Formulation buffer

Figure 12.1 Clearance of small-molecule impurities from process buffers in a formulated protein product. Trace A the NMR spectrum of a control sample containing a mixture of three components (succinate, tetraethylammonium, and tetramethylammonium) in the final formulation buffer (sodium acetate). These three components were used in the recovery process for a biopharmaceutical product. Traces B and D the proton NMR spectra of the formulated protein product. No TEA or TMA were detected, but a small amount of succinate was observed in this sample. Traces C and E the proton NMR spectra of a formulated protein product spiked with 10 jag/ml of succinate, TEA, and TMA. Traces D and E were recorded with CPMG spin-echo method to reduce the protein signals. The reduction of NMR signals from the protein allows for better observation of the small-molecule signals.

Figure 12.3 Clearance of MES in a formulated protein product. Trace A the proton NMR spectrum of a formulated protein product spiked with 8 jig/ml of MES. Trace B the proton NMR spectrum of a formulated protein product. The arrows indicate the positions where MES signals would be detected if present. Trace C the difference of traces A and B (A-B). Trace D proton NMR spectrum of 8 ftg/ml of MES in the formulation buffer. The NMR spectra in traces A, B, and D were recorded with the CPMG spin-echo method to reduce protein signals. Only the region where MES signals appear is shown.

Figure 12.6 Two sections of the proton NMR spectrum for a protein product after the protein has been removed. Only the formulation buffer (25 mM citrate) and a small amount ( 1 pg/ml) of residual Tris (a component of the penultimate buffer used in the recovery process prior to the final UF/DF formulation step) are observed after the filtration. The signal-to-noise ratio of Tris is about 70. The absence of any other significant proton NMR signals provides compelling evidence for good clearance of any other small proton-containing molecule impurities.

The CE-SDS method is a size-based separation technique generally applicable to proteins from 10 to -200—300kDa. The specificity is generally tested against the formulation buffer and any other possible contaminant proteins. There is usually no interference from the formulation buffer with the assay. For samples that contain contaminant proteins with a hydrodynamic size of 10—200kDa, the method is not specific. 

Formulation buffer vial containing 750 mg if human albumin 76 mg of sodium chloride 21 mg of sodium phosphate dibasic heptahydrate 4 mg pentetic acid 2 mg potassium phosphate monobasic 2 mg of potassium chloride lOmL of water (pH 7.1)... 

Nevertheless, the formulation buffer capacity and titratable acid content, in addition to the nature of the acid present, are perhaps the most important factors for nebulizer solutions of greater than pH 2.0. 

A pharmacokinetic comparison of both product generations, and their formulation buffers was conducted in partially heparinized Sprague-Dawley rats using a monoclonal antibody capture/activity assay. The two rFVIII concentrates exhibited a high degree of similarity in half-life, area under the curve (AUC), clearance rate, and mean retention time, predicting biological activity of rAHF-PFM in humans similar to that of rAHF. 

Small stocks of substances that have to be measured or weighed during the production process (e.g., buffers) may be kept in the production area, provided that they are not returned to the general stocks. Otherwise, dry materials used to formulate buffers, culture media, etc. should be weighed and put into solution in a contained area outside the purification and aseptic areas in order to minimize particulate contamination of the product. 

Processes for adenovirus purification typically end with concentration, formulation, and sterile filtration operations [40, 80,106]. Concentration and formulation are usually carried out in ultra-filtration units equipped with 100-300-kDA membranes [40,106]. The exact composition of the formulation buffer will depend on the intended application, mode of administration (injectable, aerosol), and required short-term and shelf stability [104,123]. A typical liquid formulation may include an aqueous buffer supplemented with cryoprotectants (e.g., sucrose) and stabilizers such as the nonionic-surfactant polysorbate-80, the chelating agent EDTA, and the oxidation inhibitors ethanol and histidine [123]. Filtration under sterile conditions is typically performed with 0.22-pm membranes [103,106]. 

Can assess aggregates over a wider range of buffer conditions than SEC. AUC-SV can be performed in low ionic strength formulation buffers as well as in higher ionic strength SEC mobile phase. AUC-SV is therefore useful for detecting differences in protein size distribution between different buffer conditions. 

Low ionic strength formulation buffers may promote the adsorption of protein onto the membrane or other surfaces. This would then necessitate the use of a higher ionic strength mobile phase buffer. The drawback to this is that the distribution of aggregates determined in the high ionic strength buffer may not be the same as that in the lower ionic strength formulation buffer. 

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