Biopharmaceuticals Characterization

Blume HH, Schug BS. Biopharmaceutical characterization of herbal medicinal products are in vivo studies necessary Eur J Drug Metabol Pharmacokinet 2000 25 41—48. 

M. Koike, S. Futaguchi, S. Takahashi, K. Sugeno, Biopharmaceutical Characterization of 450191-S, a Ring-Opened Derivative of 1,4-Benzodiazepine. II. Evidence for Reduced First-Pass Extraction by Rat Liver , Drug Metab. Dispos. 1988, 16, 609-615. 

Frick A, Moller H, Wirbitzki E. Biopharmaceutical characterization of oral immediate release drug products. In vitro/in vivo comparison of phenoxymethylpenicillin potassium, glimepiride and levofloxacin. European Journal of Pharmaceutics and Biopharmaceutics, 1998, 46 305-311. 

More recently, a comparison was made of the oral bioavailability in mice and dogs of a formulation prepared from a kava extract (WS 1490, containing 70% kavalactones), with the extract itself and the pure compound (+)-kavain. In both species, bioavailability increased in the order, pure compound, extract, and extract formulation. The data confirm the above findings on the synergistic absorption and uptake of kavalactones, and indicate that clinical data from one prepauation or formulation cannot simply be transferred to other formulations without appropriate biopharmaceutical characterization (Biber etal., 2000). 

No single analytical method will provide all the qualitative and quantitative answers during PEG-biopharmaceutical characterization. Size-exclusion/gel-filtration and SDS-PAGE may separate 1-mer from 2-mers and higher molecular weights. However, neither method usually clearly resolves positional isoforms. Ion-exchange or reverse-phase chromatography methods... 

SEC. Another liquid chromatographic technique applied in biopharmaceutical characterization is SEC. In this technique, compounds are separated based on size and shape. In contrast to RP-HPLC, SEC has a low resolving power that has limited the application of the technique. The main application of SEC in pharmaceutical analysis is the determination of the native size of a protein and the quantihcation of protein aggregates. Typically protein-friendly conditions are employed in SEC that are expected not to affect the higher order structure of the protein. 

Case history isolation and partial characterization of hementin, a compound of biopharmaceutical interest... 

The physicochemical and other properties of any newly identified drug must be extensively characterized prior to its entry into clinical trials. As the vast bulk of biopharmaceuticals are proteins, a summary overview of the approach taken to initial characterization of these biomolecules is presented. A prerequisite to such characterization is initial purification of the protein. Purification to homogeneity usually requires a combination of three or more high-resolution chromatographic steps (Chapter 6). The purification protocol is designed carefully, as it usually forms the basis of subsequent pilot- and process-scale purification systems. The purified product is then subjected to a battery of tests that aim to characterize it fully. Moreover, once these characteristics have been defined, they form the basis of many of the QC identity tests routinely performed on the product during its subsequent commercial manufacture. As these identity tests are discussed in detail in Chapter 7, only an abbreviated overview is presented here, in the form of Figure 4.5. 

A number of different techniques may be used to characterize protein-based biopharmaceutical products, and to detect any protein-based impurities that may be present in that product (Table 7.2). Analysis for non-protein-based contaminant is described in subsequent sections. 

Table 7.2 Methods used to characterize (protein-based) finished product biopharmaceuticals. An overview of most of these methods is presented over the next several sections of this chapter...

These disadvantages, along with the availability of alternative characterization methodologies, limit application of this technique in biopharmaceutical analysis. 

Key operating parameters that may change (or be optimized) throughout a product s development and approval cycle are dissolution sampling time points and dissolution limits or specifications by which the dissolution results should be evaluated. The results generated from the dissolution test need to be evaluated and interpreted based on the intended purpose of the test. If the test is used for batch-to-batch control, the results should be evaluated in regard to the established limits or specification value. If the test is being utilized as a characterization test (i.e., biopharmaceutical evaluations, formulation development studies, etc.) the results are usually evaluated by profile comparisons. 

For an extended-release dosage form, at least three test time points are chosen to characterize the in vitro drug-release profile for the routine batch-to-batch quality control for approved products. Additional sampling times may be required for formulation development studies, biopharmaceutical evaluations, and drug approval purposes. An early time... 

Analytical methods are important not only in the development and manufacture of commercial biopharmaceutical drugs, they also play a vital role in the whole drug development life cycle. Drug discovery and preclinical research require development and application of analytical methodologies to support identification, quantitation, and characterization of lead molecules. It is difficult to perform a comparative potency assay on lead molecules if one does not know how much of each is going into the assay or how pure the molecule is. Analytical methods are typically developed, qualified, and validated in step with the clinical... 

Analytical scientists will provide support for many of the activities in a biopharmaceutical company. They are responsible for characterizing the molecules in development, establishing and performing assays that aid in optimization and reproducibility of the purification schemes, and optimizing conditions for fermentation or cell culture to include product yields. Some of the characterization techniques will eventually be used in quality control to establish purity, potency, and identity of the final formulation. The techniques described here should provide the beginning of a palette from which to develop analytical solutions. 

In the development of new biopharmaceutical molecules, there is a constant need for analytical methods that provide critical information in areas that range from early characterization to routine analysis of approved products. Past experience indicates there are few projects in drug development that can be addressed by standard analytical procedures. Even well-established techniques often have to be modified to better suit the analysis of new samples. For this reason, a broad range of techniques is already an integral part of laboratories in the biopharmaceutical industry. 

Methods development starts with a relatively high number of techniques to characterize and test samples. The number of protocols is often reduced once the critical parameters and the methods that identify them have been defined. The analyst must evaluate the initial techniques with respect to their purposes. If the goal is to generate research data, the practicality of the method and its limitations are not of primary concern if the goal is to use the technique as part of a test procedure, it has to be evaluated in terms of its potential to meet full validation. Critical procedures (e.g., release testing) that cannot be validated will bring a project to an expensive halt. For these reasons, this chapter provides basic principles as well as limitations of capillary electrophoresis (CE) as applied to the analysis of real biopharmaceutical molecules. 

The MS techniques described previously for characterization of the final recombinant protein product can be applied at all stages during process development. MS might be used upstream to define clone selection, processing format, and purification steps, and downstream to characterize the final product, ascertain lotto-lot reproducibility, determine stability, and define the formulation of biopharmaceutical molecules. Presented here are some examples found either in the literature or from our own experience in which MS has been found to be a useful or necessary tool. Potential limitations of MS methods are discussed, and when appropriate, other analytical methods are mentioned that can be alternatives to MS and are also efficient tools for biopharmaceutical development. 

NMR is a remarkably flexible technique that can be effectively used to address many analytical issues in the development of biopharmaceutical products. Although it is already more than 50 years old, NMR is still underutilized in the biopharmaceutical industry for solving process-related analytical problems. In this chapter, we have described many simple and useful NMR applications for biopharmaceutical process development and validation. In particular, quantitative NMR analysis is perhaps the most important application. It is suitable for quantitating small organic molecules with a detection limit of 1 to 10 p.g/ml. In general, only simple one-dimensional NMR experiments are required for quantitative analysis. The other important application of NMR in biopharmaceutical development is the structural characterization of molecules that are product related (e.g., carbohydrates and peptide fragments) or process related (e.g., impurities and buffer components). However, structural studies typically require sophisticated multidimensional NMR experiments. 

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