Asymmetric synthesis of (S)-ethyl-4-chloro-3-hydroxybutanoate using Candida parapsilosis ATCC 7330

Asymmetric reduction of ethyl-4-chloro-3-oxobutanoate to (S)-ethyl-4-chloro-3-hydroxybutanoate in aqueous medium by resting cells of Candida parapsilosis ATCC 7330 was optimized. The influence of culture parameters (inoculum size, inoculum age and biocatalyst harvest time) and reaction parameters (co-substrate, resting cell, pH and substrate concentrations) on the asymmetric reduction were studied. It was found that these parameters significantly influenced the rate of the asymmetric reduction. Under the optimum conditions, the final concentration of (S)-ethyl-4-chloro-3-hydroxybutanoate, enantiomeric excess and the isolated yield of (S)-ethyl-4-chloro-3-hydroxybutanoate were 1.38 M (230 g/l), >99 and 95%, respectively. The space time yield was 115 mmol/lh, which is significantly higher than other whole cell biocatalysts reported so far.     

Clinal variation in carapace shape in the South American freshwater crab,Aegla uruguayana (Anomura: Aeglidae)

South America has been influenced by different geoclimatic events ever since its separation from Africa. The inland water fauna has evolved in response to the changing landscape. Currently, there are indications of variations in populations, occurring to different degrees that would indicate a clinal pattern in morphology. Among South America's fauna, the freshwater anomuran, Aegla, is an enigmatic group as a result of its endemicity and is composed of only one genus. Of all the species in this family, Aegla uruguayana has the broadest distribution. Its native habitats have been influenced by several marine transgressions during the Miocene–Quaternary Periods; thus, it is likely that their current distribution has been more recent. Its habitat spreads across a number of isolated basins and sub‐basins that display distinct degrees of isolation/connection, making clinal variation patterns in the morphology of this species possible. The present study aimed to evaluate the pattern of carapace shape variation in A. uruguayana and how it relates to the isolation and/or connection of populations from different basins and sub‐basins, allowing the determination of any extant clinal patterns. The specimens studied belong to 25 separate populations, representing all areas in which the species currently exists. A total of 523 crabs were analyzed. We identified 13 landmarks and four semi‐landmarks in the carapace. The aeglids were divided into seven size intervals to avoid an allometry effect. In each size category, shape relationships analyzed by principal component analysis suggest a geographical pattern corresponding to the distribution of the populations studied. An evaluation of covariation between body shape and geographical coordinates reveals a strong pattern and shows that population distribution had a significant effect on species morphology. Additionally, according to covariance analysis, the variation in shape was not associated with the environmental variables studied. We observed a clinal pattern throughout the species distribution, which could be attributed to genetic drift. It is possible that this process is being amplified by the geographical isolation of the basins, differences in environmental characteristics, and low dispersal ability. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 113 , 914–930.     

Sister-chromatid exchange and chromosome aberrations for 4 aliphatic epoxides in mice

Sister-chromatid exchange (SCE) and chromosome aberrations (CA) in bone marrow cells were analyzed after in vivo exposure in mice to 4 aliphatic epoxides, namely 1-naphthyl glycidyl ether (NGE), 1-naphthyl propylene oxide (NPO), 4-nitrophenyl glycidyl ether (NPGE) and trichloropropylene oxide (TCPO). These compounds were selected as being among the most mutagenic aliphatic epoxides in our previous structure-mutagenicity studies with the Ames test. There were significant dose-related increases in SCE and CA results for all 4 epoxides. The order of genotoxicity as established through SCE was NGE greater than NPO greater than NPGE approximately equal to TCPO greater than solvent control. It is of interest that Ames Salmonella results are consistent with in vivo genotoxicity for these compounds. However, only the plate test version of the Ames procedure is consistent with this order of in vivo genotoxicity and neither preincubation Ames testing results nor chemical alkylation rates would have predicted this order.     

IL-1 secretion by macrophages. Enhancement of IL-1 secretion and processing by calcium ionophores

In the present study we have demonstrated that the murine IL-1 alpha precursor lacks a cleavable signal sequence and does not undergo cotranslational translocation across microsomal membranes in vitro. Culture supernatants of the murine macrophage cell line, P388D, or from normal peritoneal macrophages collected within 0.5 to 3 h after stimulation contained the 33,000 m.w. precursor as the predominant form of IL-1 alpha. Over an 18-h period, the level of low m.w. IL-1 alpha increased as the secreted precursor was processed by extracellular and/or cell surface-associated proteolytic enzymes. The calcium ionophores A23187 and ionomycin were found to dramatically enhance the release and processing of murine and human IL-1. The rapid release of IL-1 in response to a change in the intracellular level of calcium does not appear to be caused by release of a membrane-bound form of the protein, nor is there evidence that IL-1 is packaged and released from cytoskeletal associated secretory granules. In marked contrast, calcium ionophores do not induce secretion of IL-1 from a nonmacrophage cell line that synthesizes but does not normally secrete IL-1. Our results suggest that activated macrophages possess a novel processing independent, possibly calcium-dependent, mechanism that allows for the release of the precursor forms of IL-1 alpha and IL-1 beta.     

Identification of IL-1 receptors on human monocytes

The expression and functional analysis of IL-1 beta R on human monocytes were investigated. Binding of 125I-IL-1 to human monocytes was found to be specific and saturable. Scatchard plot analysis revealed a single class of receptors with a binding constant of 600 pM and a receptor density of approximately 100 binding sites per cell. At 37 degrees C 54% of the labeled ligand was internalized over 2 h of incubation. Addition of 0.2% sodium azide to the cells reduced ligand internalization to 9% of total bound. Cross-linking studies revealed that the IL-1R in human monocytes had a Mr of 80 kDa. The addition of IL-1 to monocytes caused changes in membrane Ag expression as assessed by flow cytometric analysis. The results of this study identify IL-1 receptors on monocytes and suggest that IL-1 may act as an effector molecule for monocytes by enhancing expression of Ag correlated with cell differentiation and immune function.     

Plant regeneration from cotyledonary node explants of mungbean (Vigna radiata (L.) Wilczek)

Sexually-mature mungbean (Vigna radiata (L.) Wilczek) plants were efficiently regenerated from cotyledonary node explants. The explants were capable of directly developing multiple shoots on basal media devoid of any growth regulators. The shoot multiplication was influenced by media composition, growth regulators, age of donor seedling and explant type. The explants with both the cotyledons attached to the embryonic axis excised from 4-d-old seedlings, produced the highest number of shoots (5 or 6) in 100% of the cultures within 2 weeks on B₅ basal medium (BBM) containing BAP or 2-iP, respectively, (at 5x10^–7M) and 3% sucrose. Shoots elongated and developed better using BAP. Increasing micronutrients, carbohydrate and nitrogen levels in the medium above the original formulation of B₅ basal medium appeared to be of no benefit for increasing the number of shoots. The shoots were rooted on basal MS medium or MS containing 10^–6 of NAA, IAA or IBA. This protocol was found applicable to six other cultivars of mungbean. One hundred rooted shoots were successfully established in soil in the glasshouse, where 90% of them survived. The regenerated plants flowered precociously, but produced normal pods and viable seeds.Abbreviations BAP 6-benzylaminopurine - KIN kinetin - 2-iP 6- — -dimethylallyl aminopurine - AdS adenine sulphate - IAA indole-3-acetic acid - IBA indole-3-butyric acid - NAA 1-naphthalene acetic acid - MS Murashige and Skoog (1962) medium - B₅ Gamborg et al. (1968) medium - C medium MS salts + B₅ vitamins