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The Jurassic relict Royal fern, Osmunda regalis L., is widely distributed across temperate zones in the Northern and Southern hemispheres. Even though this species has been utilised for centuries as a medicinal plant, its phytochemical composition mainly remains unknown. As part of our ongoing research to identify potential lead compounds for future anticancer drugs, 17 natural products were characterised from the aerial parts of Osmunda regalis L. Fifteen of these compounds were identified in this species for the first time, including the six previously undescribed compounds kaempferol 3-O-(2"-O-(2"'-α-rhamnopyranosyl)-β-glucopyranosyl)-β-glucopyranoside, quercetin 3-O-(2"-O-(2"'-α-rhamnopyranosyl)-β-glucopyranosyl)-β-glucopyranoside, kaempferol 3-O-(2"-O-(2"'-α-rhamnopyranosyl-6"'-O-(E)-caffeoyl-)-β-glucopyranosyl)-β-glucopyranoside, 3-methoxy-5-hydroxy-4-olide, 4-hydroxy-3-(3'-hydroxy-4'-(hydroxyethyl)-oxotetrafuranone-5-methyl tetrahydropyranone, and 4-O-(5-hydroxy-4-oxohexanoyl) osmundalactone. The molecular structures were determined by combining several 1D and 2D NMR experiments, circular dichroism spectroscopy, and HRMS. Determination of cytotoxicity against AML MOLM-13, H9c2, and NRK cell lines showed that two isolated lactones exhibited significant cytotoxic activity. [ABSTRACT FROM AUTHOR]

A simple and miniaturized 350-kHz helium discharge for plasma mass spectrometric detection in gas chromatography (GC) has been developed. The plasma was sustained at low pressure within the end of the capillary GC column (0.32-mm i.d.) inside the ion source housing of a quadrupole mass spectrometer. This allowed direct introduction of ions from the plasma to the mass analyzer using only a repeller and electrostatic lenses to focus the ions. The plasma was sustained in only 25 mL [min.sup.-1] of helium, which was accepted by the mass spectrometer vacuum system. This low gas flow also served to enhance the energy density of the discharge and to produce a narrow spray of ions toward the mass analyzer. Due to the miniaturized nature of the plasma, it was operated at a low power level (2.0 W), and traces of oxygen were added to avoid deposition of carbon. With this new concept for GC plasma mass spectrometric detection, chlorine was successfully monitored down to the 2.2 pg [s.sup.-1] level without interference from elements like C, S, P, O, F, and N.

Introduction: Astragali radix (AR), the root of Astragalus, is an important medical herb widely used in traditional Chinese medicine. Bioactive components include isoflavones and a unique class of triterpenoid saponins (named astragalosides). Objectives: Accurate measurement of bioactive components, especially astragaloside IV, is necessary for confirming AR authenticity, quality control and future medical research. Methodology: Liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) is a suitable technique but suffers from ion suppression effects due to sample matrix. This can be corrected by using isotopic labelled internal standards, but these are not available for many phytochemicals. We explored the use of standard addition to circumvent this issue. Results: LC–MS/MS and liquid chromatography coupled with ultraviolet (LC‐UV) detection provided linear calibration curves (R2 > 0.99). LC–MS/MS provided superior selectivity and detection limits below 10 ng/mL, which was 2–3 magnitudes lower than LC‐UV detection. Precision and accuracy were overall improved by using LC–MS/MS with diluted sample extracts, resulting in an inter series coefficient of variation (CV) of 12% or less and mean recovery estimates in the 85–115% range. LC–MS/MS quantification by standard addition resulted in significantly higher concentrations of astragaloside IV measured in the samples. Concentrations calculated by standard addition were unaffected by large variation in signal response caused by matrix effects, independent of variation in slope of the standard addition curves. Conclusion: Sample dilution was helpful but not sufficient for reducing effects of ion suppression. We have shown that LC–MS/MS quantification by standard addition can be a powerful approach for accurate measurement of phytochemicals in the absence of isotopic labelled internal standards. Astragali radix is an important herb used in traditional Chinese medicine. Liquid chromatography coupled with tandem mass spectrometry can be used for quantification of astragaloside IV and isoflavones in the herb. In the absence of isotopic labelled internal standards, we applied quantification by standard addition for measurement of these phytochemicals. Calculated concentrations were then unaffected by large variation in signal response caused by matrix effects, independent of variation in slope of the standard addition curve. [ABSTRACT FROM AUTHOR]

Abstract Background Cervical intraepithelial neoplasia (CIN) grades 2 and 3 are usually grouped and treated in the same way as "high grade", in spite of their different risk to cancer progression and spontaneous regression rates. CIN2-3 is usually diagnosed in formaldehyde-fixed paraffin embedded (FFPE) punch biopsies. This procedure virtually eliminates the availability of water-soluble proteins which could have diagnostic and prognostic value. Aim To investigate whether a water-soluble protein-saving biopsy processing method followed by a proteomic analysis of supernatant samples using LC-MS/MS (LTQ Orbitrap) can be used to distinguish between CIN2 and CIN3. Methods Fresh cervical punch biopsies from 20 women were incubated in RPMI1640 medium for 24 hours at 4°C for protein extraction and subsequently subjected to standard FFPE processing. P16 and Ki67-supported histologic consensus review CIN grade (CIN2, n = 10, CIN3, n = 10) was assessed by independent gynecological pathologists. The biopsy supernatants were depleted of 7 high abundance proteins prior to uni-dimensional LC-MS/MS analysis for protein identifications. Results The age of the patients ranged from 25-40 years (median 29.7), and mean protein concentration was 0.81 mg/ml (range 0.55 - 1.14). After application of multistep identification criteria, 114 proteins were identified, including proteins like vimentin, actin, transthyretin, apolipoprotein A-1, Heat Shock protein beta 1, vitamin D binding protein and different cytokeratins. The identified proteins are annotated to metabolic processes (36%), signal transduction (27%), cell cycle processes (15%) and trafficking/transport (9%). Using binary logistic regression, Cytokeratin 2 was found to have the strongest independent discriminatory power resulting in 90% overall correct classification. Conclusions 114 proteins were identified in supernatants from fresh cervical biopsies and many differed between CIN2 and 3. Cytokeratin 2 is the strongest discriminator with 90% overall correct classifications.