On developing a process for conducting extractable-leachable assessment of components used for storage of biopharmaceuticals.

Extractables and leachables are product-related impurities that result from product contact with components such as gaskets, stoppers, storage bags, cartridges, and prefilled syringes that are used for processing, storage, and/or delivery of biopharmaceuticals. These impurities are a concern for patients due to potential effects on product quality and safety. It is possible that such an impurity could directly impact the patient or indirectly impact the patient by interacting with the protein therapeutics and forming protein adducts.

Hydrogen sulfide poisoning: clarification of some controversial issues.

Background: Hydrogen sulfide is a toxic gas about which much has been written. We discuss here several issues we believe would benefit from further clarification.

Conclusions: We conclude that: 1) Certain neurotoxic effects of exposure are probably due to a direct toxic effect on the brain, while others are almost certainly a result of hypoxia secondary to H2S-induced respiratory insufficiency; 2) pulmonary edema is a common consequence of poisoning and there is suggestive evidence of hyperactive airway responses in some individuals following brief H2S-induced unconsciousness (knockdown); 3) criteria for acceptable community levels are very different than those governing occupational standards; 4) urinary thiosulfate determinations can be useful for monitoring occupational exposure; and 5) determination of sulfide ion concentrations in blood or major organs can be useful in corroborating a diagnosis of fatal H2S toxicity, but there are many pitfalls in collecting, storing, and analyzing tissue and fluid samples.

Metal-catalyzed oxidation of histidine in human growth hormone. Mechanism, isotope effects, and inhibition by a mild denaturing alcohol.

Metal-catalyzed oxidation of proteins represents an important pathway of post-translational modification. We utilized human growth hormone (hGH), a protein with a well defined metal-binding site, to study the detailed mechanism of metal-catalyzed oxidation by ascorbate/Cu(II)/O2. Particularly His18 and His21 within the metal-binding site were oxidized, predominantly to 2-oxo-His with the incorporated oxygen originating from molecular oxygen, based on amino acid analysis, tryptic mapping, mass spectrometry, isotopic labeling, and 1H NMR.

Metal-catalyzed photooxidation of histidine in human growth hormone.

Reports on nonenzymatic oxidation of human growth hormone (hGH) have been previously limited to methionyl residues (Met14 and Met125). We report on the oxidation of a histidyl residue in hGH treated with intense light. The photooxidation process is predominately site-specific to histidine at position 21, which forms a cation-binding site along with His18 and Glu174.

A turbidimetric method to determine visual appearance of protein solutions.

An instrumental method to analyze protein solutions for visual appearance is described which is based on spectrophotometric comparison to reference suspensions with varying degrees of turbidity. This method provides a useful substitute for visual inspection of uniform opalescent suspensions in that it is more convenient and less time-consuming and has the potential to be more reproducible, accurate and objective. Established categories of opalescence based on European Pharmacopoeial reference suspensions were determined using turbidity measured as optical density in the 340-360 nm range.