On the Stability of Clathrate Hydrates Encaging Polar Guest Molecules: Contrast in the Hydrogen Bonds of Methylamine and Methanol Hydrates

Abstract

The stability of clathrate hydrates encaging highly polar guests has been investigated in order to explain the experimental observation that some amines form clathrate hydrates but alcohols act as inhibitor to hydrate formation. We choose methylamine and methanol as guest species and examine the stable structure, at which the total potential energy has a minimum value. At the local minima of those two hydrates, the potential energies of water-water and guest-water, and their hydrogen bonded networks are compared. It is found that methanol does not retain the host lattice structure, while the host-network structure is kept in the presence of methylamine. It is shown that the difference in the magnitude of the partial charge on the hydrogen atom between the hydroxyl and amino groups plays a much more significant role on the stability of both clathrate hydrates than the difference in molecular geometry. This is supported from the result of a methylamine-like model that has the same partial charges on the atoms in the hydrophilic site as methanol.     

Mechanisms of modulation of neuronal nicotinic receptors by substance P and OAG

Substance P is known to modulate neuronal nicotinicacetylcholine receptors (nAChRs) in the sympathetic nervous system.There are two conflicting proposals for the mechanism of this effect, an indirect action mediated by protein kinase C (PKC) and a direct interaction with receptor subunits. We studied the mechanisms of thiseffect in PC-12 cells. Substance P enhanced the decay of thenicotine-induced whole cell current. This effect was fast in its onsetand was not antagonized by guanosine5'-O-(2-thiodiphosphate), a G protein blocker, orstaurosporine, a nonselective PKC blocker. Staurosporine failed toreverse the inhibition by 1-oleoyl-2-acetyl-sn-glycerol (OAG), a synthetic diacylglycerol analog known to activate PKC. Theinhibitory effects of the peptide and OAG were preserved in excisedpatches, but substance P applied to the extra patch membrane wasineffective in the cell-attached patch configuration. We conclude thatsubstance P modulates neuronal nAChRs most likely by direct interactions with the receptors but independently from activation ofPKC or G proteins and that PKC does not participate in modulation by OAG.

     

Enhancement of γ-linolenic acid production by Mucor ambiguus with nonionic surfactants

Summary -Linolenic acid (GLA) production by Mucor ambiguus IFO 6742, immobilised in Biomass Support Particles (BSPs), has been investigated in a fluidized-bed fermenter in the presence of nonionic surfactants. In this system, repeated batch cultivation was achieved at higher yield and productivity than by conventional methods, since microbial lipids inlcuding GLA were significantly secreted into the culture broth and/or on the surface of the cell wall.     

Cauliflower mosaic virus gene VI causes growth suppression, development of necrotic spots and expression of defence-related genes in transgenic tobacco plants

Summary In order to study possible functions of the inclusion body matrix protein (IBMP) encoded by gene VI of cauliflower mosaic virus (CaMV), the XbaI fragment containing the gene VI of a Japanese strain of CaMV (CaMV S-Japan) was transferred to tobacco plants by Ti mediated transformation. Eight out of 18 kanamycin resistant plants (40%) expressed detectable levels of IBMP. Those transgenic plants expressing IBMP produced leaves with light green color, and their growth was suppressed as compared with control plants. Symptom-like necrotic spots also appeared on the leaves and stems of the mature transgenic plants. Furthermore, in these transgenic plants, pathogenesis-related proteins 1a, 1b and 1c were highly expressed and the activity of 1,3--glucanase was increased up to eightfold. From these results, we concluded that expression of the IBMP is associated with symptom development.     

Characterization of Vegetal Plate Cells Separated from Cytochalasin B-Treated Blastulae of the Sea Urchin, Clypeaster japonicus

Vegetal plate forming cells (VPCs) of the vegetal plate blastulae of the sea urchin, Clypeaster japonicus , had a layer of microfilaments on the basal side. The VPCs specifically protruded from the embryos after a treatment with 1 μg/ml of cytochalasin B (CB). Based on scanning electron microscopy, unlike other epithelial cells the protruded VPCs possessed neither cilium nor microvilli on their surface. The protruded VPCs were easily separated from the embryos by stirring the embryo suspension with pipette. An in vitro immunohistochemistry using a primary mesenchyme cell (PMC) surface-specific monoclonal antibody (MAb) raised against PMCs of Strongylocentrotus purpuratus showed that the MAb also specifically bound to the PMCs in mesenchyme blastulae of C. japonicus . The MAb bound in 81% of the separated VPCs in C. japonicus vegetal plate blastulae examined. However, the MAb binding occurred only after the separated VPCs were incubated in artificial sea water (ASW) for at least 1 hr. In the VPC-deprived embryos, gastrulation occurred after they were transferred to normal ASW. However, the PMCs and the spicules were not formed in these embryos.
In conclusion, a majority of the VPCs separated from the CB-treated blastulae were presumptive PMCs. These VPCs provide an excellent source of presumptive PMCs.     

Oxygen uptake rate of immobilized growing hybridoma cells

Summary The specific oxygen uptake rate of hybridoma cells immobilized in calcium alginate gel particles was measured, and the observed data was compared with those of non-immobilized cells. The uptake rate of the immobilized cells coincided with that of the non-immobilized hybridoma cells just after immobilization, but increased with cell growth. On the other hand, the cellular glucose consumption rate decreased slightly during the experiments. The increased oxygen uptake rate by immobilized cells was closely related to the formation of cell colonies in the gel particles.     

Comparative effects of a recombinant and a mutein type of granulocyte colony stimulating factor on the growth of Meth-A fibrosarcoma with 5-fluorouracil chemotherapy

The present study was designed to evaluate the effects of a recombinant human G-CSF (rhG-CSF) and a mutein G-CSF(KW-2228) on leucopenia and tumor growth in mice treated with 5-fluorouracil (5-FU). In normal mice, the number of leucocytes (white blood cell, WBC) reached the peak 12 hours after a single injection of either type of G-CSF and decreased to the normal level after 24 hours. Daily administration induced a continuous increase in the WBC count, however, administrations at intervals did not. Meth-A fibrosarcoma was subcutaneously inoculated into the backs of syngeneic BALB/c mice. The mice were treated with 5-FU alone or with G-CSFs. Chemotherapy with 5-FU alone resulted in leucopenia and an insignificant inhibition of tumor growth. The conjunctive administration of G-CSFs with 5-FU resulted in a significantly augmented inhibition of tumor growth, and leukopenia was not seen. This augmenting effect was more prominent with KW-2228.These results suggest that in 5-FU chemotherapy G-CSFs may be beneficial in restoring the number of leucocytes from leucopenic state and in augmenting the tumor inhibitory effect. Furthermore, KW-2228 may be more beneficial than the natural type rhG-CSF.     

Long-term cultivation of anchorage-independent animal cells immobilized within reticulated biomass support particles in a circulating bed fermentor

Summary Long-term cultivation of anchorage-independent animal cells immobilized within porous biomass support particles (BSPs) using a gas-stirred circulating bed fermentor (CBF) was investigated. Inoculation of mouse myeloma MPC-11 (ATCC CCL 167) cells into reticulated polyvinyl formal resin BSPs (3 × 3 × 3 mm; mean pore diameter, 60 m; porosity, 0.88) and the repeated batch culture of inoculated cells were performed under gentle circulation of BSPs, induced by sparging air from the base of the fermentor. The glucose uptake rate of cells decreased in the initial period just after the start of circulation, since a relatively large number of cells leaked from the BSPs. After that period, the uptake rate gradually increased and the leakage of cells diminished. In the meantime, when inoculated cells were incubated statically by introducing air into the upper part of the fermentor for the initial several days before circulating the BSPs, glucose consumption became very rapid and cell density in the BSPs reached at least 10⁷ cells/cm³ BSP. Thus, a long-term cultivation without significant leakage of cells and with high cell density in BSPs was successfully achieved in the CBF-BSP system.Offprint requests to: H. Yamaji     

Two monoclonal antibodies against small-cell lung cancer show existence of synergism in binding

Summary Murine IgG1 monoclonal antibodies (mAbs), ITK-2 and ITK-3, were generated against a small-cell lung cancer (SCLC) cell line. Enzyme-linked immunosorbent assay using a variety of established cell lines as substrates, immunoperoxidase staining of freshly frozen tissue sections, and fluorescence-activated cell sorter analysis of peripheral blood leukocytes showed that these mAbs recognize a part of the SCLC-associated cluster 1 antigen. In immunoprecipitation studies, both ITK-2 and ITK-3 bound to a 145-kDa glycoprotein of SCLC cell membrane extracts, as did MOC-1 and NKH-1, which both recognize the cluster 1 antigen. However, because the binding of¹²⁵I-labeled ITK-2 to SCLC cells was not inhibited by MOC-1 or NKH-1, the binding site of ITK-2 on SCLC cells appeared to be different from that of either MOC-1 or NKH-1. Unexpectedly, binding of¹²⁵I-labeled ITK-2 to SCLC cells increased in the presence of ITK-3. This ITK-3-induced increase in ITK-2 binding was due partly to an increase in the number of binding sites for ITK-2 on SCLC cells. Addition of ITK-3 may, therefore, improve the effectiveness of ITK-2-based tumor detection or therapy.     

Incorporation of fatty acids by Streptococcus mutans

In a series of investigations into the cariogenicity of Streptococcus mutans, we studied the incorporation of exogenous fatty acids with reference to glucosyltransferase secretion and membrane fatty acid changes. When cells were grown with different fatty acids, both saturated and unsaturated fatty acids were readily incorporated into the membrane lipids and were biotransformed and elongated preferentially to the longer 16- and 18-carbon-chain fatty acids. This incorporation and chain-elongation led to significant changes in fatty acids composition. By adding fatty acids to the medium, it was possible to appropriately modify the degree of unsaturation and the relative ratio between specific fatty acids in the membrane lipids of S. mutans.