Biomedical subjects

Steven M Colegate

Publications and source records attributed to Steven M Colegate.

7 recordsLinked to original sources

PubMed2005

Improved method for extraction and LC-MS analysis of pyrrolizidine alkaloids and their N-oxides in honey: application to Echium vulgare honeys.

A method for analyzing honey samples was developed that enabled the simultaneous detection and identification of pyrrolizidine alkaloids and their N-oxides. Honey samples were treated with methanol or dilute sulfuric acid and then centrifuged to remove insoluble material. Subsequent strong cation exchange, solid-phase extraction of the supernatant provided a fraction that was analyzed for the presence of pyrrolizidine alkaloids and their N-oxides using high-pressure liquid chromatography coupled to electrospray ionization mass spectrometry. The procedure was validated using extracts of Echium plantagineum and authenticated standards of pyrrolizidine alkaloids and their N-oxides from other plant sources. Of several variations of the solid-phase extraction method assessed in this study, the best combination for generic use involved the dilution of honey with 0.05 M sulfuric acid and the subsequent application of the centrifuged solution to solid-phase extraction columns at the rate of a maximum of 10 g of honey per solid-phase extraction column. The method was applied to the analysis of nine floral honeys, five of which were attributed by the apiarist to Echium vulgare. Seven of the honey samples were positive for pyrrolizidine alkaloids and N-oxides characteristic of E. vulgare.

Chromatography, High Pressure Liquid↗

PubMed2005

Pyrrolizidine alkaloids of Echium vulgare honey found in pure pollen.

The pyrrolizidine alkaloids previously identified in floral honey attributed to Echium vulgare (Boraginaceae) have been detected (8000-14 000 ppm) in pure pollen collected from the anthers of Echium vulgare. Pyrrolizidine alkaloids and/or their N-oxides were isolated from the aqueous acid extracts of pollen by use of strong cation-exchange, solid-phase extraction and identified by liquid chromatographic/mass spectrometric (LCMS) analysis. The pyrrolizidine alkaloids in the pollen are present mainly as the N-oxides. In addition to seven previously described pyrrolizidine alkaloids and/or their N-oxides (echimidine, acetylechimidine, uplandicine, 9-O-angelylretronecine, echiuplatine, leptanthine, and echimiplatine), one unidentified (echivulgarine), but previously found in honey, and two previously undescribed (vulgarine and 7-O-acetylvulgarine) pyrrolizidine alkaloids and/or their N-oxides were identified in the pollen. Tentative structures for these unidentified pyrrolizidine alkaloids are proposed on the basis of the mass spectrometric data and biogenetic considerations. The implications of these results for identifying the source and subsequent concentrations of pyrrolizidine alkaloids in honeys and commercial bee pollen are briefly discussed.

Chromatography, High Pressure Liquid↗

PubMed2004

Solid-phase extraction and LC-MS analysis of pyrrolizidine alkaloids in honeys.

Strong-cation-exchange, solid-phase extraction of pyrrolizidine alkaloids and their N-oxides from honey samples was followed by reduction of the N-oxides and subsequent analysis of total pyrrolizidine alkaloids using high-performance liquid chromatography-atmospheric pressure chemical ionization mass spectrometry. A limited survey of 63 preprocessing samples of honey, purposefully biased toward honeys attributed to floral sources known to produce pyrrolizidine alkaloids, demonstrated levels of pyrrolizidine alkaloids up to approximately 2000 parts per billion (ppb) in a sample attributed to Echium plantagineum. Up to 800 ppb pyrrolizidine alkaloids was detected in some honeys not attributed by the collector to any pyrrolizidine alkaloid-producing floral source. No pyrrolizidine alkaloids were detected in approximately 30% of the samples in this limited study, while some honeys showed the copresence of pyrrolizidine alkaloids from multiple floral sources such as E. plantagineum and Heliotropium europaeum. In addition, retail samples of blended honeys (with no labeling to suggest that pyrrolizidine alkaloid-producing floral sources were used in the blends) have been shown to contain up to approximately 250 ppb pyrrolizidine alkaloids.

Chromatography, High Pressure Liquid↗

PubMed2002

Polyhydroxy alkaloids: chromatographic analysis.

Polyhydroxy alkaloids are a burgeoning category of natural products that encompass several structural types and generally exhibit potent activity as inhibitors of glycosidases. As presently defined the group consists of monocyclic or bicyclic aLkaloids of the pyrrolidine, piperidine, pyrrolizidine, indolizidine and tropane classes, bearing two or more hydroxyl groups. These structural features render the compounds highly water soluble and frequently quite insoluble in non-hydroxylic solvents, so that their isolation and analysis by chromatographic means are consequently difficult. This problem is further confounded by the lack of a chromophore which would permit their detection by UV absorption. This review presents chromatographic techniques that have been successfully applied to the problem of isolating, purifying, detecting and analyzing polyhydroxy alkaloids.

Alkaloids↗

PubMed2002

Toxic effects of a single parenteral dose of tunicamycin in late stage pregnancy in rats.

Tunicamycin (TM) was given as a single parenteral dose at 3 levels to female rats at gestation day 15 (GD 15) and also to non-pregnant rats. At 16 h post-dosing all pregnant rats had moderate to extensive vaginal bleeding and 1/4 at each of the 2 higher doses died. The other severely affected rats, euthanized after 26-28 h, had free blood in the uterus and large decreases in red cell count (RCC), hemoglobin (Hb) and packed cell volume (PCV). The amnion of the fetuses was very easily detached from the maternal placenta by gentle manipulation. Histologically, hemorrhage, venous thrombosis and ischemic necrosis, particularly in the maternal placenta, were consistent with the gross appearance. There was no hemorrhage in any control pregnant rats. In the remaining TM-treated pregnant rats, euthanized at GD 17, there were lesser dose-related decreases in RCC, Hb and PCV, but there was no evidence of bleeding or changes in red blood cell parameters in non-pregnant rats. There was a dose-related decrease in cholesterol and GlcNAc-1-P transferase (GPT) activity and a treatment-related decrease in serum proteins in all rats. Maternal toxicity was demonstrated in pregnant rats at TM doses < 10% of a TM lethal dose in non-pregnant rats.

Animals↗

PubMedDate not supplied

Solid-phase extraction and HPLC-MS profiling of pyrrolizidine alkaloids and their N-oxides: a case study of Echium plantagineum.

Pyrrolizidine alkaloids and their N-oxides can be extracted from the dried methanolic extracts of plant material using dilute aqueous acid. The subsequent integration of solid-phase extraction (with a strong cation exchanger) of the alkaloids and N-oxides from the aqueous acid solution, together with analysis using HPLC-ESI/MS, provides a method for the simultaneous profiling of the pyrrolizidine alkaloids and their N-oxides in plant samples and the collection of useful structural data as an aid in their identification. The N-oxide character of the analytes may be confirmed by treating analytical samples with a redox resin and observing the formation of the corresponding parent pyrrolizidine alkaloids. The present case study of Echium plantagineum highlighted a higher ratio of N-oxides to the parent tertiary bases than has been previously reported. Furthermore, a higher proportion of acetylated pyrrolizidine-N-oxides was observed in the flower heads relative to the leaves. Six pyrrolizidine alkaloids or pyrrolizidine-N-oxides, not previously reported from E. plantagineum, were tentatively identified on the basis of MS and biogenetic considerations. Three of these, 3'-O-acetylintermedine/lycopsamine, leptanthine-N-oxide and 9-O-angelylretronecine-N-oxide, have been reported elsewhere, whilst three others, 3'-O-acetylechiumine-N-oxide, echimiplatine-N-oxide and echiuplatine-N-oxide, appear unreported from any other source.

Chromatography, High Pressure Liquid↗

PubMedDate not supplied

The bioactivity-guided isolation and structural identification of toxic cucurbitacin steroidal glucosides from stemodia kingii.

A histologically validated murine model for the ovine intoxication by Stemodia kingii was used as a bioassay to guide the isolation of several groups of toxins from Stemodia kingiL Two of the toxins from one group were purified sufficiently to allow structural analysis and a determination of their median lethal doses (LD50) for oral administration to mice. A combination of acid hydrolysis, elemental analysis, HPLC-MS, 1D-NMR (1H, 13C) and 2D-NMR (1H-1H COSY, 13C-1H HSQC and HMBC, and gNOESY) was used to define stemodiosides B3 and B4 as cucurbitacin steroidal glucosides. Thus stemodioside B3 is (24Z)-3 alpha-(beta-glucopyranosyloxy)-2 beta,20,27-trihydroxy- 19-(10 -9 beta)-abeo-10alpha-lanost-5,24-diene- 11-one and stemodioside B4 is (23E)-3 alpha-(beta-glucopyranosyloxy)-20,20,22,27-tetrahydroxy- 19-( 10-9 beta)-abeo- 10 alpha-lanost-5,23-diene- 11-one. The approximate oral LD50s for stemodiosides B3 and B4 in mice were estimated to be 99 and 42 mg/kg body weight, respectively.

Animals↗