Formulation of Biotech Products, Including Biopharmaceutical Considerations

formulation of biotech products, including biopharmaceutical considerations
objectives: - formulation aspects of pharmaceutical proteins
- the consideration of the biopharmaceutical issues, such as the choice of the delivery systems, the route of administration and possibilities for the site specific delivery of proteins.
microbial considerations
1- sterility 2- viral decontamination 3- pyrogen removal
sterility
- most proteins are administered parenterally have to be sterile. in general, proteins are sensitive to heat and other regularly used sterilization treatments therefore they cannot withstand autoclaving, gas sterilization, or sterilization by ionizing radiation. consequently, sterilization of the end product is not possible. therefore, protein pharmaceuticals have to be assembled under aseptic conditions.
rules in the pharmaceutical industry for aseptic manufacture
- equipment and excipients are treated separately and autoclaved, or sterilized by dry heat (> 160c), chemical treatment or gamma radiation to minimize the bioburden.
- filtration techniques are used for removal of microbacterial contaminants. the final “sterilizing” step before filling the vials is filtration through 0.2 or 0.22 mm filters.
- assembly of the product is done in classrooms with laminar airflow that is filtered through high efficiency particulate air (hepa) filters.
- additionally, the “human factor” is a major source of contamination, and operators wearing protective cloths (face masks, hats, gowns, gloves, or head-to-toe overall garments) should operate the facility. regular exchange of filters and cleaning of the room plus equipment are critical factors for success.

air flow pattern “laminar flow cleanroom"
viral decontamination
- as recombinant dna products are grown in microorganisms, these organisms should be tested for viral contaminants
- no (unwanted) viral material should be introduced during manufacturing process. excipients with a certain risk factor such as blood-derived human serum albumin should be carefully tested before use.
viral presence in the formulation process should be minimized via:
- inactive viral contaminants e.g. heat treatment(pasterurization).
- removal of viral contamination e.g.filtration (nano-filtration).



3. pyrogen removal
- pyrogens are compounds that induce fever. pyrogens can be derived from bacterial, viral or fungal sources. bacterial pyrogens are mainly endotoxins shed from gram negative bacteria. they are lipopolysaccharides.
- a general structure is shown in figure 3. the basic structure is the lipid a-moiety.
- endotoxins is their high, negative electrical charge.
- endotoxins break down when heated in the dry state. for this reason equipment and container are treated at temperatures above 160 c for prolonged periods (e.g., 30 minutes dry heat at 250 c).



figure (1) the structure of endotoxin

pyrogen removal of recombinant products derived from bacterial sources should be an integral part of preparation process. ion exchange chromatographic procedures (utilizing its negative charge) can effectively reduce endotoxin levels in solution.
- excipients used in the protein formulation should be essentially endotoxin free. for solutions “water for injection” is (freshly) distilled or produced by reverse osmosis. the aggregated endotoxins cannot pass through the reverse osmosis membrane.
- removal of endotoxins immediately before filling the final container can be accomplished by using activated charcoal or other materials with large surfaces offering hydropinghobic interactions. endotoxins can also be inactivated on utensil surfaces by oxidation (e.g., peroxide) or dry heating (e.g., 30 minutes dry heat at 250 c).



ion exchange chromatography
excipients used in parenteral formulations of biotech product
in a protein formulation a number of excipients selected to serve different purposes. this process of formulation design should be` carried out with great care to ensure therapeutic effectiveness and safe products.
the nature of the protein (e.g. lability) and its therapeutic use (e.g. multiple injection systems) can make these formulations quite complex in term of excipients profile and technology (freeze-drying, aseptic preparation).
table 1 components found in parenteral formulations of biotech products
- active ingredient
- solubility enhancers
- anti-adsorption and anti-aggregation agents
- buffer components
- preservatives and antioxidants
- osmotic agents
- lyoprotectants
- carrier system