Search results for: in vitro rooting

Authors: A. N.Ukwuani, A. K. Abdulfatah

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G. crenata is used locally for the treatment of fractured bones, wound healing and inflammatory conditions. In vitro and in vivo antioxidant activity of hydromethanolic extracts of the leaves of G. crenata were assessed. The phytochemical analysis shows the presence of phenols, flavonoids, saponins, cardiac glycosides and tannins. An in vitro quantitative analysis of phenols, flavonoids and tannins respectively were (164±1.20, 199±0.88 and 88.67±0.88 mg/100g FW). In vivo studies of hydromethanolic extract demonstrated a dose dependent increase in hepatic superoxide dismutase (1.14±0.14, 2.13±0.11, 2.55±0.11 U/mg Protein) with improvement in hepatic glutathione (6.98±0.42, 8.91±0.37, 11.07±0.46 µM/mg Protein) and Catalase (4.47±0.05, 6.24±0.02, 7.17±0.04 U/mg Protein) and Total protein (6.18±0.08, 6.69±0.18, 7.27±0.16 mg/ml) respectively at 100-300mg/kg body weight Grewia crenata leaves when compared to the control and standard drug. It can be concluded from the present findings of that G. crenata leaves possess antioxidant potential.

Keywords: Grewia crenata, antioxidant, hydromethanolic extract, in vivo, in vitro

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Authors: Kehinde T. Kareem

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The need to increase the production of cucumber has led to the use of inorganic fertilizers. This chemical affects the ecological balance of nature by increasing the nitrogen and phosphorus contents of the soil. Surface runoffs into rivers and streams cause eutrophication which affects aquatic organisms as well as the consumers of aquatic animals. Therefore, this study was carried out in the screenhouse to investigate the use of a plant growth-promoting fungus; Trichoderma longibrachiatum for the growth promotion of conventional and in-vitro propagated Ashley and Marketmoor cucumber. Before planting of cucumber, spore suspension (108 cfu/ml) of Trichoderma longibrachiatum grown on Potato dextrose agar (PDA) was inoculated into the soil. Fruits were evaluated for the presence of Trichoderma longibrachiatum using a species-specific primer. Results revealed that the highest significant plant height produced by in-vitro propagated Ashley was 19 cm while the highest plant height of in-vitro propagated Marketmoor was 19.67 cm. The yield of the conventional propagated Ashley cucumber showed that the number of fruit/plant obtained from T. longibrachiatum-fertilized plants were significantly more than those of the control. The in-vitro Ashely had 7 fruits/plant while the control produced 4 fruits/plant. In-vitro Marketmoor had ten fruits/plant, and the control had a value of 4 fruits/plant. There were no traces of Trichoderma longibrachiatum genes in the harvested cucumber fruits. Therefore, the use of Trichoderma longibrachiatum as a plant growth-promoter is safe for human health as well as the environment.

Keywords: biofertilizer, cucumber, genes, growth-promoter, in-vitro, propagation

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Authors: Titin Handayani, Endang Yuniastuti

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Kepuh (Sterculia foetida L.) is a potential plant contain mainly oil seeds that can be used as a source of alternative bioenergy and medicine. The main problem of kepuh cultivation is the limited supply of seed plants. Seeds development were very easy, but to produce fruit have to wait for approximately 5 years. The objective of this research was to obtain kepuh plants through direct in vitro regeneration. Hypocotyls and shoot tips explants were excised from sterile germinated seedlings and placed on shoot induction medium containing basal salts of Murashige and Skoog (MS) and various concentrations of plant growth regulators. The results showed that shoots induction from the apical and axillary buds on MS medium + 1.5 and 2 mg/L BAP and 0.5 and 1 mg/L IAA was growth very slowly. Increasing of BAP concentrations was increased shoot formation. The first subcultures were increased the rate of shoots growth on MS medium supplemented with 2 mg/L BAP and 0.5 mg/L IAA. The second of shoots subculture on MS medium + 1.5 to 2 mg/L BAP + 0.5 mg/L IAA was increased the number of shoots up to 4.8 in average. The best medium of shoots elongation were MS + 1 mgL-1 kinetin + 5 mg/L GA3. The highest percentage of roots (65%) occurred on MS medium with 5 mg/L IBA which average number of roots was 3.1. High percentages of survival and plants of normal appearance were obtained after five weeks of acclimatization.

Keywords: Kepuh, Sterculia foetida L, shoot multiplication, rooting, acclimatization, bioenergy, medicine

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Authors: Van Huong Do, Simon Walton, German Amaya, Madeline Batsiokis, Sally Catt, Andrew Taylor-Robinson

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Identification of the most suitable stages of bovine in vitro-derived blastocysts (early, expanded and hatching) prior to vitrification is a straightforward process that facilitates the decision as to which blastocyst stage to use for transfer of fresh and vitrified embryos. Research on in vitro evaluation of suitable stages has shown that the more advanced developmental stage of blastocysts is recommended for fresh embryo transfer while the earlier stage is proposed for embryo transfer following vitrification. There is, however, limited information on blastocyst stages using in vivo assessment. Hence, the aim of the present study was to determine the optimal stage of a blastocyst for vitrification and embryo transfer through a two-step procedure of embryo transfer followed by pregnancy testing at 35, 60 and 90 days of pregnancy. 410 good quality oocytes aspirated by the ovum pick-up technique from 8 donor cows were subjected to in vitro embryo production, vitrification and embryo transfer. Good quality embryos were selected, subjected to vitrification and embryo transfer. Subsequently, 77 vitrified embryos at different blastocyst stages were transferred to synchronised recipient cows. The overall cleavage and blastocyst rates of oocytes were 68.8% and 41.7%, respectively. In addition, the fertility and blastocyst production of 6 bulls used for in vitro fertilization was examined and shown to be statistically different (P<0.05). Results of ongoing pregnancy trials conducted at 35 days, 60 days and 90 days will be discussed. However, preliminary data indicate that individual bulls demonstrate distinctly different fertility performance in vitro. Findings from conception rates would provide a useful tool to aid selection of bovine in vitro-derived embryos for vitrification and embryo transfer in commercial settings.

Keywords: blastocyst, embryo transfer, in vitro-derived embryos, ovum pick-up, vitrification

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Authors: Hadriana Bansi, Elizabeth Wina, Toto Toharmat

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Acacia villosa is thornless shrub legume which contents high crude protein. However, the utilization of A. villosa as ruminant feed is limited by its secondary compounds. The aim of this article is to find out the maximum of substitution A. villosa in sheep ration. The nutritional evaluation consisted of in vitro two stages, in vivo, and in vitro gas production trials. The secondary compounds of A. villosa also were analyzed. Evaluating digestibility of increasing level of substitution A. villosa replacing Pennisetum purpureum was using in vitro two stages. The substitution of 30% A. villosa was compared to 100% P. purpureum by in vitro gas production technique and in vivo digestibility. The results of two stages in vitro showed that total phenol, condensed tannin, and non-protein amino acid (NPAA) were high. Substitution 15% A. villosa reached the highest digestibility for both dry matter (DM) and crude protein (CP) which were 67% and 86% respectively, but it was shown that DM and CP digestibility of substitution 30% of A. villosa was still high which were 61.82% and 75-67% respectively. The pattern of gas production showed that first 8 hours total gas production substitution of 30% A. villosa was higher than 100% P. purpureum and declined after 10 hours incubation. In vivo trials showed that substitution of 30% A. villosa significantly increased CP intake, CP digestibility, and nitrogen retention. It can be concluded that substitution A. villosa until 30% still gave the good impact even though it has high secondary compounds.

Keywords: Acacia villosa, digestibility, gas production, secondary compounds

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Authors: S. Vinotha, I. Thabrew, S. Sri Ranjani

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Hot aqueous and methanol extracts of the two selected herbal medicines such are Vellarugu Chooranam (V.C) and Amukkirai Chooranam (A.C) were examined for total phenolic and flavonoid contents and in-vitro antioxidant activity using four different methods. The total phenolic and flavonoid contents in methanol extract of V.C were found to be higher (44.41±1.26 mg GAE⁄g; 174.44±9.32 mg QE⁄g) than in the methanol extract of A.C (20.56±0.67 mg GAE⁄g;7.21±0.85 mg QE⁄g). Hot methanol and aqueous extracts of both medicines showed low antioxidant activity in DPPH, ABTS, and FRAP methods and Iron chelating activity not found at highest possible concentration. V.C contains higher concentrations of total phenolic and flavonoid contents than A.C and can also exert greater antioxidant activity than A.C, although the activities demonstrated were lower than the positive control Trolox. The in-vitro antioxidant activity was not related with the total phenolic and flavonoid contents of the methanol and aqueous extracts of both herbal medicines (A.C and V.C).

Keywords: activity, different extracts, herbal medicines, in-vitro antioxidant

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Authors: Reine Suci Wulandari, Rosa Suryantini

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Albizia (Paraserianthes falcataria) is a woody plant species that has a high economic value and multifunctional. Albizia is important timber, medicinal plants and can also be used as a plant to rehabilitate critical lands. The demand value of Albizia is increased so that the large quantities and high quality of seeds are required. In vitro propagation techniques are seed propagation that can produce more seeds and quality in a short time. In vitro cultures require growth regulators that can be obtained from biological agents such as endophytic fungi. Endophytic fungi are micro fungi that colonize live plant tissue without producing symptoms or other negative effects on host plants and increase plant growth. The purposes of this research were to isolate and identify endophytic fungi isolated from the root of Albizia and to study the effect of endophytic fungus on the growth of Albizia in vitro. The methods were root isolation, endophytic fungal identification, and inoculation of endophytic fungi to Albizia plants in vitro. Endophytic fungus isolates were grown on PDA media before being inoculated with Albizia sprouts. Incubation is done for 4 (four) weeks. The observed growth parameters were live explant percentage, percentage of explant shoot, and percentage of explant rooted. The results of the research showed that 6 (six) endophytic fungal isolates obtained from the root of Albizia, namely Aspergillus sp., Verticillium sp, Penicillium sp., Trichoderma sp., Fusarium sp., and Acremonium sp. Statistical analysis found that Trichoderma sp. and Fusarium sp. affect in vitro growth of Albizia. Endophytic fungi from the results of this research were potential as plant growth promoting. It can be applied to increase productivity either through increased plant growth and increased endurance of Albizia seedlings to pests and diseases.

Keywords: Albizia, endophytic fungi, propagation, in vitro

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Authors: Sri Wahjuningsih, Nur Ihsan, Hadiah

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In the embryo transfer program, to address the limited production of embryos in vivo, in vitro embryo production has become an alternative approach that is relatively inexpensive. One potential source of embryos that can be developed is to use immature oocytes then conducted in vitro maturation and in vitro fertilization. However, obstacles encountered were oocyte viability mammals have very limited that it cannot be stored for a long time, so we need oocyte cryopreservation. The research was conducted to know the optimal concentration use of ethylene glycol as a cryoprotectant on oocytes freezing.Material use in this research was immature oocytes; taken from abbatoir which was aspirated from follicle with diameter 2-6 mm. Concentration ethylen glycol used were 0,5 M, I M, 1,5 M and 2M. The freezing method used was conventional method combined with a five-step protocol washing oocytes from cryoprotectant after thawing. The result showed that concentration ethylen glycol have the significant effect (P<0.05) on oocytes quality after thawing and in vitro maturation. It was concluded that concentration 1,5 M was the best concentration for freezing oocytes using conventional method.

Keywords: bovine, conventional freezing, ethylen glycol, oocytes

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Authors: Sakinah Abdullah, Rosna Mat Taha

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Antioxidants are compounds that protect cells against the damaging effects of reactive oxygen species such as singlet oxygen, superoxide, peroxyl radicals, and peroxynitrite which result in oxidative stress leading to cellular damage. Natural antioxidant are in high demand because of their potential in health promotion and disease prevention and their improved safety and consumer acceptability. Plants are rich sources of natural antioxidant. Dioscorea alata L. known as 'ubi badak' in Malaysia were well known for their antioxidant content, but this plant was seasonal. Thus, tissue culture technique was used to mass propagate this plant. In the present work, a comparative study between in vitro (from tissue culture) and in vivo (from intact plant) samples of Dioscorea alata L. for their antioxidant potential by 2,2-diphenil -1- picrylhydrazyl (DPPH) radical scavenging activity method and their total phenolic and flavonoid contents were carried out. All samples had better radical scavenging activity but in vivo samples had the strongest radical scavenging activity compared to in vitro samples. Furthermore, tubers from in vivo samples showed the greatest free radical scavenging effect and comparatively greater phenolic content than in vitro samples.

Keywords: Dioscorea alata, tissue culture, antioxidant, in vivo, in vitro, DPPH

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Authors: A. Jamsheera, F. Arjmand, D. K. Mohapatra

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Small molecules binding to specific sites along DNA molecule are considered as potential chemotherapeutic agents. Their role as mediators of key biological functions and their unique intrinsic properties make them particularly attractive therapeutic agents. Keeping in view, novel dipeptide complexes Cu(II)-Val-Pro (1), Zn(II)-Val-Pro (2), Cu(II)-Ala-Pro (3) and Zn(II)-Ala-Pro (4) were synthesized and thoroughly characterized using different spectroscopic techniques including elemental analyses, IR, NMR, ESI–MS and molar conductance measurements. The solution stability study carried out by UV–vis absorption titration over a broad range of pH proved the stability of the complexes in solution. In vitro DNA binding studies of complexes 1–4 carried out employing absorption, fluorescence, circular dichroism and viscometric studies revealed the binding of complexes to DNA via groove binding. UV–vis titrations of 1–4 with mononucleotides of interest viz., 5´-GMP and 5´-TMP were also carried out. The DNA cleavage activity of the complexes 1 and 2 were ascertained by gel electrophoresis assay which revealed that the complexes are good DNA cleavage agents and the cleavage mechanism involved a hydrolytic pathway. Furthermore, in vitro antitumor activity of complex 1 was screened against human cancer cell lines of different histological origin.

Keywords: dipeptide Cu(II) and Zn(II) complexes, DNA binding profile, pBR322 DNA cleavage, in vitro anticancer activity

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Authors: Comfort Adeoye, Anthony Eneji

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Lead pollution by mining, industrial dumping, and other anthropogenic uses are corroding the environment. Efforts being made to control it include physical, chemical and biological methods. The failure of the aforementioned methods are largely due to the fact that they are cumbersome, expensive, and not eco-friendly. Some plant species can be used for remediation of these pollutants. The objective of this work is to investigate the abilities of two native weed species to remediate two lead-polluted soils: a) Battery dumpsite and, (b) Naturally occurring lead mine. Soil samples were taken from the two sites: a) Kumapayi in Ibadan, a battery dumpsite, (b) Zamfara, a natural lead mine. Screen house experiment in Complete Randomized Design (CRD) replicated three times was carried out at I.I.T.A. Unpolluted soils were collected and polluted with various rates of lead concentrations of 0, 0.1, 0.2, and 0.5%. These were planted with weed species. Plant growth parameters were monitored for twelve weeks, after which the plants were harvested. Dry weight and plant uptake of the lead were taken. Analysis of data was carried out using, Genstat, Excel and descriptive statistics. Relative concentration of lead (Pb) in the above and below ground parts of Gomphrena celusoides revealed that a higher amount of Pb is taken up in the root compared with the shoots at different levels of Pb pollution. However, lead uptake at 0.5% > 0.2% > 0.1% > Control. In essence, phytoremediation of Gomphrena is highest at soil pollution of 0.5% and its retention is greater in the root than the shoot.In S. pyramidalis, soil retention ranges from 0.1% > 0.5% > 0.2% > control. Uptake is highest at 0.5% > 0.1% > 0.2 in stem. Uptake in leaves is highest at 0.2%, but none in the 0.5% pollution. Therefore, different plant species exhibited different accumulative mode probably due to their physiological and rooting systems. Gomphrena spp. rooting system is tap root,while that of S.pyramidalis is fibrous.

Keywords: grass, lead, phytoremediation, pollution

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Authors: Afsheen Aman, Shah Ali Ul Qader

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Dextransucrase [EC 2.4.1.5] is a glucosyltransferase that catalysis the biosynthesis of a natural biopolymer called dextran. It can catalyze the transfer of D-glucopyranosyl residues from sucrose to the main chain of dextran. This unique biopolymer has multiple applications in several industries and the key utilization of dextran lies on its molecular weight and the type of branching. Extracellular dextransucrase from Leuconostoc mesenteroides is most extensively studied and characterized. Limited data is available regarding cell-bound or intracellular dextransucrase and on the characterization of dextran produced by in-vitro reaction of intracellular dextransucrase. L. mesenteroides AA1 is reported to produce extracellular dextransucrase that catalyzes biosynthesis of a high molecular weight dextran with only α-(1→6) linkage. Current study deals with the characterization of an intracellular dextransucrase and in vitro biosynthesis of low molecular weight dextran from L. mesenteroides AA1. Intracellular dextransucrase was extracted from cytoplasm and purified to homogeneity for characterization. Kinetic constants, molecular weight and N-terminal sequence analysis of intracellular dextransucrase reveal unique variation with previously reported extracellular dextransucrase from the same strain. In vitro synthesized biopolymer was characterized using NMR spectroscopic techniques. Intracellular dextransucrase exhibited Vmax and Km values of 130.8 DSU ml-1 hr-1 and 221.3 mM, respectively. Optimum catalytic activity was detected at 35°C in 0.15 M citrate phosphate buffer (pH-5.5) in 05 minutes. Molecular mass of purified intracellular dextransucrase is approximately 220.0 kDa on SDS-PAGE. N-terminal sequence of the intracellular enzyme is: GLPGYFGVN that showed no homology with previously reported sequence for the extracellular dextransucrase. This intracellular dextransucrase is capable of in vitro synthesis of dextran under specific conditions. This intracellular dextransucrase is capable of in vitro synthesis of dextran under specific conditions and this biopolymer can be hydrolyzed into different molecular weight fractions for various applications.

Keywords: characterization, dextran, dextransucrase, leuconostoc mesenteroides

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

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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

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Authors: Guem-Jae Chung, Jin-Hui Lee, Myung-Min Oh

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Environmental control of in vitro-propagated apple plantlets is required for successful acclimation to ex vitro due to its low survival rate. This study aimed to determine the proper lighting condition for ex vitro acclimation of the apple plantlets in plant factories. In vitro-propagated M9 apple plantlets treated with pre-acclimatization for 1 week were exposed to following light treatments for additional 6 weeks; 60 μmol·m⁻²·s⁻¹ (A), 100 μmol·m⁻²·s⁻¹ (B), 140 μmol·m⁻²·s⁻¹ (C), 180 μmol·m⁻²·s⁻¹ (D), 60 μmol·m⁻²·s⁻¹ → 100 μmol·m⁻²·s⁻¹ at 2 weeks (E) or 4 weeks (F), 60 μmol·m⁻²·s⁻¹ → 100 μmol·m⁻²·s⁻¹ at 2 weeks → 140 μmol·m⁻²·s⁻¹ at 4 weeks (G) and 60 μmol·m⁻²·s⁻¹ → 140 μmol·m⁻²·s⁻¹ at 4 weeks (H). Shoot height, total leaf area, soil-plant analysis development (SPAD) value, root length, fresh and dry weights of shoots and roots were measured every 2 weeks after transplanting. In addition, the photosynthetic rate was measured at 5 weeks after transplanting. At 6 weeks after transplanting, shoot height of B was significantly higher than the other treatments. SPAD value, total leaf area and root length of B and F were relatively higher than the other treatments. Root fresh weights of B, D, F, and G were relatively higher than those in the other treatments. D induced the highest value in shoot fresh weight probably due to stem hardening, but it also resulted in shoot damage in the early stage of acclimation. Photosynthetic rate at 5 weeks after the transplanting was significantly increased as the light intensity increased. These results suggest that 100 μmol·m⁻²·s⁻¹ for 6 weeks (B) or gradually increased treatment from 60 μmol·m⁻²·s⁻¹ to 140 μmol·m⁻²·s⁻¹ at 2 weeks interval (F) were the proper lighting conditions for successful acclimation of in vitro-propagated apple plantlets. Acknowledgment: This work was supported by Korea Institute of Planning and Evaluation for Technology in Food, Agriculture, Forestry and Fisheries (IPET) through Agri-Bio industry Technology Development Program, funded by Ministry of Agriculture, Food and Rural Affairs (MAFRA) (315003051SB020).

Keywords: acclimation, in vitro-propagated apple plantlets, light intensity, plant factory

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Authors: Przewodowski Włodzimierz, Przewodowska Agnieszka, Sekrecka Danuta, Michałowska Dorota

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Colloidal metal nanoparticles are widely applied in various areas and have great potential in different biotechnological applications. Their particular properties associated with both the antiseptic, antioxidant and anti aging properties as well as ability to penetrate most of the biological barriers, synergy in the absorption of nutrients and nontoxic to plants. The properties make them potentially useful in the fast and safe production of healthy, certified starting material in the production of plants exposed to many pathogenic microorganisms causing serious diseases, significantly affecting yield and causing the economic losses. In this case it is crucial to provide appropriate conditions for the production, storage and distribution of the plant material. Therefore, the aim of the proposed research was to develop and identify the influence of four colloidal metal nanoparticles on growth and proliferation of in vitro cultures of potato (Solanum tuberosum) - one of the most economically important strategic crops in the world. The research on different varieties of potato was performed by placing the explants of the in vitro cultures on sterile Murashige and Skoog (MS) type medium. The influence of the nanocolloids was evaluated using the MS medium impregnated with the examinated nanoparticles. The vigour of growth and the rate of proliferation was examinated for 6-8 weeks with both night/day-length and temperature over the ranges 8/16 h and 20–22 °C respectively. The results of our preliminary work confirmed high usefulness of the nanocolloids in the safe production of the examinated in vitro cultures.

Keywords: colloidal metal nanoparticles, in vitro cultures, potato, propagation

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Authors: Rosna Mat Taha, Sakinah Abdullah, Sadegh Mohajer, Asmah Awal

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Lycium barbarum L. (Goji) belongs to Solanaceae family and native to some areas of China. Ethnobotanical studies have shown that this plant has been consumed by the Chinese since ancient times. It has been used as medicine in providing excellent effects on cardiovascular system and cholesterol level, besides contains high antioxidant and antidiabetic properties. In the present study, some tissue culture work has been carried out to induce callus, in vitro regeneration from various explants of Goji and also some acclimatization protocols were followed to transfer the regenerated plants to soil. The main aims being to establish high efficient regeneration system for mass production and commercialization for future uses, since the growth of this species is very limited in Malaysia. The optimum hormonal regime and the most suitable and responsive explants were identified. It was found that leaves and stems gave good responses. Murashige and Skoog’s (MS) medium supplemented with 2.0 mg/L NAA and 0.5 mg/L BAP was the best for callus induction and MS media fortified with 1.0 mg/L NAA and 1.0 mg/L BAP was optimum for in vitro regeneration. The survival rates of plantlets after acclimatization was 63±1.5 % on black soil and 50±1.3 % on mixed soil (combination of black and red soil at a ratio of 2 to 1), respectively.

Keywords: callus, acclimatization, in vitro culture, regeneration

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

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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

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Authors: Asghar Abbas, Rao Zahid Abbas, Muhammad Asif Raza, Kashif Hussain

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In the current experiment, in vitro anticoccidial potential of Vitis venifera (grape seed) extract was evaluated. For this purpose, an in vitro sporulation inhibition assay was used. Collected oocysts of different Eimeria species of chicken were exposed to six different concentrations (w/v) of Vitis venifera extract (TAE) in 10% dimethylsulphoxide solution (DMSO). Dimethylsulphoxide (DMSO) and potassium dichromate solution (K₂Cr₂O₇) served as control groups. Results of the study revealed that Vitis venifera extract (TAE) showed an inhibitory effect on sporulation (%) and damage (%) of Eimeria oocysts in a dose-dependent manner as compared to both control groups. Vitis venifera extract also damaged the morphology of oocysts in terms of shape, size, and number of sporocysts.

Keywords: Vitis venifera, in vitro, Eimeria, oocysts

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Authors: Boukrouh Soumaya, Cabaraux Jean-François, Avril Claire, Noutfia Ali, Chentouf Mouad

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The introduction of marginal leguminous forage species in the diet of ruminants are of great importance. Bitter vetch is a good source of proteins, highly resistant against drought and poor soil conditions. Accordingly; two years field trials (2018/2019 and 2019-2020) were conducted to determine the digestibility of straw and grains of 17 promising bitter vetch ecotypes(Vicia ervilia) in the north of Morocco. In vitro dry and organic matter digestibility, gas production, and kinetics of fermentation of grains and straw were evaluated using gas production technique, pepsin-cellulase enzymatic digestibility of DM (CDDM)and OM (CDOM), as well as protease enzymatic CP degradation (CPD) and in vitro true digestibility, were performed using DAISYII Incubator. In vitro digestibility was performed using gas production method of (Menke et al., 1979) improved by Menke and Steingass (1988). Samples were incubated in glass syringes that contained rumen fluid and incubation solution that conserved in water bath in 39°C during 72 hours. Gas production was recorded after 2, 4, 8, 12, 24, 48, and 72 hours. Studied digestibility parameters were dry and organic matter digestibility, microbial biomass production, partitioning factor, and volatile fatty acids. Enzymatic dry matter digestibility was different (p < 0.05) among grains and straw for all ecotypes. It varied from 804.1 to 957.7 g/kg DM and 270.4 to 412.3 g/kg DM for grains and straw, respectively. Metabolizable energy varied between 11.7 to 14.3 MJ/kg DM and 2.6 to 5.0 MJ/kg DM for grains and straw, respectively. Potential gas production (A), the rate constants (c and d), and lag times of grains and straws from different bitter vetch ecotypes were different (p > 0.05). The results emphasized that in any evaluation of bitter vetch ecotypes, where straw of this legume seed is used as an animal feed, not only seed yield but also yield and quality of straw should be taken into consideration, particularly in areas where straw from this legume is considered as an important feedstuff for ruminants. Enzymatic digestibility was lower than in vitro digestibility by gaz production and by the DAISYII method because rumen fluid contains bacteria than increase digestibility. There was no difference between in vitro digestibility by gaz production and the DAISY II method. The DAISY II method can be used to increase labor efficiency in the in vitro DM digestibility analysis if gaz production is not necessary for analysis.

Keywords: bitter vetch, grains, straw, ecotype, in vitro digestibility, gaz production, enzymatic digestibility

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Authors: Roberto Cassino, Valeria Dissette, Carlo Alberto Bignozzi, Daniele Pazzi

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Background and Aims: The aim of this work was to investigate, both ‘in vitro’ and ‘in vivo’, if the new SCX technology (SiO2-Ag+Chlorex) can easily defeat infections and it is really more effective than SSD (SilverSulfaDiazine). ‘In vitro’ methods: we tested ‘in vitro’ the effectiveness of both silver materials using a pool of 5 strains: Pseudomonas Aeruginosa, Staphylococcus aureus, Escherichia Coli, Enterococcus hirae and Candida Albicans. 100 µl of this pool have been seeded on Petri dishes and kept for 24 hours in incubation at 37 C°. ‘In vivo’ methods: we enrolled patients with multiple infectious chronic wounds (according with cutting & harding criteria for infection); after a qualitative evaluation of the wounds bacterial population, taking a sample by plug, we included in the study 6 patients for a total of 10 wounds, infected by one or more of the microorganisms used for the ‘in vitro’ test. The protocol consisted of a treatment with a spray powder of SSD every 48 hours for 14 days; in case of worsening we should have to start a new treatment with a spray powder containing silicon dioxide, ionic silver and chlorexidine (SiO2-Ag+Chlorex) every 48 hours for 14 days. We evaluated the number of clinical signs of infection and the disappearance or not of the wound edge erithema. ‘In vitro’ results: SSD demonstrated a wide zone of inhibition within 24 hours, but after 5 days there was no more signs of inhibition; on the contrary SCX had a good inhibition ring that lasted more than 5 days. ‘In vivo’ results: all wounds treated with SSD got worse; the signs of infection increased and the wound edge erithema did not disappear. According with the protocol, we treated then all wounds with SCX and they all improved within the period of observation with complete disappearance of clinical signs of infection and no more wound edge erithema. Conclusions: the study demonstrated the effectiveness of SiO2-Ag+Chlorex, especially in terms of long lasting antimicrobial action. We had the same results ‘in vitro’, so that there has been a perfect correspondence between the laboratory outcomes and the clinical ones.

Keywords: chronic wounds, infections, ionic silver, SSD

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Authors: Ajay Aggarwal, Kamal Saroha, Sanju Nanda

Abstract:

Niosomes or non-ionic surfactant vesicles are microscopic lamellar structures formed on admixture of non-ionic surfactant of the alkyl or dialkyl polyglycerol ether class and cholesterol with subsequent hydration in aqueous media. They are vesicular systems similar to liposomes that can be used as carriers of amphiphilic and lipophilic drugs. Entrapment efficiency was found to be higher in case of niosome prepared with span60 than niosome prepared with tween. The amount of release was found to be in order of Span20>Tween60>Tween20>Span60. As the concentration of surfactant is increased in vitro release was increased due to high entrapment. The stability study of optimized batch revealed that particle size was increased after 3months on increasing the temperature. On the other hand entrapment efficiency was decreased on increasing the temperature.

Keywords: niosomes, vesicles, span, tween, in vitro release

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Authors: Jinfeng Zhang, Chun-Sing Lee

Abstract:

The use of different nanocarriers for delivering hydrophobic pharmaceutical agents to tumor sites has garnered major attention. Despite the merits of these nanocarriers, further studies are needed for improving their drug loading capacities (typically less than 10%) and reducing their potential systemic toxicity. So development of alternative self-carried nanodrug delivery strategies without using any inert carriers is highly desirable. In this study, we developed a self-carried theranostic curcumin (Cur) nanodrug for highly effective cancer therapy in vitro and in vivo with real-time monitoring of drug release. With a biocompatible C18PMH-PEG functionalization, the Cur nanoparticles (NPs) showed excellent dispersibility and outstanding stability in physiological environment, with drug loading capacity higher than 78 wt.%. Both confocal microscopy and flow cytometry confirmed the cellular fluorescent “OFF-ON” activation and real-time monitoring of Cur molecule release, showing its potential for cancer diagnosis. In vitro and in vivo experiments clearly show that therapeutic efficacy of the PEGylated Cur NPs is much better than that of free Cur. This self-carried theranostic strategy with real-time monitoring of drug release may open a new way for simultaneous cancer therapy and diagnosis.

Keywords: drug delivery, in vitro and in vivo cancer therapy, real-time monitoring, self-carried

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Authors: Bhuwan C. Joshi, Atish Prakash, Ajudhia N. Kalia

Abstract:

Ethnopharmacological relevance: The plant Urtica dioica L. (Urticaceae) is used in various diseases including hepatic ailments. Traditionally, the leaves and roots of the plant are used in jaundice. Objective: The aim of the present work was to evaluate hepatoprotective potential of potent antioxidant from Urtica dioica L. against CCl4 induced hepatotoxicity in-vitro and in-vivo model. Materials and methods: Antioxidant activity of hydro alcoholic extract and its fractions petroleum ether fraction (PEF), ethyl acetate fraction (EAF), n-butanol fraction (NBF) and aqueous fraction (AF) were determined by DPPH radicals scavenging assay. Fractions were subjected to in-vitro cell line study. Further, the most potent fraction (EAF) was subjected to in-vivo study. The in-vivo hepatoprotective active fraction was chromatographed on silica column to isolate the bioactive constituent(s). Structure elucidation was done by using various spectrophotometric techniques like UV, IR, 1H NMR, 13C NMR and MS spectroscopy. Results and conclusion: The ethyl acetate fraction (EAF) of Urtica. dioica L. possessed the potent antioxidant activity viz. DPPH (IC50 78.99 ± 0.17 µg/ml). The in-vitro cell line study showed EAF prevented the cell damage. The EAF significantly attenuated the increased liver enzymes activities in serum and tissue. Column chromatography of most potent antioxidant fraction (EAF) leads to the isolation of 4-hydroxy-3-methoxy cinnamic acid which is responsible for its hepatoprotective potential. Hence, the present study suggests that EAF has significant antioxidant and hepatoprotective potential on CCl4 induced hepatotoxicity in-vitro and in-vivo.

Keywords: Urtica dioica L., antioxidant, HepG2 cell line, hepatoprotective

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Authors: Abobkar Abrahem Mohamed Saad, Asma Abud Alsalam

Abstract:

Closed flowers of Maerua crassifolia were cultured on Murashige and Skoog medium supplemented with benzyl amino purine BA (1.0 mg/l). The colour of flowers changed from green to pale brown after one week. They opened after two weeks. The anthers became clear which was observed after 3 weeks. Calluses are induced from sepals after one month. 19 anthers were observed with average length of 1.9 cm. The amount of calluses increased after 40 days. These calluses were fragmented and subcultured on MS+ 2-4D (1.0 mg/l) in order to increase growth.

Keywords: in vitro, Maerua, flowers, culture

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Authors: Surajj Sarode, G. P. Vadnere, G. Vidya Sagar

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Compression coating is one of the strategies for delivering drug to the colon based on Gastrointestinal PH and transit time concept. The main aim of these formulations to develop rapidly disintegrating Zaltoprofen core tablets compression-coated with a mixture of time-dependent hydrophilic swellable polymer HPMC K 15 and PH responsive soluble polymer Chitosan and Guar gum in different ratios. The effect of the proportion of HPMC, Chitosan and Guar gum in the coat on premature drug release in upper part (Stomach and small intestine) of GIT and the amount of drug release in colon target area was studied. The formulations are carried out by using Direct Compression method. Sodium starch Glycolate used for rapid disintegration. FTIR used for Drug-Polymer Interaction studies. The prepared tablets were evaluated for hardness, thickness, friability, in-vitro disintegration, in-Vitro dissolution and in-vitro kinetic study.

Keywords: zaltoprofen, chitosan, formulation, drug delivery

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Authors: Zelikha Labbani

Abstract:

Many biotechnology methods have been used in plant breeding programs. The in vitro isolated microspore culture is the one of these methods. For durum wheat, the use of this technology has been limited for a long time due to the low number of embryos produced and also most regeneration plants are albina. The objective of this paper is to show that using isolated microspores culture on durum wheat is possible due to the development of the new methods using the new pretreatment of the microspores before their isolation and cultivation.

Keywords: isolated microspore culture, pretreatments, in vitro embryogenesis, plant breeding program

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Authors: Olaniyi Fawole, Umezuruike Opara

Abstract:

Co-products obtained from pomegranate juice processing contain high levels of polyphenols with potential high added values. From value-addition viewpoint, the aim of this study was to evaluate the stability of polyphenolic concentrations in pomegranate fruit co-products in different solvent extracts and assess the effect on the total antioxidant capacity using the FRAP, DPPH˙ and ABTS˙+ assays during simulated in vitro digestion. Pomegranate juice, marc and peel were extracted in water, 50% ethanol (50%EtOH) and absolute ethanol (100%EtOH) and analysed for total phenolic concentration (TPC), total flavonoids concentration (TFC) and total antioxidant capacity in DPPH˙, ABST˙+ and FRAP assays before and after in vitro digestion. Total phenolic concentration (TPC) and total flavonoid concentration (TFC) were in the order of peel > marc > juice throughout the in vitro digestion irrespective of the extraction solvents used. However, 50% ethanol extracted 1.1 to 12-fold more polyphenols than water and ethanol solvents depending on co-products. TPC and TFC increased significantly in gastric digests. In contrast, after the duodenal, polyphenolic concentrations decreased significantly (p < 0.05) compared to those obtained in gastric digests. Undigested samples and gastric digests showed strong and positive relationships between polyphenols and the antioxidant activities measured in DPPH, ABTS and FRAP assays, with correlation coefficients (r2) ranging between 0.930 – 0.990 whereas, the correlation between polyphenols (TPC and TFC) and radical cation scavenging activity (in ABTS) were moderately positive in duodenal digests. Findings from this study also showed that the concentration of pomegranate polyphenols and antioxidant thereof during in vitro gastro-intestinal digestion may not reflect the pre-digested phenolic concentration. Thus, this study highlights the need to provide biologically relevant information on antioxidants by providing data reflecting their stability and activity after in vitro digestion.

Keywords: by-product, DPPH, polyphenols, value addition

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Authors: Taraneh Estedlal

Abstract:

The objective of this study was to review in-vivo and in-vitro studies to compare the effectiveness of bulk-fill and conventional resin composites with regard to marginal adaptation, polymerization shrinkage, and other mechanical properties.PubMed and Scopus databases was investigated for in-vitro studies and randomized clinical trials comparing incidence of fractures, color stability, marginal adaptation, pain and discomfort, recurrent caries, occlusion, pulpal reaction, and proper proximal contacts of restorations made with conventional and bulk resins. The failure rate of conventional and flowable bulk-fill resin composites was not significantly different to sculptable bulk-fill resin composites. The objective of this study was to review in-vivo and in-vitro studies to compare the effectiveness of bulk-fill and conventional resin composites with regard to marginal adaptation, polymerization shrinkage, and other mechanical properties. PubMed and Scopus databases was investigated for in-vitro studies and randomized clinical trials comparing one of the pearlier mentioned properties between bulk-fill and control composites. Despite differences in physical and in-vitro properties, failure rate of conventional and flowable bulk-fill resin composites was not significantly different to sculptable bulk-fill resin composites.

Keywords: polymerization shrinkage, color stability, marginal adaptation, recurrent caries, occlusion, pulpal reaction

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Authors: Gajera Lalit, Shah Pranav, Shah Shailesh

Abstract:

Venlafaxine hydrochloride has a short elimination half-life of 5 ± 2 hr, and absorption window in the upper part of gastrointestinal tract. The conventional tablets need to be administered two to three times a day and possess an oral bioavailability of 45%. The purpose of this study was to formulate gastroretentive effervescent floating tablets of Venlafaxine HCl. Different grades of HPMC namely K15M, K4M, K100M and E15LV were employed as swelling polymers whereas sodium bicarbonate was employed as gas generating agent. The direct compression method was employed for the formulation of tablets. The tablets were evaluated in terms of hardness, friability, weight variation, drug content, water uptake, in-vitro floating behavior and in-vitro drug release study. All the formulations exhibited very short floating lag time of < 1 min and total floating time of 12 hr. Formulation L3 containing 25 mg and 75 mg of HPMC E15 LV and HPMC K15M respectively exhibited complete drug release within 12 hrs.

Keywords: venlafaxine HCl, hydroxyl propyl methylcellulose, floating gastro retentive tablets, in-vitro drug release, non-fickian diffusion

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Authors: Zelikha Labbani

Abstract:

Since its creation in 1964 by Guha and Maheshwari in India on Datura innoxia Mill, in vitro androgenesis has become the method of choice in the production of doubled haploid in many species. However in durum wheat, the Doubled haploid plant breeding programs remained limited due to the low production of androgenetic embryos and converting them into fertile green plants. We describe here an efficient method for inducing embryos and regenerating green plants directly from isolated microspores of durum wheat.

Keywords: Durum wheat, haploid embryos, on in vitro, pretreatment

Procedia PDF Downloads 341