Optical Configuration Analysis of Hydroxy Fatty Acids in Bacterial Lipids by Chiral Column High-Performance Liquid Chromatography

Through the adoption of a chiral stationary phase in high-performance liquid chromatography and a simple derivatization method for hydroxy fatty acids, it became easy to separate and identify the optical isomers of 2- and 3-hydroxy fatty acids composing several kinds of microbial lipids. The 2- and 3-hydroxy fatty acids were converted with dinitrophenyl isocyanate to their 3, 5-dinitrophenyl urethane derivatives (DU-derivatives), which were analyzable by HPLC using a chiral column. By varying the composition of an eluent, separation of the DU-derivatives of hydroxy fatty acids differing in optical configuration, chain length and position of hydroxyl group was achieved. The general elution orders of these DU-derivatives were determined with authentic 2- and 3-hydroxy fatty acids. Small amounts (~300 μg) of ornithine-containing lipids isolated from the Serratia marcescens strains were examined by this method to identify 3-hydroxy fatty acids of the lipids as D isomers.     

A type kappa Bence Jones protein containing a cysteinyl residue in the variable region.

The proteins precipitated with ammonium sulfate from the urine of a patient (Mat) with multiple myeloma were separated into three components by ion-exchange and gel chromatographies. Sodium dodecyl sulfate polyacrylamide gel electrophoresis, amino acid analyses, immunochemical tests, and measurement of circular dichroism showed that these components were a dimer with a disulfide bond, a stable monomer, and a variable fragment, respectively. All three protein components reacted with 5,5'-dithiobis-(2-nitrobenzoic acid) in Tris-HCl buffer at pH 8.0, indicating that they contained free sulhydryl groups. Partial reduction with dithiothreitol in the absence of denaturants yielded two SH groups per molecule from both the monomer and the dimer, and one SH group per molecule from the fragment. This indicates that the monomer of Mat protein contains a cysteinyl residue in the variable region in addition to a cysteinyl residue at the COOH terminus.The reactivities of the two SH groups of the partially reduced monomer toward iodoacetamide and iodoacetic acid were studied by polyacrylamide gel electrophoresis. The two SH groups had similar reactivities with iodacetamide, but the SH group at the COOH terminus was more reactive with iodoacetic acid than that in the variable region. The extrinsic Cotton effects of an azobenzene-2-sulfenyl group introduced into the SH group in the variable region were different from those of dye attached to the COOH terminal SH group, indicating that the two SH groups had different environments. The states of the SH groups of the intact monomer are discussed on the basis of these findings.     

Mode of protection of mice against herpes simplex virus type 2 infection by Propionibacterium

We compared various strains of Propionibacterium with regard to protection of young adult mice against lethal infection with herpes simplex virus type 2 (HSV-2). Propionibacterium acnes, P. granulosum, and P. avidum were protective, while P. acidi-propionici and P. lymphophilum were ineffective. The protective effect proved to be in the cell wall fraction. Attempts were made to elucidate possible mechanisms of the protection using both effective and ineffective strains. The results strongly suggest that induction of interferon rather than activation of macrophages and natural killer cells by Propionibacterium pretreatment plays a crucial role, directly or indirectly, in protection against infection by herpes simplex virus. Propionibacterium only moderately protected newborn mice against HSV-2 infection.     

Effect of interferon on the inhibition of malignant cell growth by human peripheral leukocytes. III. Effect of systemic administration

Human leukocyte interferon preparation (HuIFN-alpha LE) was given to the patients with cancer or with chronic hepatitis. Spontaneous tumor cell growth inhibition by human peripheral lymphocytes (STGI) and NK activity were enhanced by the systemic administration of HuIFN-alpha LE, although there were differences in the kinetics between the two activities after one time administration or by the repeated administration. This suggests that IFN acts indirectly on the tumor cells by the medium of normal lymphocytes or NK cells, and that tumor cell growth inhibition is different from NK activity.     

Isolation and characterization of hemorrhagic factors a and b from the venom of the Chinese habu snake (Trimeresurus mucrosquamatus)

Hemorrhagic factors a and b were isolated from Trimeresurus mucrosquamatus venom by Sephadex G-100, CM-Sephadex C-50 and DEAE-Sephacel column chromatographies. The hemorrhagic factors were homogeneous, as established by a single band on acrylamide gel electrophoresis, isoelectric focusing and sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Molecular weights of 15 000 and 27 000 were found for hemorrhagic factors a and b, respectively. Factor a possesses proteolytic activity hydrolyzing the His(10)-Leu(11), Tyr(16)-Leu(17) and Arg(22)-Gly(23) bonds of oxidized insulin B chain, whereas, factor b hydrolyzed only the Ala(14)-Leu(15) bond. Hemorrhagic activity of these hemorrhagic factors was inhibited by ethylenediaminetetraacetic acid, 1,10-phenanthroline or p-chloromercuribenzoate, but not by soybean trypsin inhibitor or diisopropyl fluorophosphate. The hemorrhagic factors were injected into the skin of the back of albino rabbits, and the minimum hemorrhagic dose of factors a and b was 1.7 and 2.3 μg, respectively. These purified hemorrhagic factors were not lethal at 15 μg/g in mice. Factor a hydrolyzed the Bβ chain of fibrinogen, while factor b hydrolyzed the Aα chain. Hemorrhagic factor a was shown to differ immunologically from factor b. Factors a and b produced systemic hemorrhage in internal organs such as the heart and stomach of mice. Moreover, factor b produced hemorrhage in the liver.     

Update on a titration method for measuring the interferon-producing capacity in humans

To make it possible to measure the interferon-alpha (IFN-alpha) producing capacity in a great number of healthy donors and patients, we developed the simple method (the whole blood method). For the measurement of the IFN-producing capacity, the heparinized blood was incubated with Sendai virus at 37 degrees C for 20 hours. The IFN activity of the culture supernatants was determined by the cytopathic effect inhibition assay. We measured the IFN-producing capacity in 531 healthy donors and 130 cancer patients. The results showed that the IFN-producing capacity in cancer patients was significantly lower than that in healthy donors. Although there were individual variations in the IFN-producing capacity, no age and sex differences were observed. These results indicate that this method is useful for the measurement of IFN-producing capacity in human.