Non-aqueous capillary zone electrophoresis method for the analysis of paroxetine, tamoxifen, and their main metabolites in urine

The potential of non-aqueous capillary electrophoresis (NACE) was investigated for the simultaneous separation of paroxetine, tamoxifen, and their main metabolites. Baseline separation of the studied solutes was obtained on a  μm capillary using a non-aqueous buffer composed of 18 mM ammonium acetate and 1.1% acetic acid in 80:20 (v/v) methanol-acetonitrile, with a temperature and voltage of 22 °C and 15 kV, respectively. Clomipramine was used as internal standard. Aspects such as stability of the solutions, linearity, accuracy, precision and ruggedness were examined in order to validate the proposed method. Detection limits obtained for all the studied compounds ranged between 3.0 and 7.1 μg l⁻¹. The developed method is sensitive and robust and was used to determine paroxetine, tamoxifen, and their metabolites at clinically relevant levels in human urine. Before NACE determination, the samples were purified and enriched by means of an extraction-pre-concentration step with a pre-conditioned C₁₈ cartridge. Determination of these analytes in the urine of four females urines was demonstrated.     

A Biokinetic Model to Describe Consequences of Inhibition/Stimulation in <Emphasis Type="Italic">DNA</Emphasis>-Proofreading and -Repair,Part 4

Summary. In previous papers a strongly simplified physical-mathematical (biokinetic) model has been presented, which dealt with the factors influencing the timely development of DNA mismatches dependent cells (malignant cells) in their kinetic competition to the development of normal somatic cells (i.e. cells with correct genetic information). The kinetic results have been studied by comparing them with experimental results reported in the literature upon inhibiting the organisms own enzymatic DNA-proofreading and repair machinery. In spite of the fact that the model uses fully the chances of kinetics, which allows to describe even rather complicated systems with many regulation circuits and feed back loops in a rather simple, summarizing way, it has been demonstrated that the model does not only well describe the experimentally found significant increases of mutants in cases when the DNA repair system has been inhibited, but it can also reflect cancer-development and the efficacy of classical cancer therapies like surgery or chemotherapy as well.In applying the predictions of the model as to the opposite of an inhibition of the DNA repair system, i.e., in testing the results, if the organisms own repair systems were stimulated, the model shows that there could be a chance for a new, adjuvant cancer-therapy if this concept was combined with biochemical facts and clinical findings which are reported in the literature.In continuation of this concept, the predictions of the model have been compared with findings upon cancer-therapies by apoptosis-triggerers like tamoxifen. Further, according to the fact that there exists literature by which it is demonstrated by clinical facts that it is not necessary to use living cells (e.g. from umbilical cords blood or bone-marrow) to achieve surprising therapeutic successes in cancer therapy, but also cell-free human-placenta-extracts (HPEs) can be similarily effective, it has been tried in a first preliminary analytical effort to characterize effector-substances contained therein.Received January 27, 2003; accepted (revised) March 25, 2003 Published online August 18, 2003     

Organometallic SERMs (selective estrogen receptor modulators): Cobaltifens, the (cyclobutadiene)cobalt analogues of hydroxytamoxifen

The McMurry coupling of (tetraphenylcyclobutadiene)cobalt(cyclopentadienyl) ketones, (C₄Ph₄)Co[C₅H₄C(O)R], where R = Me, 3a, or Et, 3b, with a range of substituted benzophenones furnished a series of cobaltifens, organometallic analogues of tamoxifen whereby a phenyl ring has been replaced by an organo-cobalt sandwich moiety. These systems of the general formula (η⁴-C₄Ph₄)Co[η⁵-C₅H₄C(R)C(Ar)Ar′], where R = Me or Et, and Ar = Ar′ = p-C₆H₄X where X is OH, 2a and 2b, OMe, 2c and 2d, OBn, 2e and 2f, or O(CH₂)₂NMe₂, 12a and 12b, and where Ar = C₆H₄OH and Ar′ = C₆H₄O(CH₂)₂NMe₂, 2g and 2h, have been characterised by NMR spectroscopy and/or X-ray crystallography. The effect of 2a and 2b, 2g and 2h, and 12a and 12b on the growth of MCF-7 (hormone-dependent) and MDA-MB-231 (hormone-independent breast cancer cells) was studied. The dihydroxycobaltifens 2a and 2b exhibit a strong estrogenic effect on MCF-7 cells while the aminoalkyl-hydroxycobaltifens, 2g and 2h, were found to be only slightly cytotoxic on MDA-MB-231 cells (IC₅₀ = 27.5 and 17 μM); surprisingly, however, the bis-(dimethylaminoethoxy)cobaltifens, 12a and 12b were shown to be highly cytotoxic towards both cell lines (IC₅₀ = 3.8 and 2.5 μM).     

Nonaqueous capillary electrophoresis method for the analysis of tamoxifen, imipramine and their main metabolites in urine

The viability of nonaqueous capillary electrophoresis (NACE) was investigated for the simultaneous determination of tamoxifen, imipramine and their main metabolites (4-hydroxytamoxifen and desipramine, respectively). Baseline separation of the studied solutes was obtained on a 57 cm × 75 μm capillary using a nonaqueous solution composed of 17 mM ammonium acetate and 1.25% acetic acid in 80:20 (v:v) methanol-acetonitrile, temperature and voltage 22 °C and 15 kV, respectively, and hydrodynamic injection. Paroxetine was used as internal standard. Different aspects including linearity, accuracy, ruggedness and precision was studied. Detection limits between 9.0 and 15.0 μg L⁻¹ were obtained for all the studied compounds. The developed method is simple, rapid and sensitive and has been used to determine tamoxifen, imipramine and their metabolites at clinically relevant levels in human urine. Before NACE determination, a solid phase extraction (SPE) procedure with a C₁₈ cartridge was necessary. Real determination of these analytes in three females urines were done.     

Micellar electrokinetic capillary chromatographic method for simultaneous determination of drugs used to treat advanced breast cancer

Summary A micellar electrokinetic capillary chromatography (MEKC) method has been developed for quantification of four drugs-tamoxifen, anastrozole, letrozole, and methotrexate—used to treat advanced breast cancer. Separation was performed at 25°C and 25kV, with 20mm borate buffer (pH 9.2) containing 40mm sodium dodecylsulfate as electrolyte solution. Under these conditions analyses were performed in 12 min. The linearity of the response was investigated for the concentration range 2.0–20.0 mg L⁻¹. The intra-day residual standard deviation (n=4 graphs) between the slopes of the calibration graphs was acceptable for the four drugs studied. Detection limits (signal-to-noise ratio=3) were below 1 mg L⁻¹ for all the compounds. The simplicity, precision, and sensitivity of MEKC proved suitable for quality control of pharmaceutical preparations used to treat advanced breast cancer. Six different pharmaceutical preparations, each containing one of the above-mentioned drugs, were successfully analyzed.     

Identification and quantification of tamoxifen and four metabolites in serum by liquid chromatography-tandem mass spectrometry

We have developed a method for the determination of tamoxifen (tam) and its metabolites 4-hydroxytamoxifen (4OHtam), N-demethyltamoxifen (NDtam), N-dedimethyltamoxifen (NDDtam), tamoxifen-N-oxide (tamNox), and 4-hydroxy-N-demethyltamoxifen (4OHNDtam) in 50 microl human serum. Serum proteins were precipitated with acetonitrile. Deuterated-tamoxifen (D5 tam) was added as internal standard. Sample supernatant was injected into an on-line reversed-phase extraction column coupled with a C18 analytical column and analytes were detected by tandem mass spectrometry. The lower limits of quantification were 0.25 ng/mL for 4OHtam, NDtam and tam, 1.0 ng/mL for NDDtam and tamNox. Ranges of within- and between-day variation were 2.9-15.4% and 4.4-12.9%, respectively.     

固相萃取-液相色谱串联质谱法测定城市污水中他莫昔芬和来曲唑

他莫昔芬和来曲唑常用于治疗雌激素依赖型乳腺癌。本研究建立了城市污水中他莫昔芬和来曲唑的超高效液相色谱-串联质谱(UPLC-MS/MS)联用检测方法。水样通过Oasis HLB固相萃取柱富集后串联氨基柱,先用6 mL甲醇洗脱,再用3 mL甲醇洗脱氨基柱,合并洗脱液后浓缩。使用ACQUITY UPLCTM BEH C18反相柱,流动相为0.1%甲酸-乙腈,在梯度条件下分离;目标分析物使用超高效液相色谱-电喷雾串联质谱进行测定。本方法对他莫昔芬和来曲唑的线性范围分别是1.0～100μg/L和0.1～100μg/L,相关系数R2>0.997,检出限分别为1.0和0.1 ng/L,进水和出水的3个不同水平的加标平均回收率为68.8%～103.0%,相对标准偏差小于15%。本方法可用于城市污水中相关物质的分析。     

Tamoxifen-loaded polymeric micelles: preparation, physico-chemical characterization and in vitro evaluation studies

Several samples of polymeric micelles, formed by amphiphilic derivatives of PHEA, obtained by grafting into polymeric backbone of PEGs and/or hexadecylamine groups (PHEA-PEG-C(16) and PHEA-C(16)) and containing different amount of Tamoxifen, were prepared. All Tamoxifen-loaded polymeric micelles showed to increase drug water solubility. TEM studies provided evidence of the formation of supramolecular core/shell architectures containing drug, in the nanoscopic range and with spherical shape. Samples with different amount of encapsulated Tamoxifen were subjected to in vitro cytotoxic studies in order to evaluate the effect of Tamoxifen micellization on cell growth inhibition. All samples of Tamoxifen-loaded polymeric micelles showed a significantly higher antiproliferative activity in comparison with free drug, probably attributable to fluidification of cellular membranes, caused by amphiphilic copolymers, that allows a higher penetration of the drug into tumoral cells. To gain preliminary information about the potential use of prepared micelles as Tamoxifen drug delivery systems, studies evaluating drug release ability of micelle systems in media mimicking biological fluids (buffer solutions at pH 7.4 and 5.5) and in human plasma were carried out. These studies, performed evaluating the amount of Tamoxifen that remains in solution as a function of time, showed that at pH 7.4, as well as in plasma, PHEA-C(16) polymeric micelles were able to release lower drug amounts than PHEA-PEG(5000)-C(16) ones, while at pH 5.5, the behavior difference between two kind of micelles was less pronounced.     

Trace determination of tamoxifen and 5-fluorouracil in hospital and urban wastewaters

Tamoxifen and 5-fluorouracil are widely used in cancer therapy. They are highly toxic (teratogenic, mutagenic, etc.), as are most of the anticancer drugs. Two methods were set up to analyse these drugs in wastewaters to evaluate the potential for environmental contamination by cytostatic agents. Liquid–liquid extraction followed by purification on OASIS® MCX cartridge and gas chromatography with mass spectrometry detection (GC-MS) was used for the analysis of tamoxifen. 5-Fluorouracil was extracted with an ENV+?(Isolute) cartridge (solid-phase extraction), derivatized with pentafluorobenzyl bromide (PFBBr) and detected by GC-MS. Both methods showed good recoveries (>70%), repeatability (RSD<10%) and limits of detection (LOD 6–15?ng/L). Wastewaters from a residential area, a hospital, and sewage-treatment plants (STPs) were analysed using the analytical methods developed in this study. Tamoxifen was detected in wastewaters of the hospital, residential area, and influent of STPs, but not in treated wastewaters. 5-Fluorouracil in all wastewaters was below the LOD of the analytical method.     

Systematic development and validation of RP-HPLC method for simultaneous estimation of tamoxifen and sulphoraphane with specific application for nanolipidic formulations

Tamoxifen (TAM) and Sulphoraphane (SFN) are well-known anti-estrogen drugs used for the treatment of breast cancer. Due to their synergistic therapeutic potential, their combination is preferred as it helps to minimize the drug-related toxicities and enhances therapeutic efficacy. A simple, robust and fast simultaneous reversed-phase high-performance liquid chromatography (RP-HPLC) method was developed as well validated for the analysis of both the drugs based on their particular wavelength. The separation was performed on C₁₈ analytical column with dimensions of 4.6 × 250 mm, 5 μm using mobile phase methanol: water (pH 3.5) in the ratio 70:30 and flow rate of 0.8 min/mL. Box-Behnken experimental design was used to optimized these independent variables and analyze their effect on the response variables like retention time (RT), no. of theoretical plates and tailing factor of both analytes. Method validation was carried out for establishing the specificity, linearity range, accuracy, sensitivity, robustness, precision and ruggedness. The method applicability was evaluated on different nanoformulations, i.e., solid lipid nanoparticles (SLNs), liposomes (LIPO), nanostructured lipid carriers (NLCs). The peaks of the analyte were found to be well resolved and two distinct RT were recorded for TAM and SFN. Calibration curves were found to be linear for TAM and SFN over concentration range of 6–24 μg/mL. All method validation criteria were within the range of acceptance. Relative standard deviation (%RSD) was observed to be <2% for inter- and intra-day precision. The application of developed method for estimation of drugs from the nanoformulations was suitabile for in vitro as well as in vivo studies.