Purification and characterization of HBs antigen from hepatoma huGK-14 cell line

HBs antigen was purified from the culture fluid of hepatoma huGK-14 cell line and its physico-chemical properties were studied. The purification consists of following steps: concentration of culture fluid by membrane filtration, affinity column chromatography (anti-HBs monoclonal antibody column and anti-human serum albumin antibody column), and ultracentrifugation (isopycnic centrifugation in CsCl density gradient and rate zonal centrifugation on sucrose gradient). Highly purified (purity>99%) HBs antigen was isolated with an overall yield of about 40%. The HBs antigen showed uniform spherical particles (diameter: 23.2±2.9nm) and had a specific gravity of 1.20g/cm3. The purified HBs antigen yielded, in SDS-PAGE (under reducing conditions), four protein bands with apparent molecular weights of 22,000 and 26,000 (the two major bands), and 44,000 and 47,000. The two proteins of molecular weights of 26,000 and 47,000 are likely to be glycosylated, as these were several fold reduced when the cells were cultured in the presence of Tunicamycin. Amino acid analysis, Edman degradation, carboxypeptidase digestion, and ultraviolet absorption spectrum indicated that the HBs antigen from hepatoma cells is very similar to that derived from human plasma

K. 抗菌抗カビ活性を有する水溶性銀 (1) 錯体の分子設計と合成

チオサリチル酸やチオリンゴ酸を配位子とする銀(1)錯体はバクテリア、酵母やかび類のいくつかに対して特異的な抗菌活性を示す。配位子自身には抗菌作用がなく、また銀(1)イオンには二種類のバクテリアにしか抗菌作用を示さないので、有機配位子による銀(1)錯体形成により新しい抗菌活性が発現したといえる。この事実を背景として、本研究では銀(1)-窒素結合や銀(1)-酸素結合をもつ水溶性銀(1)錯体で、幅広いスペクトルの抗菌活性もつ銀(1)錯体を、分子設計により合成することを目的とした。これまで錯体を利用した抗菌剤の開発の例は非常に少なく、また抗菌作用をきちんと証明した抗菌剤も少ないので、本研究は新規性が高く波及効果も大きい。またバクテリア、酵母だけでなくかび類にも効果が期待でき、水溶性であるので繊維、紙、木材製品への幅広い利用が考えられる。13種類のアミノ酸配位子、11種類の含カルボキシル基複素環配位子を検討した結果、アミノ酸配位子の中ではヒスチジンおよびアスパラギン酸、また含カルボキシル基複素環配位子の中ではピロリドンカルボン酸およびオキソテトラヒドロフランカルボン酸が本研究で目的とする広いスペクトルの抗菌抗カビ活性を示す水溶性銀(1)錯体を形成することがわかった

L. チオセミカルバゾン三座配位子による金属錯体の合成と抗菌活性

The reaction of Ni(OAc)_2 with N\u27-[1-(2-pyridyl) ethylidene] morpholine-4-carbothiohydrazide (HL) afforded two nickel (II) complexes, [Ni(L)_2] (1) and [Ni(L)(OAc)] (2), depending on reaction temperature and starting molar ratio; the former was a 6-coordinate, paramagnetic complex and the latter a 4-coordinate, diamagnetic complex. X-ray analysis of the complex 2 revealed that the ligands, L and OAc, were coordinated to the central metal ion to form a square-planar geometry. The complex 2 showed enhanced antimicrobial activities against selected Gram-positive bacteria, compared with those of the ligand alone, whereas the other complex 1 showed no activity

<所内学術研究成果報告>M. 抗菌抗カビ活性を有する水溶性銀 (I) 錯体の分子設計と合成

チオサリチル酸やチオリンゴ酸を配位子とする銀(I)錯体はバクテリア,酵母やかび類のいくつかに対して特異的な抗菌活性を示す。配位子自身には抗菌作用がなく,また銀(I)イオンには二種類のバクテリアにしか抗菌作用を示さないので,有機配位子による銀(I)錯体形成により新しい抗菌活性が発現したといえる。前年度に引き続き,本研究では銀(I)-窒素結合や銀(I)-酸素結合をもつ水溶性銀(I)錯体で,幅広いスペクトルの抗菌活性もつ銀(I)錯体を,分子設計により合成することを目的としている。13種類のアミノ酸配位子,11種類の含カルボキシル基複素環配位子を検討した結果,アミノ酸配位子の中ではヒスチジンおよびアスパラギン酸,また含カルボキシル基複素環配位子の中ではピロリドンカルボン酸およびオキソテトラヒドロフランカルボン酸が本研究で目的とする広いスペクトルの抗菌抗力ビ活性を示す水溶性銀(I)錯体を形成することがわかった。これらの錯体の中で4個の銀(I)錯体について,X線結晶構造を決める事が出来た。固体状態および溶液中の構造の違いを(^Cおよび^N)NMR測定からも調べた。構造と抗菌性の関係を考察した。銀(I)錯体の抗菌活性のスペクトルは銀(I)錯体の配位子交換性と密接に関係していることを明らかにした

I. <種々の複素環窒素配位子による銀 (I) および金 (I) 錯体の合成と生理活性>

(1) Two novel polymeric silver (I)-triphenylphosphine complexes with nitrogen-containing heterocycles, [Ag (1,2,3-triz) (PPh_3)_2]_n 3 and [Ag (1,2,4-triz) (PPh_3)_2]_n 4 (Htriz=triazole) were synthesized from reactions of polymeric precursors [Ag (1,2,3-triz)]_n 1 and [Ag (1,2,4-triz)]_n 2 with 3 equiv of PPh_3 in CH_2Cl_2,respectively. The polymeric precursors 1 and 2 have been obtained as a non-crystalline, colorless powder-solid : 2 showed a wide spectrum of excellent antibacterial activities, but 1 did not. The antimicrobial activities of the complexes 1-4,evaluated by MIC (minimum inhibitory concentration), were compared and key factors affecting them were discussed. The crystal structures of the complexes 3 and 4 were determined by single-crystal X-ray diffraction. The complexes 3 and 4 in the solid-state were helical polymers consisting of AgN_2P_2 cores formed by bridging triazolate anions and two PPh_3 ligands. The complexes 3 and 4 were also fully characterized by elemental analyses, TG/DTA and FT-IR in the solid-state and by various solution NMR (^P, ^Ag, ^1H and ^C) spectroscopies and molecular weight measurements in solution. (2) Two novel gold (I)-triphenylphosphine complexes with nitrogen-containing heterocycles, [Au (1,2,3-L) (PPh_3)] 1 and [Au (1,2,4-L) (PPh_3)]_2・xH_2O (x=0.5-1.0) 2 (HL=triazole) were synthesized from stoichiometric reactions of a precursor complex [AuCl (PPh_3)] with HL in acetone in the presence of aqueous NaOH, and isolated as colorless needles and cubic crystals, respectively. The crystal structures of 1 and 2 were determined by single-crystal X-ray diffraction. 1 and 2 were also fully characterized by complete elemental analyses, TG/DTA and FT-IR in the solid-state and by solution NMR (^P, ^1H and ^C) spectroscopies and solution molecular-weight measurements. 1 consisted of a monomeric 2-coordinate AuNP core both in the solid-state and in solution, while, in contrast, 2 comprised a dimeric (AuNP)_2 core through an Au-Au bond in the solid-state, but a monomeric AuNP core in solution. Within the two gold (I) complexes composed of very closely related nitrogen-containing heterocycles and a common bulky PPh_3,ligand, it was found that the aggregation through the Au…Au interaction in 2 was overruled in 1. The molecular structures of 1 and 2 were also compared with those of the corresponding silver (I) analogs, [Ag (1,2,3-L) (PPh_3)_2]_n 3 and [Ag (1,2,4-L) (PPh_3)_2]_n 4,the molecular structures of both of which have been recently determined as helical polymers in the solid-state. (3) Polymeric silver (I) imidazolate [Ag (imd)]_n (1; Himd=imidazole, C_3H_4N_2) showing a wide spectrum of excellent antimicrobial activities against bacteria, yeast and mold, which has been obtained as non-crystalline, colorless powder, was successfully converted to molecular, crystalline complex [Ag (imd) (PPh_3)_3] 2 by a reaction of 1 with 3 equivalents of PPh_3 in CH_2Cl_2. This novel complex 2 was not obtained from reactions in solutions of the precursors [AgCl (PPh_3)_3] and/or [AgCl(PPh_3)_2]_2 with Himd in the presence of NaOH or with sodium salt of imd. On the contrary, the corresponding gold (I) complex [Au (imd) (PPh_3)] 3 was formed by a reaction of the precursor [AuCl (PPh_3)] and Himd in the presence of NaOH. From a synthetic viewpoint, these facts show the ordering of ligand replacement in these complexes to be Ag-P>>Ag-Cl>Ag-N, while Au-P>>Au-N>Au-Cl. This ordering also exhibits the different reactivity of silver (I) and gold (I) complexes with biological ligands such as proteins, enzymes, nucleic acids and amino acids. The complex 2 was fully characterized by complete elemental analyses, TG/DTA, FT-IR, solution molecular weight measurement and various solution NMR (^1H, ^C, ^P and ^Ag) spectroscopies. The complex 2 in solution was present in a rapid equilibrium with the dissociated species of two PPh_3 ligands, i. e., [Ag (imd) (PPh_3)]. The ^P NMR measurements at low temperature showed that (i) all PPh_3 ligands in 2 in CHCl_3 solution migrated at room temperature between the two isotopes ^Ag and ^Ag, in addition to the dissociation of two PPh_3 ligands, and (ii) there were present the 4-coordinate tetrahedral complex 2 and the dissociated complex [Ag (imd) (PPh_3)] in solution

Problems in three Japanese drug users with Human Immunodeficiency Virus infection

Numbers of individuals infected with Human Immunodeficiency Virus (HIV) are increasing in Japan. The majority of them are Men who have sex with men and a part of them take drugs as ‘Sex drug’ at their sexual intercourse. Especially, Amyl nitrite, Methamphetamine, 5-methoxy-N, N-diisopropyltryptamine (5-MeO-DIPT Foxy), and 3, 4- methylenedioxy- methamphetamine (MDMA Ecstasy) are used, and they sometimes cause the physical and mental disorders. However, the actual drug inducing troubles among Japanese HIV-infected drug users had not yet been discussed enough. In this report, we describe three cases with HIV infection a case developed severe neuroleptic malignant syndrome (NMS) after taking 5-MeO-DIPT, a case with persistent convulsion due to multiple drug intake and a case with rhabdomyolysis due to the nonsubjective methamphetamine intake. Through these cases, we raise and discuss several underlying problems associated with drug use among HIV-infected individuals

Complete Sequencing and Pan-Genomic Analysis of Lactobacillus delbrueckii subsp. bulgaricus Reveal Its Genetic Basis for Industrial Yogurt Production

Lactobacillus delbrueckii subsp. bulgaricus (Lb. bulgaricus) is an important species of Lactic Acid Bacteria (LAB) used for cheese and yogurt fermentation. The genome of Lb. bulgaricus 2038, an industrial strain mainly used for yogurt production, was completely sequenced and compared against the other two ATCC collection strains of the same subspecies. Specific physiological properties of strain 2038, such as lysine biosynthesis, formate production, aspartate-related carbon-skeleton intermediate metabolism, unique EPS synthesis and efficient DNA restriction/modification systems, are all different from those of the collection strains that might benefit the industrial production of yogurt. Other common features shared by Lb. bulgaricus strains, such as efficient protocooperation with Streptococcus thermophilus and lactate production as well as well-equipped stress tolerance mechanisms may account for it being selected originally for yogurt fermentation industry. Multiple lines of evidence suggested that Lb. bulgaricus 2038 was genetically closer to the common ancestor of the subspecies than the other two sequenced collection strains, probably due to a strict industrial maintenance process for strain 2038 that might have halted its genome decay and sustained a gene network suitable for large scale yogurt production