Analysis of antitumour [1,1-bis(aminomethyl)cyclohexane]platinum(II) complexes derived from spiroplatin by high-performance liquid chromatography with differential pulse amperometric detection

A reversed-phase high-performance liquid chromatographic analysis was developed for aqua[1,1-bis(aminomethyl)cyclohexane]sulphatoplatinum(II) (spiroplatin) and its hydrolysis and oligomerization products. The platinum complexes were detected by differential pulse amperometry at a hanging mercury drop electrode, at -540 mV vs. Ag/AgCl. The limit of detection was 0.05 microM. Aqueous solutions of spiroplatin appeared to contain the diaqua, monoaquamonosulphato, monoaquamonochloro, dichloro and hydroxo-bridged dimer complexes of [1,1-bis(aminomethyl)cyclohexane]platinum(II) in mutual equilibrium. The equilibrium shifts after dilution in infusion fluids were studied. Detection of these platinum(II) complexes in untreated human plasma ultrafiltrate and urine demonstrated the selectivity of the described analysis.     

A high performance liquid chromatography – inductively coupled plasma-mass spectrometry interface employing desolvation for speciation studies of platinum in chemotherapy drugs

A novel interface between high performance liquid chromatography (HPLC) and inductively coupled plasma-mass spectrometry (ICP-MS) is described. The eluent from the HPLC is nebulised into a heated cyclone spray-chamber and the solvent removed using a Nafion membrane drier, held at 75 degrees C, and a cryogenic condenser. The condenser consists of 6 Peltier heat pumps connected to liquid cooled aluminium blocks. At a nebuliser gas flow rate of 0.6 l min(-1), the membrane drier removes 58% of the vapour and the Peltier condenser 75% of the remaining vapour, i.e. a total desolvation efficiency of 89%. This enables the use of HPLC solvents which otherwise would destabilise the ICP, e.g. 100% acetonitrile or methanol, and permits the use of solvent gradients with minimal baseline drift. The system has been applied to the determination of platinum species in an organoplatinum drug used for chemotherapy in human plasma ultrafiltrate of patients treated with this new drug (JM-216). The limit of detection for platinum species has been 0.6 ng nl(-1) (i.e. 120 pg of Pt) and several species have been separated with good resolution.     

Validated method for the determination of platinum from a liposomal source (SPI-77) in human plasma using graphite furnace Zeeman atomic absorption spectrometry

A sensitive analytical method based on flameless atomic absorption spectrometry with Zeeman correction has been validated for the quantitative determination in human plasma of platinum originating from cisplatin in a liposomal source, SPI-77. The performance of the method was acceptable over a sample concentration range of 0. 125-1.25 micromol platinum/L and the lower limit of quantification was determined to be 1.25 micromol platinum/L in undiluted clinical samples. The performance data of the assay were investigated using both a calibration curve with carboplatin in plasma ultrafiltrate and diluted human plasma samples spiked with SPI-77. The recoveries, between-day and the within-day precisions of both methods of calibration were not significantly different allowing carboplatin ultrafiltrate calibration standards to be used to quantify platinum derived from SPI-77 in human plasma. Apparently, the liposomal formulation had no significant influence on the determination of platinum. The usefulness of the presented method was demonstrated in a phase I clinical and pharmacokinetic study. In addition, in vitro experiments were carried out to determine the distribution of SPI-77 in blood. The results indicated that platinum from SPI-77 mainly concentrates in plasma and that binding to and/or endocytosis in red blood cells is negligible.     

Validated method for the determination of platinum from a liposomal source (SPI-77) in human plasma using graphite furnace Zeeman atomic absorption spectrometry

A sensitive analytical method based on flameless atomic absorption spectrometry with Zeeman correction has been validated for the quantitative determination in human plasma of platinum originating from cisplatin in a liposomal source, SPI-77. The performance of the method was acceptable over a sample concentration range of 0.125–1.25 μmol platinum/L and the lower limit of quantification was determined to be 1.25 μmol platinum/L in undiluted clinical samples. The performance data of the assay were investigated using both a calibration curve with carboplatin in plasma ultrafiltrate and diluted human plasma samples spiked with SPI-77. The recoveries, between-day and the within-day precisions of both methods of calibration were not significantly different allowing carboplatin ultrafiltrate calibration standards to be used to quantify platinum derived from SPI-77 in human plasma. Apparently, the liposomal formulation had no significant influence on the determination of platinum. The usefulness of the presented method was demonstrated in a phase I clinical and pharmacokinetic study. In addition, in vitro experiments were carried out to determine the distribution of SPI-77 in blood. The results indicated that platinum from SPI-77 mainly concentrates in plasma and that binding to and/or endocytosis in red blood cells is negligible.     

Sensitive inductively coupled plasma mass spectrometry assay for the determination of platinum originating from cisplatin, carboplatin, and oxaliplatin in human plasma ultrafiltrate

We present a highly sensitive, rapid method for the determination of platinum originating from the anticancer agents cisplatin, carboplatin, and oxaliplatin in human plasma ultrafiltrate. The method is based on the quantification of platinum by inductively coupled plasma mass spectrometry and allows quantification of 7.50 ng l-1 platinum in only 150 microl of matrix. Sample pretreatment involves dilution of samples with 1% HNO3. Validation fulfilled the most recent FDA guidelines for bioanalytical method validation. Validated ranges of quantification were 7.50 ng l-1 to 1.00x10(5) ng l-1 in plasma ultrafiltrate for all three platinum compounds. The assay is now successfully used to support pharmacokinetic studies in cancer patients treated with cisplatin, carboplatin, or oxaliplatin.     

HG/LT-GC/ICP-MS coupling for identification of metal(loid) species in human urine after fish consumption

Human urine samples after fish consumption have been investigated by low-temperature gas chromatography with inductively coupled plasma mass spectrometric detection after sample derivatization by hydride generation (HG/LT-GC/ICP-MS). This analytical technique enabled the identification of organometal(loid) compounds in human urine; species of the six elements germanium, arsenic, selenium, tin, antimony, and mercury were determined. Three different organic selenium species, two germanium species, seven arsenic species, four tin species, five antimony species, and one species of mercury were found; 18 of the 22 species detected could be identified. The relative detection limits ranged between 2 and 12 pg x L(-1) (x=element). These organometal(loid) compounds probably build up in the human body under the influence of micro-organisms, in the presence of hydrogen sulfide and methane, in the human intestine.     

Determination of penicillamine, penicillamine disulfide and penicillamine-glutathione mixed disulfide by high-performance liquid chromatography with electrochemical detection

Methodology is described for the simultaneous determination of D-penicillamine, penicillamine disulfide and the penicillamine-glutathione mixed disulfide, as well as glutathione and glutathione disulfide, in human plasma, erythrocytes and urine. The various thiols and disulfides are separated by reversed-phase ion-pairing liquid chromatography with detection by an electrochemical detector with dual gold/mercury amalgam electrodes in series. The thiols are detected at the downstream electrode; the disulfides are reduced at the upstream electrode and then detected as the thiols at the downstream electrode. Detection limits (at a signal-to-noise ratio of 2.0) are in the picomole range for 20 microliters of injected solution for all compounds except penicillamine disulfide, which has a detection limit of 600 pmol in 20 microliters. A convenient method is described for preparation of the penicillamine-glutathione mixed disulfide by thiol/disulfide exchange with standardization of the solution by 1H NMR spectroscopy.     

The determination of platinum in sea water by adsorptive cathodic stripping voltammetry

The catalytic effect of the formazone complex with platinum on the development of hydrogen at a mercury electrode is used to provide a very sensitive method of determining dissolved platinum in fresh and sea water by adsorptive cathodic stripping voltammetry. The optimal analytical conditions include the addition of 0.012% (w/v) formaldehyde, 0.0015% (w/v) hydrazine and 0.5 M sulphuric acid to the sea water. The complex is adsorbed for 1–20 min at ?0.925 V on the hanging mercury drop electrode, followed by a differential-pulse scan in a negative direction. The hydrogen reduction peak catalysed by the platinum complex appears at ?1.045 V. The limit of detection is 0.04 pM Pt with an adsorption time of 10 min, which is quite sufficient to determine dissolved platinum in sea water. The interference of surface-active compounds is eliminated by ultraviolet irradiation of the acidified sample. The method is applied to several sea-water samples. Comparative experiments showed that stripping chronopotentiometry is not sufficiently sensitive to determine platinum in sea water without preconcentration.     

Integration of microchip electrophoresis with electrochemical detection using an epoxy-based molding method to embed multiple electrode materials

This paper describes the use of epoxy-encapsulated electrodes to integrate microchip-based electrophoresis with electrochemical detection. Devices with various electrode combinations can easily be developed. This includes a palladium decoupler with a downstream working electrode material of either gold, mercury/gold, platinum, glassy carbon, or a carbon fiber bundle. Additional device components such as the platinum wires for the electrophoresis separation and the counter electrode for detection can also be integrated into the epoxy base. The effect of the decoupler configuration was studied in terms of the separation performance, detector noise, and the ability to analyze samples of a high ionic strength. The ability of both glassy carbon and carbon fiber bundle electrodes to analyze a complex mixture was demonstrated. It was also shown that a PDMS-based valving microchip can be used along with the epoxy-embedded electrodes to integrate microdialysis sampling with microchip electrophoresis and electrochemical detection, with the microdialysis tubing also being embedded in the epoxy substrate. This approach enables one to vary the detection electrode material as desired in a manner where the electrodes can be polished and modified as is done with electrochemical flow cells used in liquid chromatography.     

Dual Electrode Liquid Chromatography-Electrochemical Detection (LCEC) for Platinum-Derived Cancer Chemotherapy Agents

Abstract

Trace methods of analysis and speciation have now been developed for a number of platinum derived anti-cancer chemotherapeutic agents, drugs such as: cis-dichloro diammine platinum (CDDP), cis-diammine-1,1-cyclobutane dicarboxylate platinum (CBDCA), and cis-dichloro-trans-dihydroxy diisopropylamine platinum (CHIP). It is possible to utilize parallel dual electrode operations for all three of these Pt derivatives, with overall improved analyte specificity and identification. At the same time, these approaches provide calibration plots of detector sensitivity as a function of the particular working electrode potentials in use via dual electrode LCEC. These response ratios as a function of the applied potentials then become quite unique for individual Pt compounds. By suitably selecting the operating electrode potentials in parallel operation, it is possible to alter the detectability of individual Pt analytes and to drastically vary the resultant LCEC chromatograms. The overall analyte selectivity possible via dual electrode LCEC surpasses that thus far possible via LC-polarographic reduction or single electrode LCEC operations. Glassy carbon as well as gold/mercury electrodes can readily be used for some of these Pt derivatives. These overall trace methods of analysis and speciation for the Pt anti-cancer agents have also been applied to plasma samples spiked with known levels of each drug. It is also possible to utilize these single or dual electrode approaches for the analysis of each of these Pt derivatives in cancer patients undergoing chemotherapy treatment.     

Determination of cisplatin and related platinum complexes in plasma ultrafiltrate and urine by high-performance liquid chromatography with on-line radioactivity detection

A reversed-phase ion-pair chromatographic method with on-line radioactivity detection for the simultaneous determination of 195mPt-labelled cisplatin and related platinum complexes has been developed. With this system a good resolution of various radiolabelled platinum complexes can be achieved. The detection limit of the radioactivity detector is 10 ng of cisplatin (specific activity of 15 MBq/mg cisplatin) per millilitre of urine or plasma ultrafiltrate. The detector response is independent of both the chemical structure of the platinum complexes and the matrix composition of the samples. This method may serve as a reference system for other high-performance liquid chromatographic systems with less specific and sensitive detectors.     

HPLC determination of cis-diamminedichloroplatinum(II) in plasma and urine with UV detection and column-switching

A method for determining cis-diamminedichloroplatinum(II) (CDDP), an anticancer drug, in plasma and urine by HPLC with UV detection and column-switching has been developed. Typical conditions were as follows. An apparatus was composed of two columns, two pumps, a UV detector, a sample injector with a 100 microL loop, a switching valve, a column oven and a recorder. A Rheodyne model 7125 sample injector was used as the switching valve. A precolumn (4.6 mm ID x 25 cm) was packed with MCI GEL CK10S (a strong cation exchanger), and an analytical column (4.6 mm ID x 5 cm) was packed with MCI GEL CDR10 (a strong anion exchanger). Both columns were connected in series via the switching valve. The CDDP-containing fraction of the effluent from the precolumn was loaded to the analytical column by column-switching and the effluent from the analytical column was monitored at 210 nm. An eluent of 0.3 M sodium dihydrogen phosphate was pumped at a flow rate of 1 mL/min and the columns were maintained at 40 degrees C. CDDP was eluted at about 11 min and the identity of the peak of CDDP on the chromatogram was confirmed by its 3-dimensional chromatogram and analysis of platinum in the column effluent. Under the conditions described above, a linear relationship was obtained between peak height and concentration of CDDP up to 100 microM. Correlation efficients were 0.998 for plasma and 0.999 for urine. The detection limit was 0.1 microM for CDDP in both plasma and urine (S/N = 3,0.005 AUFS). The reproducibility was within 3% for 10 determinations.(ABSTRACT TRUNCATED AT 250 WORDS)     

Speciation analysis for mercury in gas condensates by capillary gas chromatography with inductively coupled plasma mass spectrometric detection

A method allowing species-selective determination of atomic mercury, non-polar dialkylated mercury compounds,polar monoalkylated species and inorganic mercury complexes in natural gas condensates was developed. Inductively coupled plasma mass spectrometry was employed as a detection method for capillary gas chromatography and compared with microwave induced plasma atomic emission detection for the analysis of hydrocarbon-rich matrices. The method was based on two consecutive injections allowing comprehensive speciation analysis. First a sample aliquot was diluted with toluene and analysed for Hg0 and individual dialkylmercury compounds. Then, another aliquot was butylated with a Grignard reagent for the species specific determination of Hg(II) and monoalkylated mercury species. The detection limits were down to 0.08 pg level.     

Cyclic voltammetric investigation of the reduction of tin(ii) chloride in acetonitrile

Cyclic voltammetric data collected at mercury and platinum electrodes are presented for the reduction of tin(II) chloride in acetonitrile and acetonitrile—tetramethylammonium chloride solutions. The data taken on mercury generally confirm the nature of the electrode processes postulated from polarographic and spectrophotometric data. Voltammetric data taken on platinum suggest the production of a soluble tin species via a le- reduction of the SnCl₃ -ion.     

Electrochemical investigations of cationictrans-haloarylisocyanidebis(tertiaryphosphino)platinum (II) complexes

Summary The electrochemical behaviour of a series of cationic platinum(II) isocyanide complexes has been studied in acetonitrile. All the tested compounds are oxidized at a platinum electrode via a two-electron process and reduced at a platinum or mercury electrode via two successive one-electron steps. The anodic step involves the formation of platinum(IV) complexes. The main reduction product formed in correspondence to the first cathodic process is a stable dimer platinum(I) containing bridging isocyanide ligands. Platinum(0) species are formed in the subsequent reduction step.     

The determination of cysteamine in physiological fluids by HPLC with electrochemical detection

Cysteamine, an amino thiol, was separated by rapid isocratic cation exchange chromatography and detected by electrochemical oxidation at a platinum electrode maintained at +0.45 V relative to an Ag/AgCl reference electrode. Eluent pH and electrode working potentials were optimized and the effects of alternative buffers and organic modifiers have been examined. On column sensitivity for cysteamine was 1.5 pmol at a signal-to-noise ratio of 5. Although the specificity was good, plasma samples required maximal sensitivity whereas urine samples required greater selectivity, which was achieved by use of lower working potentials. Cysteamine concentrations were determined in serial samples of plasma and urine from volunteers who had received a single oral dose of 200 mg of the drug. Cysteamine was rapidly oxidized in vivo, and detection required prior reduction with dithiothreitol before analysis.     

Post-column reaction detection of biotin in human plasma ultrafiltrate based on laser-induced fluorescence energy transfer in the far-red spectral region

High performance liquid chromatography followed by post-column reaction detection in the far-red spectral region provides added sensitivity and selectivity. A homogeneous fluorescence energy transfer assay in the competitive mode based on the binding of biotin and streptavidin was developed as an on-line post-column reaction detection system. The labels used for energy transfer were R-Phycoerythrin conjugated to biotin and Cyanine 5 labeled with streptavidin. The energy transfer peak was measured at 670 nm and excitation was achieved using the 488 nm line of an argon ion laser. The biotin concentration in plasma ultrafiltrate ranged from 0.024 to 6.12 ng/mL (n = 6). The precision of the two controls, 0.24 and 2. 44 ng/mL, was found to be 18.70% and 9.92% relative standard deviation respectively. Accuracy was 10.47% and 1.95% difference from spiked, respectively (n = 6). The limit of detection was 21.70 pg/mL (8.90 x 10(-11)M) calculated based on a factor of 2x the standard deviation of the blank (n = 6). The correlation coefficient for the calibration curve was found to be 0.9995. Recovery from plasma ultrafiltrate at 2.44 ng/mL was 103.40% (n = 6). Detection selectivity was indicated by the absence of background fluorescence in six different plasma samples collected from six individual donors. Endogenous levels were detected in two of the six pools of plasma ultrafiltrates.     

Validated assay for the simultaneous quantification of total vincristine and actinomycin‐D concentrations in human EDTA plasma and of vincristine concentrations in human plasma ultrafiltrate by high‐performance liquid chromatography coupled with tandem mass spectrometry

A sensitive, specific and efficient high‐performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) assay for the simultaneous determination of total vincristine and actinomycin‐D concentrations in human plasma and an assay for the determination of unbound vincristine are presented. Electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI) and heated electrospray ionization (H‐ESI) were tested as ionization interfaces. For reasons of robustness ESI was chosen followed by tandem mass spectrometry (ESI‐MS/MS). For the plasma assay a 30 µL aliquot was protein precipitated with acetonitrile/methanol (50:50, v/v) containing the internal standard vinorelbine and 10 µL volumes were injected onto the HPLC system. To determine unbound vincristine, ultrafiltrate was produced from plasma using 30 kDa centrifugal filter units. The plasma ultrafiltrate was mixed with methanol (50:50, v/v), internal standard vinorelbine was added and 20 µL aliquots were injected onto the HPLC system. Separation was achieved on a 50 × 2.1 mm i.d. Xbridge C₁₈ column using 1 mM ammonium acetate/acetonitrile (30:70, v/v) adjusted to pH 10.5 with ammonia, run in a gradient with methanol at a flow rate of 0.4 mL/min. HPLC run time was 6 min. The assay quantifies in plasma vincristine from 0.25 to 100 ng/mL and actinomycin‐D from 0.5 to 250 ng/mL using plasma sample volumes of only 30 µL. Vincristine in plasma ultrafiltrate can be quantified from 1 to 100 ng/mL. Validation results demonstrate that vincristine and actinomycin‐D can be accurately and precisely quantified in human plasma and plasma ultrafiltrate with the presented methods. The assays are now in use to support clinical pharmacological studies in children treated with vincristine and actinomycin‐D. Copyright © 2009 John Wiley & Sons, Ltd.     

A simple method for methylmercury,inorganic mercury and ethylmercury determination in plasma samples by high performance liquid chromatography–cold-vapor-inductively coupled plasma mass spectrometry

A simple and sensitive method with a fast sample preparation procedure is proposed for the determination of mercury species in plasma/serum. The method combines online high-performance liquid chromatography separation, Hg cold-vapor formation and inductively coupled plasma mass spectrometry detection. Prior to analysis, plasma (250 μL) was accurately pipetted into 15 mL conical tubes. Then, an extractant solution containing mercaptoethanol, L-cysteine and HCl was added to the samples following sonication for 10 min. Quantitative mercury extraction was achieved with the proposed procedure. Separation of mercury species was accomplished in less than 8 min on a C8 reverse phase column with a mobile phase containing 3% v/v methanol + 97% v/v (0.5% v/v 2-mercaptoethanol + 0.05% v/v formic acid). The method detection limits were found to be 12 ng L⁻¹, 5 ng L⁻¹ and 4 ng L⁻¹ for inorganic mercury, ethylmercury and methylmercury, respectively. Method accuracy is traceable to Standard Reference Material (SRM) 966 Toxic Metals in Bovine Blood from NIST. Additional validation was provided by the analysis of a secondary reference serum sample from the INSQ-Canada. Finally, the method was successfully applied for the speciation of mercury in plasma samples collected from volunteers exposed to methylmercury through fish consumption. For the first time to our knowledge, levels of different species of Hg in plasma samples from riverside populations exposed to MeHg were determined.