Search results for: in vitro metabolites

Authors: Adnan A. Kadi, Nasser S. Al-Shakliah, Haitham Al-Rabiah

Abstract:

Zorifertinib is a novel, potent, oral, a small molecule used to treat non-small cell lung cancer (NSCLC). zorifertinib is an Epidermal Growth Factor Receptor (EGFR) inhibitor and has good blood–brain barrier permeability for (NSCLC) patients with EGFR mutations. zorifertinibis currently at phase II/III clinical trials. The current research reports the characterization and identification of in vitro, in vivo and reactive intermediates of zorifertinib. Prediction of susceptible sites of metabolism and reactivity pathways (cyanide and GSH) of zorifertinib were performed by the Xenosite web predictor tool. In-vitro metabolites of zorifertinib were performed by incubation with rat liver microsomes (RLMs) and isolated perfused rat liver hepatocytes. Extraction of zorifertinib and it's in vitro metabolites from the incubation mixtures were done by protein precipitation. In vivo metabolism was done by giving a single oral dose of zorifertinib(10 mg/Kg) to Sprague Dawely rats in metabolic cages by using oral gavage. Urine was gathered and filtered at specific time intervals (0, 6, 12, 18, 24, 48, 72,96and 120 hr) from zorifertinib dosing. A similar volume of ACN was added to each collected urine sample. Both layers (organic and aqueous) were injected into liquid chromatography ion trap mass spectrometry(LC-IT-MS) to detect vivozorifertinib metabolites. N-methyl piperizine ring and quinazoline group of zorifertinib undergoe metabolism forming iminium and electro deficient conjugated system respectively, which are very reactive toward nucleophilic macromolecules. Incubation of zorifertinib with RLMs in the presence of 1.0 mM KCN and 1.0 Mm glutathione were made to check reactive metabolites as it is often responsible for toxicities associated with this drug. For in vitro metabolites there were nine in vitro phase I metabolites, four in vitro phase II metabolites, eleven reactive metabolites(three cyano adducts, five GSH conjugates metabolites, and three methoxy metabolites of zorifertinib were detected by LC-IT-MS. For in vivo metabolites, there were eight in vivo phase I, tenin vivo phase II metabolitesofzorifertinib were detected by LC-IT-MS. In vitro and in vivo phase I metabolic pathways wereN- demthylation, O-demethylation, hydroxylation, reduction, defluorination, and dechlorination. In vivo phase II metabolic reaction was direct conjugation of zorifertinib with glucuronic acid and sulphate.

Keywords: in vivo metabolites, in vitro metabolites, cyano adducts, GSH conjugate

Procedia PDF Downloads 169

Authors: Deepanjali Gurav, Kun Qian

Abstract:

In-vitro metabolic diagnosis relies on designed materials-based analytical platforms for detection of selected metabolites in biological samples, which has a key role in disease detection and therapeutic evaluation in clinics. However, the basic challenge deals with developing a simple approach for metabolic analysis in bio-samples with high sample complexity and low molecular abundance. In this work, we report a designer plasmonic nanoshells based platform for direct detection of small metabolites in clinical samples for in-vitro metabolic diagnostics. We first synthesized a series of plasmonic core-shell particles with tunable nanoshell structures. The optimized plasmonic nanoshells as new matrices allowed fast, multiplex, sensitive, and selective LDI MS (Laser desorption/ionization mass spectrometry) detection of small metabolites in 0.5 μL of bio-fluids without enrichment or purification. Furthermore, coupling with isotopic quantification of selected metabolites, we demonstrated the use of these plasmonic nanoshells for disease detection and therapeutic evaluation in clinics. For disease detection, we identified patients with postoperative brain infection through glucose quantitation and daily monitoring by cerebrospinal fluid (CSF) analysis. For therapeutic evaluation, we investigated drug distribution in blood and CSF systems and validated the function and permeability of blood-brain/CSF-barriers, during therapeutic treatment of patients with cerebral edema for pharmacokinetic study. Our work sheds light on the design of materials for high-performance metabolic analysis and precision diagnostics in real cases.

Keywords: plasmonic nanoparticles, metabolites, fingerprinting, mass spectrometry, in-vitro diagnostics

Procedia PDF Downloads 111

Authors: H. Chassaigne, S. Gioria, J. Lobo Vicente, D. Carpi, P. Barboro, G. Tomasi, A. Kinsner-Ovaskainen, F. Rossi

Abstract:

Emerging approaches in the area of exposure to nanomaterials and assessment of human health effects combine the use of in vitro systems and analytical techniques to study the perturbation of the proteome and/or the metabolome. We investigated the modification in the cytoplasmic compartment of the Balb/3T3 cell line exposed to gold nanoparticles. On one hand, the proteomic approach is quite standardized even if it requires precautions when dealing with in vitro systems. On the other hand, metabolomic analysis is challenging due to the chemical diversity of cellular metabolites that complicate data elaboration and interpretation. Differentially expressed proteins were found to cover a range of functions including stress response, cell metabolism, cell growth and cytoskeleton organization. In addition, de-regulated metabolites were annotated using the HMDB database. The "omics" fields hold huge promises in the interaction of nanoparticles with biological systems. The combination of proteomics and metabolomics data is possible however challenging.

Keywords: data processing, gold nanoparticles, in vitro systems, metabolomics, proteomics

Procedia PDF Downloads 478

Authors: Ennus Tajuddin Tamboli, Madhukar Garg, Mohd. Mujeeb, Sayeed Ahmad

Abstract:

An efficient protocol for simultaneous production of forskolin and rosmarinic acid in in vitro callus derived from the leaves of Coleus forskohlii Briq. has been developed. MS media was used for the establishment of cultures and NAA + 6-BA (1.0 ppm) was found best for callus growth. The callus was further subjected to treatment with various elicitor/precursors viz. chitosan, thidiazuron and methyl jasmonate to observe their effect on production of biomass and accumulation of secondary metabolites. The content of forskolin and rosmarinic acid were estimated by HPTLC, in comparison to natural explant which showed 2 fold and 10 fold rise in forskolin and rosmarinic acid content, respectively. Methy1 jasmonate 50 µM was found best for production of forskolin, whereas thidiazuron showed best results in the yield of rosmarinic acid, separately in static culture. However, combined treatment in suspension culture showed moderated effect for increase in secondary metabolites but the biomass increased significantly as compared to static culture.

Keywords: plant tissue culture, secondary metabolites, coleus, forskolin, rosmarinic acid, HPTLC

Procedia PDF Downloads 315

Authors: Madhusmita Dehingia, Supriyo Sen, Bhuwan Bhaskar, Tulsi Joishy, Mojibur R. Khan

Abstract:

Human gut microbes and their metabolites are important for maintaining homeostasis in the gut and are responsible for many metabolic and immune mediated diseases. In the present study, we determined the profiles of the gut metabolites of five different ethnic groups (Bodo, Tai-Phake, Karbi, Tea tribe and Tai-Aiton) of Assam. Fecal metabolite profiling of the 39 individuals belonging to the ethnic groups was carried out using Gas chromatography – Mass spectrometry (GC-MS), and comparison was performed among the tribes for common and unique metabolites produced within their gut. Partial Least Squares Discriminant Analysis (PLS-DA) of the metabolites suggested that the individuals grouped according to their ethnicity. Among the 66 abundant metabolites, 12 metabolites were found to be common among the five ethnic groups. Additionally, ethnicity wise some unique metabolites were also detected. For example, the tea tribe of Assam contained the tea components, Aniline and Benzoate more in their gut in comparison to others. Metabolites of microbial origin were also correlated with the already published metagenomic data of the same ethnic group and functional analysis were carried out based on human metabolome database.

Keywords: ethnicity, gut microbiota, GC-MS, metabolites

Procedia PDF Downloads 395

Authors: Nouf Alourfi

Abstract:

This study examines the metabolism of amitriptyline (AMI) and verapamil (VER) using a KNIME-based method. KNIME improved workflow is an open-source data-analytics platform that integrates a number of open-source metabolomics tools such as CFMID and MetFrag to provide standard data visualisations, predict candidate metabolites, assess them against experimental data, and produce reports on identified metabolites. The use of this workflow is demonstrated by employing three types of liver microsomes (human, rat, and Guinea pig) to study the in vitro metabolism of the two drugs (AMI and VER). This workflow is used to create and treat UPLC-MS (Orbitrap) data. The formulas and structures of these drugs' metabolites can be assigned automatically. The key metabolic routes for amitriptyline are hydroxylation, N-dealkylation, N-oxidation, and conjugation, while N-demethylation, O-demethylation and N-dealkylation, and conjugation are the primary metabolic routes for verapamil. The identified metabolites are compatible to the published, clarifying the solidity of the workflow technique and the usage of computational tools like KNIME in supporting the integration and interoperability of emerging novel software packages in the metabolomics area.

Keywords: KNIME, CFMID, MetFrag, Data Analysis, Metabolomics

Procedia PDF Downloads 92

Authors: Aleya Ferdausi, Meriel Jones, Anthony Halls

Abstract:

The Amaryllidaceae genus Narcissus contains secondary metabolites, which are important sources of bioactive compounds such as pharmaceuticals indicating that their biological activity extends from the native plant to humans. Transcriptome analysis (RNA-seq) is an effective platform for the identification and functional characterization of candidate genes as well as to identify genes encoding uncharacterized enzymes. The biotechnological production of secondary metabolites in plant cell or organ cultures has become a tempting alternative to the extraction of whole plant material. The biochemical pathways for the production of secondary metabolites require primary metabolites to undergo a series of modifications catalyzed by enzymes such as cytochrome P450s, methyltransferases, glycosyltransferases, and acyltransferases. Differential gene expression analysis of Narcissus was obtained from two conditions, i.e. field and in vitro callus. Callus was obtained from modified MS (Murashige and Skoog) media supplemented with growth regulators and twin-scale explants from Narcissus cv. Carlton bulb. A total of 2153 differentially expressed transcripts were detected in Narcissus bulb and in vitro callus, and 78.95% of those were annotated. It showed the expression of genes involved in the biosynthesis of alkaloids were present in both conditions i.e. cytochrome P450s, O-methyltransferase (OMTs), NADP/NADPH dehydrogenases or reductases, SAM-synthetases or decarboxylases, 3-ketoacyl-CoA, acyl-CoA, cinnamoyl-CoA, cinnamate 4-hydroxylase, alcohol dehydrogenase, caffeic acid, N-methyltransferase, and NADPH-cytochrome P450s. However, cytochrome P450s and OMTs involved in the later stage of Amaryllidaceae alkaloids biosynthesis were mainly up-regulated in field samples. Whereas, the enzymes involved in initial biosynthetic pathways i.e. fructose biphosphate adolase, aminotransferases, dehydrogenases, hydroxyl methyl glutarate and glutamate synthase leading to the biosynthesis of precursors; tyrosine, phenylalanine and tryptophan for secondary metabolites were up-regulated in callus. The knowledge of probable genes involved in secondary metabolism and their regulation in different tissues will provide insight into the Narcissus plant biology related to alkaloid production.

Keywords: narcissus, callus, transcriptomics, secondary metabolites

Procedia PDF Downloads 114

Authors: Gayatri Nahak, Satyajit Kanungo, Rajani Kanta Sahu

Abstract:

Holarrhena antidysenterica Linn. (Apocynaceae) is a typical Indian medicinal plant popularly known as “Indrajav”. Traditionally the plant has been considered a popular remedy for the treatment of dysentery, diarrhea, intestinal worms and the seeds of this plant are also used as an anti-diabetic remedy. In the present study axillary shoot multiplication, callus induction and shoot regeneration from callus culture were obtained on Murashige and Skoog (MS) medium supplemented with different concentrations and combinations of plant growth regulators. Then in vivo and in vitro grown healthy plants were selected for study of antioxidant activity through DPPH and OH methods. Significantly higher antioxidant activity and phenol contents were observed in vitro raised plant in comparison to in vivo plants. The findings indicated the greater amount of phenolic compounds leads to more potent radical scavenging effect as shown in in vitro raised plant in comparison to in vivo plants which showed the ability to utilize tissue culture techniques towards development of desired bioactive metabolites from in vitro culture as an alternative way to avoid using endangered plants in pharmaceutical purposes.

Keywords: Holarrhena antidysenterica, in vitro, in vivo, antioxidant activity

Procedia PDF Downloads 479

Authors: H. Heydari, B. Konuklugil, P. Proksch

Abstract:

Marine-derived fungi can produce interesting bioactive secondary metabolites that can be considered the potential for drug development. Turkey is a country of a peninsula surrounded by the Black Sea at the north, the Aegean Sea at the west, and the Mediterranean Sea at the south. Despite the approximately 8400 km of coastline, studies on marine secondary metabolites and their biological activity are limited. In our ongoing search for new natural products with different bioactivities produced by the marine-derived fungi, we have investigated secondary metabolites of Turkish collection of the marine sea slug (Peltodoris atromaculata) associated fungi Hypocrea nigricans collected from Seferihisar in the Egean sea. According to the author’s best knowledge, no study was found on this fungal species in terms of secondary metabolites. Isolated from ethyl acetate extract of the culture of Hypocrea nigricans were (isodihydroauroglaucin,tetrahydroauroglaucin and dihydroauroglaucin. The structures of the compounds were established based on an NMR and MS analysis. Structural elucidation of another isolated secondary metabolite/s continues.

Keywords: Hypocrea nigricans, isolation, marine fungi, secondary metabolites

Procedia PDF Downloads 134

Authors: Muhammad Aasim, Mehmet Karataş, Fatih Erci, Şeyma Bakırcı, Ecenur Korkmaz, Burak Kahveci

Abstract:

Water hyssop (Bacopa monnieri L. Wettst.) is an important medicinal aquatic/semi aquatic plant native to India where it is used in traditional medicinal system. The plant contains bioactive compounds mainly Bacosides which are the main ingridient of commercial drug available as memory enhancer tonic. The local name of water hyssop is Brahmi and brahmi based drugs are available against for curing chronic diseases and disorders Alzheimer’s disease, anxiety, asthma, cancer, mental illness, respiratory ailments, and stomach ulcers. The plant is not a cultivated plant and collection of plant from nature make palnt threatened to endangered. On the other hand, low seed viability and availability make it difficult to propagate plant through traditional techniques. In recent years, plant tissue culture techniques have been employed to propagate plant for its conservation and production for continuous availability of secondary metabolites. On the other hand, application of nanoparticles has been reported for increasing biomass, in vitro regeneration and secondary metabolites production. In this study, silver nanoparticles (AgNPs) were applied at the rate of 2, 4, 6, 8 and 10 ppm to Murashihe and Skoog (MS) medium supplemented with 1.0 mg/l Benzylaminopurine (BAP), 3.0% sucrose and 0.7% agar. Leaf explants of water hyssop were cultured on AgNPs containing medium. Shoot induction from leaf explants were relatively slow compared to medium without AgNPs. Multiple shoot induction was recorded after 3-4 weeks of culture comapred to control that occured within 10 days. Regenerated shoots were rooted successfully on MS medium supplemented with 1.0 mg/l IBA and acclimatized in the aquariums for further studies.

Keywords: Water hyssop, Silver nanoparticles, In vitro, Regeneration, Secondary metabolites

Procedia PDF Downloads 146

Authors: Yao-Te Yen, Chao-Hsin Cheng, Meng-Shun Huang, Shan-Zong Cyue

Abstract:

'Poppy-seed defense' has been a serious problem all over the world, that is because the opiates metabolites in urine are difficult to distinguish where they come from precisely. In this research, a powerful analytic method has been developed to trace the opiates metabolites in urine by Gas-Chromatography Isotope Ratio Mass Spectrometry (GC-IRMS). In order to eliminate the interference of synthesis to heroin or metabolism through human body, opiates metabolites in urine and sized heroin were hydrolyzed to morphine. Morphine is the key compound for tracing between opiates metabolites and seized heroin in this research. By matching δ13C and δ15N values through morphine, it is successful to distinguish the opiates metabolites coming from heroin or medicine. We tested seven heroin abuser’s metabolites and seized heroin in crime sites, the result showed that opiates metabolites coming from seized heroin, the variation of δ13C and δ15N for morphine are within 0.2 and 2.5‰, respectively. The variation of δ13C and δ15N for morphine are reasonable with the result of matrix match experiments. Above all, the uncertainty of 'Poppy-seed defense' can be solved easily by this analytic method, it provides the direct evidence for judge to make accurate conviction without hesitation.

Keywords: poppy-seed defense, heroin, opiates metabolites, isotope ratio mass spectrometry

Procedia PDF Downloads 214

Authors: Richard I. Licayan, Aisle Janne B. Dagpin, Romeo M. Del Rosario, Nenita D. Palmes

Abstract:

Hiptage benghalensis, Antigonon leptopus, Macroptillium atropurpureum, and Dioscorea bulbifera L. are herbal weeds that have been used by traditional healers in rural communities in the Philippines as medicine. In this study, the basic pharmacological components of the crude secondary metabolites extracted from the four herbal weeds and their in vitro antioxidant properties was investigated to provide baseline data for the possible development of these metabolites in pharmaceutical products. Qualitative screening of the secondary metabolites showed that alkaloids, tannins, saponins, steroids, and flavonoids were present in their leaf extracts. All of the plant extracts showed varied antioxidant activity. The greatest DPPH radical scavenging activity was observed in H. begnhalensis (84.64%), followed by A. leptopus (68.21%), M. atropurpureum (26.62%), and D. bulbifera L. (19.04%). The FRAP assay revealed that H. benghalensis had the highest antioxidant activity (8.32 mg/g) while ABTS assay showed that M. atropurpureum had the strongest scavenging ability of free radicals (0.0842 mg Trolox/g). The total flavonoid content (TFC) analysis showed that D. bulbifera L. had the highest TFC (420.35 mg quercetin per gram-dried material). The total phenolic content (TPC) of the four herbal weeds showed large variations, between 26.56±0.160 and 55.91±0.087 mg GAE/g dried material. The plant leaf extracts arranged in increasing values of TPC are H. benghalensis (26.565) < A. leptopus (37.29) < D. bulbifera L. (46.81) < M. atropurpureum (55.91). The obtained results may support their use in herbal medicine and as baseline data for the development of new drugs and standardized phytomedicines.

Keywords: antioxidant properties, total flavonoids, total phenolics, creeping herbal weeds

Procedia PDF Downloads 709

Authors: Maria P. Geneva, Ira V. Stancheva, Marieta G. Hristozkova, Roumiana D. Vasilevska-Ivanova, Mariana T. Sichanova, Janet R. Mincheva

Abstract:

Hyssopus officinalis L., Lamiaceae, commonly called hyssop, is an aromatic, semi-evergreen, woody-based, shrubby perennial plant. Hyssop is a good expectorant and antiviral herb commonly used to treat respiratory conditions such as influenza, sinus infections, colds, and bronchitis. Most of its medicinal properties are attributed to the essential oil of hyssop. The study was conducted to evaluate the influence of inoculation with arbuscular mycorrhizal fungi of in vitro propagated hyssop plants on the: activities of antioxidant enzymes superoxide dismutase, catalase, guaiacol peroxidase and ascorbate peroxidase; accumulation of non-enzymatic antioxidants total phenols and flavonoid, water-soluble soluble antioxidant metabolites expressed as ascorbic acid; the antioxidant potential of hyssop methanol extracts assessed by two common methods: free radical scavenging activity using free stable radical (2,2-diphenyl-1-picrylhydrazyl, DPPH• and ferric reducing antioxidant power FRAP in flowers and leaves. The successfully adapted to field conditions in vitro plants (survival rate 95%) were inoculated with arbuscular mycorrhizal fungi (Claroideoglomus claroideum, ref. EEZ 54). It was established that the activities of enzymes with antioxidant capacity (superoxide dismutase, catalase, guaiacol peroxidase and ascorbate peroxidase) were significantly higher in leaves than in flowers in both control and mycorrhized plants. In flowers and leaves of inoculated plants, the antioxidant enzymes activity were lower than in non-inoculated plants, only in SOD activity, there was no difference. The content of low molecular metabolites with antioxidant capacity as total phenols, total flavonoids, and water soluble antioxidants was higher in inoculated plants. There were no significant differences between control and inoculated plants both for FRAP and DPPH antioxidant activity. According to plant essential oil content, there was no difference between non-inoculated and inoculated plants. Based on our results we could suggest that antioxidant capacity of in vitro propagated hyssop plant under conditions of cultivation is determined by the phenolic compounds-total phenols and flavonoids as well as by the levels of water-soluble metabolites with antioxidant potential. Acknowledgments: This study was conducted with financial support from National Science Fund at the Bulgarian Ministry of Education and Science, Project DN06/7 17.12.16.

Keywords: antioxidant enzymes, antioxidant metabolites, arbuscular mycorrhizal fungi, Hyssopus officinalis L.

Procedia PDF Downloads 290

Authors: Ouzid Yasmina, Aiche-Iratni Ghenima, Harchaoui Lina, Saadoun Noria, Houali Karim

Abstract:

Medicinal plants are an important source of bioactive molecules with biological activities such as anticancer, antioxidant, anti-inflammatory, antibacterial, antimitotic.... These molecules include alkaloids, polyphenols and terpenes. The latter can be extracted by different solvents, namely: water, ethanol, methanol, butanol, acetone... This is why it seemed interesting to us to evaluate in vitro the antimitotic and genotoxic effect of these secondary metabolites contained in the aqueous extract of the leaves of Peganum harmala L. by the Allium cepa L. test on meristematic cells by calculating the mitotic parameters (The mitotic index, the aberration index and the limit value of cytotoxicity).A spectrophotometric determination of secondary metabolites, namely alkaloids and flavonoids in the aqueous extract of this essence, was performed. As a result, the alkaloid content is estimated to be 28.42 μg EC/mg extract, and the flavonoid content is 12.52 μg EQ/mg extract. The determination of the mitotic index revealed disturbances in cell division with a highly significant difference between the negative control (distilled water) and the different samples (aqueous extracts, colchicine and quecetin). The exposure of meristematic cells to our samples resulted in a large number of chromosomal, nuclear and cellular aberrations with an aberration index reaching 16.21±1.28% for the 4mg/ml aqueous extract and 11.71±3.32% for the 10mg/ml aqueous extract. The limit value of cytotoxicity revealed that our samples are sublethal on Allium cepa L. meristematic cells.

Keywords: allium cepa l., antimitotic and genotoxic effect, aqueous leaf extract, laghouat (algeria), peganum harmala l., secondary metabolites

Procedia PDF Downloads 58

Authors: Chi-Jung Chung, Chao-Hsiang Chang, Chiu-Shong Liu, Sheng-Wei Li, Mu-Chi Chung, Ting-Jie Wen, Hui-Ling Lee

Abstract:

Cigarette smoke contains volatile organic compounds (VOCs), such as acrylamide, 1,3-butadiene, and benzene. This study aimed to explore the associations between the urinary levels of cotinine and VOC metabolites and the risk of urothelial carcinoma (UC). A hospital-based case–control study involving two groups matched on the basis of age ( ± 3 years) and gender was designed. UC was clinically diagnosed through urological examinations and pathologically verified. Smoking-related information was collected through questionnaires and face-to-face interviews with all study participants. Urine samples were collected for the analysis of the urinary levels of VOC metabolites, cotinine, and 8-hydroxydeoxygua- nosine (8-OHdG), which was selected as a proxy of oxidative stress. Multiple logistic regressions were applied to estimate the risk of UC. The urinary cotinine and 8-OHdG levels of the UC group were higher than those of the control group. The urinary levels of VOC metabolites, including N-acetyl-S-(2-carbamoylethyl)-L-cysteine (AAMA), N- acetyl-S-(2-carbamoyl-2-hydroxyethyl)-L-cysteine, N-acetyl-S- (4- hydroxy-2-buten-1-yl)-Lcysteine-3, trans, trans-muconic acid (t,t- MA), and S-phenylmercapturic acid (SPMA) increased as the urinary levels of cotinine increased. Relevant dose-response relationships between the risk of UC risk and the urinary levels of AAMA , t,t-MA, SPMA, and 8-OHdG were found after adjusting for potential risk factors. The UC risk of participants with high urinary levels of cotinine, AAMA, t,t-MA, SPMA, and 8-OHdG were 3.5–6-fold higher than those of other participants. Increased urinary levels of VOC metabolites were associated with smoking-related UC risk. The development of UC should be explored in large-scale in vitro or in vivo studies with the repeated measurement of VOC metabolites.

Keywords: volatile organic compound, urothelial carcinoma, cotinine, 8-hydroxydeoxyguanosine

Procedia PDF Downloads 117

Authors: Miguel Mellado, Cecilia Zapata

Abstract:

Goats can be exploited in harsh environments due to their capacity to adjust to limited quantity and quality forage sources. In these environments, leguminous trees can be used as supplementary feeds as foliage and fruits of these trees can contribute to maintain or improve production efficiency in ruminants. The objective of this study was to determine the nutritional value of three leguminous trees heavily selected by goats in a xerophyllous shrubland. Chemical composition and in vitro dry matter disappearance (IVDMD) of leaves and pods from leucaena (Leucaena leucocephala), mesquite (Prosopis laevigata) and huisache (Acacia farnesiana) is presented. Crude protein (CP) ranged from 17.3% for leaves of huisache to 21.9% for leucaena. The neutral detergent fiber (NDF) content ranged from 39.0 to 40.3 with no difference among fodder threes. Across tree species, mean IVDMD was 61.6% for pods and 52.2% for leaves. IVDMD for leaves was highest (P < 0.01) for leucaena (54.9%) and lowest for huisache (47.3%). Condensed tannins in an acetonic extract were highest for leaves of huisache (45.3 mg CE/g DM) and lowest for mesquite (25.9 mg CE/g DM). Pods and leaves of huisache presented the highest number of secondary metabolites, mainly related to hydrobenzoic acid and flavonols; leucaena and mesquite presented mainly flavonols and anthocyanins. It was concluded that leaves and pods of leucaena, mesquite and huisache constitute valuable forages for ruminant livestock due to its low fiber, high CP levels, moderate in vitro fermentation characteristics and high mineral content. Keywords: Fodder tree; ruminants; secondary metabolites; minerals; tannins

Keywords: fodder tree, ruminants, secondary metabolites, minerals, tannins

Procedia PDF Downloads 115

Authors: Fadila Mohamed Mahmoud., Derkaoui I., Krimi Z.

Abstract:

Date palm is a plant which presents a very good adaptation to the difficult conditions of the environment in particular to the drought and saline stress even at high temperatures. This adaptation is related on the biology of the plant and to the presence of a microflora endophyte which live inside its tissues. Fifteen endophytics fungi isolated from date palm were tested in vitro in the presence of various NaCl concentrations to select halotolerantes isolates. These same endophytes were tested for their colonizing capacity by the description of the production of secondary metabolites more particularly the enzymes (pectinases, proteases, and phosphorylases), and the production of antibiotics and growth hormones. Significant difference was observed between the isolates with respect to the tests carried out.

Keywords: Date palm, Halotolerantes, endophyte, Secondary metabolites.

Procedia PDF Downloads 487

Authors: Marco Clericuzio, Alfredo Vizzini

Abstract:

The secondary metabolites of Russulales (one of the main orders of phylum Basidiomycota), have been studied. They are mainly terpenoids, with sesquiterpenes being the most common ones, but also triterpenoids and prenylated phenols have been isolated. We found that classes of specific compounds seem to be often allied to systematic groupings, so that they may have chemotaxonomic significance. Moreover, the ecological implications of such metabolites, as well as their biological activities, are discussed. Lately, we have focused our attention on the anti-arthropod activity of Russula metabolites, in particular on the toxicity against mites and other crop pests.

Keywords: chemotaxonomy, fungi, insecticidal activity, russulales, terpenoids

Procedia PDF Downloads 129

Authors: Raviv Harris, Maggie Levy

Abstract:

Natural product-based pesticides may serve as an alternative to the traditional synthetic pesticides, which have a potentially damaging effect, both to human health and for the environment. Along with plants, microorganisms are a prospective source of such biological pesticides. A unique and active strain of P. aphidis (designated isolate L12, Israel 2004), an epiphytic and non-pathogenic basidiomycete yeast, was isolated in our lab from strawberry leaves. P. aphidis L12 secretions were found to inhibit broad range of plant pathogens. This work demonstrates that metabolites isolated from the biocontrol agent P. aphidis (isolate L12) can inhibit varied fungal and bacterial phytopathogens. Biologically active metabolites were extracted from P. aphidis biomass, using the organic solvent ethyl acetate. The antimicrobial activity of the extract was demonstrated, both in vitro and in planta. Using disk diffusion assays, the following inhibition zones were obtained: 43cm² for Pseudomonas syringae pv. tomato, 28.5cm² for Xanthomonas campestris pv. vesicatoria, 59cm² for Clavibacter michiganensis subsp. michiganensis, 34cm² for Erwinia amylovora and 34cm² for Agrobacterium tumefaciens. Additionally, strong inhibitory activity of the extract against fungi mycelial growth was established, with IC₅₀ values of 606µg ml⁻¹ for Botrytis cinerea, 221µg ml⁻¹ for Pythium spp., 519µg ml⁻¹ for Rhizoctonia solani, 455µg ml⁻¹ for Sclerotinia sclerotiorum, 2270µg ml⁻¹ for Fusarium oxysporum f. sp. lycopersici, and 2038µg ml⁻¹ for Alternaria alternata. The results of the in planta experiments demonstrated a dose-dependent reduction in disease infection. Significant inhibition of B. cinerea lesions on tomato plants was obtained when a spore suspension of this pathogen was treated with extract concentrations higher than 4.2mg ml⁻¹. Concentration of 7mg ml⁻¹ caused a reduction of over 95% in the lesion size of B. cinerea on tomato plants. The strong antimicrobial activity demonstrated both in vitro and in planta against varied phytopathogens, may indicate that the extracted antimicrobial metabolites have potential to serve as natural pesticides in the field.

Keywords: antimicrobial, B. cinerea, metabolites, natural pesticides, P. aphidis

Procedia PDF Downloads 209

Authors: Rami Al Jafar, Paul Elliott, Konstantinos K. Tsilidis, Abbas Dehghan

Abstract:

Although Ramadan fasting is a ritual that is practiced every year by millions of Muslims, studies on Ramadan fasting are still scarce. To the best of our knowledge, none of the previous studies comprehensively explored the effect on the metabolic profile. In London Ramadan Fasting Study, blood samples were collected from 81 participants before and 10-14 days after Ramadan. Blood samples were analysed using nuclear magnetic resonance (NMR) spectroscopy which covers 249 metabolites. Mixed-effects models were used to analyse collected data and assess the effect of Ramadan fasting on the metabolic profile. Our observational study involved 85 individuals with a mean age of 45.2 years, and 53.1% of them were males. After Ramadan, forty metabolites were affected significantly by Ramadan fasting. Most of these metabolites were metabolites ratios (24), and the rest were three Glycolysis, three ketone bodies, nine Lipoprotein subclasses and one inflammation marker. This study suggests that Ramadan fasting is significantly associated with changes in the metabolic profile. However, the changes are assumed to be temporary, and the long-term effect of these changes is unknown.

Keywords: metabolic profile, Ramadan fasting, metabolites, intermittent fasting

Procedia PDF Downloads 139

Authors: Jackson Ishara, Ariel Buzera, Gustave N. Mushagalusa, Ahmed R. A. Hammam, Judith Munga, Paul Karanja, John Kinyuru

Abstract:

Numerous mushroom bioactive metabolites, including polysaccharides, eritadenine, lignin, chitosan, mevinolin, and astrakurkurone have been studied in life-threatening conditions and diseases such as diabetes, cardiovascular, hypertension, cancer, DNA damage, hypercholesterolemia, and obesity attempting to identify natural therapies. These bioactive metabolites have shown potential as antiviral and immune system strengthener natural agents through diverse cellular and physiological pathways modulation with no toxicity evidence, widely available, and affordable. In light of the emerging literature, this paper compiles the most recent information describing the molecular mechanisms that underlie the nutraceutical potentials of these mushroom metabolites suggesting their effectiveness if combined with existing drug therapies. The findings raise hope that these mushroom bioactive metabolites may be utilized as natural therapies considering their therapeutic potential while anticipating further research designing clinical trials and developing new drug therapies while encouraging their consumption as a natural adjuvant in preventing and controlling life-threatening conditions and diseases.

Keywords: bioactive metabolites, food functionality, health-threatening conditions, mushrooms, nutraceutical

Procedia PDF Downloads 72

Authors: Maged El-Sayed Mohamed

Abstract:

Plant tissue culture practices are presented nowadays as the most promising substitute to a whole plant in the terms of secondary metabolites production. They offer the advantages of high production, tunability and they have less effect on plant ecosystems. Lantana camara is a weed, which is common all over the world as an ornamental plant. Weeds can adapt to any type of soil and climate due to their rich cellular machinery for secondary metabolites’ production. This characteristic is found in Lantana camara as a plant of very rich diversity of secondary metabolites with no dominant class of compounds. Aim: This trait has encouraged the author to develop tissue culture experiments for Lantana camara to be a platform for production and manipulation of secondary metabolites through biotransformation. Methodology: The plant was collected in its flowering stage in September 2014, from which explants were prepared from shoot tip, auxiliary bud and leaf. Different types of culture media were tried as well as four phytohormones and their combinations; NAA, 2,4-D, BAP and kinetin. Explants were grown in dark or in 12 hours dark and light cycles at 25°C. A metabolic profile for the produced callus was made and then compared to the whole plant profile. The metabolic profile was made using GC-MS for volatile constituents (extracted by n-hexane) and by HPLC-MS and capillary electrophoresis-mass spectrometry (CE-MS) for non-volatile constituents (extracted by ethanol and water). Results: The best conditions for the callus induction was achieved using MS media supplied with 30 gm sucrose and NAA/BAP (1:0.2 mg/L). Initiation of callus was favoured by incubation in dark for 20 day. The callus produced under these conditions showed yellow colour, which changed to brownish after 30 days. The rate of callus growth was high, expressed in the callus diameter, which reached to 1.15±0.2 cm in 30 days; however, the induction of callus delayed for 15 days. The metabolic profile for both volatile and non-volatile constituents of callus showed more simple background metabolites than the whole plant with two new (unresolved) peaks in the callus’ nonvolatile constituents’ chromatogram. Conclusion: Lantana camara callus production can be itself a source of new secondary metabolites and could be used for biotransformation studies due to its simple metabolic background, which allow easy identification of newly formed metabolites. The callus production gathered the simple metabolic background with the rich cellular secondary metabolite machinery of the plant, which could be elicited to produce valuable medicinally active products.

Keywords: capillary electrophoresis-mass spectrometry, gas chromatography, metabolic profile, plant tissue culture

Procedia PDF Downloads 344

Authors: Kristine Vike-Jonas, Susana V. Gonzalez, Olav L. Bakkerud, Karoline S. Gjelstad, Shazia N. Aslam, Øyvind Mikkelsen, Alexandros Asimakopoulos

Abstract:

P-hydrobenzoic acid esters (parabens), paraben metabolites, and triclocarban (TCC) are a group of synthetic antimicrobials classified as endocrine disrupting chemicals (EDCs) and emerging pollutants. The aim of this study was to investigate the levels of these compounds in sediment near the effluent of a wastewater treatment plant (WWTP) in the Trondheim Fjord, Norway. Paraben, paraben metabolites, and TCC are high volume production chemicals that are found in a range of consumer products, especially pharmaceuticals and personal care products (PCPs). In this study, six parabens (methyl paraben; MeP, ethyl paraben; EtP, propyl paraben; PrP, butyl paraben; BuP, benzyl paraben; BezP, heptyl paraben; HeP), four paraben metabolites (4-hydroxybenzoic acid; 4-HB, 3,4-dihydroxybenzoic acid; 3,4-DHB, methyl protocatechuic acid; OH-MeP, ethyl protocatechuic acid; OH-EtP) and TCC were determined by ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) in 64 sediment samples from 10 different locations outside Trondheim, Norway. Of these 11 target analytes, four were detected in 40 % or more of the samples. The sum of six parabens (∑Parabens), four paraben metabolites (∑Metabolites) and TCC in sediment ranged from 4.88 to 11.56 (mean 6.81) ng/g, 52.16 to 368.28 (mean 93.89) ng/g and 0.53 to 3.65 (mean 1.50) ng/g dry sediment, respectively. Pearson correlation coefficients indicated that TCC was positively correlated with OH-MeP, but negatively correlated with 4-HB. To the best of the author’s knowledge, this is the first time parabens, paraben metabolites and TCC have been reported in the Trondheim Fjord.

Keywords: parabens, liquid chromatography, sediment, tandem mass spectrometry

Procedia PDF Downloads 98

Authors: Sonu Gandhi, Neena Capalash, Prince Sharma, C. Raman Suri

Abstract:

A sensitive chemiluminescence immunoassay (CIA) for heroin and its major metabolites is reported. The method is based on the competitive reaction of horseradish peroxidase (HRP)-labeled anti-MAM antibody and free drug in spiked urine samples. A hapten-protein conjugate was synthesized by using acidic derivative of monoacetyl morphine (MAM) coupled to carrier protein BSA and was used as an immunogen for the generation of anti-MAM (monoacetyl morphine) antibody. A high titer of antibody (1:64,0000) was obtained and the relative affinity constant (Kaff) of antibody was 3.1×107 l/mol. Under the optimal conditions, linear range and reactivity for heroin, mono acetyl morphine (MAM), morphine and codeine were 0.08, 0.09, 0.095 and 0.092 ng/mL respectively. The developed chemiluminescence inhibition assay could detect heroin and its metabolites in standard and urine samples up to 0.01 ng/ml.

Keywords: heroin, metabolites, chemiluminescence immunoassay, horse radish peroxidase

Procedia PDF Downloads 240

Authors: Uftah Ali M. Shushni, Viola Stuppec, Ulrike Lindequist

Abstract:

Because of the evolving resistance of microorganisms to existing antibiotics, there is an increasing need for new antibiotics not only in human but also in veterinary medicine. As part of our ongoing work on the secondary metabolites produced by marine fungi, the organic extract of the culture filtrate of an Ascomycetous fungus, which was found on driftwood collected from the coast of the Greifswalder Bodden, Baltic Sea, Germany displayed antimicrobial activity against some fish and human pathogenic bacteria. Bioactivity-guided column chromatographic separation led to the isolation of 6-Deoxybostrycoidin. The structure was determined from the interpretation of spectroscopic data (UV, MS, and NMR). 6-Deoxybostrycoidin exhibited in vitro activity against Bacillus subtilis, Staphylococcus aureus and Flexibacter maritimus with minimal inhibitory concentrations of 25, 12.5 and 12.5 μg/ml respectively.

Keywords: marine fungi, fish pathogenic bacteria, microorganism, medicine

Procedia PDF Downloads 502

Authors: Anna Perrone, Federico Martinelli

Abstract:

Olive fruit is well-known for the high content in secondary metabolites with high interest at nutritional, nutraceutical, antioxidant, and healthy levels. The content of secondary metabolites in olive at harvest may be affected by different water regimes, with significant effects on olive oil composition and quality and, consequently, on its healthy and nutritional features. In this work, a summary of several research studies dealing with the biosynthesis of healthy and nutraceutical metabolites of the secondary metabolism in olive fruit will be reported. The phytochemical findings have been correlated with the expression of key genes involved in polyphenol, terpenoid, and carotenoid biosynthesis and metabolism in response to different development stages and water regimes. Flavonoids were highest in immature fruits, while anthocyanins increased at ripening. In epicarp tissue, this was clearly associated with an up-regulation of the UFGT gene. Olive fruits cultivated under different water regimes were analyzed by metabolomics. This method identified several hundred metabolites in the ripe mesocarp. Among them, 46 were differentially accumulated in the comparison between rain-fed and irrigated conditions. Well-known healthy metabolites were more abundant at a higher level of water regimes. Increased content of polyphenols was observed in the rain-fed fruit; particularly, anthocyanin concentration was higher at ripening. Several secondary metabolites were differentially accumulated between different irrigation conditions. These results showed that these metabolic approaches could be efficiently used to determine the effects of agronomic treatments on olive fruit physiology and, consequently, on nutritional and healthy properties of the obtained extra-virgin olive oil.

Keywords: olea europea, anthocyanins, polyphenols, water regimes

Procedia PDF Downloads 117

Authors: Changde Zhang, Qiang Zhang, Shilong Zheng, Jiawang Liu, Shanchun Guo, Qiu Zhong, Guangdi Wang

Abstract:

Fulvestrant was approved by FDA to treat breast cancer as a selective estrogen receptor downregulator (SERD) with intramuscular injection administration. ZB716, a fulvestarnt-3 boronic acid, is an SERD with comparable anticancer effect to fulvestrant, but could produce good pharmacokinetic properties under oral administration with mice or rat models. To understand why ZB716 produced much better oral bioavailability, it was proposed that the boronic acid blocked the phase II direct biotransformation with the hydroxyl group on the 3 position of the aromatic ring on fulvestrant. In this study, ZB716 or fulvestrant was incubated with human liver microsome and oxidation cofactor NADPH in vitro. Their metabolites after oxidation were profiled with the Q-Exactive, a high-resolution mass spectrometer. The result showed that ZB716 blocked the forming of hydroxyl groups on its benzene ring except for the oxidation of C-B bond forming fulvestrant in its metabolites, and the concentration of fulvestrant with one more hydroxyl group found in the metabolites from incubation with fulvestrant was about 34 fold high as that formed from incubation with ZB716. Compared to fulvestrant, ZB716 is expected to be much difficult to be further bio-transformed into more hydrophilic compounds, to be difficult excreted out of blood system, and to have longer residence time in blood, which can lead to higher oral bioavailability. This study provided evidence to explain the high bioavailability of ZB716 after oral administration from the perspective of its difficulty of oxidation, a phase I biotransformation, on positions on its aromatic ring.

Keywords: biotransformation, fulvestrant, metabolite profiling, ZB716

Procedia PDF Downloads 229

Authors: Sajjad ur Rahman, Mariam Azam, Mukarram Bashir, Seemal Javaid, Aoun Muhammad, Muhammad Tahir, Jawad, Hannan Khan, Muhammad Zohaib

Abstract:

Partially fermented soya hulls and wheat bran through Saccharomyces cerevisiae (DL-22 S/N) substantiated as a natural source for quality milk production. Saccharomyces cerevisiae (DL-22 S/N) were grown under in-vivo conditions and processed through two-step fermentation with substrates. The extra pure metabolites (XPM) were dried and processed for maintaining 1mm mesh size particles for supplementation of pelleted feed. Two groups of a cow (Holstein Friesian) having 8 animals of similar age and lactation were given the experimental concentrates. Group A was fed daily with 12gm of XPM and 22% protein-pelleted feed, while Group B was provided with no metabolites in their feed. In thirty-nine days of trial, improvement in the overall health, body score, milk protein, milk fat, ash, and solid not fat (SNF), yield, and incidence rate of mastitis was observed. The collected data revealed an improvement in milk production of 2.02 liter/h/d. However, a reduction (3.75%) in the milk fats and an increase in the milk SNF was around 0.58%. The ash content ranged between 6.4-7.5%. The incidence of mastitis was reduced to less than 2%.

Keywords: microbial metabolites, Saccharomyces cerevisiae, milk production, fermentation, post-biotic metabolites, immunity

Procedia PDF Downloads 63

Authors: O. Kahraman, M. S. Alatas, O. B. Citil

Abstract:

In determination of values of feeds which, are used in ruminant nutrition, different methods are used like in vivo, in vitro, in situ or in sacco. Generally, the most reliable results are taken from the in vivo studies. But because of the disadvantages like being hard, laborious and expensive, time consuming, being hard to keep the experiment conditions under control and too much samples are needed, the in vitro techniques are more preferred. The most widely used in vitro techniques are two-staged digestion technique and gas production technique. In vitro gas production technique is based on the measurement of the CO2 which is released as a result of microbial fermentation of the feeds. In this review, the factors affecting the results obtained from in vitro gas production technique (Hohenheim Feed Test) were discussed. Some factors must be taken into consideration when interpreting the findings obtained in these studies and also comparing the findings reported by different researchers for the same feeds. These factors were discussed in 3 groups: factors related to animal, factors related to feeds and factors related with differences in the application of method. These factors and their effects on the results were explained. Also it can be concluded that the use of in vitro gas production technique in feed evaluation routinely can be contributed to the comprehensive feed evaluation, but standardization is needed in this technique to attain more reliable results.

Keywords: In vitro, gas production technique, Hohenheim feed test, standardization

Procedia PDF Downloads 558

Authors: Valentina L. Kouznetsova, Jonathan W. Wang, Igor F. Tsigelny

Abstract:

Metabolomics has become a rising field of research for various diseases, particularly cancer. Increases or decreases in metabolite concentrations in the human body are indicative of various cancers. Further elucidation of metabolic pathways and their significance in cancer research may greatly spur medicinal discovery. We analyzed the metabolomics profiles of lung cancer. Thirty-three metabolites were selected as significant. These metabolites are involved in 37 metabolic pathways delivered by MetaboAnalyst software. The top pathways are glyoxylate and dicarboxylate pathway (its hubs are formic acid and glyoxylic acid) along with Citrate cycle pathway followed by Taurine and hypotaurine pathway (the hubs in the latter are taurine and sulfoacetaldehyde) and Glycine, serine, and threonine pathway (the hubs are glycine and L-serine). We studied interactions of the metabolites with the proteins involved in cancer-related signaling networks, and developed an approach to metabolomics biomarker use in cancer diagnostics. Our analysis showed that a significant part of lung-cancer-related metabolites interacts with main cancer-related signaling pathways present in this network: PI3K–mTOR–AKT pathway, RAS–RAF–ERK1/2 pathway, and NFKB pathway. These results can be employed for use of metabolomics profiles in elucidation of the related cancer proteins signaling networks.

Keywords: cancer, metabolites, metabolic pathway, signaling pathway

Procedia PDF Downloads 368