Nanoscale Three-Dimensional Imaging of Drug Distributions in Single Cells via Laser Desorption Post-Ionization Mass Spectrometry.

Exploring the three-dimensional (3D) drug distribution within a single cell at nanoscale resolution with mass spectrometry imaging (MSI) techniques is crucial in cellular biology, yet it remains a great challenge due to limited lateral resolution, detection sensitivities, and reconstruction problems. Herein, a microlensed fiber laser desorption post-ionization time-of-flight mass spectrometer (MLF-LDPI-TOFMS) was developed for the 3D imaging of two anticancer drugs within single cells at a 500 × 500 × 500 nm3 voxel resolution. Nanoscale desorption was obtained with a microlensed fiber (MLF), and a 157 nm post-ionization laser was introduced to enhance the ionization yield. Furthermore, a new type of alignment method for 3D reconstruction was developed on the basis of our embedded uniform circular polystyrene microspheres (PMs). Our findings demonstrate that this 3D imaging technique has the potential to provide information about the 3D distributions of specific molecules at the nanoscale level.

Direct and comprehensive analysis of dyes based on integrated molecular and structural information via laser desorption laser postionization mass spectrometry.

Laser desorption laser postionization time-of-flight mass spectrometry (LDPI-TOFMS) was employed for direct analysis and determination of typical basic dyes. It was also used for the analysis and comprehensive understanding of complex materials such as blue ballpoint pen inks. Simultaneous emergences of fragmental and molecular information largely simplify and facilitate unambiguous identification of dyes via variable energy of 266nm postionization laser. More specifically, by optimizing postionization laser energy with the same energy of desorption laser, the structurally significant results show definite differences in the fragmentation patterns, which offer opportunities for discrimination of isomeric species with identical molecular weight. Moreover, relatively high spectra resolution can be acquired without the expense of sensitivity. In contrast to laser desorption/ionization mass spectrometry (LDI-MS), LDPI-MS simultaneously offers valuable molecular information about dyes in traces, solvents and additives about inks, thereby offering direct determination and comprehensive understanding of blue ballpoint inks and giving a high level of confidence to discriminate the complicated evidentiary samples. In addition, direct analysis of the inks not only allows the avoidance of the tedious sample preparation processes, significantly shortening the overall analysis time and improving throughput, but allows minimized sample consumption which is important for rare and precious samples.

Identification of DW532 as a novel anti-tumor agent targeting both kinases and tubulin.

AIM: 7,8-Dihydroxy-4-(3-hydroxy-4-methoxyphenyl)-2H-chromen-2-one (DW532) is one of simplified analogues of hematoxylin that has shown broad-spectrum inhibition on tyrosine kinases and in vitro anti-cancer activities. The aim of this study was to identify DW532 as a agent targeting both kinases and tubulin, and to investigate its anti-cancer and anti-angiogenesis activities. METHODS: In vitro tyrosine kinases activity was examined with ELISA, and tyrosine kinases activity in cells was evaluated with Western blot analysis. Tubulin turbidity assay, surface plasmon resonance and immunofluorescence technique were used to characterize the tubulin inhibitory activity. Cell proliferation was examined with SRB assay, and cell apoptosis and cell cycle distribution were analyzed with Annexin-V/PI staining and flow cytometry. Tube formation, aortic ring and chick chorioallantoic membrane assays were used to evaluate the anti-angiogenesis efficacy. RESULTS: DW532 inhibited EGFR and VEGFR2 in vitro kinase activity (the IC50 values were 4.9 and 5.5 µmol/L, respectively), and suppressed their downstream signaling. DW532 dose-dependently inhibited tubulin polymerization via direct binding to tubulin, thus disrupting the mitotic spindle assembly and leading to abnormal cell division. In a panel of human cancer cells, DW532 (1 and 10 µmol/L) induced G2/M phase arrest and cell apoptosis, which subsequently resulted in cytotoxicity. Knockdown of BubR1 or Mps1, the two core proteins of the spindle assembly checkpoint dramatically decreased DW532-induced cell cycle arrest in MDA-MB-468 cells. Moreover, treatment with DW532 potently and dose-dependently suppressed angiogenesis in vitro and in vivo. CONCLUSION: DW532 is a dual inhibitor against tubulin and tyrosine kinases, and deserves further development as a novel anti-cancer agent.

HPLC determination of anethole trithione and its application to pharmacokinetics in rabbits.

To evaluate the relative bioavailability of anethole trithione (ATT) from self-microemulsifying drug delivery system (SMEDDS) and tablet, a sensitive, accurate and reliable liquid chromatography method was developed and validated to determine ATT in rabbit plasma. Chromatographic separation was performed on a Diamonsil C18 column by using a mixture of methanol-water (90:10, v/v) delivered at a flow rate of 1.0 ml/min. The wavelength was set at 348 nm and mifepristone was used as the internal standard. A linear relationship for ATT was found in the range of 0.5-32 ng/ml. The mean extraction recoveries of ATT determined over three concentrations were 84.7+/-5.8, 92.3+/-3.4 and 89.9+/-5.1%. After administration of SMEDDS and tablets to rabbits, significant differences were found in main pharmacokinetic parameters of Tmax, Cmax and AUC(0-infinity) between these two formulations, and a 2.5-fold enhancement of relative bioavailability of ATT was observed from the SMEDDS compared with tablets.