title: Photolysis of recombinant human insulin in the solid state: formation of a dithiohemiacetal product at the C-terminal disulfide bond.
abstract: OBJECTIVE
  Exposure of protein pharmaceuticals to light can result in chemical and physical modifications, potentially leading to loss of potency, aggregation, and/or immunogenicity. To correlate these potential consequences with molecular changes, the nature of photoproducts and their mechanisms of formation must be characterized. The present study focuses on the photochemical degradation of insulin in the solid state.
  
  
  METHODS
  Solid insulin was characterized by solid-state NMR, polarized optical microscopy and scanning electron microscopy; various insulin preparations were exposed to UV light prior to product analysis by mass spectrometry.
  
  
  RESULTS
  UV-exposure of solid human insulin results in photodissociation of the C-terminal intrachain disulfide bond, leading to formation of a CysS(•) thiyl radical pair which ultimately disproportionates into thiol and thioaldehyde species. The high reactivity of the thioaldehyde and proximity to the thiol allow the formation of a dithiohemiacetal structure. Dithiohemiacetal is formed during the UV-exposure of both crystalline and amorphous insulin.
  
  
  CONCLUSIONS
  Dithiohemiacetals represent novel structures generated through the photochemical modification of disulfide bonds. This is the first time that such structure is identified during the photolysis of a protein in the solid state.
authors: Department of Pharmaceutical Chemistry, University of Kansas, 2095 Constant Avenue, Lawrence, Kansas 66047, USA.
authors: Olivier
authors: O
authors: Mozziconacci
authors: Jessica
authors: J
authors: Haywood
authors: Eric M
authors: EM
authors: Gorman
authors: Eric
authors: E
authors: Munson
authors: Christian
authors: C
authors: Schöneich
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003460:Crystallization; D041961:Deuterium Exchange Measurement; D004220:Disulfides / Q000737:chemistry / Q000528:radiation effects*; D004355:Drug Stability; D005609:Free Radicals / Q000737:chemistry / Q000528:radiation effects; D006801:Humans; D007328:Insulin / Q000737:chemistry / Q000528:radiation effects*; D009682:Magnetic Resonance Spectroscopy; D008855:Microscopy, Electron, Scanning; D008969:Molecular Sequence Data; D010782:Photolysis; D011994:Recombinant Proteins / Q000737:chemistry / Q000528:radiation effects*; D021241:Spectrometry, Mass, Electrospray Ionization; D013438:Sulfhydryl Compounds / Q000737:chemistry / Q000528:radiation effects*; D014466:Ultraviolet Rays; D015032:Zinc / Q000032:analysis
chemical_list: D004220:Disulfides; D005609:Free Radicals; D007328:Insulin; D011994:Recombinant Proteins; D013438:Sulfhydryl Compounds; D015032:Zinc