Direct evidence for co‐binding of cisplatin and cadmium to a native zinc‐ and cadmium‐containing metallothionein

Cisplatin is widely used to treat a number of cancers, and its covalent binding to DNA is believed to cause cell death; however, the roles of cisplatin–protein interactions in the mechanisms of action, toxicity, and resistance of the drug largely remain to be elucidated. Here, we investigate the interactions of cisplatin and a native rabbit metallothionein (MT), containing 1.4% zinc and 7.9% cadmium, using nanospray tandem quadrupole time‐of‐flight mass spectrometry (MS) and size‐exclusion high‐performance liquid chromatography with inductively coupled plasma MS. At near‐neutral pH conditions, reactions between cisplatin and MT resulted in the formation of complexes that contained Cd₄–Pt_n–MT (n = 1–7). While zinc was displaced by cisplatin, both platinum and cadmium were bound to the same MT molecule. This is the first report to provide direct evidence for the co‐binding of cadmium and platinum to MT, which suggests that the mechanism of the binding of cisplatin to the native MT may not be through the displacement of cadmium as previously proposed. A tandem MS investigation into the binding sites of the platinum and cadmium to MT showed platinum‐ and cadmium‐related fragments, such as (PtS₂C₂H₇N)⁺ and (CdS₃C₅H₁₇N₂)⁺, demonstrating the platinum–cysteine and cadmium–cysteine binding. In addition, detection of Cd₄–Pt₇–MT demonstrated more than ten metals bound to a single MT molecule. This finding was extended to the binding of MT with a five‐fold excess of CdCl₂. As many as 14 metal atoms (13 cadmium and one zinc) were detected bound to a single MT molecule, the complexes being Cd_x–Zn–MT (x = 5–13). The high binding capacity of MT for cadmium and platinum is consistent with the role of MT in reduction of metal toxicity and its involvement in drug resistance. Copyright © 2003 John Wiley & Sons, Ltd.     

Therapeutic drug monitoring of mitotane: Analytical assay and patient follow‐up

Adrenocortical carcinoma (ACC) is an aggressive malignancy of the adrenal gland. Mitotane (o ,p' ‐DDD) is the most effective chemotherapy for ACC. According to the literature, mitotane plasma trough concentrations within 14–20 mg L⁻¹ are correlated with a higher response rate with acceptable toxicity. Therapeutic drug monitoring (TDM) of mitotane is therefore recommended. The aim of this study was to propose a robust and simple method for mitotane quantification in plasma. The validation procedures were based on international guidelines. Sample preparation consisted of a single protein precipitation with methanol using 100 μL of plasma. The supernatant was submitted to liquid chromatography coupled with ultra‐violet detection at 230 nm. Mitotane retention time was 7.1 min. The limit of detection was 0.1 mg L⁻¹ and the limit of quantification was 0.78 mg L⁻¹. The assay demonstrated a linear range of 0.78–25 mg L⁻¹ with correlation coefficients (r ²) at 0.999. Inter‐ and intra‐assay precision was <4.85%. Evaluation of accuracy showed a deviation <13.69% from target concentration at each quality control level. This method proved easy and rapid to perform mitotane TDM and required a small volume of sample. It was successfully applied to routine TDM in our laboratory.     

Voltammetric Studies of Cadmium‐ and Zinc‐Containing Metallothioneins at Nafion‐Coated Mercury Thin Film Electrodes

Voltammetric studies of rabbit liver metallothioneins (MTs, containing both Zn and Cd ions) and Zn₇‐MT were carried out at Nafion‐coated mercury film electrodes (NCMFEs). The accumulation of MT molecules into the NCMFEs enhances the voltammetric signals and the electrostatic interaction between the Nafion membrane and MT facilitates facile electron transfer reactions. Two well‐defined redox waves, with reduction potential (E_pc) values at ?0.740 and ?1.173 V, respectively, were observed. The peak at E_pc =?0.740 V is attributable to the reduction of the Cd‐MT complex, whereas that at E_pc=?1.173 V was assigned to the reduction of the Zn‐MT complex. Zn₇‐MT exhibits only one redox wave with E_pc=?1.198 V. The NCMFE was found to be more advantageous than thin mercury film electrode (MFE), because the pristine metal ions in MTs (e.g., Cd²⁺ and/or Zn²⁺) are not significantly replaced by Hg²⁺. The NCMFE is also complementary to Nafion‐coated bismuth film electrode in that it has a greater hydrogen overpotential, which allows the reduction of the Zn‐MT complex to be clearly observed. Moreover, intermetallic compound formation between Cd and Zn appears to be less serious at NCMFEs. Consequently, the amounts of Cd and Zn deposited into the electrode upon the reduction reactions can be quantified more accurately.     

Synthesis and self‐assembly of thermosensitive double‐hydrophilic poly(N‐vinylcaprolactam)‐b‐poly(N‐vinyl‐2‐pyrrolidone) diblock copolymers

We report on novel diblock copolymers of poly(N‐vinylcaprolactam) (PVCL) and poly(N‐vinyl‐2‐pyrrolidone) (PVPON) (PVCL‐b‐PVPON) with well‐defined block lengths synthesized by the MADIX/reversible addition‐fragmentation chain transfer (RAFT) process. We show that the lower critical solution temperatures (LCST) of the block copolymers are controllable over the length of PVCL and PVPON segments. All of the diblock copolymers dissolve molecularly in aqueous solutions when the temperature is below the LCST and form spherical micellar or vesicular morphologies when temperature is raised above the LCST. The size of the self‐assembled structures is controlled by the molar ratio of PVCL and PVPON segments. The synthesized homopolymers and diblock copolymers are demonstrated to be nontoxic at 0.1–1 mg mL^?1 concentrations when incubated with HeLa and HEK293 cancer cells for various incubation times and have potential as nanovehicles for drug delivery. © 2014 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2014 , 52, 2725–2737     

Study of the reaction methyl 4‐nitrobenzene‐sulfonate + Cl− in mixed hexadecyltrimethyl‐ammonium chloride‐triton X‐100 micellar solutions

The reaction methyl 4‐nitrobenzenesulfonate + Cl^? was studied in hexadecyltrimethylammonium chloride (CTAC) in the absence and presence of 0.1 M NaCl, as well as in mixed CTAC/Triton X‐100 (polyoxyethylene(9.5)octylphenyl ether) aqueous micellar solutions with CTAC molar fractions of 0.9, 0.8, 0.7, and 0.6. Conductivity measurements were used to obtain critical micellar concentrations and micellar ionization degrees of the various micellar reaction media. From these data, thermodynamic information on the cationic/nonionic mixed micellar solutions was obtained. Micellar effects on the observed rate constant were explained by pseudophase kinetic models. The estimated second‐order rate constants in the micellar pseudophase of the different micellar reaction media showed that pure CTAC and mixed CTAC/Triton X‐100 micelles, at the high cationic surfactant molar fractions studied, provide reaction sites of similar characteristics at the interfacial region. This was in agreement with previous structural studies carried out on mixed CTAC/Triton X‐100 micellar solutions. © 2002 Wiley Periodicals, Inc. Int J Chem Kinet 35: 45–51, 2003     

Bioaccumulation of metals and induction of metallo‐ thioneins in selected tissues of common carp (Cyprinus carpio L.) co‐exposed to cadmium,mercury and lead

The concentrations of mercury (Hg), cadmium (Cd) and lead (Pb) at various exposure periods were determined in the gill, kidney, liver and muscle of common carp (Cyprinus carpio L.) co‐exposed to 1.0 µg ml^?1 each of Cd²⁺, Hg²⁺ and Pb²⁺ for up to 10 days. Metallothionein fractions (MTs) in these organs were characterized using the hyphenated technique of size‐exclusion chromatography (SEC) and inductively coupled plasma mass spectrometry (ICP‐MS). After 10 days of exposure, maximum toxic metal concentrations of Hg, Cd and Pb were 10.7 (gill), 0.145 (kidney) and 0.112 µg g^?1_dryweight (gill), respectively. The pattern of accumulation of Hg and Pb was in the order gill > kidney > liver > muscle. In the case of Cd, accumulation was in the order kidney > gill > liver > muscle. Cd and Hg binding MTs were significantly induced in the gill, kidney and liver of all the exposure groups in comparison with the control group (p < 0.05), and the amounts of them increased with the longer exposure time. Despite the higher intracellular Hg concentration and the stronger Hg? SH binding affinity, the amount of Cd‐binding MTs was much higher than that of Hg‐binding MTs. The results indicate that MT synthesis in these organs was clearly metal‐specific. MTs in gill may be used as a bio‐marker to detect the metal pollution caused by Hg and Cd. Zinc and copper binding MTs in the organs of the exposed fish were also increased. This may be due to the MTs' important role in the homeostatic regulation of essential metals and their protective role against the acute toxicity of non‐essential metals. Even though there was considerable accumulation of lead in the organs of the exposed fish, Pb‐binding MT synthesis was non‐significant. Copyright © 2006 John Wiley & Sons, Ltd.     

Synthesis of linear and star‐like poly(ε‐caprolactone)‐b‐poly{γ‐2‐[2‐(2‐methoxy‐ethoxy)ethoxy]ethoxy‐ε‐caprolactone} amphiphilic block copolymers using zinc undecylenate

Linear and star‐like amphiphilic diblock copolymers were synthesized by the ring‐opening polymerization of ε‐caprolactone and γ‐2‐[2‐(2‐methoxyethoxy)ethoxy]ethoxy‐ε‐caprolactone monomers using zinc undecylenate as a catalyst. These polymers have potential applications as micellar drug delivery vehicles, therefore the properties of the linear and 4‐arm star‐like structures were examined in terms of their molecular weight, viscosity, thermodynamic stability, size, morphology, and drug loading capacity. Both the star‐like and linear block copolymers showed good thermodynamic stability and degradability. However, the star‐like polymers were shown to have increased stability at lower concentrations with a critical micelle concentration (CMC) of 5.62 × 10^?4 g L^?1, which is less than half the concentration of linear polymer needed to form micelles. The star‐like polymeric micelles showed smaller sizes when compared with their linear counterparts and a higher drug loading capacity of doxorubicin, making them better suited for drug delivery purposes. © 2016 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016 , 54, 3601–3608     

Cancer‐associated pH‐responsive tetracopolymeric micelles composed of poly(ethylene glycol)‐b‐poly(L‐histidine)‐b‐poly(L‐lactic acid)‐b‐poly(ethylene glycol)

To create a novel vector for specifically delivering anticancer therapy to solid tumors, we used diafiltration to synthesize pH‐sensitive polymeric micelles. The micelles, formed from a tetrablock copolymer [poly(ethylene glycol)‐b‐poly(L ‐histidine)‐b‐poly(L ‐lactic acid)‐b‐poly(ethylene glycol)] consisted of a hydrophobic poly(L ‐histidine) (polyHis) and poly(L ‐lactic acid) (PLA) core and a hydrophilic poly(ethylene glycol) (PEG) shell, in which we encapsulated the model anticancer drug doxorubicin (DOX). The robust micelles exhibited a critical micellar concentration (CMC) of 2.1–3.5 µg/ml and an average size of 65–80 nm pH 7.4. Importantly, they showed a pH‐dependent micellar destabilization, due to the concurrent ionization of the polyHis and the rigidity of the PLA in the micellar core. In particular, the molecular weight of PLA block affected the ionization of the micellar core. Depending on the molecular weight of the PLA block, the micelles triggering released DOX at pH 6.8 (i.e. cancer acidic pH) or pH 6.4 (i.e. endosomal pH), making this system a useful tool for specifically treating solid cancers or delivering cytoplasmic cargo in vivo. Copyright © 2008 John Wiley & Sons, Ltd.     

Pharmacokinetics of 2,3,5,4′‐tetrahydroxystilbene‐2‐O‐β‐D‐glucoside in rat using ultra‐performance LC‐quadrupole TOF‐MS

2,3,5,4′‐Tetrahydroxystilbene‐2‐O‐β‐D‐glucoside (THSG) from Polygoni multiflori has been demonstrated to possess a variety of pharmacological activities, including antioxidant, anti‐inflammatory and hepatoprotective activities. Ultra‐performance LC‐quadrupole TOF‐MS with MS Elevated Energy data collection technique and rapid resolution LC with diode array detection and ESI multistage MSⁿ methods were developed for the pharmacokinetics, tissue distribution, metabolism, and excretion studies of THSG in rats following a single intravenous or oral dose. The three metabolites were identified by rapid resolution LC‐MSⁿ. The concentrations of the THSG in rat plasma, bile, urine, feces, or tissue samples were determined by ultra‐performance LC‐MS. The results showed that THSG was rapidly distributed and eliminated from rat plasma. After the intravenous administration, THSG was mainly distributing in the liver, heart, and lung. For the rat, the major distribution tissues after oral administration were heart, kidney, liver, and lung. There was no long‐term storage of THSG in rat tissues. Total recoveries of THSG within 24 h were low (0.1% in bile, 0.007% in urine, and 0.063% in feces) and THSG was excreted mainly in the forms of metabolites, which may resulted from biotransformation in the liver.     

New pressure‐assisted sweeping on‐line preconcentration for highly polar environmentally relevant nitrosamines: Part 2. Cationic and anionic surfactants with zero‐flow capillaries

We recently introduced a pressure‐assisted sweeping‐reversed migration‐EKC (RM‐EKC) method for preconcentration of neutral polar N‐nitrosamines with low affinity for the micellar phase. The type of surfactant and phase ratio are dominant factors in dictating the magnitude of interactions between analyte and micellar phase, thus four surfactants (anionic and cationic) with a range of functionalities (SDS, ammonium perfluorooctanoate (APFO), bile salts, and cetyltrimethylammonium chloride (CTAC)) were evaluated for sweeping‐RM‐EKC of highly polar N‐nitrosamines. All gave acceptable results for sweeping‐RM‐EKC when used in high concentrations (≥200 mM) with low EOF. While no single surfactant was superior by all measures, all but the bile salts had useful performance characteristics. APFO showed the narrowest peak widths and highest number of theoretical plates, though two species co‐migrated at all concentrations (25–300 mM); SDS and the cationic surfactant CTAC also showed good separation characteristics and could resolve all peaks, but CTAC had wider separation window. Various types of capillaries coated for EOF control were compared for use with anionic and cationic surfactants. A commercial zero‐EOF capillary coated with a polymer bearing sulfonic acid functional groups showed superior EOF suppression and reproducibility of migration time with all surfactants.     

Synthesis,micellization and gelation of temperature‐responsive star‐shaped block copolymers

A series of amphiphilic temperature‐responsive star‐shaped poly(D,L‐lactic‐co‐glycolic acid)‐b‐methoxy poly(ethylene glycol) (PLGA‐mPEG) block copolymers with different arm numbers were synthesized via the arm‐first method. Gel permeation chromatography data confirmed that star‐shaped PLGA‐mPEG copolymers had narrow polydispersity index, indicating the successful formation of star‐shaped block copolymers. Indirectly, the ¹H NMR spectra in two kinds of solvents and dye solubilization method had confirmed the formation of core‐shell micelles. Further, core‐shell micelles with sizes of about 30–50 nm were directly observed by transmission electron microscopy. Subsequently, the micellar sizes and distributions as a function of concentrations and temperature were measured. At various copolymer concentrations, individual micelles with size of 20–40 nm and grouped micelles with size of 600–700 nm were found. Micellar mechanism of star‐shaped block copolymers in aqueous solution was simultaneously discussed. In addition, sol–gel transition of star‐shaped block copolymers in water was also investigated via the inverting test method. The critical gel temperature (CGT) and critical gel concentration (CGC) values of two‐arm, three‐arm and four‐arm copolymer solutions were markedly higher than ones of one‐arm copolymer. Moreover, the same CGC values of copolymer solution with different molecular weight and the same arm composition were ~15 wt %, and CGT values increased from ~38 to ~47°C with increasing arm numbers. Finally, the temperature‐dependent micellar packing gelation mechanism of star‐shaped block copolymer was schematically illustrated. Copyright © 2013 John Wiley & Sons, Ltd.     

Temperature‐induced spontaneous sol–gel transitions of poly(D,L‐lactic acid‐co‐glycolic acid)‐b‐poly(ethylene glycol)‐b‐poly(D,L‐lactic acid‐co‐glycolic acid) triblock copolymers and their end‐capped derivatives in water

The spontaneous hydrogel formation of a sort of biocompatible and biodegradable amphiphilic block copolymer in water was observed, and the underlying gelling mechanism was assumed. A series of ABA‐type triblock copolymers [poly(D,L ‐lactic acid‐co‐glycolic acid)‐b‐poly(ethylene glycol)‐b‐poly(D,L ‐lactic acid‐co‐glycolic acid)] and different derivatives end‐capped by small alkyl groups were synthesized, and the aqueous phase behaviors of these samples were studied. The virgin triblock copolymers and most of the derivatives exhibited a temperature‐dependent reversible sol–gel transition in water. Both the poly(D,L ‐lactic acid‐co‐glycolic acid) length and end group were found to significantly tune the gel windows in the phase diagrams, but with different behaviors. The critical micelle concentrations were much lower than the associated critical gel concentrations, and an intact micellar structure remained after gelation. A combination of various measurement techniques confirmed that the sol–gel transition with an increase in the temperature was induced not simply via the self‐assembly of amphiphilic polymer chains but also via the further hydrophobic aggregation of micelles resulting in a micelle network due to a large‐scale self‐assembly. The coarsening of the micelle network was further suggested to account for the transition from a transparent gel to an opaque gel. © 2006 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 45: 1122–1133, 2007     

Association of hydrophobically α,ω‐end‐capped poly(2‐methyl‐2‐oxazoline) in water

The α,ω‐end‐capped poly(2‐methyl‐2‐oxazoline) (C_n‐POXZ‐C_n) have been synthesized by a one‐pot process using cationic ring‐opening polymerization with an appropriate initiator and terminating agent. The polymers bearing different alkyl groups C₁₂ and C₁₈ have molecular weight in the range of 2.4 × 10³ to 14 × 10³ with a small polydispersity index. The solution behavior of the free chains has been analyzed in a nonselective solvent, dichloromethane, by small‐angle neutron scattering and dynamic light scattering. These amphiphilic polymers associate in water to form flower‐like micellar structures. Critical micelle concentrations, investigated by fluorescence technique, are in the range of 0.03–0.5 g L^?1 and are dependent on the hydrophilic/lipophilic balance. The structural properties of the aggregates have also been investigated by viscometry. Intrinsic viscosities of these polymers are in the same range as that of the precursors poly(2‐methyl‐2‐oxazoline) (POXZ) and mono‐functionalized polymers. Large viscosity increase corresponding to intermicellar bridging was observed in the vicinity of the micelle overlap concentration. Addition of hydroxypropyl β‐cyclodextrin (HβCD) has dissociated the aggregates and the intrinsic viscosities of the HβCD‐end‐capped chains have become comparable with the ones of POXZ precursor chains. © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 48: 2477–2485, 2010     

Synthesis and Anti‐tumor Evaluation of Novel C‐37 Modified Derivatives of Gambogic Acid

Gambogic acid (GA, 1 ), the most prominent representative of Garcinia natural products, has been reported to be a promising anti‐tumor agent. In order to further explore the structure‐activity relationship of GA and discover novel GA derivatives as anti‐tumor agents, 17 novel C‐37 modified derivatives of GA were synthesized and evaluated for their in vitro anti‐tumor activities against A549, HCT‐116, BGC‐823, HepG2 and MCF‐7 cancer cell lines. Among them, 11 compounds were found to be more potent than GA against some cancer cell lines. Notably, compound 8 was almost 5–10 folds more active than GA against A549 and BGC‐823 cell lines with the IC₅₀ values of 0.12 µmol·L^?1 and 0.57 µmol·L^?1, respectively. Chemical modification at C‐37 position of GA by introducing of hydrophilic amines could lead to increased activity and improved drug‐like properties. These findings will enhance our understanding of the structure‐activity relationship (SAR) of GA and lead to the discovery of novel GA derivatives as potential anti‐tumor agents.     

Formation of Core‐Shell‐Corona Micellar Complexes through Adsorption of Double Hydrophilic Diblock Copolymers into Core‐Shell Micelles

Summary: The complexation between polystyrene‐block‐poly(acrylic acid) (PS‐b‐PAA) micelles and poly(ethylene glycol)‐block‐poly(4‐vinyl pyridine) (PEG‐b‐P4VP) is studied, and a facile strategy is proposed to prepare core‐shell‐corona micellar complexes. Micellization of PS‐b‐PAA in ethanol forms spherical core‐shell micelles with PS block as core and PAA block as shell. When PEG‐b‐P4VP is added into the core‐shell micellar solution, the P4VP block is absorbed into the core‐shell micelles to form spherical core‐shell‐corona micellar complexes with the PS block as core, the combined PAA/P4VP blocks as shell and the PEG block as corona. A model is suggested to characterize the core‐shell‐corona micellar complexes.

Schematic formation of core‐shell‐corona (CSC) micellar complexes by adsorption of PEG‐b‐P4VP into core‐shell PS‐b‐PAA micelles.     

Effective Cancer Cell Killing by Hydrophobic Nanovoid‐Enhanced Cavitation under Safe Low‐Energy Ultrasound

β‐Cyclodextrin (β‐CD)‐capped mesoporous silica nanoparticles with hydrophobic internal nanovoids were prepared and used for effective cancer cell killing in synergistic combination with low‐energy ultrasound (≤1.0 W cm^?2, 1 MHz). The water‐dispersible nanoparticles with hydrophobic internal nanovoids can be taken up by cancer cells and subsequently evoke a remarkable cavitation effect under irradiation with mild low‐energy ultrasound (≤1.0 W cm^?2, 1 MHz). A significant cancer cell killing effect was observed in cancer cells and in a mouse xenograft tumor model treated with the nanoagents together with the low‐energy ultrasound, showing a distinct dependence on the concentration of nanoagents and ultrasound intensity. By contrast, an antitumor effect was not observed when either low‐energy ultrasound or nanoagents were applied alone. These findings are significant as the technique promises a safe, low‐cost, and effective treatment for cancer therapy.     

Validation of a liquid chromatography‐triple quadrupole mass spectrometric method for the determination of 5‐nitro‐5′‐hydroxy‐indirubin‐3′‐oxime (AGM‐130) in human plasma and its application to microdose clinical trial

A liquid chromatography–triple quadrupole mass spectrometric (LC‐MS/MS) method was developed and validated for the determination of 5‐nitro‐5′‐hydroxy‐indirubin‐3′‐oxime (AGM‐130) in human plasma to support a microdose clinical trial. The method consisted of a liquid–liquid extraction for sample preparation and LC‐MS/MS analysis in the positive ion mode using TurboIonSpray^TM for analysis. d₃‐AGM‐130 was used as the internal standard. A linear regression (weighted 1/concentration) was used to fit calibration curves over the concentration range of 10–2000 pg/mL for AGM‐130. There were no endogenous interference components in the blank human plasma tested. The accuracy at the lower limit of quantitation was 96.6% with a precision (coefficient of variation, CV) of 4.4%. For quality control samples at 30, 160 and 1600 pg/mL, the between run CV was ≤5.0 %. Between‐run accuracy ranged from 98.1 to 101.0%. AGM‐130 was stable in 50% acetonitrile for 168 h at 4°C and 6 h at room temperature. AGM‐130 was also stable in human plasma at room temperature for 6 h and through three freeze–thaw cycles. The variability of selected samples for the incurred sample reanalysis was ≤12.7% when compared with the original sample concentrations. This validated LC‐MS/MS method for determination of AGM‐130 was used to support a phase 0 microdose clinical trial. Copyright © 2015 John Wiley & Sons, Ltd.     

Synthesis and Anti‐tumor Evaluation of B‐ring Modified Caged Xanthone Analogues of Gambogic Acid

Gambogic acid (GA, 1 ), the most prominent member of Garcinia natural products, has been reported to be a promising anti‐tumor agent. Previous studies have suggested that the planar B ring and the unique 4‐oxa‐tricyclo[4.3.1.0^3,7]dec‐2‐one caged motif were essential for anti‐tumor activity. To further explore the structure‐activity relationship (SAR) of caged Garcinia xanthones, two new series of B‐ring modified caged GA analogues 13a – 13e and 15a – 15e were synthesized utilizing a Claisen/Diel‐Alder cascade reaction. Subsequently, these compounds were evaluated for their in vitro anti‐tumor activities against A549, MCF‐7, SMMC‐7721 and BGC‐823 cancer cell lines by MTT assay. Among them, 13b – 13e exhibited micromolar inhibition against several cancer cell lines, being approximately 2–4 fold less potent in comparison to GA. SAR analysis revealed that the peripheral gem‐dimethyl groups are essential for maintaining anti‐tumor activity and substituent group on C1 position of B‐ring has a significant effect on potency, while modifications at C‐2, C‐3 and C‐4 positions are relatively tolerated. These findings will enhance our understanding of the SAR of Garcinia xanthones and lead to the development of simplified analogues as potential anti‐tumor agents.     

Approaching over 10 000‐fold sensitivity increase in chiral capillary electrophoresis: Cation‐selective exhaustive injection and sweeping cyclodextrin‐modified micellar electrokinetic chromatography

A novel and simple method that combines an online concentration technique with an enantioseparation technique for capillary electrophoresis—namely, cation‐selective exhaustive injection and sweeping cyclodextrin‐modified micellar electrokinetic chromatography (CSEI‐sweeping CD‐modified MEKC)—realizes the effective enantioseparation of cationic analytes while keeping a significant increase of detection sensitivity. This technique consists of a slight modification of the basic CSEI‐sweeping MEKC. The main idea is to simply add an anionic CD as a chiral selector into the micellar buffer including sodium dodecyl sulfate, but not to change any other buffers in order to preserve the online concentration mechanism. When applied to analysis of the street drug, methamphetamine, the method achieved not only a baseline enantioseparation but also limits of detection (LODs; S/N = 3) of 70–90 pg/mL (ppt) for each isomer. This translates to a more than 10 000‐fold improvement compared to the LODs by the usual injection method. The present technique, which was made from a slight modification of CSEI‐sweeping MEKC, would give an attractive approach that is applicable to almost any analytes for which CSEI‐sweeping MEKC is applicable; all that is required is the selection of an appropriate anionic CD to be added to the micellar buffer.     

Automated determination of venlafaxine in human plasma by on‐line SPE‐LC‐MS/MS. Application to a bioequivalence study

A new automated SPE‐LC‐ESI‐MS/MS method was developed and validated to quantify venlafaxine in human plasma using fluoxetine as an internal standard. The analytes were automatically extracted from plasma by C18 SPE cartridges, separated on a C8 RP column and analyzed by MS in the multiple reaction‐monitoring (MRM) mode. The method has a chromatographic run time of 4.0 min and a linear calibration curve over the range of 0.25–200 ng/mL (r >0.997). The between‐run precisions, based on the percent RSD for replicate quality controls (0.75; 80, and 200 ng/mL), were < 8.5% for all concentrations. The between‐run accuracies, based on the percent relative error, were < 4.0%. This method was successfully employed in a bioequivalence study of two venlafaxine capsule formulations (test formulation from Eurofarma (Brazil) and Efexor XR, reference formulation, from Wyeth‐Whitehall, Brazil) in 48 healthy volunteers of both sexes who received a single 150 mg dose of each formulation. More than 3000 samples were analyzed eliminating the analyst's exposure to hazardous organic solvents normally employed in off‐line liquid–liquid extractions. The 90% confidence interval (CI) of the individual ratio geometric mean for Test/Reference was 91.6–103.4% for AUC_{0–48 h} and 102.2–112.6% for C_max. Since both 90% CI for AUC_{0–48 h} and C_max were included in the 80–125% interval proposed by the US Food and Drug Administration (FDA) and the Brazilian National Health Surveillance Agency (ANVISA), the test formulation was considered bioequivalent to Efexor XR according to both the rate and extent of absorption.