Employing Two-stage Derivatisation and GC-MS to Assay for Cathine and Related Stimulant Alkaloids across the Celastraceae.

INTRODUCTION: Catha edulis (qat, khat, mirra) is a woody plant species that is grown and consumed in East Africa and Yemen for its stimulant alkaloids cathinone, cathine and norephedrine. Two Celastraceae species, in addition to qat, have been noted for their stimulant properties in ethnobotanical literature. Recent phylogenetic reconstructions place four genera in a clade sister to Catha edulis, and these genera are primary candidates to search for cathine and related alkaloids. OBJECTIVE: Determine if cathine or related alkaloids are present in species of Celastraceae other than Catha edulis. METHODS: Leaf samples from 43 Celastraceae species were extracted in water followed by basification of the aqueous extract and partitioning with methyl-t-butyl ether to provide an alkaloid-enriched fraction. The extract was derivatised in a two-stage process and analysed using GC-MS for the presence of cathine. Related alkaloids and other metabolites in this alkaloid-enriched fraction were tentatively identified. RESULTS: Cathinone, cathine and norephedrine were not detected in any of the 43 Celastraceae species assayed other than Catha edulis. However, the phenylalanine- or tyrosine-derived alkaloid phenylethylamine was identified in five species. Nine species were found to be enriched for numerous sterol- and terpene-like compounds. CONCLUSION: These results indicate that cathine is unique to Catha edulis, and not the compound responsible for the stimulant properties reported in related Celastraceae species. Copyright © 2017 John Wiley & Sons, Ltd.

Lupeol and its esters: NMR, powder XRD data and in vitro evaluation of cancer cell growth

ABSTRACT The triterpene lupeol (1) and some of its esters are secondary metabolites produced by species of Celastraceae family, which have being associated with cytotoxic activity. We report herein the isolation of 1, the semi-synthesis of eight lupeol esters and the evaluation of their in vitro activity against nine strains of cancer cells. The reaction of carboxylic acids with 1 and DIC/DMAP was used to obtain lupeol stearate (2), lupeol palmitate (3) lupeol miristate (4), and the new esters lupeol laurate (5), lupeol caprate (6), lupeol caprilate (7), lupeol caproate (8) and lupeol 3',4'-dimethoxybenzoate (9), with high yields. Compounds 1-9 were identified using FT-IR, 1H, 13C-NMR, CHN analysis and XRD data and were tested in vitro for proliferation of human cancer cell activity. In these assays, lupeol was inactive (GI50> 250µg/mL) while lupeol esters 2 -4 and 7 - 9 showed a cytostatic effect. The XRD method was a suitable tool to determine the structure of lupeol and its esters in solid state. Compound 3 showed a selective growth inhibition effect on erythromyeloblastoid leukemia (K-562) cells in a concentration-dependent way. Lupeol esters 4 and 9 showed a selective cytostatic effect with low GI50 values representing promising prototypes for the development of new anticancer drugs.

Biogeography of the Malagasy Celastraceae: Multiple independent origins followed by widespread dispersal of genera from Madagascar.

Of the 97 currently recognized genera of Celastraceae, 19 are native to Madagascar, including six endemics. In this study we conducted the most thorough phylogenetic analysis of Celastraceae yet completed with respect to both character and taxon sampling, and include representatives of five new endemic genera. Fifty-one new accessions, together with 328 previously used accessions of Celastrales, were sampled for morphological characters, two rDNA gene regions, and two plastid gene regions. The endemic Malagasy genera are resolved in two separate lineages-Xenodrys by itself and all other endemic genera in a clade that also includes four lineages inferred to have dispersed from Madagascar: Brexia madagascariensis (Mascarene Islands, coastal Africa), Elaeodendron (West Indies, Africa to New Caledonia), and Pleurostylia (Africa to New Caledonia). Of the 12 extant Malagasy Celastraceae lineages identified, eight are clearly of African origin. The origins of the remaining four lineages are less clear, but reasonable possibilities include America, Eurasia, Africa, southern India, Malesia, and Australia. Based on 95% credible age intervals from fossil-calibrated molecular dating, all 12 extant Malagasy Celastraceae lineages appear to have arisen following dispersal after the separation of Madagascar from other landmasses within the last 70 million years.

An artifact caused by undersampling optimal trees in supermatrix analyses of locally sampled characters.

Empirical and simulated examples are used to demonstrate an artifact caused by undersampling optimal trees in data matrices that consist mostly or entirely of locally sampled (as opposed to globally, for most or all terminals) characters. The artifact is that unsupported clades consisting entirely of terminals scored for the same locally sampled partition may be resolved and assigned high resampling support-despite their being properly unsupported (i.e., not resolved in the strict consensus of all optimal trees). This artifact occurs despite application of random-addition sequences for stepwise terminal addition. The artifact is not necessarily obviated with thorough conventional branch swapping methods (even tree-bisection-reconnection) when just a single tree is held, as is sometimes implemented in parsimony bootstrap pseudoreplicates, and in every GARLI, PhyML, and RAxML pseudoreplicate and search for the most likely tree for the matrix as a whole. Hence GARLI, RAxML, and PhyML-based likelihood results require extra scrutiny, particularly when they provide high resolution and support for clades that are entirely unsupported by methods that perform more thorough searches, as in most parsimony analyses.

Applying plant DNA barcodes to identify species of Parnassia (Parnassiaceae).

DNA barcoding is a technique to identify species by using standardized DNA sequences. In this study, a total of 105 samples, representing 30 Parnassia species, were collected to test the effectiveness of four proposed DNA barcodes (rbcL, matK, trnH-psbA and ITS) for species identification. Our results demonstrated that all four candidate DNA markers have a maximum level of primer universality and sequencing success. As a single DNA marker, the ITS region provided the highest species resolution with 86.7%, followed by trnH-psbA with 73.3%. The combination of the core barcode regions, matK+rbcL, gave the lowest species identification success (63.3%) among any combination of multiple markers and was found unsuitable as DNA barcode for Parnassia. The combination of ITS+trnH-psbA achieved the highest species discrimination with 90.0% resolution (27 of 30 sampled species), equal to the four-marker combination and higher than any two or three marker combination including rbcL or matK. Therefore, matK and rbcL should not be used as DNA barcodes for the species identification of Parnassia. Based on the overall performance, the combination of ITS+trnH-psbA is proposed as the most suitable DNA barcode for identifying Parnassia species. DNA barcoding is a useful technique and provides a reliable and effective mean for the discrimination of Parnassia species, and in combination with morphology-based taxonomy, will be a robust approach for tackling taxonomically complex groups. In the light of our findings, we found among the three species not identified a possible cryptic speciation event in Parnassia.

Phylogeny of Celastraceae subfamily Hippocrateoideae inferred from morphological characters and nuclear and plastid loci.

The phylogeny of Celastraceae subfamily Hippocrateoideae (∼ 100 species and 19 genera in the Old and New World tropics) was inferred using morphological characters together with plastid (matK, trnL-F) and nuclear (ITS and 26S rDNA) genes. The subfamily is easily recognized by the synapomorphies of transversely flattened, deeply lobed capsules and seeds with membranous basal wings or narrow stipes together with bisexual, 5-merous flowers that generally have an extrastaminal disk and three stamens. Hippocrateoideae, like Salacioideae, are inferred to have an Old World origin. The narrow stipes of Neotropical species that are water-dispersed are inferred to be derived within the subfamily from ancestral species with wind-dispersed winged seeds. Helictonema, a monotypic genus endemic to tropical Africa, has a small, white, spongy aril that is located at the base of the seed wing and appears to be unique within Hippocrateoideae. Our inference that Helictonema is sister to the remaining members of the subfamily, considered in the context of Sarawakodendron being sister to Salacioideae, suggests that small arils and capsular fruit were primitive within both subfamilies. The aril became dramatically enlarged within Salacioideae, in which the fruits are berries, and lost entirely within Hippocrateoideae, in which the fruits are transversely flattened capsules. All five Old World taxa of Prionostemma and all eight currently recognized species within Simirestis are transferred to Pristimera, one South African variety of Pristimera is raised to species level, and all three taxa in Pristimera subgenus Trochantha are transferred to the new genus Trochantha.

[Population structure of endangered Monimopetalum chinense and its relationships with environmental factors].

Based on the field investigation data obtained from the typical plots of four community types, i. e. , secondary shrub, Phyllostachys edulis forest, Cunninghamia lanceolata forest, and Pinus massoniana forest, in the Zongli Village of Qimen County in Anhui Province, this paper studied the mean basal diameter and structure of Monimopetalum chinense population, and the effects of environmental factors on the population characteristics. The results showed that the mean basal diameter of M. chinense in the communities was in the order of P. edulis forest > P. massoniana forest > C. lanceolata forest > secondary shrub, and significantly larger in the two former forests than in the others (P < 0.05). The population structure of M. chinense also differed with habits. In secondary shrub and P. massoniana forest, the structure was a aptypical pyramid-like form, suggesting that the population was stable; in P. edulis forest, it was a spindle type, indicating that the population was at the early stage of declining; whereas in C. lanceolata forest, it was a typical pyramid-like form, with most young individuals in the population. The survival curve of the whole population belonged to Deevey II, suggesting that the population was in developing tendency with no declination. M. chinense preferred the sites with low altitude, high soil moisture and organic matter contents, gentle slope, and high coverage of tree layer; while frequent human disturbance decreased its natural regeneration and stability. Based on the results obtained, some preliminary protection suggestions were proposed.