Use of pneumatic nebulization and laser ablation–inductively coupled plasma–mass spectrometry to study the distribution and bioavailability of an intraperitoneally administered Pt-containing chemotherapeutic drug

Quadrupole-based inductively coupled–mass spectrometry (ICP-MS) with pneumatic nebulization as a means of sample introduction was employed for quantification of platinum in blood and tissue samples of rats with peritoneal carcinomatosis, receiving intraperitoneal treatment with the Pt-containing chemotherapeutic drug oxaliplatin, and in the perfusate solution used for this purpose. The Pt levels were measured for various treatment conditions, i.e., with and without supporting treatment with the drug bevacizumab and at two different temperatures. Limits of detection obtained for platinum in blood and tissue samples were 0.3 and 2.0 pg g_,⁻¹ respectively. Evaluation of drug penetration into the tumor, under different conditions of treatment, was carried out via laser ablation–ICP-MS. Quantitative mapping of the Pt distribution in tissue sections of rat was attempted relying on gelatin standards. The results show an influence of the temperature at which the treatment is carried out, while supporting administration of the drug bevacizumab did not seem to affect the results.     

Identification of a metallothionein in Synechococcus by capillary electrophoresis hyphenated with inductively coupled plasma mass spectrometry.

A home-made system hyphenating capillary electrophoresis with an inductively coupled plasma mass spectrometer (CE-ICP-MS) for cadmium speciation of protein-binding and free cadmium ions in solution is presented. The CE-ICP-MS interface consisted of an acrylic block with an internal volume ca. 20 microL in which a platinum electrode, a capillary column, and a connection to an ICP nebulizer were inserted. A make-up electrolyte solution containing 50 mmol L(-1) Tris-HCl buffer solution (pH 9.0) was continuously flowed through the interface to the ICP nebulizer. The separation of free Cd ions, Cd-cysteine, and Cd bounded to metallothionein (MT) isoforms from rabbit liver was carried out by capillary electrophoresis, and the analytes were detected by ICP-MS. The feasibility to isolate metallothionein compounds extracted from the cyanobacterium Synechococcus PCC7942 was demonstrated. The Cd binding proteins were induced in Synechococcus PCC7942 and further analyzed by CE ICP-MS.     

Metallomics in drug development: characterization of a liposomal cisplatin drug formulation in human plasma by CE–ICP–MS

A capillary electrophoresis inductively coupled plasma mass spectrometry method for separation of free cisplatin from liposome-encapsulated cisplatin and protein-bound cisplatin was developed. A liposomal formulation of cisplatin based on PEGylated liposomes was used as model drug formulation. The effect of human plasma matrix on the analysis of liposome-encapsulated cisplatin and intact cisplatin was studied. The presence of 1 % of dextran and 4 mM of sodium dodecyl sulfate in HEPES buffer was demonstrated to be effective in improving the separation of liposomes and cisplatin bound to proteins in plasma. A detection limit of 41 ng/mL of platinum and a precision of 2.1 % (for 10 μg/mL of cisplatin standard) were obtained. Simultaneous measurements of phosphorous and platinum allows the simultaneous monitoring of the liposomes, liposome-encapsulated cisplatin, free cisplatin and cisplatin bound to plasma constituents in plasma samples. It was demonstrated that this approach is suitable for studies of the stability of liposome formulations as leakage of active drug from the liposomes and subsequent binding to biomolecules in plasma can be monitored. This methodology has not been reported before and will improve characterization of liposomal drugs during drug development and in studies on kinetics.

Quantification of phosphorus in DNA using capillary electrophoresis hyphenated with inductively coupled plasma mass spectrometry

We have analyzed phosphorus in an enzymatically digested DNA molecule using capillary electrophoresis (CE) hyphenated with inductively coupled plasma mass spectrometry (ICP-MS). The DNA concentration was quantified by the phosphorus value obtained in the CE-ICP-MS analysis. The CE-ICP-MS measurement, for which the interface device AIF-01 equipped three layered nebulizer was adopted, was achieved with limited μL/min nebulizing without loss of sample in the vaporizing chamber. The samples of nucleotides and free phosphate were separated well in the CE-ICP-MS measurement, and the calibration curve (0.1-10μg/mL) of the phosphorus showed a linear (R(2)=0.999) increase in intensity. After digestion of the 100-bp double-strand DNA sample to deoxyribonucleotide-5'-monophosphates (dNMPs) by phosphodiesterase-I, phosphorus was detected by CE-ICP-MS without further purification steps. In this study, we applied two calculation schemes of DNA analysis using a dNMP concentration obtained from CE-ICP-MS. Comparative CE-ICP-MS analysis with DNA digested to dNMPs showed that the assay gave an equal value obtained from the total DNA quantification using fluorescence detection. The detection limits of the DNA sample obtained from these species and phosphorus in nucleotides using CE-ICP-MS were 3.1-26ng/mL. These LOD values were equal to the conventional fluorescence determination of DNA.     

Design and development of a low cost,high performance UV digester prototype: Application to the determination of trace elements by stripping voltammetry

A low cost, high performance ultraviolet digester was developed and tested in our laboratory. Its main features include limited costs (around 600 € for all the parts) and high efficiency toward total organic carbon (TOC) abatement. As a result the TOC of typical natural waters is reduced to less than 10% of its initial value with half an hour treatment. Easiness of use, temperature control and flexibility in sample size (up to 25 mL) and number (up to twelve 12 mL test tube) are other important characteristics of the developed UV digester.This apparatus was tested as a pretreatment method for the determination of the total content of trace elements such as copper, uranium and platinum in natural waters by cathodic adsorptive stripping voltammetry. The voltammetric technique was chosen because it is strongly interfered by the presence of even low levels of natural organic complexing molecules. As a result, a 30 minute UV treatment yielded results non distinguishable from ICP-MS data.     

Elimination of chloride interference on arsenic speciation in ion chromatography inductively coupled mass spectrometry using an octopole collision/reaction system

Anion-exchange chromatography with inductively coupled plasma mass spectrometry (ICP-MS) is often used for the speciation of arsenic (As). In this work, either He or H₂ was introduced to the octopole collision/reaction cell to eliminate chloride (Cl⁻) interferences during As speciation by ICP-MS. Polyatomic species, ⁴⁰Ar³⁵Cl and ³⁸Ar³⁷Cl, which are formed in high chloride matrices interfere with the ICP-MS detection of ⁷⁵As. These interferences were reduced or eliminated by introducing He or H₂ to the collision/reaction cell, with some loss in sensitivity when compared to the standard mode (no gas). For example, the sensitivity of As(V) was 30.4 and 17.7% of that observed in standard mode when introducing He and H₂, respectively. Chloride interference was completely eliminated using a flow rate of 3.0 mL min^− 1 with H₂ as a reaction gas with detection limits in the range of 0.3-0.6 μg L^− 1. The developed method was applied to determination of arsenic species in waters containing high concentrations of chloride by following a simple procedure and without modification of the ICP-MS instrument.     

Pyrohydrolysis of carbon nanotubes for Br and I determination by ICP-MS

Bromine and iodine determination was performed in carbon nanotubes (CNTs) by inductively coupled plasma mass spectrometry (ICP-MS) after sample preparation using pyrohydrolysis. Samples of CNTs (up to 500 mg) were mixed with 750 mg of V₂O₅ and heated at 950 °C during 12.5 min in a quartz tube under water vapor and air. The main operational conditions of pyrohydrolysis (carrier gas, absorbing solution, heating time, sample mass and use of V₂O₅) were evaluated. Accuracy was evaluated using certified reference materials (CRM) with similar matrix and also by comparison of results obtained after digestion of samples by microwave-induced combustion (MIC) and determination by ICP-MS. Agreement with CRM values was higher than 97% for Br and better than 96% in comparison with reference values (MIC/ICP-MS) of Br and I in CNTs samples. The limit of detection of the method for Br and I determination by ICP-MS was 0.05 and 0.004 μg g^? 1, respectively. Using a relatively simple and low cost pyrohydrolysis apparatus up to four samples can be processed per hour. The pyrohydrolysis sample preparation procedure is easy to be performed and provide a clean solution for analysis by ICP-MS, which is very attractive for Br and I control in CNTs.     

Microwave-assisted solid phase extraction prior to ICP-MS determination of Pd and Pt in environmental and biological samples

A sensitive and selective microwave-assisted solid phase extraction procedure coupled to inductively coupled plasma-mass spectrometry (ICP-MS) is proposed for palladium (Pd) and platinum (Pt) quantification in environmental and biological samples. Pd and Pt were quantitatively retained on commercial thioureido propyl functionalised silica gel packed inside a home-made glass microcolumn, and later eluted with 0.5% thiourea solution under microwave irradiation, followed by ICP-MS determination. The main variables affecting the procedural stages (i.e., sorption and desorption) and ICP-MS determination were optimised. The best conditions found were: (a) sorption: sample acidity, 1?M HCl; sample flow rate, 3?mL?min^?1; (b) desorption: microwave radiation, power 800?W; eluent concentration, 0.5% thiourea; eluent flow rate, 0.5?mL?min^?1; (c) ICP-MS determination: nebuliser feeding, free aspiration (0.3?mL?min^?1); internal standard, Rh (5?µg?L^?1). Analyte recoveries were higher than 90% and concentration factors up to 90 and 92 were achieved for Pd and Pt, respectively. Depending on the conditions, the methodological limits of detection were down to 0.2?ng?L^?1 for both analytes and repeatability, expressed as RSD%, varied between 1.3 and 11.0%. A method selectivity evaluation showed that most of the ICP-MS interferents were either quantitatively separated or more than 86% eliminated, except for Cu (elimination efficiency around 30%). Finally, the method was successfully used to determine Pd in certified reference materials (i.e. human urine and serum) and Pd and Pt in PM₁₀ airborne particulate matter fractions.     

On-line precipitation–dissolution in knotted reactor for thermospray flame furnace AAS for determination of ultratrace cadmium

A simple, environmentally friendly, cost-effective and sensitive method was developed for the determination of trace cadmium in rice and water by using flow injection (FI) on-line precipitation–dissolution in a knotted reactor (KR) as a preconcentration scheme for thermospray flame furnace atomic absorption spectrometry (TS-FF-AAS). The preconcentration was achieved by online merging the sample solution and the precipitating reagent in a KR and subsequently eluting the resultant precipitate of cadmium hydroxide with 1 mol/L HNO₃. The eluant was then introduced into TS-FF-AAS for the determination. A self-assembled FI system was employed to hyphenate the KR system with TS-FF-AAS. Under optimal chemical and instrumental conditions, a limit of detection of 0.04 μg/L and a sensitivity enrichment factor of 34 for cadmium was obtained with a total initial sample volume of 4 mL. The proposed method was applied to the determination of cadmium in certified reference rice and water samples with analytical results in good agreement with their certified values. Real rice samples and real water samples were also determined by the proposed method, with analytical results confirmed by inductively coupled plasma mass spectroscopy (ICP-MS).     

Microcalorimetric studies of the effects of artesunate liposomes on the metabolism of <Emphasis Type="Italic">Escherichia coli</Emphasis> during growth

A thermal dynamic model of nanoformulations entrapped in artesunate liposomes was established and biological thermodynamics was applied for investigation of the drug formulations. Effects of artesunate liposomes on the growth metabolism of Escherichia coli were studied by microcalorimetry. The results showed that (1) Comparison of artesunate and artesunate liposomes, the thermogenesis curves of E. coli were significant different in the metabolic process: lag phase (AB), log phase (BC), stationary phase (CD), and decline phase (DE); (2) Linear fit of the data of total metabolic heat of E. coli effected by different concentration artesunate (1–300 μg), the equation can be obtained as follows: Y = 364720.61−1075.25x, R = 0.9985; Linear fit of the data of total metabolic heat of E. coli effected by different concentration artesunate liposomes (30–120 μg), the linear equation can be obtained as follows: Y = 54251.5765−35.71122x, R = 0.98345; (3) The half inhibitory concentration I _C50 was 50.05 μg/mL, the relative sensitivity was obviously different; (4) Artesunate liposomes having better sustained release properties as compare to artesunate.     

Glutathione modified CdTe quantum dots as a label for studying DNA interactions with platinum based cytostatics

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, λ_em = 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 (R² = 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 (R² = 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.     

Rapid solid‐phase extraction coupled with GC–MS method for the determination of venlafaxine in rat plasma: Application to the drug–drug pharmacokinetic interaction study of venlafaxine combined with fluoxetine

A rapid and sensitive gas chromatography with mass spectrometry method for the determination of venlafaxine in rat plasma has been developed and applied to a drug–drug interaction study of fluoxetine on pharmacokinetics of venlafaxine in rats. Rat plasma was spiked with 2% aqueous ammonia before subjected to preactivated C₁₈ solid‐phase extraction columns and eluted with methanol. No endogenous interferences were observed under optimal condition. The calibration curve was linear (R ² = 0.9994) in the range of 10–1000 ng/mL. The quantification limit of venlafaxine in rat plasma was 10 ng/mL. The accuracy was in the range of 85–110%, and the extraction recovery was no less than 50%. Both the intra‐ and interday precision were 5.0–10.7%. The concentration–time curve showed that plasma concentrations of the coadministration group (group B) were higher than that of single dose group (group A). Both values of C _max (0.069 mg/L) and AUC_0→∞ (0.291 mg h/L) in group B were statistically greater than that of C _max (0.046 mg/L) and AUC_0→∞ (0.181 mg·h/L) in group A (P  < 0.05). The results indicated that a significant effect of fluoxetine was shown on the pharmacokinetics of venlafaxine, suggesting that drug–drug interactions are of concern for the treatment of depression with the combined use of venlafaxine and fluoxetine.     

Estimation of platinum in environmental water samples with solid phase extraction technique using inductively coupled plasma mass spectrometry

A solid phase extraction technique for the determination of platinum(IV) at trace levels by inductively coupled plasma mass spectromA solid phase extraction technique for the determination of platinum(IV) at trace levels by inductively coupled plasma mass spectrometry (ICP-MS) was developed. The method was based on retention of platinum in a sample on silica gel modified with aminepropyl groups. The retention of platinum(IV) from the sample solution and the recovery of platinum with 1.0 mol L⁻¹ thiourea solution were quantitative. The relative standard deviation (RSD) was calculated as 5% (n = 7) at the 10 ng L⁻¹ level. The enrichment factor was found to be (50-fold) for 250 mL of water sample. Under optimum conditions, the method detection limit (MDL) was found to be 1 ng L⁻¹ for platinum in water matrices. Recoveries of Pt from spike addition to atmospheric water samples were quantitative (80–95%). The present method was used for the determination of platinum in precipitation, throughfall and runoff water samples.      

Comparison of no gas and He/H2 cell modes used for reduction of isobaric interferences in selenium speciation by ion chromatography with inductively coupled plasma mass spectrometry

Inductively coupled plasma mass spectrometry (ICP-MS) used for the detection of most common selenium isotopes ⁷⁸Se (23.8%, abundance) and ⁸⁰Se (49.6%, abundance) is interfered in the presence of ³⁸Ar⁴⁰Ar⁺ and ⁴⁰Ar⁴⁰Ar⁺ in argon (Ar) gas. To address this issue, ICP-MS with an octopole reaction system (ORS) was explored for the detection of selenite and selenate, which was separated by anion-exchange chromatography. Results indicate that it was possible to detect ⁷⁸Se using no collusion gas, while to detect ⁸⁰Se a H₂ as the collusion/reaction gas was recommended since the background and noise were significantly reduced using H₂ as the gas. The selenium speciation interested was separated on a new column (G3154/101A, Agilent technologies) within 5 min using a mobile phase containing 10 mM NH₄H₂PO₄ and 20 mM NH₄NO₃ at pH 6.5. Linear plots were obtained in a concentration range of 1–200 μg/L with detection limits less than of 0.4 μg/L for ⁸⁰Se (IV) and 0.6 μg/L for ⁸⁰Se (VI) using H₂ as the reaction gas. Finally, the proposed method was used in the determination of selenium in water and soil.     

Validation of a high‐performance liquid chromatography method for the determination of (−)‐α‐bisabolol from particulate systems

A reversed‐phase high performance liquid chromatography method has been developed and validated for determination and quantitation of the natural sesquiterpene (−)‐α‐bisabolol. Furthermore the application of the method was done by characterization of chitosan milispheres and liposomes entrapping Zanthoxylum tingoassuiba essential oil, which contains appreciable amount of (−)‐α‐bisabolol. A reversed‐phase C₁₈ column and gradient elution was used with the mobile phase composed of (A) acetonitrile–water–phosphoric acid (19:80:1) and (B) acetonitrile. The eluent was pumped at a flow rate of 0.8 mL/min with UV detection at 200 nm. In the range 0.02–0.64 mg/mL the assay showed good linearity (R² = 0.9999) and specificity for successful identification and quantitation of (−)‐α‐bisabolol in the essential oil without interfering peaks. The method also showed good reproducibility, demonstrating inter‐day and intra‐day precision based on relative standard deviation values (up to 3.03%), accuracy (mean recovery of 100.69% ± 1.05%) and low values of detection and quantitation limits (0.0005 and 0.0016 mg/mL, respectively). The method was also robust for showing a recovery of 98.81% under a change of solvent in standard solutions. The suitability of the method was demonstrated by the successful determination of association efficiency of the (−)‐α‐bisabolol in chitosan milispheres and liposomes. Copyright © 2009 John Wiley & Sons, Ltd.     

Stealth Liposomes (PEGylated) Containing an Anticancer Drug Camptothecin: In Vitro Characterization and In Vivo Pharmacokinetic and Tissue Distribution Study

Numerous attempts to overcome the poor water solubility of cam ptothecin (CPT) by various nano drug delivery systems are described in various sources in the literature. However, the results of these approaches may be hampered by the incomplete separation of free CPT from the formulations, and this issue has not been investigated in detail. This study aimed to promote the solubility and continuous delivery of CPT by developing long-lasting liposomes using various weights (M.W. 2000 and 5000 Daltons) of the hydrophilic polymer polyethylene glycol (PEG). Conventional and PEGylated liposomes containing CPT were formulated via the lipid film hydration method (solvent evaporation) using a rotary flash evaporator after optimising various formulation parameters. The following physicochemical characteristics were investigated: surface morphology, particle size, encapsulation efficiency, in vitro release, and formulation stability. Different molecular weights of PEG were used to improve the encapsulation efficiency and particle size. The stealth liposomes prepared with PEG₅₀₀₀ were discrete in shape and with a higher encapsulation efficiency (83 ± 0.4%) and a prolonged rate of drug release (32.2% in 9 h) compared with conventional liposomes (64.8 ± 0.8% and 52.4%, respectively) and stealth liposomes containing PEG₂₀₀₀ (79.00 ± 0.4% and 45.3%, respectively). Furthermore, the stealth liposomes prepared with PEG₅₀₀₀ were highly stable at refrigeration temperature. Significant changes were observed using various pharmacokinetic parameters (mean residence time (MRT), half-life, elimination rate, volume of distribution, clearance, and area under the curve) of stealth liposomes containing PEG₂₀₀₀ and PEG₅₀₀₀ compared with conventional liposomes. The stealth liposomes prepared with PEG₅₀₀₀ showed promising results with a slow rate of release over a long period compared with conventional liposomes and liposomes prepared with PEG₂₀₀₀, with altered tissue distribution and pharmacokinetic parameters.     

Modulation of the physicochemical state of interior agents to prepare controlled release liposomes

To prepare controlled release liposomes, freeze-drying of double emulsions (FDE) was employed to entrap a model drug, topotecan, which has been reported to tend to leak rapidly from vesicles. In addition, hydrogenated soy phosphatidylcholine (HSPC), N-(carbonyl-methoxypolyethyleneglycol2000)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine (PEG-PE), and cholesterol (3:1:1, mass ratio) were used as the vesicle lipid. Different inner aqueous phases (W1) containing topotecan together with a variety of chemicals, such as citrate, sulfate, and divalent copper ions, were used to prepare W1/O/W2 double emulsions, which were then freeze-dried to obtain dry products. Upon rehydration, the dry products formed topotecan-entrapping unilamelar liposomes with an encapsulation efficiency of 80% and a mean diameter of less than 200 nm. The in vitro release experiments demonstrated that the drug release of topotecan-entrapping FDE liposomes could be successfully controlled through altering the state of the incorporated drug by means of protonation, precipitation, or forming a transition metal-complex. Specifically, the formulation of 300 mM CuSO₄ had a drug release half-life of 36 h. This novel method is a promising way to prepare controlled release liposomes that are more suitable for therapeutic application. In addition, the liposome formation mechanism was discussed based on micrographs and the size of the double emulsions and vesicles, as well as the small angle X-ray scattering pattern and transmission electron microscopy images.     

Speciation analysis of platinum antitumoral drugs in impacted tissues

Chemical compounds containing platinum have been employed since 1978 as drugs to beat certain type of tumours. Nevertheless, besides of their exceptional antitumoral properties, these drugs also have important deleterious side effects, such as, nephrotoxicity and ototoxicity.A study of Pt accumulation and a speciation analysis has been performed by ICP-MS in samples from kidney and inner ear in a controlled population of Wistar rats treated with, either, cisplatin, carboplatin or oxaliplatin. The results on Pt accumulation point out to drug structure and not only to Pt content as the responsible for the alteration of organ functionality.Speciation studies in the samples from kidney and inner ear were performed coupling two-dimensional liquid chromatography (2D-LC) to ICP-MS. Size exclusion (SEC) and anion exchange fast protein liquid chromatography (FPLC) was employed for 2D orthogonal separation. After these separations, free drug peaks were not observed in any of the samples.The binding of Pt to biomolecules was demonstrated by SEC and, independently of the drug used, Pt eluted as two main bands with molecular weights of 12 kDa and 25-65 kDa for inner ear samples, and as two different bands with 20 kDa and 50-60 kDa in the samples from kidney. However, the relative band intensity presented important differences for the three drugs. Using the same chromatographic conditions, it was shown that a metallothionein (MT) standard eluted in the same position as some of the cytosolic Pt-biomolecules.High Pt-containing fractions eluting from the SEC column were analysed by anion exchange FPLC after a preconcentration step. Among the different preconcentration methods tested, sample focusing on the head of the FPLC column shows main advantages. In this way, the separation by 2D chromatography of the high molecular Pt-species has been considerably improved.     

Inductively coupled plasma mass spectrometric analysis of the total amount of platinum in DNA extracts from peripheral blood mononuclear cells and tissue from patients treated with cisplatin

We present a highly sensitive method for the determination of platinum (Pt) in DNA extracts of peripheral blood mononuclear cells (PBMCs) and tissue samples from patients treated with cisplatin. The method is based on the measurement of Pt by inductively coupled plasma mass spectrometry (ICP-MS) and allows quantification of Pt-DNA adducts in PBMCs isolated from 10 mL blood and 1 mg tissue. The lower limit of quantification is 0.75 pg Pt or 7.5 fg Pt μg⁻¹ DNA when using 100 μg DNA. The method proved to be accurate and precise. The results obtained using the ICP-MS method were in good agreement with results from the alternative ³²P-postlabelling assay. The ICP-MS method was, however, more sensitive and proved to be less laborious. The advantages of the presented ICP-MS technique were demonstrated by the analysis of PBMCs, normal gastric tissue, and gastric tumour tissue of patients treated with cisplatin.