Fast, ultra-trace detection of juvenile hormone III from mosquitoes using mass spectrometry.

In the present work, a new protocol for fast separation and quantification of JH III from biological samples using liquid chromatography coupled to electrospray tandem mass spectrometry is described. In particular, the proposed protocol improves existing methodologies by combining a limited number of sample preparation steps with fast LC-MS/MS detection, providing lower limits of detection and demonstrated matrix effect control, together with high inter and intraday reproducibility. A limit of detection of 8pg/mL (0.32pg on column) was achieved, representing a 15-fold gain in sensitivity with respect to previous LC-MS based protocols. The performance of the LC-MS/MS protocol is comparable to previously described JH III quantitation protocol based on fluorescence detection, with the added advantage that quantification is independent of the availability of fluorescent tags that are often unavailable or show quite diverse responses on a batch-to-batch basis. Additionally, a detailed description of the JH III fragmentation pathway is provided for the first time, based on isolation of the molecular ion and their intermediate fragments using in-source MS/MS, MS/MS(n) and FT-ICR MS/MS measurements. The JH III workflow was evaluated as a function of developmental changes, sugar feeding and farnesoic acid stimulation in mosquitoes and can be applied to the detection of other juvenile hormones.

Crystal structure of methyl N-ferro-cenyl-carbamate.

The asymmetric unit of the title compound, [Fe(C5H5)(C7H8NO2)], contains two independent mol-ecules consisting of a ferrocenyl moiety and a nitro-gen-bound methyl carbamate. These units are almost perpendicular to each other, making dihedral angles of 87.74 (9) and 87.32 (8)°. In each independent mol-ecule, the cyclo-penta-dienyl rings deviate slightly from an eclipsed conformation and lie virtually parallel [dihedral angles = 1.42 (15) and 0.49 (13)°]. In the crystal, mol-ecules are linked by N-H⋯O hydrogen bonds into chains along the a-axis direction.

Identification of a reduction product of aristolochic acid: implications for the metabolic activation of carcinogenic aristolochic acid.

Aristolochic acids are nephrotoxic and carcinogenic natural products that have been implicated both in endemic nephropathy in the Balkan region and in ailments caused by ingestion of herbal remedies. Aristolochic acids are metabolized to active intermediates that bind to DNA. In this study, reduction of aristolochic acid I with zinc in acetic acid afforded a new product that was characterized as 9-methoxy-7-methyl-2H-1,3-oxazolo[5',4'-10,9]phenanthro[3,4-d]-1,3-dioxolane-5-carboxylic acid, designated as aristoxazole, along with the expected aristolactam I. This new compound is a condensation product of aristolochic acid and acetic acid that may be related to the aristolochic acid-DNA adducts. The proposed mechanism of formation of aristoxazole involves nucleophilic attack of acetic acid on the nitrenium ion of aristolochic acid I. On the basis of these studies, a route to the metabolic activation of aristolochic acids and formation of adducts with DNA in in vitro systems is proposed and discussed.

Bilirubin present in diverse angiosperms.

BACKGROUND AND AIMS: Bilirubin is an orange-yellow tetrapyrrole produced from the breakdown of heme by mammals and some other vertebrates. Plants, algae and cyanobacteria synthesize molecules similar to bilirubin, including the protein-bound bilins and phytochromobilin which harvest or sense light. Recently, we discovered bilirubin in the arils of Strelitzia nicolai, the White Bird of Paradise Tree, which was the first example of this molecule in a higher plant. Subsequently, we identified bilirubin in both the arils and the flowers of Strelitzia reginae, the Bird of Paradise Flower. In the arils of both species, bilirubin is present as the primary pigment, and thus functions to produce colour. Previously, no tetrapyrroles were known to generate display colour in plants. We were therefore interested in determining whether bilirubin is broadly distributed in the plant kingdom and whether it contributes to colour in other species. METHODOLOGY: In this paper, we use HPLC/UV and HPLC/UV/electrospray ionization-tandem mass spectrometry (HPLC/UV/ESI-MS/MS) to search for bilirubin in 10 species across diverse angiosperm lineages. PRINCIPAL RESULTS: Bilirubin was present in eight species from the orders Zingiberales, Arecales and Myrtales, but only contributed to colour in species within the Strelitziaceae. CONCLUSIONS: The wide distribution of bilirubin in angiosperms indicates the need to re-assess some metabolic details of an important and universal biosynthetic pathway in plants, and further explore its evolutionary history and function. Although colour production was limited to the Strelitziaceae in this study, further sampling may indicate otherwise.

Animal pigment bilirubin discovered in plants.

The bile pigment bilirubin-IXalpha is the degradative product of heme, distributed among mammals and some other vertebrates. It can be recognized as the pigment responsible for the yellow color of jaundice and healing bruises. In this paper we present the first example of the isolation of bilirubin in plants. The compound was isolated from the brilliant orange-colored arils of Strelitzia nicolai, the white bird of paradise tree, and characterized by HPLC-ESMS, UV-visible, (1)H NMR, and (13)C NMR spectroscopy, as well as comparison with an authentic standard. This discovery indicates that plant cyclic tetrapyrroles may undergo degradation by a previously unknown pathway. Preliminary analyses of related plants, including S. reginae, the bird of paradise, also revealed bilirubin in the arils and flowers, indicating that the occurrence of bilirubin is not limited to a single species or tissue type.

Thin-layer chromatography and electrospray mass spectrometry coupled using a surface sampling probe.

A combined surface sampling probe/electrospray emitter was used for the direct readout of thin-layer chromatography plates by electrospray mass spectrometry. The technique was demonstrated with reversed-phase C18 plates using a three-dye mixture composed of methylene blue, crystal violet, and rhodamine 6G for positive ion mode detection and a separate dye mixture containing fluorescein, naphthol blue black, and fast green FCF for negative ion mode detection. Acquisition of mass spectra of components of individual bands on the plate was shown by manual stepping to and sampling from specific locations within the bands. Computer-controlled scanning of development lanes on the plate was illustrated by using multiple ion monitoring in both positive and negative ion modes. Commercial TLC plates were used and no post-separation processing other than drying of the plates was required prior to mass spectrometric analysis. Readout resolution, the limits of scan speed, detection levels, TLC phase, and eluting solvents were investigated and discussed.