Filtration sterilizing

Microorganisms in liquids and gases can be removed by microfiltration hence, air supplied to aerobic fermentors can be sterilized in this way. Membrane filters are often used for the sterilization of liquids, such as culture media for fermentation (especially for tissue culture), and also for the removal of microorganisms from various fermentation products, the heating of which should be avoided. 

In selecting a membrane material, its pH compatibility and wettability should be considered. Some hydrophobic membranes require prewetting with a low-surface-tension solvent such as alcohol, whereas cartridges containing membranes are often presterilized using gamma irradiation. Such filter systems do not require assembly and steam sterilization. 

Naturally, the size of the membrane pores should depend on the size of microbes to be filtered. For example, membranes with 0.2 pm pores are often used to sterilize culture media, while membranes with 0.45 pm pores are often used for the removal of microbes from culture products. It should be noted here that the so-called pore size of a membrane is the size of largest pores on the membrane surface. The sizes of the pores on the surface are not uniform moreover, the pore sizes usually decrease with increasing depth from the membrane surface. Such a pore size gradient will increase the total filter capacity, as smaller microbes and particles are captured within the inner pores of the membrane. 

The so-called bubble point of a membrane - a measure of the membrane pore size - can be determined by using standard apparatus. When determining the bubble point of small, disk-shaped membrane samples (47 mm in diameter), the membrane is supported from above by a screen. The disk is then flooded with a liquid, so that a pool of liquid is left on top. Air is then slowly introduced from 

The degree of removal of microbes of a certain size by a membrane is normally expressed by the reduction ratio, R. For example, if a membrane of a certain pore size is fed 107 microbes per cm2 and stops them all except one, the value of log reduction ratio log R is 7. It has been shown [8] that a log-log plot of R (ordinate) against the bubble points (abscissa) of a series of membranes will produce a straight line with a slope of 2. 

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Steric stabilization Sterilant Sterilants Sterile filling Sterile filtration... 

The separation of cells from the culture media or fermentation broth is the first step in a bioproduct recovery sequence. Whereas centrifugation is common for recombinant bacterial cells (see Centrifugal separation), the final removal of CHO cells utilizes sterile-filtration techniques. Safety concerns with respect to contamination of the product with CHO cells were addressed by confirming the absence of cells in the product, and their relative noninfectivity with respect to immune competent rodents injected with a large number of CHO cells. 

Adsorption of t-PA to process equipment surfaces consisting of either stainless steel or glass was minimized by adding the detergent polyoxyethylene sorbitan monooleate (Tween 80) to the semm-free culture conditioned media at 0.01% (vol/vol). The equipment was also rinsed, before use, with phosphate buffered saline (PBS) containing 0.01% Tween 80. Hydrophilic, plastic equipment was used whenever possible. AH buffers were sterile filtered. Sterile filtration of Hquids and gases is usually carried out using 0.2 or 0.45 p.m filters. 

The earliest commercially available filters were manufactured in two pore sizes 0.45 and 0.8 pm. The 0.45 pm-rated membranes were considered to be stefilizing-grade filters and were successfully used in the sterile filtration of pharmaceuticals and parenterals. The membrane filters were qualified using Serratia marcescens a standard bacterium, having dimensions of 0.6 x 1 pm. However, in the late 1960s it became apparent that the matrix of the 0.45 pm-rated filters could be penetrated by some pseudomonad-like organisms (1). For sterile filtration apphcations in the 1990s, 0.2 pm-rated membranes are the industry standard in the manufacture of sterile parenterals and pharmaceuticals. 

Sterile Filtration of Gases. Primary appHcations for sterile gas filtration are the sterilization of fermentor inlet air, fermentor vent gas, vents on water for injection tanks, and vacuum break filters during lyophilization. Operational and process considerations apply. Typically, the membrane in gas... 

Formulation Sterile filtration/ dispensing Freeze drying... 

Then, 1.3 ml of glycerine are mixed with 0.5 ml of a 25% solution of methyl p-hydroxy-benzoate in ethanol, and 50 ml of distilled water are added. To the produced mixture are, after sterile filtration, added 10 ml of the stock solution 1, 2.5 ml of the stock solution 2 and 10 ml of the stock solution 3, after which 3.0 ml of sterile 0.1 N sodium hydroxide are added, and the mixture is filled up with sterile distilled water to a volume of 100 ml. The insulin will be precipitated amorphously by the admixture of the sodium hydroxide, and the produced suspension acquires the pH value of 7. It will contain approximately 1 gamma zinc per insulin unit. 

Downstream Processing Microfiltration plays a significant role in downstream processing of fermentation products in the pharmaceutical and bioprocessing industry. Examples are clarification of fermentation broths, sterile filtration, cell recycle in continuous fermentation, harvesting mammahan cells, cell washing, mycelia recovery, lysate recovery, enzyme purification, vaccines, and so forth. 

Sterile filtration, with subsequent aseptic filling, is common because of the heat sensitivity of many drugs. Those drug solutions that can withstand heat should be terminally autoclave sterilized after filling, since this best assures product sterility. 

The USP also recommends the use of biological indicators, whenever possible, to monitor all sterilization methods except sterile filtration. Biological indicators are generally of two types. If a product to be sterilized is a liquid, microorganisms are added directly to carefully identified representative samples of the product. When this is not practical, as with solids or equipment to be sterilized, the culture is added to strips of filter paper. The organism chosen varies with the method of sterilization. 

All production processes, such as ampoule washing and sterilization, solution filtration, equipment set-up and operation, sorting, and freeze-drier cleaning and operation, should be covered in detail in a procedure manual to ensure that all operations are understood as well as carried out properly and uniformly. Cleaning, sterilization, sterile filtration, filling, and aseptic processing operations must be validated. 

In general, aqueous ophthalmic solutions are manufactured by methods that call for the dissolution of the active ingredient and all or a portion of the excipients into all or a portion of the water and the sterilization of this solution by heat or by sterilizing filtration through sterile depth or membrane filter media into a sterile receptacle. If incomplete at this point, this sterile solution is then mixed with the additional required sterile components, such as previously sterilized solutions of viscosity-imparting agents, preservatives, and so on, and the batch is brought to final volume with additional sterile water. 

Aqueous suspensions are prepared in much the same manner, except that before bringing the batch to final volume with additional sterile water, the solid that is to be suspended is previously rendered sterile by heat, by exposure to ethylene oxide or ionizing radiation (gamma or electrons), or by dissolution in an appropriate solvent, sterile filtration, and aseptic crystallization. The sterile solid is then added to the batch, either directly or by first dispersing the solid in a small portion of the batch. After adequate dispersion, the batch is brought to final volume with sterile water. Because the eye is... 

Particular attention should be paid to nonstandard production technologies including nonstandard methods of sterilization, sterile filtration and aseptic processing, lyophilization, microencapsulation, and certain critical mixing and coating operations. With such processes pilot-scale manufacture may not be predictive of industrial scale manufacture, and data on three full-scale production batches may be required in the application. 

Where sterilization by filtration is used, more information will be required. The maximum acceptable bioburden limit prior to sterilization filtration is to be... 

Where terminal processing is not possible, the justification for alternative sterilization methods will be included in the EPAR, or at least a statement to the effect that sterile filtration/aseptic processing will be used. Presterilization bioburden issues that arose during the assessment will be included in the EPAR. 

Sugg and Hehre43 also obtained precipitin reactions with dextran or with sterile filtrates of sucrose broth cultures of L. mesenteroides (designated for convenience strain A) and not only anti-Leuconostoc sera, but also pneumococcus Types II, XII and XX antisera. Leuconostoc organisms cultured on D-glucose broth neither stimulated the production of dextran-reactive antibodies in rabbits, nor absorbed dextran-reactive antibodies from sera, as did organisms cultured on sucrose. Absorption with the homologous bacteria (Leuconostoc, pneumococcus Types II,... 

After its purification, sterile filtration and aseptic filling, human urokinase is normally freeze-dried. Because of its heat stability, the final product may also be heated to 60 °C for up to 10 h in an effort to inactivate any undetected viral particles present. The product utilized clinically contains both molecular mass forms, with the higher molecular mass moiety predominating. Urokinase can also be produced by techniques of animal cell culture utilizing human kidney cells or by recombinant DNA technology. 

Bacillus anthracis-denved antigens found in a sterile filtrate of cultures of this microorganism. 

Antigen found in the sterile filtrate of B. anthracis Purified capsular polysaccharide of... 

Sterile filtration and aseptic filling into final product container... 

Sterilant gases, ethylene oxide, 10 664 Sterile filtration, 12 138 in fermentation, 11 36 Sterilizability... 

Liquid product is fed to the BFS machine from a holding tank or vessel. The pathway is sterilized in place prior to receiving product, and product is sterilized by means of in-line sterilizing-grade filters. There is usually more than one stage of sterile filtration required on the product pathway. 

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