compounds have significant anticancer activity against human cancer cell lines. YH12 is found to be significantly more active than cisplatin against cisplatin-resistant ovary cell line A2780(cisR).

AP 5280 is a novel polymer-conjugated platinum anticancer agent showing promising in vitro and in vivo activity against solid tumors. The aim of this study was to develop a parenteral pharmaceutical dosage form for phase I clinical trials. AP 5280 drug substance was characterized by using a wide range of analytical techniques and showed excellent solubility in water. However, as aqueous solutions of AP 5280 proved to be labile upon sterilization by moist heat, it was decided to develop a lyophilized dosage form. Initially, glass vials were used as primary packaging, but this led to a high breakage rate, which could be completely prevented by the use of CZ resin vials. Stability studies to date show that the lyophilized product in glass vials is stable for at least 12 months when stored at 2-8 degrees C in the dark and the lyophilized product in CZ resin vials is stable for at least 6 months under these conditions. Photostability testing revealed photolability of AP 5280 drug substance and lyophilized product in both types of primary container, necessitating storage in the dark. The first clinical experiences indicate that the proposed formulation is fully applicable for use in the clinical setting.

Platinum complexes which are known to react preferentially with guanine (G) and adenine (A) bases of oligonucleotides can be used as tools to analyze their tertiary structures and eventually to cross-link them. However, this requires efficient methods to allow the identification and quantification of the corresponding adducts which have so far been developed only for oligodeoxyribonucleotides. Maxam-Gilbert type digestions cannot be used for RNAs and HPLC techniques would require too large amounts of expensive material for separation and further characterization. We report a method to determine platination sites on oligoribonucleotides based on the cleavage activity of ribonucleases T1 and U2. To test the method, these enzymes were first used under conditions of limited digestion on 5-mer oligoribonucleotides platinated at a single defined purine. The phosphodiester bond on the 3' side of platinated G or A appeared fully resistant to cleavage by ribonuclease T1 or U2, respectively. An inhibitory effect was also observed due to neighboring platinated purines, which decreases with their distance (-2, -1, +1, +2) from the cleavage site and with the enzyme concentration. The method allowed the identification and quantification of the platination sites of a 17-mer oligoribonucleotide, based on the analysis of the mixture of monoplatinated adducts.

A rapid and sensitive RP-HPLC method with UV detection for routine control of oxaliplatin in a pharmaceutical formulation (Eloxatin) was developed. Quantitation was accomplished with the internal standard method. The procedure was validated by linearity (correlation coefficient=0.999948), accuracy, robustness and intermediate precision. Experimental design was used during validation to calculate method robustness and intermediate precision. For robustness test three factors were considered: percentage v/v of acetonitrile, flow rate and temperature; an increase in the flow rate results in a decrease of the drug found concentration, while the percentage of organic modifier and temperature have no important effect on the response. For intermediate precision measure the considered variables were: analyst, equipment and days. The RSD value (2.27%, n=24) indicated a good precision of the analytical method.

Cis-dichloroamine (4-amino-2, 2, 6, 6-tetramethylpiperidine-1-oxyl) platinum (II) (abbreviated to HJ5) is a novel platinum anticancer compound which has prospects for research and application. In order to establish a conventional analytical method, an HPLC has been studied. Under the conditions of acetonitrile-water (10:90, volume ratio) as mobile phase, detection wavelength of 210 nm, a Phenomenex column C18(150 mm x 3.9 mm i.d., 10 microns). The linear equation, recovery and accuracy of the method were determined with external standard method. The HPLC has satisfactory resolution between the peaks of HJ5 and impurities. The peak areas of HJ5 were linear to its amounts detected and the detection limit was 0.07 microgram. The hydration of HJ5 can be inhibited effectively by NaCl solution. In a 9 g/L NaCl solution, the stability can be kept for more than 2 hours which is enough for analysis.

We have investigated whether or not the cellular content of reactive platinum, aside from total cellular and DNA-bound platinum, is a measure of the growth inhibitory potential of a given platinum complex. Human MCF-7 breast cancer cells, after treatment with cisplatin [cis-diamminedichloroplatinum(II)] and several 1,2-diphenylethylenediamineplatinum(II) complexes at a fixed dose of 3 microM, were analyzed for their contents of platinum in total cells, isolated nuclei, chromosomal DNA, and the cellular pool of reactive platinum, and compared with ED50-values. Platinum was measured by atomic absorption. Reactive platinum was identified after its reaction with calf thymus DNA that had been added to the cells before their lysis. The amounts of platinum binding to chromosomal DNA were related to previously established ED50-values, and such a correlation could not be found for platinum in total cells, nuclei, and, especially, reactive platinum. The observed differences in the platinum contents of DNA were referred to variations in the rate of adduct formation rather than repair because two representative platinum complexes were indistinguishable by their effects on the chloramphenicol acetyltransferase (EC 2.3.1.28) transfection system. One of the other platinum complexes accumulated, showing an increased growth inhibition in support of this interpretation with regard to the other platinum complexes. During prolonged treatment of MCF-7 cells with the platinum(II) complexes, pools of reactive platinum were found to persist even after drug depletion in the culture medium. This suggested a hitherto unrecognized cellular storage and availability of reactive platinum.

Matrix-assisted laser desorption ionization (MALDI) time-of-flight mass spectrometry has been used to characterize the reaction products of the 18-mer deoxyribonucleotide d(AACGGTTAACCGTTAATT) with [Pt(NH3)3(H2O)]2+ and cis-[Pt(NH3)2(H2O)2]2+. Characteristic peaks corresponding to different monofunctional adducts (18-mer+n[Pt(NH3)3]) (n = 1, 2, 3 and 4) have been observed with the triamino-monoaqua complex. With the diamino-diaqua cis-Pt complex, formation of a chelate (18-mer+[Pt(NH3)2]) involving two adjacent guanines has been demonstrated. A good correlation between MALDI and polyacrylamide gel electrophoresis results is observed.

A survey of investigations performed over the last decade concerning analytical, clinical and pharmacological data on cancer chemotherapy with Pt-based drugs is reported. From this standpoint discussion focuses on clinical studies aimed at evaluating therapeutic response and toxicity during regional and systemic treatment of cisplatin against solid tumors in adults as well as in children. Concurrent treatments with locoregional hyperthermia in the case of limb tumors or with radiotherapy in the case of lung carcinoma are also dealt with.

A sensitive postcolumn derivatization/UV detection system has been developed for HPLC analysis of antitumor divalent and quadrivalent platinum complexes. It is based on the derivatization of platinum complexes by reaction with sodium bisulfite to corresponding product(s) which has enhanced absorptivity at 280-300 nm. Platinum complexes examined in this study were cisplatin, carboplatin and oxaliplatin (divalent platinum complexes) and oxoplatin and tetraplatin (quadrivalent ones). The proposed detection system was sensitive to all these complexes. Under the detection conditions optimized for individual complexes, the HPLC gave linear relationships between the complex concentration and the peak height. Detection limits at 290 nm with 100 microliters injection were 20 nM for cisplatin, 40 nM for oxoplatin, 60 nM for carboplatin and tetraplatin and 100 nM for oxaliplatin (S/N = 3 at 0.005 AUFS). The proposed system was successfully applied for the determination of cisplatin and oxoplatin in plasma and urine. Pharmacokinetic behavior of oxoplatin and its reduced product cisplatin following a single intravenous injection of oxoplatin in rabbits has been discussed.

The purpose of this study was to determine the relationship between platinum-DNA adducts in leukemic cells and the success of remission induction therapy in adult patients with acute nonlymphocytic leukemia (ANLL). Freshly isolated cells from pre-treatment bone marrow aspirates of 14 patients were incubated with cisplatin in vitro and the amount of platinum bound to DNA was determined using a competitive ELISA procedure and an antibody directed against platinum-modified DNA. Platinum-DNA adduct levels discriminated well between complete remission (CR) and remission failure (RF) patients, treated with mitoxantrone, cytosine arabinoside +/- carboplatin.

A sensitive method is described for measuring cisplatin and some possible metabolites. The method combines reversed-phase ion-pairing liquid chromatography (LC) with inductively coupled plasma mass spectrometry (ICP-MS) for platinum-specific detection. Separation conditions for cisplatin hydrolysis products and the reaction products of cisplatin with methionine, cysteine, and glutathione have been investigated with sodium dodecylsulfate or heptanesulfonate as the ion-pairing agent. The detection limit for cisplatin was found to be 0.1 ng, corresponding to a concentration detection limit of 1 ng/ml when using an injection volume of 100 microliters. This study has demonstrated the usefulness of LC-ICP-MS for cisplatin metabolism studies.

The use of ICP-MS and its various sample introduction techniques has great potential in extending the time scale of study for the pharmacokinetics of platinum containing anti-tumour drugs. Electrothermal vaporization ICP-MS can be used to measure very small samples with very low concentrations. Speciation studies of platinum in tissues can be carried out with HPLC-ICP-MS over longer time periods. Finally, LA-ICP-MS is potentially useful for studying the spatial distribution of platinum in tissues and tumours.

The stability of the five-membered chelate ring of the cisplatin analogue [meso-1,2-bis(2,6-dichloro-4- hydroxyphenyl)ethylenediamine]dichloroplatinum(II) was investigated under typical cell culture conditions (IMEM-Richter's medium with 10% fetal calf serum, 37 degrees C). For this purpose, the platinum compound was radiolabeled with tritium in the meta position of the aromatic ring by an acid-catalyzed tritium-exchange reaction, and a reversed-phase HPLC assay with radiochemical detection was developed to monitor for the presence of the free diamine ligand in the cell culture medium. A gradual increase in radioactivity attributed to the free diamine was found in medium containing the dichloroplatinum(II) complex (ca. 25% after 24 h), indicating that the diamine ligand was being released from the metal atom. When 1 mM glutathione (GSH) was included in the incubation medium, the amount of free diamine nearly doubled after 24 h, while the amount of radioactivity attributed to serum protein-platinum adducts decreased relative to incubations without GSH. On the other hand, the omission of serum from the incubations resulted in a dramatic decrease in the amount of radioactivity eluting under the diamine peak, while the concentrations of the two methionine-Pt adducts, which formed in a 1:1 ratio, rose. Through the use of liquid secondary ion mass spectroscopy, the two methionine-Pt adducts were identified as monomethionine metabolites of the title compound, whereby the two chloride ligands have been replaced by the amino acid. These compounds are probably diastereomers since the sulfur of methionine can coordinate to platinum with equal probability either cis or trans to the R-configured benzylamine carbon. On the basis of the chemical shifts of the MeS groups in the 250-MHz 1H NMR, it is concluded that a S,N-five-membered chelate ring is present in these methionine-Pt adducts.

The compatibility of ionic strength gradient with solvent-generated ion-exchange chromatography on an octadecylsilica sorbent was proven for a mobile phase containing octanesulphonate. Only a slight baseline shift was observed during the gradient of the phosphate buffer, even at 210 nm. An equilibration time of 3 min between the runs was sufficient to obtain retention times with a reproducibility better than 1%. The compounds separated were cisplatin, carboplatin and related neutral and cationic platinum(II) complexes, including transplatin and the aquation products of cisplatin.

Quantification and identification of platinum drugs and their metabolites in biological samples has always been difficult because these compounds are thermally unstable, non-volatile and insoluble. We have demonstrated that electrospray ionization mass spectrometry can be a valuable technique for direct mass spectral analysis of platinum anticancer agents and for obtaining structural information as a result of fragmentation. Full-scan spectra were obtained with approximately 10 pmol samples. These results show the potential of applying this technique in pharmacokinetic studies of platinum anticancer drugs.

Ormaplatin is a racemic, platinum(IV), anti-cancer drug which is currently involved in phase 1 clinical trials. Characterizing the purity of ormaplatin represents an analytical challenge for several reasons. These include the lack of solution stability of ormaplatin, its process impurities and solution decomposition products, the strong concentration and pH dependence of various decomposition pathways and solution equilibria and the absence of chromatographic reproducibility. Two independent types of aqueous decomposition pathways have been observed. The first pathway involves the generation of soluble PtIV containing species which remain in equilibrium with the parent material in solution. The second pathway involves the generation of PtII containing materials which tend to be less soluble. The most interesting material in this latter class is probably the chloride bridged mixed valence material which has been characterized structurally by X-ray diffraction. Liquid chromatography and sample handling procedures have been developed which allow for the accurate determination of bulk drug purity, as well as, an understanding of numerous reversible equilibria.