Formulation development of an antifibrin monoclonal antibody radiopharmaceutical.

These studies have shown that formulation development of a monoclonal antibody radio-immunoscintigraphy agent is a challenging task involving a number of issues related to the radiochemistry of labeling as well as the stability of the antibody. Through a systematic approach, as described in this study, a stable and efficacious product of high quality can be developed in a rational and efficient manner. In developing an optimized formulation of Tc-99m-antifibrin Fab' for use in immunoscintigraphy of DVT, a number of critical components were examined in regard to technetium-labeling chemistry and the ability of the formulation to support the long-term stability of the product. It was found that the addition of glucarate as a transfer ligand, SnCl2 as a reducing agent, and neutral-to-acidic pH of the solution were essential for optimum radiolabeling of 0.5 mg of antifibrin Fab' to a desired activity of 25 mCi of Tc-99m. The lyophilization of the final product was also required to further stabilize both the antifibrin Fab' fragment and the reducing agent. The addition of carbohydrate as bulking agent and lyoprotectant and inclusion of EDTA as a chelating agent further improved the performance of the formulations, resulting in products with long shelf-life. Many of the principles described in this study are not only useful in developing a technetium-based immunoscintigraphic agent but are also applicable to other immunopharmaceuticals, including products involving delivery of radionuclides, drugs, and toxins for immunotherapy of cancer and other diseases.

Near-infrared spectroscopic determination of residual moisture in lyophilized sucrose through intact glass vials.

A rapid, noninvasive, and nondestructive method for determining moisture in sealed freeze-drying vials is described. The method, based on near-infrared spectrometry, used a novel fiber-optic diffuse-reflectance probe to make remote reflectance measurements from 1100 to 2500 nm through the bottom of glass vials. The correlation of the method to results obtained by Karl Fischer analysis was good (r2 = 0.97). The moisture content of sucrose, a common cryoprotectant, was determined with an error of 0.27% using a single sample scan.

Hygroscopicity of cefazolin sodium: application to evaluate the crystallinity of freeze-dried products.

The hygroscopicity behavior of the pentahydrate, monohydrate, amorphous, and dehydrated forms of solid cefazolin sodium (CEZ) was studied under different relative humidity (RH) conditions. Between 42 and 86% RH, the pentahydrate (alpha form), the monohydrate, and their dehydrated forms absorbed atmospheric moisture equivalent to their hydrate numbers. The pentahydrate demonstrated a hysteresis effect at 15 and 31% RH. On the other hand, the water content of the amorphous form increased linearly with increases in RH. The noncrystalline state was maintained below 56% RH. For the dehydrated alpha form there was a distinct birefringence when viewed under polarizing light, the X-ray diffraction pattern was weak and diffuse, and the infrared (IR) spectra were discernibly different from that of the amorphous form. The freeze-dried CEZ showed hygroscopic behavior almost similar to that of the dehydrated alpha form. Two-component mixtures of various CEZ forms showed a linear relationship between the water content and the mixing ratio when stored at 31, 42, and 56% RH. From the hygroscopicity data, the crystallinity of freeze-dried CEZ could be estimated as the percentage of the dehydrated alpha form.

Estimation of the degree of crystallinity of cefazolin sodium by X-ray and infrared methods.

The pentahydrate (alpha form) of cefazolin sodium (CEZ) exhibited sharp X-ray diffraction peaks, while the dehydrated alpha form showed weak but distinct diffraction peaks. As expected the amorphous form exhibited a diffuse and halo diffraction pattern. The X-ray procedure to estimate the degree of crystallinity of CEZ was based upon the measurement of the total scattering and the scattering from the crystalline region of the drug. The major difference in the infrared (IR) spectra among the three forms of CEZ was the absence of a spectral band at 1542 cm-1 in the amorphous form. The IR procedure was based upon the measurement of the peak percentage area ratio between the bands at 1542 and 1760 cm-1, where the latter was used as a normalizing peak. The degree of crystallinity of CEZ samples, obtained by either freeze-drying aqueous CEZ solutions or storing the crystalline forms under different humidity conditions, was determined by these two methods. Although the correlation of results by the two methods was good, the X-ray procedure appears to be superior since it can differentiate among the three solid CEZ forms, whereas IR could distinguish between only crystalline and amorphous CEZ, reproducibly.