Some Leading Edge Prospects in Lyophilization
Some Leading Edge Prospects in Lyophilization
Summer 2003
Professor Louis Rey, Ph.D., Chemin de Verdonnet 2, CH 1010 Lausanne, Switzerland
The development of Lyophilization/Freeze-Drying is almost a century old venture since the 1906 publication of Bordas and d?Arsonval. In the first part of the 20th century, pioneering work in the field has been carried out by many remarkable scientists, such as Altman, Gersh, Greaves, Flosdorf, Henaff and others, but it is mainly under the pressure of servicing the second world war battle fields that Lyophilization took its marks with freeze-dried plasma and later under the impetus of the Nobel Laureate Ernst Chain with freeze-dried Penicillin. As any new technology, the field of freeze-drying opened more and more and became strongly rooted in the biological, pharmaceutical, food and cosmetic industries. In the early sixties, we did propose to resort to dry from the frozen state many new products in the chemical field and even for radioactive wastes. Thus, the new technology diversified and, as it is often the case, some cross fertilization took place between the different applications bringing new opportunities in already ?classical? areas. The purpose of this article is precisely to give an overall survey of this evolution and attempt to identify the mainstreams of further diversification of Lyophilization. As such, it was compulsory to review a certain number of scientific and industrial areas which lie outside of the main focus of pharmacy.
As this is a most general case in science, Lyophilization has been propelled in the last decades by a push-pull phenomenon between products and technology. New products fulfill new needs, open new markets but, in turn, request new developments in process and industrialization. Sometimes they are purely genuine, sometimes they are just a consequence of the spin-off of other technologies. This is the reason why I shall switch from basic science to engineering and from technology to consumer acceptance without following a strict rationality.
I. Biological Products and Pharmaceuticals
1. Some Basic Issues
For a very long time, Freeze-Drying has been essentially concerned with the preservation of unstable biochemicals and, more particularly, of injectables. The very first issue has been, then, to secure their sterility and safeguard their potency during processing, storage and reconstitution. At the onstart most products were freeze-dried as such, either as a natural substance, such are blood plasma, biosynthetic isolates or antibiotics or else, for vaccines, as controlled concentrates of inactivated and/or living cells resulting from selected culture fermentation broths.
Often the product was bulky and resistant (like blood plasma) and could be processed directly. However, as more and more new advances were made, many products became difficult to treat and required the incorporation of a whole set of additives: cryoprotectors, free-radical scavengers, stabilizers and so on. Today, and with the rocketing development of biotechnology, more and more potent and fragile compounds are produced and, most often, their activity is such that they represent only a few milligrams in the basic formulation. There is, thus, an obvious need to incorporate the active substance into a solid matrix which will prevent it to fly away during drying and keep it secluded in a confined stable environment during storage. This means that, on top of the already quoted cryoprotectors, stabilizers and the like, different bulking agents have to be added like sucrose, mannitol, lactose ?, the behavior of which might have a major impact on the properties of the active substance. This is particularly true for proteins where a given steric configuration should be preserved during the whole process and withstand the stress of freezing and drying. Carpenter, Pikal and others have shown that developing a glassy matrix during freezing could substantially reduce the osmotic and mechanical strains to the molecules. This, unfortunately, might also be contrary to the requirements of the drying process itself which is easier to carry from a non amorphous state. Thus, it might be necessary to mitigate between the conventional freezing and what we called in 1960 a "thermal treatment," most often referred to today as an annealing process: double freezing with intermediate rewarming.
Depending upon the products, there might be very narrow margins to that exercise and the end-points? temperatures. Velocities of cooling and rewarming need to be known very accurately by previous laboratory determinations such as Differential Thermal Analysis (DTA) or Differential Scanning Calorimetry (DSC), Low Temperature Electric Impedance measurements, velocity of crystallization in the supercooled state ?
Moreover, in this context of highly diluted active substances of powerful potency (like for instance Botulinum toxine) there might be a strong interference between the product and the container-closure system. Glass manufacturers have pioneered in the field showing that a Type I tubing vial might interact with the solution prior to freezing, and this is often in an irreversible way. For instance, it might adsorb more than 50 percent of an active protein (tests have been made with a nicotinic acetylcholine receptor) and equally leach substantial amounts of undesirable glass components (sodium, calcium, boron, aluminium ?) into the solution which may also dissolve the inner surface of the container. To obviate those problems, a glass manufacturer developed the Type I Plus vials in which, thanks to a special high temperature plasma process (PIVD: Plasma Impulse Chemical Vapor Deposition), a very thin coating (100-200 nm) of pure silica is deposited and strongly bonded to the glass surface (something like a quartz glazing of the inner wall of the vial) which becomes then totally neutral. Much interest is given today to this new process, at a time where Regulating Agencies, and especially FDA, are deeply concerned by the effect on brain and liver of aluminium released from glass in very young infant vaccines. Silica coating affords then an excellent protection, still better than some new cyclo olefin polymers (such as Topas 6013) which could be also used to manufacture the entire vial.
What is true for the container in the "container closure system" is equally true for the closure alone and, essentially, for the elastomeric stoppers, generally butyl-rubber, which cap the vial. Fran DeGrazio from the WEST PHARMACEUTICAL SERVICES has done a lot of research in this field and studied many alternatives to their formulation trying to keep down the extractables (essentially volatile) and leachables and prevent adsorption of oils, waxes, polymers and others on the freeze-dried plug.
Another critical issue in the same field is the determination of the optimum residual moisture, understanding that it could result from the drying process itself obviously, but also from release by the stoppers of water picked up during the sterilization process. Fran DeGrazio, Maninder Hora, and others did show that this phenomenon definitely influences the storage ability of the freeze-dried products which also depends, among other issues, upon the quality of the fit between the vial and the stopper which, in turn, depends upon how tight the manufacturers can guarantee the dimensional tolerance of their products.
Water is, indeed, a recurrent and unavoidable issue in the whole Lyophilization process. Formally regulated by the different responsible Agencies, the residual moisture is, indeed, a floating, nebulous concept which is quite difficult to grasp with exactitude. Joan May from FDA, who has been a leading authority in the field for so many years, knows quite well that neither Karl Fischer titration, neither Thermogravimetry nor Equilibrium Water Vapor Pressure Determination could bring the right answer. Moreover, the first two techniques being destructive, they prevent any follow-up of the "fate" of the water in the product - with time. They titrate the "total water," as a chemical, without discriminating between the part which is free to move and exchange between the stopper and the cake and the one, bound to the cake, which is often essential to maintain within the freeze-dried active substance, the tridimensional structure which is at the root of its potency. The Equilibrium Water Vapor Pressure that we have introduced, solves the follow-up issue since it is a non-intrusive, non destructive technique but it gives an "indirect" reading of the water content. Finally, we know that, in the dry product itself and without any interference from the container-closure system, there are movements of water with time and, that, during storage, the ratio between "free" and the different types of "bound" water changes and might impact the final potency.
These are only limited glimpses at the basic issues that any professional has to challenge when performing freeze-drying of biological products and pharmaceuticals.
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