Recent Advances in the use of Immobilized Lipases Directed Toward the Asymmetric Syntheses of Complex Molecules

Water-insoluble compounds can be substrates for enzymatic reactions when lipases are immobilized properly and suitable organic solvents are used. In this review, three type of lipase immobilization method and their application to the asymmetric syntheses of complex molecules are described. Lipases immobilized with Celite or synthetic prepolymers such as urethane prepolymer and photo-crosslinkable resin prepolymer have been applied for the kinetic resolution of many kinds of water-insoluble substrate.

Phospholipid-lipase aggregates with ether linkages are novel and have been found to function effectively as immobilized lipases in asymmetric hydrolysis or esterification reactions in water-saturated organic solvent. The phospholipid-lipase aggregates are considered to have a stacked bilayer based on X-ray diffraction analysis structure of the lipid in the crystalline phase.     

Replication and cytopathic effects of ovine lentivirus strains in alveolar macrophages correlate with in vivo pathogenicity.

Ruminant lentiviruses share genomic sequences and biologic properties with human immunodeficiency viruses. Four ovine lentivirus strains were assessed for cytopathic effects and virus replication. Lentivirus isolate H/24 produced high virus titers and lysis of synovial cells but replicated slowly and caused no fusion of alveolar macrophages. Lentivirus isolates 84/28 and 85/14 produced low virus titers, less syncytia, and limited or no cell lysis in synovial cells and macrophages. In contrast, ovine lentivirus isolate 85/34 produced early peak virus titers and caused rapid fusion and lysis of both macrophages and synovial cells. Ovine lentivirus isolates which were cytopathic for macrophages induced lymphoproliferative disease when inoculated into lambs.     

Electron microscopic observations on the formation of elastic fibres in cultures of aortic medial cells and adventitial cells

The formation of elastic fibres was observed in the cultured cells derived from the tunica media and the tunica adventitia of mouse aorta. Bundles of myofilaments with dense bodies were abundantly observed in the cytoplasm of the cultured medial cells, and numerous bundles of microfibrillar components were present in the intercellular spaces. Fine granules of approximately 50 nm in diameter were observed in the bundles of microfibrillar components. It was supposed that these fine granules of elastin fused with each other and formed elastic aggregates and then formed large elastic clumps. Numerous bundles of microfibrillar components were also present in the intercellular spaces of the cultured adventitial cells. Elastic aggregates were scarcely observed in the bundles of microfibrillar components. However, large elastic clumps as observed in the medial cell culture could not be found in the adventitial cell culture. It is suggested that the formation of large elastic clumps might be related to the sheet structures or lamellae of elastic fibres in the tunica media.     

Effects of Nutrient Limitation and {gamma}-Irradiation on Tracheary Element Differentiation and Cell Division in Single Mesophyll Cells of Zinnia elegans

The effects of nutrient limitation and -irradiation on trachearyelement differentiation and cell division were investigatedusing single cells isolated from the mesophyll of Zinnia elegans.When the phosphate concentration of the medium was reduced to10 µM (1/50 of Fukuda and Komamine's medium, 1980a), thefrequency of cell division during 4 days of culture decreased,while the frequency of tracheary element differentiation wasunaffected. -Irradiation with a dose of 92 Gy at 36 h of culturepreferentially and thoroughly suppressed cell division withoutreducing the number of tracheary elements formed. The appearanceof secondary cell wall thickenings was delayed by irradiation,but synchrony was maintained. Thus the Zinnia system previouslyreported [Fukuda and Komamine (1980a) Plant Physiol. 65: 57]was improved to give a more useful system for the study of cytodifferentiation,in which tracheary element formation occurred from single cellswithout cell division. ¹Present address: Biological Institute, Faculty of Science,Tohoku University, Sendai, Miyagi 980, Japan. (Received November 28, 1985; Accepted February 22, 1986)     

Fine-granular cationic iron colloid. Its preparation, physicochemical characteristics and histochemical use for the detection of ionized anionic groups

In order to obtain distinct and reliable information concerning the localization of ionized anionic groups in tissues, fine-granular cationic ferric hydroxide colloid solution (Fe-Cac-f) was newly devised. This can be obtained by boiling a mixture of ferric chloride and ammonium cacodylate solutions. The colloid particles of Fe-Cac-f are about 1.0 nm in size, i.e., one-fifth of the size of ferric cacodylate colloid (Fe-Cac; Seno et al. 1983a). As with Fe-Cac, Fe-Cac-f particles in the pH range of 1.6-7.6 carry a positive electric charge, but the latter show a better permeation of tissues. Using the Prussian blue reaction, Fe-Cac-f gives a distinct deep-blue color and can be used for the detection of anionic groups of acid mucopolysaccharides and proteins by light microscopy. It is also useful for detecting the exact sites of ionized anionic groups in deep tissue areas using electron microscopy.     

A new method of the immunohistochemical detection of cellular antigens for light and electron microscopy

Summary A new immunohistochemical method for light and electron microscopy of tissue- and cell-specific antigens by using ferric colloid-labeled antibody is presented. The antibodies labeled with the cationic cacodylate ferric colloid are stable and bind specifically to the target antigens to show clearly the site of antigens in tissue sections and on free cells by Prussian blue reaction for light microscopy and by the specific figure of electron opaque ferric colloid particles for electron microscopy. The staining procedure is very simple and it gives clear picture. So the method will be of beneficial for general laboratory use in immuno-histochemical researches.     

The types of bioreactors used for shoots and embryos

Plant regenerated organs such as shoots, bulbs, microtubers, corms, embryos, etc. have been successfully proliferated in the bioreactor. The use of a bioreactor leads to the development of technology suitable for large scale plant propagation. The basic construction and characteristics of various types of bioreactor systems are reviewed in relation to shoot and embryo cultures. A pilot scale 500 liter bioreactor system was applied to the production of large scale Stevia rebaudiana shoots.Abbreviations DW dry weight - EC electrical conductivity - FW fresh weight - ORP oxidation-reduction potential     

Mass propagation of shoots of Stevia rebaudiana using a large scale bioreactor

A procedure for the mass propagation of multiple shoots of Stevia rebaudiana is described. Isolated shoot primordia were used as the inoculum to obtain clusters of shoot primordia. Such clusters were grown in a 500 liter bioreactor to obtain shoots. A total of 64.6 Kg of shoots were propagated from 460 g of the inoculated shoot primordia. These shoots were easily acclimatized in soil.     

Interrelationship between lignin deposition and the activities of peroxidase isoenzymes in differentiating tracheary elements of Zinnia

In a culture system in which single cells isolated from the mesophyll of Zinnia elegans L. differentiate to tracheary elements (TEs), two inhibitors of phenylalanine ammonia-lyase (EC 4.3.1.5), L-α-aminooxy-β-phenylpropionic acid (AOPP) at 10 μM inhibited lignification without reducing the number of TEs formed. These inhibitors caused intracellular changes in peroxidase (EC 1.11.1.7) activities. The inhibitors increased the activity of peroxidases bound to the cell walls and especially the activity of peroxidase bound ionically to the cell walls. In contrast, the activity of extracellular peroxidase decreased. There were five isoenzymes, P1-P5, in the ionically bound peroxidase of cultured Zinnia cells. Among the isoenzymes, P4 and P5 appeared to be specific for TE differentation. Treatment with AOPP and AIP resulted in increases in the activities of P2, P4 and P5 isoenzymes, with the most prominent increase in P5 activity. The addition of lignin precursors, including coniferyl alcohol, to the AOPP-treated cells restored lignification, and suppressed the alteration of peroxidase isoenzyme patterns caused by AOPP. The relationship between the wall-bound peroxidases and lignification during TE differentiation is discussed in the light of these results.