Histo-morphological analysis of rice callus cultures reveals differential regeneration response with varying media combinations

Genetic transformation of most indica rice (Oryza sativa) cultivars is hampered by poor in vitro culture performance and low regeneration potential. Histological study of primary calli can provide substantial information on their regeneration potential and can be used for early grading of calli expected to develop plantlets on regeneration media. The study was aimed to undertake histological analysis of primary calli derived from mature seeds of five indica rice cultivars viz. KSK-133, KS-282, Shaheen Basmati, Super Basmati, and DilRosh in order to assess their regeneration potential on different media combinations supplemented with various hormone concentrations (N6 + 2 mg/L 2,4-Dichlorophenoxyacetic acid; N6 + 2 mg/L 2–4 D + 2 mg/L Benzylaminopurine and MS + 2 mg/L 2,4-D). Calli with regeneration capability were subjected to histological assays by examining toulidine blue stained 5–8 μm thin sections for the presence of meristematic zones exhibiting embryogenic callus features. Based on our observations, formation of embryoids or embryoid-like structures was pronounced in KSK-133 and KS-282 calli. However, DilRosh, Super Basmati and Shaheen Basmati did not show these characteristic features. Three-week-old calli of all rice cultivars were transferred into regeneration medium (MS + 2 mg/L BAP + 1 mg/L Naphthaleneacetic acid). KSK-133 and KS-282 showed the highest regeneration potential (81% and 76%, respectively). These data were supported by histological observations where characteristic embryogenic units (EU) were noticed in these genotypes. These meristematic regions displayed high mitotic activity and stained relatively dark. The embryogenic calli cells were found heavily cytoplasmic with prominent nuclei and were located on the callus surface or inside surrounded by parenchymal cells.

     

Two new protease-inhibiting glycosphingolipids from Buddleja crispa

Crispins A (1) and B (2), two new glycosphingolipids, were isolated from the whole plant Buddleja crispa, along with three known compounds: alpha-amyrin, linoleic acid, and stigmasterol. Their structures were elucidated by chemical and spectroscopic techniques. Both 1 and 2 showed significant inhibitory activity against alpha-chymotrypsin in a concentration-dependent manner.     

Isolation and enzyme-inhibition studies of the chemical constituents from Ajuga bracteosa

Bractin A (=(2S,3S,4R,5E)-2-{[(2R)-2-hydroxydodecanoyl]amino}triacont-5-ene-1,3,4-triol; 1) and bractin B (=(2S,3S,4R,5E,8E)-2-{[(2R)-2-hydroxyhexacosanoyl]amino}pentadeca-5,8-diene-3,4,15-triol 1-O-beta-D-glucopyranoside; 2), new sphingolipids, and bractic acid (=(5Z,10Z,15Z)-2-decyl-4,7,8,12,13,17,18-heptahydroxy-20,23-dioxopentacosa-5,10,15-trienoic acid; 3), a long-chain polyhydroxy acid, were isolated from the whole plant Ajuga bracteosa along with four known diterpenoids 4-7. Their structures were deduced by spectral studies including 1D- and 2D-NMR spectroscopy. Compounds 1-3 displayed inhibitory potential against enzyme lipoxygenase, while compounds 4-7 inhibited cholinesterase enzymes in a concentration-dependent manner with IC(50) values in the range 10.0-33.0, 14.0-35.2, and 10.0-19.0 microM for lipoxygenase, acetylcholinesterase, and butyrylcholinesterase, respectively. Lineweaver-Burk, and Dixon plots, and their secondary replots indicated that all compounds exhibit non-competitive type of inhibition with K(i) values in the range of 9.5-35.2, 15.2-36.0, and 11.6-20.5 microM, for lipoxygenase, acetylcholinesterase, and butyrylcholinesterase, respectively.     

Staurosporine-induced cell death in salmonid cells: the role of apoptotic volume decrease, ion fluxes and MAP kinase signaling

Apoptotic cell death in mammalian models is frequently associated with cell shrinkage. Inhibition of apoptotic volume decrease (AVD) is cytoprotective, suggesting that cell shrinkage is an important early event in apoptosis. In salmonid hepatoma and gill cells staurosporine induced apoptosis, as assessed by activation of effector caspases, nuclear condensation, and a decrease of mitochondrial membrane potential (MMP), and these changes were accompanied by cell shrinkage. The Cl⁻ transport inhibitor DIDS and the K⁺ channel inhibitor quinidine prevented AVD, but only DIDS inhibited apoptosis. Other Cl⁻ flux inhibitors, as well as a pan-caspase inhibitor, did not prevent cell shrinkage, but still prevented caspase activation. Furthermore, regulatory volume decrease (RVD) under hypotonic conditions was not facilitated, but diminished in apoptotic cells. Since all transport inhibitors used blocked RVD, but only DIDS and quinidine inhibited AVD, the ion transporters involved in both processes are apparently not identical. In addition, our data indicate that inhibition of Cl⁻ fluxes rather than blocking cell shrinkage or K⁺ fluxes is important for preventing apoptosis. In line with this, inhibition of MAP kinases reduced RVD and not AVD, but still diminished caspase activation. Finally, we observed that MAP kinases were activated upon staurosporine treatment and that at least activation of ERK was prevented when AVD was inhibited.     

The effects of reaction conditions on the production of γ-cyclodextrin from tapioca starch by using a novel recombinant engineered CGTase

A novel mutant enzyme namely H43T CGTase can produce up to 39% γ-cyclodextrin (γ-CD) compared to the native enzyme which produces only 10% γ-CD. The effect of the reaction conditions on γ-CD production was studied using this mutant CGTase. The effects of substrate–buffer combination, starch pretreatment and concentration, pH, additives and finally the use of a debranching enzyme improved the γ-CD ratio further. The tapioca–acetate pair gave the highest conversion (16% conversion) among four types of starch and four buffer system combinations. Gelatinized starch was preferred compared to raw tapioca starch in producing a high percentage of γ-CD and conversion rate. Higher pH especially pH 8–9 led to a higher proportion of γ-CD, and was relatively more apparent when the concentration of starch was increased. Forty-six percent γ-CD was produced using 2.5% gelatinized tapioca starch at pH 8. Pullulanase enzyme was found to be useful in reducing the viscosity of tapioca starch paste thus increasing the efficiency of utilization of starch by CGTase by at least 20- to 30-fold. Up to 48% γ-CD can be produced when 4% pullulanase-pretreated tapioca starch was reacted with the CGTase mutant. It was also found that the supplementation of the reaction mixture with glucose, toluene, or cyclododecanone improved the γ-CD yield by 42.2, 46.4, 43.4, and 43.4%, respectively. All the parameters involved have been shown to affect the product specificity of the mutant H43T CGTase transglycosylation mechanism.     

Selenium in chemistry and biochemistry in comparison to sulfur

What makes selenoenzymes--seen from a chemist's view--so special that they cannot be substituted by just more analogous or adapted sulfur proteins? This review compiles and compares physicochemical properties of selenium and sulfur, synthetic routes to selenocysteine (Sec) and its peptides, and comparative studies of relevant thiols and selenols and their (mixed) dichalcogens, required to understand the special role of selenium in selenoproteins on the atomic molecular level. The biochemically most relevant differences are the higher polarizability of Se- and the lower pKa of SeH. The latter has a strikingly different pH-dependence than thiols, with selenols being active at much lower pH. Finally, selected typical enzymatic mechanisms which involve selenocysteine are critically discussed, also in view of the authors' own results.     

Bioconcentration of some macrominerals in soil,forage and buffalo hair continuum: A case study on pasture irrigated with sewage water

The present study aimed to evaluate the bioaccumulation of some macrominerals in grazing buffaloes fed forage irrigated with sewage water or canal water. In particular, the transfer of sodium (Na), magnesium (Mg), potassium (K) and calcium (Ca) from soil to plant and in turn to animals was evaluated under sub-tropical environmental conditions. Samples of soil, forage and buffalo hair were collected and digested by wet method. Sodium and K concentrations were significantly higher in the soil but lower in the forages; however, Mg and Ca concentrations in both soil and forages were higher. The correlation between soil, forage and hair showed an imbalanced flow of Na, Mg and K and a balanced flow of Ca from soil to forage and then to animals. Based on the findings, the highest rates of transfer of minerals were found for sewage water treatment, whereas lowest rates were found for canal water treatment, except for Na. As the transfer of minerals depends on their bioavailability, the highest values may be due to the high rates of mineral uptake by plants. Thus, the high transfer rate of some elements by plants could become toxic in future causing detrimental effect to grazing livestock.     

Post-exposure temperature influence on the toxicity of conventional and new chemistry insecticides to green lacewing Chrysoperla carnea (Stephens) (Neuroptera: Chrysopidae)

Chrysoperla carnea (Stephens) is an important biological control agent currently being used in many integrated pest management (IPM) programs to control insect pests. The effect of post-treatment temperature on insecticide toxicity of a spinosyn (spinosad), pyrethroid (lambda cyhalothrin), organophosphate (chlorpyrifos) and new chemistry (acetamiprid) to C. carnea larvae was investigated under laboratory conditions. Temperature coefficients of each insecticide tested were evaluated. From 20 to 40 °C, toxicity of lambda cyhalothrin and spinosad decreased by 2.15- and 1.87-fold while toxicity of acetamiprid and chlorpyrifos increased by 2.00 and 1.79-fold, respectively. The study demonstrates that pesticide effectiveness may vary according to environmental conditions. In cropping systems where multiple insecticide products are used, attention should be given to temperature variation as a key factor in making pest management strategies safer for biological control agents. Insecticides with a negative temperature coefficient may play a constructive role to conserve C. carnea populations.