Simultaneous quantification of Pacific ciguatoxins in fish blood using liquid chromatography–tandem mass spectrometry

Ciguatera fish poisoning (CFP) is a food intoxication caused by exposure to ciguatoxins (CTXs) in coral reef fish. Rapid analytical methods have been developed recently to quantify Pacific-CTX-1 (P-CTX-1) in fish muscle, but it is destructive and can cause harm to valuable live coral reef fish. Also fish muscle extract was complex making CTX quantification challenging. Not only P-CTX-1, but also P-CTX-2 and P-CTX-3 could be present in fish, contributing to ciguatoxicity. Therefore, an analytical method for simultaneous quantification of P-CTX-1, P-CTX-2, and P-CTX-3 in whole blood of marketed coral reef fish using sonication, solid-phase extraction (SPE), and liquid chromatography tandem mass spectrometry (LC-MS/MS) was developed. The optimized method gave acceptable recoveries of P-CTXs (74–103 %) in fish blood. Matrix effects (6–26 %) in blood extracts were found to be significantly reduced compared with those in muscle extracts (suppressed by 34–75 % as reported in other studies), thereby minimizing potential for false negative results. The target P-CTXs were detectable in whole blood from four coral reef fish species collected in a CFP-endemic region. Similar trends in total P-CTX levels and patterns of P-CTX composition profiles in blood and muscle of these fish were observed, suggesting a relationship between blood and muscle levels of P-CTXs. This optimized method provides an essential tool for studies of P-CTX pharmacokinetics and pharmacodynamics in fish, which are needed for establishing the use of fish blood as a reliable sample for the assessment and control of CFP.

Ultrafast extreme ultraviolet holography: dynamic monitoring of surface deformation

We demonstrate femtosecond time-resolved dynamic Gabor holography using highly coherent extreme ultraviolet light generated by high harmonic upconversion of a femtosecond laser. By reflecting this light from an impulsively heated surface, we implement a simple and robust single-reflection geometry for phase-sensitive holographic detection at extreme UV wavelengths. Using this setup, we study the ultrafast deformation and subsequent acoustic oscillations within a thin metal film. These measurements exhibit subpicometer spatial sensitivity in the vertical dimension.     

Synthesis of docetaxel and butitaxel analogues through kinetic resolution of racemic beta-lactams with 7-O-triethylsilylbaccatin III

The kinetic resolution of racemic cis-4-phenyl- and cis-4-tert-butyl-3-hydroxy-beta-lactam derivatives with 7-O-triethylsilylbaccatin III yielded paclitaxel and butitaxel analogues with high diastereoselectivity. The results demonstrated that the tert-butyldimethylsilyl protecting group at the C3-hydroxy group of the beta-lactams provided optimum kinetic resolution in comparison with the sterically less demanding triethylsilyl group and the larger triisopropylsilyl group. In addition, it was found that the C4 beta-lactam substituents also influenced diastereoselectivity. The C4 tert-butyl-beta-lactams provided better diastereoselectivity than the corresponding C4 phenyl beta-lactams.     

Synthetic 3-O-methylmannose-containing polysaccharides (sMMPs): design and synthesis

With the hope of mimicking the chemical and biological properties of natural 3-O-methylmannose-containing polysaccharides (MMPs), synthetic 3-O-methylmannose-containing polysaccharides (sMMPs) were designed and synthesized in a convergent manner. With little modification of the Mukaiyama glycosidation, high alpha-selectivity (>50:1 approximately >20:1) and yields (79 approximately 74%) were achieved for the key glycosidation steps. The exceptionally high alpha-selectivity observed was shown to be consequent to the selective anomerization of beta- to alpha-anomer under the glycosidation conditions. This glycosidation is well suited for a highly convergent oligosaccharide synthesis, particularly because of excellent chemical yields even when using approximately equal-sized donors and acceptors in an approximately 1:1 molar ratio. An iterative reaction sequence allowed the growing oligosaccharide to double in size after each cycle and led to an efficient synthesis of sMMP 8-, 12-, and 16-mers 18-20.     

Synchrotron FTIR microspectroscopic analysis of the effects of anti-inflammatory therapeutics on wound healing in laminectomized rats

Peridural scarring, or the excessive formation of scar tissue following spinal surgery, is one of the important contributing factors that result in persistent pain and disability in many individuals who have undergone elective back surgery. Treatment with anti-inflammatory agents following surgery may reduce oxidative stress and scarring, leading to a reduction in post-operative pain. We are using a surgical rat model to test the hypothesis that post-surgical inflammation and oxidative stress following laminectomy can be reduced by systemic administration of L-2-oxo-thiazolidine-4-carboxylate (OTC) and quercetin. OTC is a cysteine precursor required for the synthesis of glutathione, an important antioxidant. Quercetin is a flavenoid with anti-oxidant properties, found in fruits and vegetables. Synchrotron FTIR microspectroscopy data has been collected on OTC, quercetin and saline (control)-treated post-surgery animals, sacrificed at 3 and 21 days (n = 6 per age and treatment group). This paper presents preliminary IR results, supported by immunocytochemistry, on the heterogenous distribution of biological components present in the healing tissue. The data collected on animals sacrificed at 3 and 21 days post-surgery will be combined in the future with data from animals sacrificed 63 days after surgery (representing a third time point) to evaluate the efficacy of the different treatments. Initial statistical analysis of ED1 immunohistochemistry results indicates a decrease in the number of activated macrophages 21 days post-surgery in the OTC-treated animals compared with the saline controls.     

A Series of Collaborations between Various Pharmaceutical Companies and Regulatory Authorities Concerning the Analysis of Biomolecules Using Capillary Electrophoresis: Additional Instruments/Buffer

An international project team (including members from US, Canada and UK) was formed from a number of interested biopharmaceutical companies and regulatory authorities to conduct a cross-organisation collaboration exercise. The results of the first comparison with eight different organisations that used instruments of the same equipment model, the same reagents, and the same methodology has been reported previously [1]. This report represents the addition of other instruments using a different run buffer. The relative migration times were different, as expected, prohibiting a direct comparison between companies. The within-organisation variability was low for both relative migration time (<0.34% RSD% for all companies save one) and the peak area (<5% RSD% for all companies save one) when measuring the purity of a representative IgG sample. The apparent molecular weight of bovine serum albumin was measured with good precision (less than 10% RSD% across all companies) to the theoretical value when all data is utilized (67.5 kDa compared to 66.4 kDa). For a representative IgG sample, the three main components, IgG Light Chain, IgG Non-glycosylated Heavy Chain, and IgG Heavy Chain, could not be separated, specifically the IgG Non-glycosylated Heavy Chain and IgG Heavy Chain. When the IgG Non-glycosylated Heavy Chain and IgG Heavy Chain were combined for all organisations, the fractional peak area for the IgG Light Chain and IgG Non-glycosylated Heavy Chain + IgG Heavy Chain peak also showed excellent agreement, with less than 7.5 and 3.5% RSD%, respectively. The value of this exercise is in demonstrating the reliability of CE for the determination of apparent size of biopharmaceutical proteins. This underpins the appropriate use of such CE data in support of regulatory submissions.

     

Total synthesis of paecilospirone

Neutrality is the best policy: Key features of the first total synthesis of paecilospirone include an anti-selective, lactate-derived aldol reaction, and a double deallylation/spirocyclization conducted at neutral pH to construct the sensitive hydroxy-substituted benzannulated spiroacetal (see scheme; Bn=benzyl, TBS=tert-butyldimethylsilyl, TES=triethylsilyl).