Sei-ichi Manabe

The Particle Size Of Hepatitis C Virus Estimated By Filtration Through Microporous Regenerated Cellulose Fibre

To estimate the particle size of hepatitis C virus (HCV), a major causative agent of post-transfusion non-A, non-B hepatitis, we filtered plasma or serum samples through microporous cellulose fibres with different pore sizes. The amount of HCV particles in samples before and after filtration was determined by a quantitative reverse transcriptase polymerase chain reaction (PCR) method. Since there is no quantitative biological assay for HCV, except for that in chimpanzees, the HCV titre obtained from the PCR method was used in an equation constructed previously for application to filtration experiments with a flavivirus which is distantly related to HCV. The particle was estimated to be between 30 and 38 nm in diameter, although the possibility remained that larger HCV particles or HCV aggregates with a diameter of more than 39 nm might exist. Double-step filtration through microporous cellulose fibres with a pore size of 35 nm reduced the HCV content to below levels detectable by our PCR method, indicating that it is possible to eliminate HCV particles by simple filtration techniques.

A novel method for removal of human immunodeficiency virus: Filtration with porous polymeric membranes

Abstract. We propose a new method to rid solutions of a virus by using a novel regenerated multilayered structured cellulose membrane (BMM). When the filtrate of human immunodeficiency virus (HIV) preparation was obtained through BMM it showed no infectivity. Electron microscopic observation revealed that HIV was completely caught by the multilayers of the BMM. Conveniently, BMM was seldomly found to adsorb protein molecules and also to have a high filtration rate. These characteristics may have a use in the removal of other variously sized pathogenic agents from plasma.

Rejection of single stranded and double stranded DNA by porous hollow fiber membranes

Abstract The permeation of DNA through regenerated cellulose hollow fibers that have a mean pore diameter of 15 nm was investigated in the absence of protein at a transmembrane pressure of 200 mmHg at 25°C. The membranes can reject both double stranded DNA and single stranded DNA to some extent when the DNA concentration of the feed solution is 10–100 ppm. It was found that single stranded DNA can permeate through the membranes preferentially over double stranded DNA. This is explained by the higher flexibility of the single stranded DNA over the double stranded DNA and is not mainly due to a different distribution of molecular weights between the single stranded DNA and the double stranded DNA. The circular dichroism spectra of feed and permeate solutions suggest that the DNA was not denatured by permeation through the membranes.

Sieving study of chromatin and histone-DNA complex by porous hollow fiber membranes

Abstract Permeation of chromatin isolated from human HeLa S3 cells and histone-DNA complex through regenerated cellulose hollow fibers was investigated as a model study of DNA removal for drug manufacturing using membrane technology. It was found that the permeation of histone-DNA and chromatin through the membranes having a mean pore diameter of 15 nm was substantially lower than the permeation of free DNA which did not complex with proteins based on the concentration determination of the feed and permeate solutions using UV spectroscopy. Direct observation to determine the existence of chromatin and histone-DNA complex in the permeate solution was also performed using atomic force microscopic imaging. No chromatin and chromatin-like structures were found in the examination of 10 areas of 1.0×1.0 μm on the mica which had been adsorbed from the solution permeated through the membranes. Thus, the membranes having a mean pore diameter of 15 nm most likely reject the chromatin and histone-DNA complex that are observed to have an apparent width of 25–30 nm based on atomic force microscopic imaging.

Filtration of PrP

A protease resistant isoform of the prion protein (PrPSc) is found in transmissible spongiform encephalopathies; Kuru, Creutzfeldt-Jakob disease (CJD), and Gerstmann-Straussler-Scheinker syndrome (GSS) in humans, and scrapie in various animal species (Prusiner, 1982). Experimental inoculation of PrPSc into laboratory animals causes spongiform encephalopathy after long incubation periods. Infectivity of PrPSc is highly resistant for chemico-physical inactivation procedures, such as formalin fixation for many years and heating at 121°C for 4 hours (Tateishi et al., 1980). Here we report the effective removal of PrPSc using validatable filter BMMR.

Novel membrane separation method using liquid and porous polymeric membranes

Abstract A novel method is proposed to effectively separate a substance from its aqueous solution by using emulsion type liquid membranes (emulsion particles) and porous polymeric membranes. The validity of the proposed separation method is confirmed by a model experiment of ethanol (or acetone) separation from its aqueous solution revealing that (1) the interfacial tension between emulsion particles and matrix can be kept constant, irrespective of the transport of EtOH into emulsion particles, (2) the mobility of emulsion particles can be controlled by adding a surfactant and imposing an electrostatic gradient, (3) the emulsion particles can be separated from the matrix by filtration through a polymeric porous membrane, and (4) using a carrier may result in a higher separation factor than that attained under therodynamic equilibrium conditions. A continuous system for separation of tryptophan from its aqueous solution was designed based on the above novel method and a considerably high separation factor was obtained.

Change in Supermolecular Structure of Nylon 66 Fiber with Time Elapsed

ナイロン66繊維の放置過程における超分子構造, 特に結晶および無定型領域内部の分子鎖の凝集構造の経時変化について検討した.ナイロン66繊維は室内および室外に約1000日放置した.その間の超分子構造の変化は広角X線回折, 干渉顕微鏡, 動的粘弾性によって評価した.その結果, 放置時間の経過に伴ってナイロン66繊維の結晶領域においては, 結晶は大きくなり, その完全度も高くなる.また配向度は小さくなる.無定型領域においては, 放置時間の経過に伴って, 分子鎖はいったん緊張状態になり, その後緩和する.さらに900日を経過すると分子鎖の充?状態の密な部分が新たに出現する.これらの変化は室外放置試料において著しい.

Theoretical Requirements for The Integrity Test Method and Its Practical Application

Recently, process validation of virus inactivation/removal has been requested in the purification process of biopharmaceutical products, by the regulatory agencies. Consequently, various kinds of virus inactivation/removal methods have been developed and introduced to the production process of biopharmaceuticals.

Characteristics of plasma filtration by microporous regenerated cellulose hollow fiber as virus-removal membrane.

The Bemberg Microporous Membrane (BMM) is composed of microporous regenerated cellulose hollow fiber and has been developed for virus removal from plasma. When fresh plasma was appiled on the BMM-30 (average pore size 30nm), 93 percent of albumin was passed through. However, more than 50 percent of IgM and Factor VIII were trapped in the hollow fiber. On the other hand with the BMM-50 filtration, 70 percent of IgM and Factor VIII were recovered in the filtrate. The double filtration by BMM-50 modules was carried out for HB virus positive fresh frozen plasma. Trapping of plasma components occurred only at the first BMM filtration and there was virtually no trapping at the second BMM filtration. HBV-DNA in Dane particles could be effectively removed by the double BMM-50 filtration. The residual amount of HBV-DNA in the first filtrate became under the detectable level after the second filtration. Thus, HBV-free plasma with sufficient amounts of plasma components could be obtained by the double filtration with the BMM-50 modules.

DELUSTRING MECHANISMS OF CELLULOSE DIACETATE FILM DURING HEAT-TREATMENT WITH HOT WATER

The delustring phenomena, observed often during heat-treatment of cellulose diacetate (CDA) in hot water, were studied in connection with the molecular characteristics of CDA and the thermal motion of CDA molecules. The phenomena accompany with decreases in (1) density, (2) weight remaining, and (3) storage modulus, and disappeared by treatment of film under high pressure or by dipping it into good solvents for CA with relatively low degree of acetic acid content (AC). The delustring phenomena can be attributed to a change from uniformly packed state of CDA molecules in the original film to highly heterogeneously packed state by dissolution and/or swelling of CA molecules of low acetyl content and of low molecular weight and CA molecules having the groups of sulfonic ester and carboxylic acid as impurities during the course of heat-treatment in hot water. Such a change is closely associated with the microbrownian motion of main chains in the amorphous region. Water molecules act selectively on the CA molecules of low acetyl content and of low molecular weight as a dissolving, swelling, and plastisizing agent.