Background:Gelsemium elegans Benth(G.elegans)is a poisonous perennial evergreen vine plant that has been applied in livestock production and veterinary clinical practice.Early studies found that the toxicity of G.eleg...Background:Gelsemium elegans Benth(G.elegans)is a poisonous perennial evergreen vine plant that has been applied in livestock production and veterinary clinical practice.Early studies found that the toxicity of G.elegans showed significant gender differences in rats,but the underlying reasons for this difference are still not well understood.Methods:In order to explore whether the gender differences in the toxicity of G.elegans are related to pharmacokinetic differences,based on the previous pharmacokinetic study of multiple components of G.elegans in male rats,this study used HPLC-MS/MS method established in the laboratory to conduct a pharmacokinetic study of multiple alkaloids in the plasma of female rats after a single gavage administration of G.elegans(dose of 0.1 g/kg).Results:Through detection,17 alkaloid components in the plasma of female rats were identified,and the pharmacokinetic parameters of 11 of these alkaloids were calculated.We find that in female rats.The T_(max)values were generally less than 0.5 h,and the T_(1/2)values exceeded 3 h,with the longest reaching up to 32.80 h half elimination time.Additionally,the C_(max)and AUC results indicated that female rats had generally higher absorption and exposure levels for most alkaloids.Conclusion:These results suggest that the reason for the differences in the toxicology of G.elegans may be related to the absorption and exposure of gelsemidine-type alkaloids in animals.展开更多
Two new benzofuran lignan glycosides, gelsemiunoside A and B, were isolated from the whole plant of Gelsemium elegans Benth. Their structures were elucidated on the basis of spectroscopic evidence. Furthermore, gelsem...Two new benzofuran lignan glycosides, gelsemiunoside A and B, were isolated from the whole plant of Gelsemium elegans Benth. Their structures were elucidated on the basis of spectroscopic evidence. Furthermore, gelsemiunoside A and B were shown a potent cytotoxic activity by suppressing the proliferation of A375-S2 cells.展开更多
Low dose remedies are widely administered in medicine. We used Tele-Stereo-EEG and the hippocampal slice preparation to measure physiological effects of orally given Coffea D6 (40 mg/kg), Gelsemium D4 (10 mg/kg) and V...Low dose remedies are widely administered in medicine. We used Tele-Stereo-EEG and the hippocampal slice preparation to measure physiological effects of orally given Coffea D6 (40 mg/kg), Gelsemium D4 (10 mg/kg) and Veratrum D6 (30 mg/kg) in rats. Adult rats were implanted with electrodes positioned stereotactically into four brain regions. Changes in field potentials were transmitted wirelessly. After frequency analysis data from 6 - 8 animals were averaged. For in vitro testing, preparations were superfused directly on hippocampal slices. Stimulation of Schaffer Collaterals by single stimuli (SS) or theta burst stimulation (TBS) resulted in stable population spike amplitudes. All three low dose preparations produced decreases of spectral power. Statistically significant changes were observed in delta, theta and alpha2 spectral power. In the hippocampal slice preparation Coffea facilitated signal transfer presumably by enhancing glutamate AMPA receptor transmission. Gelsemium showed a similar effect, but only after single shock stimulation. Opposite to this, attenuation of the electric pathway was recognized after theta burst stimulation due to AMPA receptor and glutamate metabotropic II receptor mediated transmission. Veratrum was able to attenuate glutamatergic due to receptor-mediated signalling sensitive to AMPA and NMDA. The results strongly speak in favour of the existence of biologically active molecules in these low dose preparations.展开更多
The modified Hofmann degradation of koumine(1) has proved abortive. The struture and stereochemistry of the main product(3) were deduced by spectral methods and confirmed by X-ray diffration analysis. From the roots o...The modified Hofmann degradation of koumine(1) has proved abortive. The struture and stereochemistry of the main product(3) were deduced by spectral methods and confirmed by X-ray diffration analysis. From the roots of Gelsemium elegans, dihydrokounine was first isolated as a natural product.展开更多
Objective:To evaluate the anti-hepatocellular carcinoma(HCC)activity of total alkaloids from Gelsemium elegans Benth.(TAG)in vivo and in vitro and to elucidate their potential mechanisms of action through transcriptom...Objective:To evaluate the anti-hepatocellular carcinoma(HCC)activity of total alkaloids from Gelsemium elegans Benth.(TAG)in vivo and in vitro and to elucidate their potential mechanisms of action through transcriptomic analysis.Methods:TAG extraction was conducted,and the primary components were quantified using high-performance liquid chromatography(HPLC).The effects of TAG(100,150,and 200μg/mL)on various tumor cells,including SMMC-7721,HepG2,H22,CAL27,MCF7,HT29,and HCT116,were assessed.Effects of TAG on HCC proliferation and apoptosis were detected by colony formation assays and cell stainings.Caspase-3,Bcl-2,and Bax protein levels were detected by Western blotting.In vivo,a tumor xenograft model was developed using H22 cells.Totally 40 Kunming mice were randomly assigned to model,cyclophosphamide(20 mg/kg),TAG low-dose(TAG-L,0.5 mg/kg),and TAG high-dose(TAG-H,1 mg/kg)groups,with 10 mice in each group.Tumor volume,body weight,and tumor weight were recorded and compared during 14-day treatment.Immune organ index were calculated.Tissue changes were oberseved by hematoxylin and eosin staining and immunohistochemistry.Additionally,transcriptomic and metabolomic analyses,as well as quatitative real-time polymerase chain reaction(RT-qPCR),were performed to detect mRNA and metabolite expressions.Results:HPLC successfully identified the components of TAG extraction.Live cell imaging and analysis,along with cell viability assays,demonstrated that TAG inhibited the proliferation of SMMC-7721,HepG2,H22,CAL27,MCF7,HT29,and HCT116 cells.Colony formation assays,Hoechst 33258 staining,Rhodamine 123 staining,and Western blotting revealed that TAG not only inhibited HCC proliferation but also promoted apoptosis(P<0.05).In vivo experiments showed that TAG inhibited the growth of solid tumors in HCC in mice(P<0.05).Transcriptomic analysis and RT-qPCR indicated that the inhibition of HCC by TAG was associated with the regulation of the key gene CXCL13.Conclusion:TAG inhibits HCC both in vivo and in vitro,with its inhibitory effect linked to the regulation of the keygene CXCL13.展开更多
Gelsemium elegans(G.elegans)is an extremely poisonous plant that is widely distributed in southern China and southeastern Asia.G.elegans poisoning events occur frequently in southern China,and are therefore an urgent ...Gelsemium elegans(G.elegans)is an extremely poisonous plant that is widely distributed in southern China and southeastern Asia.G.elegans poisoning events occur frequently in southern China,and are therefore an urgent public health problem requiring multidisciplinary action.However,the toxic components and toxicological mechanisms remain unclear.Here,we describe a systematic investigation on the toxic components of G.elegans,resulting in the isolation and identification of 120 alkaloids.Based on acute toxicity screening,the structure-toxicity relationship of Gelsemium alkaloids was proposed for the first time.Moreover,gelsedine-and humantenine-type alkaloids were detected in the clinical blood sample,and were confirmed to be causative in the poisoning.The most toxic compound,gelsenicine(1),had selective inhibitory effects toward ventral respiratory group(VRG)neurons in the medulla,which is the main brain region controlling respiration in the central nervous system.Gelsenicine(1)strongly inhibited the firing of action potentials in VRG neurons through its ability to stimulate GABA_(A)receptors,the main receptors involved in inhibitory neurotransmission.Application of GABA_(A)receptor antagonists successively reversed action potential firing in gelsenicine(1)-treated VRG neurons.Importantly,the GABA_(A)receptor antagonists securinine and flumazenil significantly increased the survival of poisoned animals.Our findings provide insight into the components and mechanisms of G.elegans toxicity,and should assist the development of effective emergency treatments for G.elegans poisoning.展开更多
Gelsemium elegans Benth alkaloids are the main components of G.elegans and can cause acute toxicosis or even death.Although several studies have reported methods for detecting G.elegans alkaloids,a high-throughput and...Gelsemium elegans Benth alkaloids are the main components of G.elegans and can cause acute toxicosis or even death.Although several studies have reported methods for detecting G.elegans alkaloids,a high-throughput and environmental-friendly strategy for detection of multiple G.elegans alkaloids has not been realized.In this work,a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry method was developed for rapid detection of G.elegans alkaloids in human plasma and urine for diagnosis of poisoning.Multiple matrices and crys-tal spotting methods were evaluated to obtain stable and high peak intensities without“sweet spot”.We verified the methodology and obtained excellent results.The matrix effects with different dilutions were compared and good recoveries and a low relative standard deviation were obtained with a 40-fold dilution.This method could shorten the analysis time and greatly reduce the consumption of chemical solvents.Furthermore,it could be applied to quan-titative assessment of G.elegans alkaloid poisoning incidents.展开更多
Objective To develop an efficient preparative method for the separation of Gelsemium alkaloids from Gelsemium elegans. Methods High-speed counter-current chromatography （HSCCC） with several two-phase solvent systems...Objective To develop an efficient preparative method for the separation of Gelsemium alkaloids from Gelsemium elegans. Methods High-speed counter-current chromatography （HSCCC） with several two-phase solvent systems was investigated for the separation of Gelsemium alkaloids. The purity and structure identification of the purified compounds were performed with HPLC and NMR spectra, respectively. Results In a single operation, 206.6 mg of crude alkaloid sample was separated to yield 28.7 mg of koumine, 24.9 mg of gelsemine, 26.9 mg of humantenine, and 7.2 mg of gelsevirine, with the purities of 97.8%, 95.4%, 97.4%, and 93.5%, respectively. Conclusion A preparative HSCCC method is successfully established for the separation of four Gelsemium alkaloids from G. elegans with a modified two-phase solvent system com posed of n-hexane-ethyl acetate-ethanol-O. 5% triethylamine-H2O （3：5：3：4）.展开更多
钩吻生物碱是引起钩吻中毒的主要成分,严重威胁人民群众的生命健康。该文建立了分散固相萃取-高效液相色谱串联质谱(high performance liquid chromatography tandem mass spectrometry,HPLC-MS/MS)法同时快速分析食品中10种钩吻生物碱...钩吻生物碱是引起钩吻中毒的主要成分,严重威胁人民群众的生命健康。该文建立了分散固相萃取-高效液相色谱串联质谱(high performance liquid chromatography tandem mass spectrometry,HPLC-MS/MS)法同时快速分析食品中10种钩吻生物碱的方法。食品样品经1%(体积分数)HCl超声提取后调pH值至11~12,加入乙腈涡旋振荡提取后,以NaCl辅助分层离心,上清液经无水Na_(2)SO_(4)和C18脱水净化后,离心所得上清液吹干复溶,在电喷雾正离子模式(ESI^(+))下,采用多反应监测模式,以0.1%氨水-10 mmol/L乙酸铵溶液和乙腈为流动相,经Hilic(2.1 mm×100 mm×1.7μm)柱梯度洗脱分离测定,采用基质标曲外标法定量。结果表明,钩吻生物碱的基质效应为13.1%~3692.7%,在0.5~10.0 ng/mL线性良好,相关系数(R)在0.995以上,检出限和定量限分别为0.075~0.225 ng/g和0.250~0.750 ng/g,在4种食品基质中3个水平的加标回收率为79.3%~118.7%,相对标准偏差为0.1%~12.1%(n=6)。该方法操作简单、高效、灵敏度高,对误食钩吻引起的急性中毒事件诊断并采取必要措施保障生命安全具有重要现实意义。展开更多
Gelsemium elegans(G.elegans)(2 n=2 x=16)is genus of flowering plants belonging to the Gelsemicaeae family.Here,a high-quality genome assembly using the Oxford Nanopore Technologies(ONT)platform and high-throughput chr...Gelsemium elegans(G.elegans)(2 n=2 x=16)is genus of flowering plants belonging to the Gelsemicaeae family.Here,a high-quality genome assembly using the Oxford Nanopore Technologies(ONT)platform and high-throughput chromosome conformation capture techniques(Hi-C)were used.A total of 56.11 Gb of raw GridION X5 platform ONT reads(6.23 Gb per cell)were generated.After filtering,53.45 Gb of clean reads were obtained,giving 160 x coverage depth.The de novo genome assemblies 335.13 Mb,close to the 338 Mb estimated by k-mer analysis,was generated with contig N50 of 10.23 Mb.The vast majority(99.2%)of the G.elegans assembled sequence was anchored onto 8 pseudo-chromosomes.The genome completeness was then evaluated and 1338 of the 1440 conserved genes(92.9%)could be found in the assembly.Genome annotation revealed that 43.16%of the G.elegans genome is composed of repetitive elements and 23.9%is composed of long terminal repeat elements.We predicted 26,768 protein-coding genes,of which 84.56%were functionally annotated.The genomic sequences of G.elegans could be a valuable source for comparative genomic analysis in the Gelsemicaeae family and will be useful for understanding the phylogenetic relationships of the indole alkaloid metabolism.展开更多
Objective: To develop a powerful integrated strategy based on liquid chromatography coupled with mass spectrometry(LC-MS) systems for the comprehensive characterization and quantification of multiple components of her...Objective: To develop a powerful integrated strategy based on liquid chromatography coupled with mass spectrometry(LC-MS) systems for the comprehensive characterization and quantification of multiple components of herbal medicines.Methods: Firstly, different mobile phase additives, analysis time, and MS acquisition modes were orthogonally tested with liquid chromatography coupled with quadrupole time-of-flight mass spectrometry(LC-QTOF/MS) in order to detect as many components of Gelsemium elegans as possible with high peak intensity. Secondly, several data mining strategies, including database searching, diagnostic ion filtering and neutral loss filtering, were utilized to perform chemical profiling. Subsequently, this study focused on the quantification and validation of the performance of a liquid chromatography-triple mass spectrometry(LC-Qq Q/MS) assay based on derivative multiple reaction monitoring(De MRM).Results: A total of 147 components from G. elegans were characterized, among them 116 nontarget components were reported for the first time. A sensitive and reproducible LC-Qq Q/MS method was successfully developed and validated for the simultaneous relative quantification of 41 components of G. elegans.This LC-Qq Q/MS method was then applied to compare the contents of components in the roots, stems and leaves.Conclusion: The present integrated strategy would significantly contribute to chemical studies on herbal medicine, and its utility could be extended to other research fields, such as metabolomics, quality control,and pharmacokinetics.展开更多
目的探讨胆汁制钩吻总碱(胆钩吻总碱)体外抗肿瘤效果,旨在为钩吻减毒后存效的研究提供科学依据。方法通过CCK-8检测观察胆钩吻总碱对人结肠HCT-116细胞、人神经胶质瘤U87细胞、人肝癌HepG2细胞、人肺癌A549细胞的增殖作用,进一步以结肠...目的探讨胆汁制钩吻总碱(胆钩吻总碱)体外抗肿瘤效果,旨在为钩吻减毒后存效的研究提供科学依据。方法通过CCK-8检测观察胆钩吻总碱对人结肠HCT-116细胞、人神经胶质瘤U87细胞、人肝癌HepG2细胞、人肺癌A549细胞的增殖作用,进一步以结肠癌HCT-116为研究对象,以不同浓度胆钩吻总碱(50、100、200μg·mL-1)干预HCT-116细胞,通过流式细胞技术检测其对细胞周期阻滞的影响;Annexin V FITC/PI流式细胞术检测HCT-116细胞凋亡情况;进一步从蛋白水平检测凋亡相关蛋白表达。结果在钩吻总生物碱IC 50浓度下,胆钩吻总碱对U87、A549、HepG2、HCT-116肿瘤细胞的增殖抑制率均高于钩吻总生物碱组,并且各组之间的差异具有统计学意义(P<0.01)。与空白组相比,不同浓度的胆钩吻总碱处理(50、100、200μg·mL-1)可有效降低HCT-116细胞的集落形成,并将细胞周期阻滞在G 2/M期。胆钩吻总碱还能引发结肠癌HCT-116细胞的晚期凋亡,并对凋亡相关蛋白Bax、Bcl-2、caspase-3起到调控作用。结论胆钩吻总碱可以通过调控结肠癌HCT-116细胞周期和凋亡相关蛋白的表达来抑制其增殖。展开更多
基金supported by the National Natural Science Foundation of China(No.31972737).
文摘Background:Gelsemium elegans Benth(G.elegans)is a poisonous perennial evergreen vine plant that has been applied in livestock production and veterinary clinical practice.Early studies found that the toxicity of G.elegans showed significant gender differences in rats,but the underlying reasons for this difference are still not well understood.Methods:In order to explore whether the gender differences in the toxicity of G.elegans are related to pharmacokinetic differences,based on the previous pharmacokinetic study of multiple components of G.elegans in male rats,this study used HPLC-MS/MS method established in the laboratory to conduct a pharmacokinetic study of multiple alkaloids in the plasma of female rats after a single gavage administration of G.elegans(dose of 0.1 g/kg).Results:Through detection,17 alkaloid components in the plasma of female rats were identified,and the pharmacokinetic parameters of 11 of these alkaloids were calculated.We find that in female rats.The T_(max)values were generally less than 0.5 h,and the T_(1/2)values exceeded 3 h,with the longest reaching up to 32.80 h half elimination time.Additionally,the C_(max)and AUC results indicated that female rats had generally higher absorption and exposure levels for most alkaloids.Conclusion:These results suggest that the reason for the differences in the toxicology of G.elegans may be related to the absorption and exposure of gelsemidine-type alkaloids in animals.
基金sponsored by the Scientific Research Foundation for the doctoral Scholars (Q.C.Zhao,No.20031040),Liaoning,China.
文摘Two new benzofuran lignan glycosides, gelsemiunoside A and B, were isolated from the whole plant of Gelsemium elegans Benth. Their structures were elucidated on the basis of spectroscopic evidence. Furthermore, gelsemiunoside A and B were shown a potent cytotoxic activity by suppressing the proliferation of A375-S2 cells.
文摘Low dose remedies are widely administered in medicine. We used Tele-Stereo-EEG and the hippocampal slice preparation to measure physiological effects of orally given Coffea D6 (40 mg/kg), Gelsemium D4 (10 mg/kg) and Veratrum D6 (30 mg/kg) in rats. Adult rats were implanted with electrodes positioned stereotactically into four brain regions. Changes in field potentials were transmitted wirelessly. After frequency analysis data from 6 - 8 animals were averaged. For in vitro testing, preparations were superfused directly on hippocampal slices. Stimulation of Schaffer Collaterals by single stimuli (SS) or theta burst stimulation (TBS) resulted in stable population spike amplitudes. All three low dose preparations produced decreases of spectral power. Statistically significant changes were observed in delta, theta and alpha2 spectral power. In the hippocampal slice preparation Coffea facilitated signal transfer presumably by enhancing glutamate AMPA receptor transmission. Gelsemium showed a similar effect, but only after single shock stimulation. Opposite to this, attenuation of the electric pathway was recognized after theta burst stimulation due to AMPA receptor and glutamate metabotropic II receptor mediated transmission. Veratrum was able to attenuate glutamatergic due to receptor-mediated signalling sensitive to AMPA and NMDA. The results strongly speak in favour of the existence of biologically active molecules in these low dose preparations.
文摘The modified Hofmann degradation of koumine(1) has proved abortive. The struture and stereochemistry of the main product(3) were deduced by spectral methods and confirmed by X-ray diffration analysis. From the roots of Gelsemium elegans, dihydrokounine was first isolated as a natural product.
基金Supported by the Innovation Project of Guangxi Graduate Education(No.YCSW2023388)Guangxi University of Chinese Medicine PhD Start-up Fund Project(No.2020BS034)。
文摘Objective:To evaluate the anti-hepatocellular carcinoma(HCC)activity of total alkaloids from Gelsemium elegans Benth.(TAG)in vivo and in vitro and to elucidate their potential mechanisms of action through transcriptomic analysis.Methods:TAG extraction was conducted,and the primary components were quantified using high-performance liquid chromatography(HPLC).The effects of TAG(100,150,and 200μg/mL)on various tumor cells,including SMMC-7721,HepG2,H22,CAL27,MCF7,HT29,and HCT116,were assessed.Effects of TAG on HCC proliferation and apoptosis were detected by colony formation assays and cell stainings.Caspase-3,Bcl-2,and Bax protein levels were detected by Western blotting.In vivo,a tumor xenograft model was developed using H22 cells.Totally 40 Kunming mice were randomly assigned to model,cyclophosphamide(20 mg/kg),TAG low-dose(TAG-L,0.5 mg/kg),and TAG high-dose(TAG-H,1 mg/kg)groups,with 10 mice in each group.Tumor volume,body weight,and tumor weight were recorded and compared during 14-day treatment.Immune organ index were calculated.Tissue changes were oberseved by hematoxylin and eosin staining and immunohistochemistry.Additionally,transcriptomic and metabolomic analyses,as well as quatitative real-time polymerase chain reaction(RT-qPCR),were performed to detect mRNA and metabolite expressions.Results:HPLC successfully identified the components of TAG extraction.Live cell imaging and analysis,along with cell viability assays,demonstrated that TAG inhibited the proliferation of SMMC-7721,HepG2,H22,CAL27,MCF7,HT29,and HCT116 cells.Colony formation assays,Hoechst 33258 staining,Rhodamine 123 staining,and Western blotting revealed that TAG not only inhibited HCC proliferation but also promoted apoptosis(P<0.05).In vivo experiments showed that TAG inhibited the growth of solid tumors in HCC in mice(P<0.05).Transcriptomic analysis and RT-qPCR indicated that the inhibition of HCC by TAG was associated with the regulation of the key gene CXCL13.Conclusion:TAG inhibits HCC both in vivo and in vitro,with its inhibitory effect linked to the regulation of the keygene CXCL13.
基金supported by the Guangdong Basic and Applied Basic Research Foundation(Nos.2024B1515040014,2022B1515130007,2023B1515040015,and 2023A1515030012,China)the National Natural Science Foundation of China(Nos.82373750,82293681(82293680),82204236,and 82371175)+1 种基金the Science and Technology Key Project of Guangdong Province(Nos.2020B1111110004 and 2023A0505050121,China)the Science and Technology Project in Guangzhou(Nos.2024B03J1286 and 202102070001,China).
文摘Gelsemium elegans(G.elegans)is an extremely poisonous plant that is widely distributed in southern China and southeastern Asia.G.elegans poisoning events occur frequently in southern China,and are therefore an urgent public health problem requiring multidisciplinary action.However,the toxic components and toxicological mechanisms remain unclear.Here,we describe a systematic investigation on the toxic components of G.elegans,resulting in the isolation and identification of 120 alkaloids.Based on acute toxicity screening,the structure-toxicity relationship of Gelsemium alkaloids was proposed for the first time.Moreover,gelsedine-and humantenine-type alkaloids were detected in the clinical blood sample,and were confirmed to be causative in the poisoning.The most toxic compound,gelsenicine(1),had selective inhibitory effects toward ventral respiratory group(VRG)neurons in the medulla,which is the main brain region controlling respiration in the central nervous system.Gelsenicine(1)strongly inhibited the firing of action potentials in VRG neurons through its ability to stimulate GABA_(A)receptors,the main receptors involved in inhibitory neurotransmission.Application of GABA_(A)receptor antagonists successively reversed action potential firing in gelsenicine(1)-treated VRG neurons.Importantly,the GABA_(A)receptor antagonists securinine and flumazenil significantly increased the survival of poisoned animals.Our findings provide insight into the components and mechanisms of G.elegans toxicity,and should assist the development of effective emergency treatments for G.elegans poisoning.
基金supported by the National Natural Science Foundation of China(Grant No.32372448)Guangdong Basic and Applied Basic Research Foundation,China(Grant No.2023A1515012605)+2 种基金the Science and Technology Program of Guangdong Administration for Market Regulation,China(Grant No.2023CS01)the Science and Technology Program of National General Customs Administration of China(Grant No.2022HK108)the Science and Technology Program of Shantou City,China(Grant No.STKJ2023024).
文摘Gelsemium elegans Benth alkaloids are the main components of G.elegans and can cause acute toxicosis or even death.Although several studies have reported methods for detecting G.elegans alkaloids,a high-throughput and environmental-friendly strategy for detection of multiple G.elegans alkaloids has not been realized.In this work,a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry method was developed for rapid detection of G.elegans alkaloids in human plasma and urine for diagnosis of poisoning.Multiple matrices and crys-tal spotting methods were evaluated to obtain stable and high peak intensities without“sweet spot”.We verified the methodology and obtained excellent results.The matrix effects with different dilutions were compared and good recoveries and a low relative standard deviation were obtained with a 40-fold dilution.This method could shorten the analysis time and greatly reduce the consumption of chemical solvents.Furthermore,it could be applied to quan-titative assessment of G.elegans alkaloid poisoning incidents.
文摘Objective To develop an efficient preparative method for the separation of Gelsemium alkaloids from Gelsemium elegans. Methods High-speed counter-current chromatography （HSCCC） with several two-phase solvent systems was investigated for the separation of Gelsemium alkaloids. The purity and structure identification of the purified compounds were performed with HPLC and NMR spectra, respectively. Results In a single operation, 206.6 mg of crude alkaloid sample was separated to yield 28.7 mg of koumine, 24.9 mg of gelsemine, 26.9 mg of humantenine, and 7.2 mg of gelsevirine, with the purities of 97.8%, 95.4%, 97.4%, and 93.5%, respectively. Conclusion A preparative HSCCC method is successfully established for the separation of four Gelsemium alkaloids from G. elegans with a modified two-phase solvent system com posed of n-hexane-ethyl acetate-ethanol-O. 5% triethylamine-H2O （3：5：3：4）.
基金financially supported by Hunan Provincial Natural Science Foundation of China(grant 2017JJ1017)National Key R&D Program of China(grant 2017YFD0501403)+1 种基金National Natural Science Foundation of China(grant 31400275)Hunan Provincial Natural Science Foundation of China(2018JJ2172).
文摘Gelsemium elegans(G.elegans)(2 n=2 x=16)is genus of flowering plants belonging to the Gelsemicaeae family.Here,a high-quality genome assembly using the Oxford Nanopore Technologies(ONT)platform and high-throughput chromosome conformation capture techniques(Hi-C)were used.A total of 56.11 Gb of raw GridION X5 platform ONT reads(6.23 Gb per cell)were generated.After filtering,53.45 Gb of clean reads were obtained,giving 160 x coverage depth.The de novo genome assemblies 335.13 Mb,close to the 338 Mb estimated by k-mer analysis,was generated with contig N50 of 10.23 Mb.The vast majority(99.2%)of the G.elegans assembled sequence was anchored onto 8 pseudo-chromosomes.The genome completeness was then evaluated and 1338 of the 1440 conserved genes(92.9%)could be found in the assembly.Genome annotation revealed that 43.16%of the G.elegans genome is composed of repetitive elements and 23.9%is composed of long terminal repeat elements.We predicted 26,768 protein-coding genes,of which 84.56%were functionally annotated.The genomic sequences of G.elegans could be a valuable source for comparative genomic analysis in the Gelsemicaeae family and will be useful for understanding the phylogenetic relationships of the indole alkaloid metabolism.
基金supported by National Key R&D Program of Intergovernmental Key Projects (Grant No: 2018YFE0101700)National Natural Science Foundation of China (Grant No. 31972737)。
文摘Objective: To develop a powerful integrated strategy based on liquid chromatography coupled with mass spectrometry(LC-MS) systems for the comprehensive characterization and quantification of multiple components of herbal medicines.Methods: Firstly, different mobile phase additives, analysis time, and MS acquisition modes were orthogonally tested with liquid chromatography coupled with quadrupole time-of-flight mass spectrometry(LC-QTOF/MS) in order to detect as many components of Gelsemium elegans as possible with high peak intensity. Secondly, several data mining strategies, including database searching, diagnostic ion filtering and neutral loss filtering, were utilized to perform chemical profiling. Subsequently, this study focused on the quantification and validation of the performance of a liquid chromatography-triple mass spectrometry(LC-Qq Q/MS) assay based on derivative multiple reaction monitoring(De MRM).Results: A total of 147 components from G. elegans were characterized, among them 116 nontarget components were reported for the first time. A sensitive and reproducible LC-Qq Q/MS method was successfully developed and validated for the simultaneous relative quantification of 41 components of G. elegans.This LC-Qq Q/MS method was then applied to compare the contents of components in the roots, stems and leaves.Conclusion: The present integrated strategy would significantly contribute to chemical studies on herbal medicine, and its utility could be extended to other research fields, such as metabolomics, quality control,and pharmacokinetics.
文摘目的探讨胆汁制钩吻总碱(胆钩吻总碱)体外抗肿瘤效果,旨在为钩吻减毒后存效的研究提供科学依据。方法通过CCK-8检测观察胆钩吻总碱对人结肠HCT-116细胞、人神经胶质瘤U87细胞、人肝癌HepG2细胞、人肺癌A549细胞的增殖作用,进一步以结肠癌HCT-116为研究对象,以不同浓度胆钩吻总碱(50、100、200μg·mL-1)干预HCT-116细胞,通过流式细胞技术检测其对细胞周期阻滞的影响;Annexin V FITC/PI流式细胞术检测HCT-116细胞凋亡情况;进一步从蛋白水平检测凋亡相关蛋白表达。结果在钩吻总生物碱IC 50浓度下,胆钩吻总碱对U87、A549、HepG2、HCT-116肿瘤细胞的增殖抑制率均高于钩吻总生物碱组,并且各组之间的差异具有统计学意义(P<0.01)。与空白组相比,不同浓度的胆钩吻总碱处理(50、100、200μg·mL-1)可有效降低HCT-116细胞的集落形成,并将细胞周期阻滞在G 2/M期。胆钩吻总碱还能引发结肠癌HCT-116细胞的晚期凋亡,并对凋亡相关蛋白Bax、Bcl-2、caspase-3起到调控作用。结论胆钩吻总碱可以通过调控结肠癌HCT-116细胞周期和凋亡相关蛋白的表达来抑制其增殖。
