Corticosteroidogenesis in the hypertrophic adrenal gland produced by betamethasone 17, 21-dipropionate in the rat fetus

Betamethasone 17, 21-dipropionate (BMDP), a potent glucocorticoid, produces adrenal hypertrophy in the rat fetus. The present study was performed to investigate the possible alterations of corticosteroidogenesis due to endogeneous substrates or exogenous pregnenolone in the incubation of homogenates of fetal hypertrophic adrenals caused by BMDP given to pregnant rats at day 19 of pregnancy.The corticosteroidal products and those levels per mg homogenate in an incubate of the hypertrophic adrenal homogenate did not differ from those of a normal adrenal. No accumulations of abnormal precursors or intermediates were found in the incubates of the hypertrophic adrenals. It is concluded from these findings that no qualitative alterations in the pathway of corticosteroidogenesis occurred in the hypertrophic adrenal glands caused by BMDP in the rat fetus. When the calculation was done per adrenal gland, the content of corticosterone in the incubate of the homogenate of the hypertrophic adrenal was remarkably higher than that found in a normal gland. This finding was compatible with the significant increase of the plasma corticosterone concentration in the fetuses with the adrenal hypertrophy caused by BMDP.     

Change in the ultraviolet spectrum of solubilized Ca2+-dependent ATPase from sarcoplasmic reticulum due to binding with Ca2+ ions.

Solubilized sarcoplasmic reticulum (SSR) was prepared by solubilizing fragmented sarcoplasmic reticulum (FSR) with a nonionic detergent (C12E8) then displacing the detergent with Tween 80, using a DEAE-cellulose column. The UV absorption of SSR decreased reversibly at about 286 and 292 nm on removal of free Ca2+ ions, while no change in the fluorescence spectrum was detectable. On the other hand, the fluorescence intensity of FSR decreased 3-4% on removal of free Ca2+ ions, as previously reported by Dupont [(1976) Biochem. Biophys. Res. Commun. 71, 544-550]. The UV absorption of FSR increased reversibly at about 270-280 nm on removal of free Ca2+ ions, but the rate of the change was very slow (k = about 0.1 min-1).     

Ibaraki Virus,an Agent of Epizootic Disease of Cattle Resembling Bluetongue

Replication of Ibaraki virus was not inhibited by 5-iodo-2′-deoxyuridine, indicating that the virus is an RNA virus. The virus was resistant to ether, chloroform and deoxycholate, sensitive to trypsin, very labile at acidic pH but stable at pH 6.4 or higher, and was resistant to repeated freezing and thawing. The virus was readily inactivated at 56 C or higher, was fairly stable at 37 C, and very stable at 4 C, while it rapidly lost infectivity when stored frozen at —20 C. The virus was readily sedimented by centrifugation at 40 000Xg for 60 min. It readily passed through membrane filters of 200 mμ pore size, passed through 100 μfilters but only with some titer loss and did not through 50 mμ filters. In these tests, the bluetongue virus used as a control behaved in the same manner as Ibaraki virus. These findings provide additional evidence for the similarity of Ibaraki virus to bluetongue virus which had been previously demonstrated on the basis of seasonal incidence, symptomatology and pathology of the diseases caused by these viruses and the behavior of the viruses in cell cultures, embryonated eggs and laboratory animals. The present study, however, provided no evidence for any serological relation between these two viruses. More Information is needed to reach a final decision on the classification of Ibaraki virus, particularly regarding the morphology of the virion, the doublestrandedness of the viral RNA and other basic features.     

Control of the growth of Alicyclobacillus acidoterrestris in industrialized orange juice using rosemary essential oil and nisin

The use of rosemary essential oil (RO) and its combination with nisin (RO+N) in preventing the multiplication of Alicyclobacillus acidoterrestris in orange juice was evaluated. The minimum inhibitory and bactericidal concentrations (MIC and MBC) for RO were both 125 μg ml⁻¹ while RO+N displayed a synergistic effect. The use of RO and RO+N at concentrations of 1, 4 and 8× MIC in orange juice for 96 h was evaluated in terms of their sporicidal effectiveness. With regard to the action against A. acidoterrestris spores, RO at 8× MIC was sporostatic, whereas RO+N at 1× MIC was sporicidal. Morphological changes in the structure of the micro-organism after treatment were also observed by microscopy. Furthermore, flow cytometric analysis showed that most cells were damaged or killed after treatment. In general, the antioxidant activity after addition of RO+N decreased with time. The results demonstrate that using the combination of RO and nisin can prevent the A. acidoterrestris growth in orange juice.     

Involvement of the Binuclear Copper Site in the Proteolytic Activity of Polyphenol Oxidase

Plant polyphenol oxidase (PPO) is apt to degrade during andeven after purification. We developed a method to stabilizePPO by 0.3 M NaCl, 0.1% (w/v) Tween 20, and 50% (w/v) ethyleneglycol at pH 6.5. The protein slowly degraded by itself whenthe stabilizing reagents were removed. Ascorbate and/or H₂O₂accelerated the degradation. The ascorbate-induced degradationwas inhibited by catalase, suggesting that H₂O₂ is generatedthrough reduction of PPO by ascorbate. It is likely that dissolvedoxygen is converted to peroxide through two-electron reductionby the reaction center of PPO, binuclear Cu site, and a Fenton-typereaction occurred on it. This understanding was supported bythe finding that the H₂O₂-induced degradation was inhibitedby metal-chelators as well as by polyphenolic substrate of PPO.Considering the postulated mechanism of the self-degradationof PPO, we re-examined the degradation of the 23-kDa proteinof PSII by PPO [Kuwabara et al. (1997) Plant Cell Physiol. 38:179]. The obtained results suggested that the 23-kDa proteintriggers the active oxygen production by the binuclear Cu site,probably as reductant, and receives the radical species preferentiallyto the polypeptide moiety of PPO. (Received April 15, 1999; Accepted July 21, 1999)     

Sequences of cDNAs for human salivary and pancreatic α-amylases

The nucleotide sequences of the cloned human salivary and pancreatic α-amylase cDNAs correspond to the continuous mRNA sequences of 1768 and 1566 nucleotides, respectively. These include all of the amino acid coding regions. Salivary cDNA contains 200 bp in the 5′-noncoding region and 32 in the 3′-noncoding region. Pancreatic cDNA contains 3 and 27 bp of 5′- and 3′-noncoding regions, respectively. The nucleotide sequence humology of the two cDNAs is 96% in the coding region, and the predicted amino acid sequences are 94% homologous.Comparison of the sequences of human α-amylase cDNAs with those previously obtained for mouse α-amylase genes (Hagenbuchle et al., 1980; Schibler et al., 1982) showed the possibility of gene conversion between the two genes of human α-amylase.     

Degradation of aromatic amino acids by Rhodococcus erythropolis

A Gram-positive Rhodococcus erythropolis strain S1 was shown to assimilate aromatic amino acids such as L-phenylalanine, L-tyrosine, L-tryptophan, D-phenylalanine, D-tyrosine and D-tryptophan, which were utilized not only as the sole carbon source but also as a suitable nitrogen source. The highest growth on these aromatic amino acids occurred at a temperature of 30°C. L-Phenylalanine, L-tyrosine and L-tryptophan degradative pathways would appear to be independent, and to be induced alternatively. The strain S1 also showed the ability to assimilate peptides which consisted of only L-phenylalanine and L-tyrosine.     

Application of a new slot comb to electrophoretic analysis.

In the present experiment, a new slot comb was designed in order to form a wide and sloped sample well on the stacking gel of electrophoresis. Using this slot comb, a gradient of the reagent layer of interest can be easily formed transversely on the gel that is perpendicular to the direction of electrophoresis. Thus, the protein sample overlaid on the agent migrates across the gradient layer during electrophoresis to produce a continuous electrophoretic band reflecting the interaction between the protein and reagent. This new slot comb (tentatively called slope comb) was applied to the following two experiments. In the first experiment, in combination with this comb and a reducing agent, 2-mercaptoethanol, the reducing steps of cross-linked axonemal proteins with o-iodosobenzoic acid (OIBA) were analyzed electrophoretically, enabling visualization of the reducing pattern of each axonemal protein in a single experiment. The results obtained indicate that alpha and beta tubulins are cross-linked differently by OIBA. In the second experiment, the formation of the Ca2+ gradient layer using this slope comb could electrophoretically differentiate the Ca2+ sensitivity of three Ca(2+)-binding proteins.     

Gelation mechanism of agarose

The non-Newtonian behavior and dynamic viscoelasticity of a series of aqueous solutions of agarose were measured with a rheogoniometer. The flow curve, at 25°, of agarose solution approximated to plastic behavior at 0.1, 0.13, and 0.15% concentrations. Gelation occurred at concentration of 0.13% at low temperature (0°). The dynamic modulus of agarose showed a very high value at low temperature, and increased with an increase in temperature, showing a maximum value at 30°, then it decreased. In the presence of NaCl, KCl, CaCl₂, and MgCl₂ for a solution of agarose at 0.08% concentration, the transition temperature, at which dynamic modulus decreased rapidly, was observed at 60°. Gelation was also observed at low temperature (0°) in acid and alkaline range after reaching pH values of 2.3 and 9.5, respectively, by addition of 100m HCl, H₂SO₄, NaOH, and Ca(OH)₂ to a 0.08% agarose solution. A possible mode of intra- and inter-molecular hydrogen bonding within and between the agarose molecules in aqueous solution is proposed.