Screening of housekeeping genes in Gelsemium elegans and expression patterns of genes involved in its alkaloid biosynthesis / 生物工程学报

Gelsemium elegans is a traditional Chinese herb of medicinal importance, with indole terpene alkaloids as its main active components. To study the expression of the most suitable housekeeping reference genes in G. elegans, the root bark, stem segments, leaves and inflorescences of four different parts of G. elegans were used as materials in this study. The expression stability of 10 candidate housekeeping reference genes (18S, GAPDH, Actin, TUA, TUB, SAND, EF-1α, UBC, UBQ, and cdc25) was assessed through real-time fluorescence quantitative PCR, GeNorm, NormFinder, BestKeeper, ΔCT, and RefFinder. The results showed that EF-1α was stably expressed in all four parts of G. elegans and was the most suitable housekeeping gene. Based on the coexpression pattern of genome, full-length transcriptome and metabolome, the key candidate targets of 18 related genes (AS, AnPRT, PRAI, IGPS, TSA, TSB, TDC, GES, G8H, 8-HGO, IS, 7-DLS, 7-DLGT, 7-DLH, LAMT, SLS, STR, and SGD) involved in the Gelsemium alkaloid biosynthesis were obtained. The expression of 18 related enzyme genes were analyzed by qRT-PCR using the housekeeping gene EF-1α as a reference. The results showed that these genes' expression and gelsenicine content trends were correlated and were likely to be involved in the biosynthesis of the Gelsemium alkaloid, gelsenicine.

Difference of alkaloid components between old stems and tender stems of Gelsemium elegans / 中国中药杂志

This study aimed to comprehensively assess the difference of alkaloid components between old stems and tender stems of Gelsemium elegans by using ultra high-performance liquid chromatography coupled with photo-diode array and quadrupole time-of-flight mass spectrometry( UPLC-Q-TOF/MS~E) and high-performance liquid chromatography coupled with UV detector( HPLC-UV). Firstly,the different components in old stems and tender stems were analyzed by UHPLC-Q-TOF/MSEcombined with principal component analysis( PCA) and orthogonal partial least squares discriminant analysis( OPLS-DA),respectively. As a result,17 major different components were found. At the same time,the distribution of these alkaloids in old stems and tender stems was determined,and the alkaloids with higher polarity are relatively higher in the tender stems,while the old stems are in the opposite case. In addition,three main components in the G. elegans were quantified by HPLC-UV. The results showed that the contents of koumine and humantenmine in old stems were higher than those in tender stems,and the content of gelsemine in tender stems was relatively high. This study systematically evaluated the differences of alkaloids between the old stems and tender stems of G. elegans,and quantified the main three alkaloids. It laid the foundation of the safe and effective application of G. elegans.

Study on molecular identification of water extracts from Gelsemium elegans and Lonicera japonica and its close species by specific PCR amplification / 中国中药杂志

<p><b>OBJECTIVE</b>To explore the new method of discriminating Gelsemium elegans from Lonicera japonica and its close species by using specific PCR amplification.</p><p><b>METHOD</b>Thirteen samples of the different G. elegans materials and 58 samples of L. japonica, L. macranthoides and L. dasystyla were collected. The total DNA of the samples were extracted, and the DNA of G. elegans, L. japonica and L. macranthoides water extracts were extracted. PsbA-rnnH sequence from G. elegans was amplified by PCR and sequenced unidirectionally, ClustulW was used to align psbA-trnH sequences of the G. elegans and L. japonica and its close species from GenBank database.</p><p><b>RESULT</b>All samples were amplified by PCR with specific primer, DNA from G. elegans would be amplified 97 bp whereas PCR products from all of Lonicera samples had not bands.</p><p><b>CONCLUSION</b>Specific PCR amplification can be used to identify G. elegans from L. japonica and its close species successfully and is an efficient molecular marker for authentication of G. elegans and L. japonica and its close species.</p>

Chemical constituents of aerial parts of Gelsemium elegans / 中国中药杂志

<p><b>OBJECTIVE</b>To study the chemical constituents from the aerial parts of Gelsemium elegans.</p><p><b>METHOD</b>Compounds were isolated and purified by repeated column chromatography, as well as semiprep arative HPLC, and their structures were identified by physicochemical properties and spectroscopic methods, such as NMR and MS.</p><p><b>RESULT</b>Sixteen compounds were obtained and identified from G. elegans, including nine alkaloids: koumine (1), gelsenicine (2), 19-(Z)-akuammidine (3), gelsemoxonine(4), gelsemin (5), gelsevirine (6), humantenine (7), 11-methoxygelsemamide (8) and gelegamine D (9). Three megastigmane glycosides: (3R, 5S, 6S, 7E, 9R)-megastigman-7-ene-3, 5, 6, 9-tetrol-9-O-beta-D-glucopyranoside (10), (6R, 7E, 9R)-9-hydroxy-4, 7-megastigmadien-3-one-9-O-[alpha-L-arabinopyranosyl-(1 --> 6)-beta-D-glucopyranoside] (11) and (6S, 7E, 9R) -6, 9-dihydroxy-4, 7-megastigmadien-3-one-9-O-[alpha-L-arabinopyranosyl-(1 --> 6) -beta-D-glucopyranoside] (12). Two flavone C-glycosides: orientin (13) and isorientin (14); one iridoid glycoside, sweroside (15) and one fructoside, n-butyl-alpha-D-fructofuranoside (16).</p><p><b>CONCLUSION</b>Compounds 10-16 were isolated from the genus Gelsemium for the first time.</p>

Non-alkaloid constituents of Gelsemium elegans / 中国中药杂志

<p><b>OBJECTIVE</b>To study the non-alkaloid chemical constituents of Gelsemium elegans.</p><p><b>METHOD</b>Compounds were isolated and purified by repeated column chromatography, and their structures were elucidated by spectroscopic methods.</p><p><b>RESULT</b>Ten compounds were isolated and their structures were identified as tamarixin (1), tamarixetin 3-O-beta-D-galactopyranoside (2), scopolin (3), scopoletin (4), uradine (5), caffeic acid (6), caffeic acid ethyl ester (7), ferulic acid ethyl ester (8), ethyl-alpha-D-fructofuranoside (9), and ethyl-beta-D-fructopyranoside (10).</p><p><b>CONCLUSION</b>Compounds 1-3,5-10 are firstly isolated from this plant and compounds 1, 2, and 5-10 are isolated from the genus Gelsemium for the first time.</p>

Antineoplastic effect of koumine in mice bearing H22 solid tumor / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the antitumor effects of koumine in mice bearing H22 solid tumor and its effect on the immune system of the mice.</p><p><b>METHODS</b>The changes in spleen and tumor weights and blood cell count were observed after koumine treatment in BALB/c athymic mice bearing H22 solid tumor, using normal saline solution and 5-Fu as the controls.</p><p><b>RESULTS</b>Koumine significantly inhibited the tumor growth in a dose-dependent manner. The spleen index and blood cell counts in koumine group showed no significant differences from those in the saline control group, but higher than those in 5-Fu group.</p><p><b>CONCLUSION</b>Koumine can significantly inhibit the growth of H22 solid tumor without obvious inhibitory effect on the immune system in mice.</p>