Ante- and post-mortem cellular injury dynamics in hybrid poplar foliage as a function of phytotoxic O3 dose.

After reaching phytotoxic levels during the last century, tropospheric ozone (O3) pollution is likely to remain a major concern in the coming decades. Despite similar injury processes, there is astounding interspecific-and sometimes intraspecific-foliar symptom variability, which may be related to spatial and temporal variation in injury dynamics. After characterizing the dynamics of physiological responses and O3 injury in the foliage of hybrid poplar in an earlier study, here we investigated the dynamics of changes in the cell structure occurring in the mesophyll as a function of O3 treatment, time, phytotoxic O3 dose (POD0), leaf developmental stage, and mesophyll layer. While the number of Hypersensitive Response-like (HR-like) lesions increased with higher O3 concentrations and POD0, especially in older leaves, most structural HR-like markers developed after cell death, independent of the experimental factors. The pace of degenerative Accelerated Cell Senescence (ACS) responses depended closely on the O3 concentration and POD0, in interaction with leaf age. Changes in total chlorophyll content, plastoglobuli and chloroplast shape pointed to thylakoid membranes in chloroplasts as being especially sensitive to O3 stress. Hence, our study demonstrates that early HR-like markers can provide reasonably specific, sensitive and reliable quantitative structural estimates of O3 stress for e.g. risk assessment studies, especially if they are associated with degenerative and thylakoid-related injury in chloroplasts from mesophyll.

[Effect of apigenin in combination with oxymatrine on non-small cell lung cancer and mechanism].

This study explores the effect of apigenin(APG), oxymatrine(OMT), and APG+OMT on the proliferation of non-small cell lung cancer cell lines and the underlying mechanisms. Cell counting kit-8(CCK-8) assay was used to detect the vitality of A549 and NCI-H1975 cells, and colony formation assay to evaluate the colony formation ability of the cells. EdU assay was employed to examine the proliferation of NCI-H1975 cells. RT-qPCR and Western blot were performed to detect the mRNA and protein expression of PLOD2. Molecular docking was carried out to explore the direct action ability and action sites between APG/OMT and PLOD2/EGFR. Western blot was used to study the expression of related proteins in EGFR pathway. The viability of A549 and NCI-H1975 cells was inhibited by APG and APG+OMT at 20, 40, and 80 µmol·L~(-1) in a dose-dependent manner. The colony formation ability of NCI-H1975 cells was significantly suppressed by APG and APG+OMT. The mRNA and protein expression of PLOD2 was significantly inhibited by APG and APG+OMT. In addition, APG and OMT had strong binding activity with PLOD2 and EGFR. In APG and APG+OMT groups, the expression of EGFR and proteins in its downstream signaling pathways was significantly down-regulated. It is concluded that APG in combination with OMT could inhibit non-small lung cancer, and the mechanism may be related to EGFR and its downstream signaling pathways. This study lays a new theoretical basis for the clinical treatment of non-small cell lung cancer with APG in combination with OMT and provides a reference for further research on the anti-tumor mechanism of APG in combination with OMT.

Embellicines C-E: Macrocyclic Alkaloids with a Cyclopenta[b]fluorene Ring System from the Fungus Sarocladium sp.

Macrocyclic alkaloids with a cyclopenta[b]fluorene ring system are a relatively young structural class of fungal metabolites, with the first members reported in 2013. Bioassay-guided fractionation of a Sarocladium sp. (fungal strain MSX6737) led to a series of both known and new members of this structural class (1-5), including the known embellicine A (1), three new embellicine analogues (2, 4, and 5), and a semisynthetic acetylated analogue (3). The structures were identified by examining both high-resolution electrospray ionization mass spectrometry data and one-dimensional and two-dimensional NMR spectra. The relative configurations of these molecules were established via 1H-1H coupling constants and nuclear Overhauser effect spectroscopy, while comparisons of the experimental electronic circular dichroism (ECD) spectra with the time-dependent density functional theory ECD calculations were utilized to assign their absolute configurations, which were in good agreement with the literature. These alkaloids (1-5) showed cytotoxic activity against a human breast cancer cell line (MDA-MB-231) that ranged from 0.4 to 4.8 µM. Compounds 1 and 5 were also cytotoxic against human ovarian (OVCAR3) and melanoma (MDA-MB-435) cancer cell lines.

Natural Alkaloids as Multi-Target Compounds towards Factors Implicated in Alzheimer's Disease.

Alzheimer's disease (AD) is the most common cause of dementia in elderly people; currently, there is no efficient treatment. Considering the increase in life expectancy worldwide AD rates are predicted to increase enormously, and thus the search for new AD drugs is urgently needed. A great amount of experimental and clinical evidence indicated that AD is a complex disorder characterized by widespread neurodegeneration of the CNS, with major involvement of the cholinergic system, causing progressive cognitive decline and dementia. The current treatment, based on the cholinergic hypothesis, is only symptomatic and mainly involves the restoration of acetylcholine (ACh) levels through the inhibition of acetylcholinesterase (AChE). Since the introduction of the Amaryllidaceae alkaloid galanthamine as an antidementia drug in 2001, alkaloids have been one of the most attractive groups for searching for new AD drugs. The present review aims to comprehensively summarize alkaloids of various origins as multi-target compounds for AD. From this point of view, the most promising compounds seem to be the ß-carboline alkaloid harmine and several isoquinoline alkaloids since they can simultaneously inhibit several key enzymes of AD's pathophysiology. However, this topic remains open for further research on detailed mechanisms of action and the synthesis of potentially better semi-synthetic analogues.

SWATH-MS-Based Proteomics Reveals the Regulatory Metabolism of Amaryllidaceae Alkaloids in Three Lycoris Species.

Alkaloids are a class of nitrogen-containing alkaline organic compounds found in nature, with significant biological activity, and are also important active ingredients in Chinese herbal medicine. Amaryllidaceae plants are rich in alkaloids, among which galanthamine, lycorine, and lycoramine are representative. Since the difficulty and high cost of synthesizing alkaloids have been the major obstacles in industrial production, particularly the molecular mechanism underlying alkaloid biosynthesis is largely unknown. Here, we determined the alkaloid content in Lycoris longituba, Lycoris incarnata, and Lycoris sprengeri, and performed a SWATH-MS (sequential window acquisition of all theoretical mass spectra)-based quantitative approach to detect proteome changes in the three Lycoris. A total of 2193 proteins were quantified, of which 720 proteins showed a difference in abundance between Ll and Ls, and 463 proteins showed a difference in abundance between Li and Ls. KEGG enrichment analysis revealed that differentially expressed proteins are distributed in specific biological processes including amino acid metabolism, starch, and sucrose metabolism, implicating a supportive role for Amaryllidaceae alkaloids metabolism in Lycoris. Furthermore, several key genes collectively known as OMT and NMT were identified, which are probably responsible for galanthamine biosynthesis. Interestingly, RNA processing-related proteins were also abundantly detected in alkaloid-rich Ll, suggesting that posttranscriptional regulation such as alternative splicing may contribute to the biosynthesis of Amaryllidaceae alkaloids. Taken together, our SWATH-MS-based proteomic investigation may reveal the differences in alkaloid contents at the protein levels, providing a comprehensive proteome reference for the regulatory metabolism of Amaryllidaceae alkaloids.

Two New 4-Hydroxy-2-pyridone Alkaloids with Antimicrobial and Cytotoxic Activities from Arthrinium sp. GZWMJZ-606 Endophytic with Houttuynia cordata Thunb.

Two new 4-hydroxy-2-pyridone alkaloids furanpydone A and B (1 and 2), along with two known compounds N-hydroxyapiosporamide (3) and apiosporamide (4) were isolated from the endophytic fungus Arthrinium sp. GZWMJZ-606 in Houttuynia cordata Thunb. Furanpydone A and B had unusual 5-(7-oxabicyclo[2.2.1]heptane)-4-hydroxy-2-pyridone skeleton. Their structures including absolute configurations were determined on the basis of spectroscopic analysis, as well as the X-ray diffraction experiment. Compound 1 showed inhibitory activity against ten cancer cell lines (MKN-45, HCT116, K562, A549, DU145, SF126, A-375, 786O, 5637, and PATU8988T) with IC50 values from 4.35 to 9.72 µM. Compounds 1, 3 and 4 showed moderate inhibitory effects against four Gram-positive strains (Staphylococcus aureus, methicillin-resistant S. aureus, Bacillus Subtilis, Clostridium perfringens) and one Gram-negative strain (Ralstonia solanacarum) with MIC values from 1.56 to 25 µM. However, compounds 1-4 showed no obvious inhibitory activity against two Gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa) and two pathogenic fungi (Candida albicans and Candida glabrata) at 50 µM. These results show that compounds 1-4 are expected to be developed as lead compounds for antibacterial or anti-tumor drugs.

Anti-Alzheimer's Natural Products Derived from Plant Endophytic Fungi.

Alzheimer's is the most common cause of dementia worldwide and seriously affects patients' daily tasks. Plant endophytic fungi are known for providing novel and unique secondary metabolites with diverse activities. This review focuses primarily on the published research regarding anti-Alzheimer's natural products derived from endophytic fungi between 2002 and 2022. Following a thorough review of the literature, 468 compounds with anti-Alzheimer's-related activities are reviewed and classified based on their structural skeletons, primarily including alkaloids, peptides, polyketides, terpenoids, and sterides. The classification, occurrences, and bioactivities of these natural products from endophytic fungi are summarized in detail. Our results provide a reference on endophytic fungi natural products that may assist in the development of new anti-Alzheimer's compounds.

Untargeted profiling of secondary metabolites and phytotoxins associated with stemphylium blight of lentil.

MAIN CONCLUSION: Stemphylium botryosum alters lentil secondary metabolism and differentially affects resistant and susceptible genotypes. Untargeted metabolomics identifies metabolites and their potential biosynthetic pathways that play a crucial role in resistance to S. botryosum. The molecular and metabolic processes that mediate resistance to stemphylium blight caused by Stemphylium botryosum Wallr. in lentil are largely unknown. Identifying metabolites and pathways associated with Stemphylium infection may provide valuable insights and novel targets to breed for enhanced resistance. The metabolic changes following infection of four lentil genotypes by S. botryosum were investigated by comprehensive untargeted metabolic profiling employing reversed-phase or hydrophilic interaction liquid chromatography (HILIC) coupled to a Q-Exactive mass spectrometer. At the pre-flowering stage, plants were inoculated with S. botryosum isolate SB19 spore suspension and leaf samples were collected at 24, 96 and 144 h post-inoculation (hpi). Mock-inoculated plants were used as negative controls. After analyte separation, high-resolution mass spectrometry data was acquired in positive and negative ionization modes. Multivariate modeling revealed significant treatment, genotype and hpi effects on metabolic profile changes that reflect lentil response to Stemphylium infection. In addition, univariate analyses highlighted numerous differentially accumulated metabolites. By contrasting the metabolic profiles of SB19-inoculated and mock-inoculated plants and among lentil genotypes, 840 pathogenesis-related metabolites were detected including seven S. botryosum phytotoxins. These metabolites included amino acids, sugars, fatty acids and flavonoids in primary and secondary metabolism. Metabolic pathway analysis revealed 11 significant pathways including flavonoid and phenylpropanoid biosynthesis, which were affected upon S. botryosum infection. This research contributes to ongoing efforts toward a comprehensive understanding of the regulation and reprogramming of lentil metabolism under biotic stress, which will provide targets for potential applications in breeding for enhanced disease resistance.

An epipolythiodioxopiperazine alkaloid and diversified aromatic polyketides with cytotoxicity from the Beibu Gulf coral-derived fungus Emericella nidulans GXIMD 02509. / 北部湾珊瑚来源构巢裸胞壳菌GXIMD 02509ä¸­å ·æœ‰ç»†èƒžæ¯’æ€§çš„ä¸€ä¸ªå¤šç¡«ä»£äºŒé ®å“Œå—ªç”Ÿç‰©ç¢±å’Œç»“æž„å¤šæ ·çš„èŠ³é¦™èšé ®ç±»åŒ–åˆç‰©.

Marine microorganisms, especially marine fungi, have historically proven their value as a prolific source for structurally novel and pharmacologically active secondary metabolites (Deshmukh et al., 2018; Carroll et al., 2022). The corals constitute a dominant part of reefs with the highest biodiversity, and harbor highly diverse and abundant microbial symbionts in their tissue, skeleton, and mucus layer, with species-specific core members that are spatially partitioned across coral microhabitats (Wang WQ et al., 2022). The coral-associated fungi were very recently found to be vital producers of structurally diverse compounds, terpenes, alkaloids, peptides, aromatics, lactones, and steroids. They demonstrate a wide range of bioactivity such as anticancer, antimicrobial, and antifouling activity (Chen et al., 2022). The genetically powerful genus Emericella (Ascomycota), which has marine and terrestrial sources, includes over 30 species and is distributed worldwide. It is considered a rich source of diverse secondary metabolites with antimicrobial activity or cytotoxicity (Alburae et al., 2020). Notably, Emericella nidulans, the sexual state of a classic biosynthetic strain Aspergillus nidulans, was recently reported as an important source of highly methylated polyketides (Li et al., 2019) and isoindolone-containing meroterpenoids (Zhou et al., 2016) with unusual skeletons.

Discovery and Biosynthetic Origin of Quinolizidomycins A and B, Two Quinolizidine Alkaloids from Streptomyces sp. KIB-1714.

Quinolizidomycins A (1) and B (2), two unprecedented quinolizidine alkaloids featuring a tricyclic 6/6/5 ring system, were isolated from Streptomyces sp. KIB-1714. Their structures were assigned by detailed spectroscopic data analyses and X-ray diffraction. Stable isotope labeling experiments suggested that compounds 1 and 2 are derived from lysine, ribose 5-phosphate, and acetate units, which indicates an unprecedented manner of assembly of the quinolizidine (1-azabicyclo[4.4.0]decane) scaffold in quinolizidomycin biosynthesis. Quinolizidomycin A (1) was active in an acetylcholinesterase inhibitory assay.

Revealing evolution of tropane alkaloid biosynthesis by analyzing two genomes in the Solanaceae family.

Tropane alkaloids (TAs) are widely distributed in the Solanaceae, while some important medicinal tropane alkaloids (mTAs), such as hyoscyamine and scopolamine, are restricted to certain species/tribes in this family. Little is known about the genomic basis and evolution of TAs biosynthesis and specialization in the Solanaceae. Here, we present chromosome-level genomes of two representative mTAs-producing species: Atropa belladonna and Datura stramonium. Our results reveal that the two species employ a conserved biosynthetic pathway to produce mTAs despite being distantly related within the nightshade family. A conserved gene cluster combined with gene duplication underlies the wide distribution of TAs in this family. We also provide evidence that branching genes leading to mTAs likely have evolved in early ancestral Solanaceae species but have been lost in most of the lineages, with A. belladonna and D. stramonium being exceptions. Furthermore, we identify a cytochrome P450 that modifies hyoscyamine into norhyoscyamine. Our results provide a genomic basis for evolutionary insights into the biosynthesis of TAs in the Solanaceae and will be useful for biotechnological production of mTAs via synthetic biology approaches.

Bioactivities and mechanism of action of securinega alkaloids derivatives reported prior to 2022.

Securinega alkaloids are indolizidine alkaloids extracted from the leaf and root of an Asian plant, Securinega suffruticosa. Since its discovery in 1956 by Russian scientists, numerous studies have been conducted on securinega alkaloids and their derivatives as bioactive agents. In this review, published work on the bioactivities and the mechanism of action of securinega alkaloids and their derivatives is addressed. References were obtained through for example, the Web of Science, Science Direct, Pubmed and Google Scholar. Research into the synthesis of securinega alkaloids and their derivatives lacking activity assessment has been excluded. Comprehensive reviews show that securinega alkaloids and their derivatives exhibit a wide range of activities among which antineoplastic activity and nervous system related activity were reported although the mechanisms of action remain in part unknown. The other activities such as induction of differentiation, reversal of multi-drug resistance, cardiovascular system related activity, anti-inflammatory, adjuvant agent and anti-pathogenic activity are also reviewed. We found that modification at the C12, C14, and C15 sites on securinine improves the antitumor activity, while derivatives in which a bivalent mimetic is linked to the C15 site is beneficial for differentiation induction activity and reversal of P-glycoprotein mediated drug resistance. The most related pathways involved in the bioactivity of securinega alkaloids and their derivatives are JAK/STAT, PI3K/AKT/mTOR and MAPK. A perspective and expectation concerning the research of securinega alkaloids is presented at the end of this article. This review indicates directions around which constant endeavor could be valuable for researchers in the near future.

Synthesis of novel glycerol-fluorinated triazole derivatives and evaluation of their phytotoxic and cytogenotoxic activities.

The control of weeds in agriculture is mainly conducted with the use of synthetic herbicides. However, environmental and human health concerns and increased resistance of weeds to existing herbicides have increased the pressure on researchers to find new active ingredients for weed control which present low toxicity to non-target organisms, are environmentally safe, and can be applied at low concentrations. It is herein described the synthesis of glycerol-fluorinated triazole derivatives and evaluation of their phytotoxic and cytogenotoxic activities. Starting from glycerol, ten fluorinated triazole derivatives were prepared in four steps. The assessment of them on Lactuca sativa revealed that they present effects on phytotoxic and cytogenotoxic parameters with different degrees of efficiency. The compounds 4a, 4b, 4d, 4e, 4i, and 4j have pre-emergent inhibition behavior, while all the investigated compounds showed post emergent effect. Mechanism of action as clastogenic, aneugenic, and epigenetic were observed in the lettuce root meristematic cells, with alterations as stick chromosome, bridge, delay, c-metaphase, and loss. It is believed that glycerol-fluorinated triazole derivatives possess a scaffold that can be explored towards the development of new chemicals for the control of weed species.

Synthesis, biological activity and mechanism of action of novel allosecurinine derivatives as potential antitumor agents.

Cancer with low survival rates is the second main cause of death among all diseases in the world and consequently, effective antineoplastic agents are urgently needed. Allosecurinine is a plant-derived indolicidine securinega alkaloid shown bioactivity. The object of this study is to investigate synthetic allosecurinine derivatives with considerable anticancer capacity against nine human cancer cell lines as well as mechanism of action. We synthesized twenty-three novel allosecurinine derivatives and evaluated their antitumor activity against nine cancer cell lines for 72 h by MTT and CCK8 assays. FCM was applied to analyze the apoptosis, mitochondrial membrane potential, DNA content, ROS production, CD11b expression. Western blot was selected to analyze the protein expression. Structure-activity relationships were established and potential anticancer lead BA-3 which induced differentiation of leukemia cells towards granulocytosis at low concentration and apoptosis at high concentration was identified. Mechanism studies showed that mitochondrial pathway mediated apoptosis within cancer cells with cell cycle blocking was induced by BA-3. In addition, western blot assays revealed that BA-3 induced expression of the proapoptotic factor Bax, p21 and reduced the levels of antiapoptotic protein such as Bcl-2, XIAP, YAP1, PARP, STAT3, p-STAT3, and c-Myc. Collectively, BA-3 was a lead compound for oncotherapy at least in part, through the STAT3 pathway. These results were an important step in further studies on allosecurinine-based antitumor agent development.

Biotechnological interventions and production of galanthamine in Crinum spp.

Genus Crinum L. is a member of the Amaryllidaceae family having beautiful, huge, ornamental plants with umbels of lily-like blooms that are found in tropical and subtropical climates all over the world. For thousands of years, Crinum has been used as a traditional medicine to treat illnesses and disorders. Numerous distinct alkaloids of the Amaryllidaceae group, whose most well-known properties include analgesic, anticholinergic, antitumor, and antiviral, have recently been discovered by phytochemical analyses. However, because of decades of overexploitation for their economically significant bioactive ingredients and poor seed viability and germination rates, these plants are now threatened in their native environments. Because of these factors, researchers are investigating micropropagation techniques to optimize phytochemicals in vitro. This review's objective is to offer details on the distribution, phytochemistry, micropropagation, in vitro galanthamine synthesis, and pharmacology which will help to design biotechnological techniques for the preservation, widespread multiplication, and required secondary metabolite production from Crinum spp. KEY POINTS: • Botanical description and phytochemical profile of Crinum spp. • In vitro micropropagation method of Crinum sp. • Bioactive compound galanthamine isolation techniques and its pharmacological properties.

The SlDOG1 Affect Biosynthesis of Steroidal Glycoalkaloids by Regulating GAME Expression in Tomato.

Steroidal alkaloids (SAs) and steroidal glycoalkaloids (SGAs) are common constituents of plant species belonging to the Solanaceae family. However, the molecular mechanism regulating the formation of SAs and SGAs remains unknown. Here, genome-wide association mapping was used to elucidate SA and SGA regulation in tomatoes: a SlGAME5-like glycosyltransferase (Solyc10g085240) and the transcription factor SlDOG1 (Solyc10g085210) were significantly associated with steroidal alkaloid composition. In this study, it was found that rSlGAME5-like can catalyze a variety of substrates for glycosidation and can catalyze SA and flavonol pathways to form O-glucoside and O-galactoside in vitro. The overexpression of SlGAME5-like promoted α-tomatine, hydroxytomatine, and flavonol glycoside accumulation in tomatoes. Furthermore, assessments of natural variation combined with functional analyses identified SlDOG1 as a major determinant of tomato SGA content, which also promoted SA and SGA accumulation via the regulation of GAME gene expression. This study provides new insights into the regulatory mechanisms underlying SGA production in tomatoes.

Structure-function analysis of CYP719As involved in methylenedioxy bridge-formation in the biosynthesis of benzylisoquinoline alkaloids and its de novo production.

Benzylisoquinoline alkaloids (BIAs) are a type of secondary metabolite with clinical application value. (S)-stylopine is a special BIA which contains methylenedioxy bridge structures. CYP719As could catalyze the methylenedioxy bridge-formation on the A or D rings of protoberberine alkaloids, while displaying significant substrate regiospecificity. To explore the substrate preference of CYP719As, we cloned and identified five CyCYP719A candidates from Corydalis yanhusuo. Two CyCYP719As (CyCYP719A39 and CyCYP719A42) with high catalytic efficiency for the methylenedioxy bridge-formation on the D or A rings were characterized, respectively. The residues (Leu 294 for CyCYP719A42 and Asp 289 for CyCYP719A39) were identified as the key to controlling the regioselectivity of CYP719As affecting the methylenedioxy bridge-formation on the A or D rings by homology modeling and mutation analysis. Furthermore, for de novo production of BIAs, CyCYP719A39, CyCYP719A42, and their mutants were introduced into the (S)-scoulerine-producing yeast to produce 32 mg/L (S)-stylopine. These results lay a foundation for understanding the structure-function relationship of CYP719A-mediated methylenedioxy bridge-formation and provide yeast strains for the BIAs production by synthetic biology.

Tinospora sinensis (Lour.) Merr alkaloid rich extract induces colon cancer cell death via ROS mediated, mTOR dependent apoptosis pathway: "an in-vitro study".

BACKGROUND: Colorectal cancer (CRC) is the second most mortality rate causing disease after lung cancer. Though there is a significant improvement in the treatment schedule offered to CRC. However, there is no notable decrease in terms of cases as well as death rate. Hence, there is an urgent need to discover novel cancer therapeutics to treat CRC. Since ancient times, the use of phytochemicals has drawn huge attention as chemo-preventive and chemotherapeutic agents. Earlier studies on Tinospora sinensis (TS) revealed the cytotoxic effect on human colorectal carcinoma (HCT-116) cells, yet the mechanism is to be uncovered. Therefore, the present study was designed to study the cell death mechanism of TS in HCT-116 cells. METHOD: Different extracts such as n-hexane, ethyl acetate, and ethanol extracts from the root part of TS were prepared using a cold maceration process. The extracts were screened against cancer cell lines by methyl thiazoldiphenyltetrazolium bromide (MTT) assay. From the result, the most active extract was subjected to gas chromatography-mass spectrometry (GC-MS) and Fourier-Transform infrared spectroscopy (FTIR) analyses to identify the major constituents. Finally, the mechanism of cytotoxicity to cancer cells for the most active extract was evaluated using various experiments such as cell cycle analysis, Annexin-V assay, and Western blot. RESULTS: The results from the MTT assay indicated that the n-hexane extract of TS inhibits the growth of HCT-116 cells more effectively than other cancer cells like Henrietta Lacks cervical cancer cells (Hela), and Michigan cancer foundation-breast cancer (MCF-7). The GC-MS and FT-IR analyses revealed the presence of alkaloids in the n-hexane extract and were responsible for the apoptosis activity in HCT-cells via reactive oxygen species (ROS) generation, and phosphoinositide 3-kinase (PI3K)/ protein Kinase B (Akt)/ mammalian target of rapamycin (mTOR) down-regulation. CONCLUSION: This study concludes that this finding is unique of its kind, and for the first time. The anticancer effect of TS root is specific to colon cancer cells (HCT-116). This distinctive finding helps the researchers to investigate further, and to identify a novel source for anti-colon cancer drug candidates in near future.

[Screening of housekeeping genes in Gelsemium elegans and expression patterns of genes involved in its alkaloid biosynthesis].

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.

Development of an HPLC-MS/MS Method for the Determination of Alkaloids in Lupins.

Lupin alkaloids (LAs) represent a class of toxic secondary metabolites in plants, in particular in Lupinus spp.; they are produced as a defense mechanism due to their strong bitter taste and are very dangerous for human and animals. In this work, a sensitive and reliable high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) analytical method for the identification and quantification of thirteen lupin alkaloids was developed and validated according to FDA guidelines. Efficient extraction and clean-up steps, carried out by solid-phase extraction, were finely tuned on the basis of the characteristics of the analytes and lupin samples, providing good selectivity with minimized matrix interference. The effectiveness of the method was proven by the satisfactory recovery values obtained for most of the analytes and a matrix effect ≤23% for all tested levels. In addition, a sensitive and reliable determination of the target compounds was obtained; LOQs were between 1 and 25 µg Kg-1, i.e., below the requested maximum levels (<200 mg Kg-1). The method was applied to evaluate the LAs profile in different batches of raw L. albus L. samples, varying in size and across farming treatments.