Detection of High Levels of Pyrrolizidine-N-oxides in the Endangered Plant Cryptantha crassipes (Terlingua Creek Cat's-eye) Using HPLC-ESI-MS

ArticleinPhytochemical Analysis 22(6):532-40 · November 2011with21 Reads
DOI: 10.1002/pca.1314 · Source: PubMed
Abstract
A previous investigation of pyrrolizidine alkaloids produced by nine species of Cryptantha identified at least two chemotypes within the genus. Other research has postulated that pyrrolizidine-N-oxide concentrations increase as the growing conditions become harsher, particularly with respect to water availability. Cryptantha crassipes is an endangered plant with a very limited distribution range within a dry, harsh Texan ecosystem. To determine the pyrrolizidine alkaloid (and their N-oxides) profile and concentrations in Cryptantha crassipes. Methanolic extracts of Cryptantha crassipes were partitioned into dilute sulphuric acid and the alkaloids concentrated using strong cation exchange, solid-phase extraction columns. Extracts were analysed using reversed-phase high-pressure liquid chromatography coupled to electrospray ionisation ion trap mass spectrometry. The N-oxides of lycopsamine and intermedine were the major pyrrolizidine alkaloids detected in Cryptantha crassipes. Smaller to trace amounts of other pyrrolizidine alkaloids observed were: the 7- and 3'-acetylated derivatives and the 1,2-dihydro analogs of lycopsamine-N-oxide and/or intermedine-N-oxide; a pair of unidentified N-oxides, isobaric with lycopsamine-N-oxide; and the N-oxides of leptanthine, echimiplatine, amabiline, echiumine and dihydroechiumine. Only trace amounts, if any, of the parent free base pyrrolizidine alkaloids were detected. The concentration of pyrrolizidine alkaloids was estimated to be 3-5% of the dry weight of milled leaves, or 10-50 times the levels previously reported for similar chemotypes. The high levels of the N-oxides of lycopsamine and intermedine establish the genus chemotype of the endangered Cryptantha crassipes and support earlier data linking high levels of N-oxides to dry, harsh growing conditions.
    • "Re-extraction of the residues yielded less than 1% PAs, confirming that the extraction efficiencies were greater than 99% with the second extraction for the plant materials. Based on these findings, the routine method for PA analysis in plants was reduced to two hours compared to previous methods taking several hours or days depending on the number of re-extractions of the plants (Colegate et al. 2005; Molyneux et al. 2011; Williams et al. 2011). "
    [Show abstract] [Hide abstract] ABSTRACT: Pyrrolizidine alkaloids (PAs) are a class of naturally occurring compounds produced by many flowering plants around the World. Their presence as contaminants in food systems has become a significant concern in recent years. For example, PAs are often found as contaminants in honey through pollen transfer. A validated method was developed for the quantification of four pyrrolizidine alkaloids and one pyrrolizidine alkaloid N-oxide in plants and honey grown and produced in British Columbia. The method was optimised for extraction efficiency from the plant materials and then subjected to a single-laboratory validation to assess repeatability, accuracy, selectivity, LOD, LOQ and method linearity. The PA content in plants ranged from1.0 to 307.8 µg/g with repeatability precision between 3.8 and 20.8% RSD. HorRat values were within acceptable limits and ranged from 0.62 to 1.63 for plant material and 0.56-1.82 for honey samples. Method accuracy was determined through spike studies with recoveries ranging from 84.6 to 108.2% from the raw material negative control and from 82.1-106.0 % for the pyrrolizidine alkaloids in corn syrup. Based on the findings in this single-laboratory validation, this method is suitable for the quantitation of lycopsamine, senecionine, senecionine N-oxide, heliosupine and echimidine in common comfrey (Symphytum officinale), tansy ragwort (Senecio jacobaea), blueweed (Echium vulgare) and hound's tongue (Cynoglossum officinale) and for PA quantitation in honey and found that PA contaminants were present at low levels in BC honey.
    Full-text · Article · Oct 2015
    • "The N-oxides of this class are polar, water soluble, non-volatile and thermally labile. Thus, the application of LC-MS to the analysis of PA N-oxides has become a widely applied method [64,177,199,209,210], for example, in honey samples. "
    [Show abstract] [Hide abstract] ABSTRACT: Among the diversity of secondary metabolites which are produced by plants as means of defence against herbivores and microbes, pyrrolizidine alkaloids (PAs) are common in Boraginaceae, Asteraceae and some other plant families. Pyrrolizidine alkaloids are infamous as toxic compounds which can alkylate DNA und thus cause mutations and even cancer in herbivores and humans. Almost all genera of the family Boraginaceae synthesize and store this type of alkaloids. This review reports the available information on the present status (literature up to early 2014) of the pyrrolizidine alkaloids in the Boraginaceae and summarizes the topics structure, distribution, chemistry, chemotaxonomic significance, and biological properties.
    Full-text · Article · Mar 2014
    • "The CI mass spectrum of the alkaloid showed [M+H] + at m/z = 316 for the formula [ indicating the loss of an oxygen atom, a typical behavior of alkaloid N-oxides [4, 7]. Furthermore, we found prominent peaks at m/z = 156 and 138, which are characteristic of the 1,2-unsaturated pyrrolizidine unit [5]. By comparing the 1 H and 13 C NMR data of the alkaloid with those available for the five presently known pyrrolizidine-N-oxides of the formula C 15 H 25 NO 6 (intermedine-N-oxide 1 [18, 19], lycopsamine-N-oxide 2 [20, 21], echinatine-N-oxide 3 [21, 22], rinderine-N-oxide 4, and indicine-N-oxide 5 [18, 21]), we found a very good accordance only with the data of 1, which had previously been obtained by unambiguous total synthesis [18] (Table 1 ). "
    [Show abstract] [Hide abstract] ABSTRACT: The pyrrolizidine alkaloid intermedine-N-oxide was isolated from Cerinthe glabra for the first time. Due to the questionable assignments of NMR signals of this compound in previous literature reports ab initio calculations of the
    Article · May 2012
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