title: Formation of amide- and imide-linked degradation products between the peptide drug oxytocin and citrate in citrate-buffered formulations.
abstract: Citric acid is widely used to buffer pharmaceutical formulations including protein pharmaceuticals. In accelerated stability studies of the small cyclic peptide oxytocin, we have noted that additional degradation products form when oxytocin is formulated in citrate that do not form in other common buffers such as acetate and phosphate. Using high-pressure liquid chromatography combined with high-resolution and tandem mass spectrometry, we identified these degradation products as amide- and imide-linked adducts of oxytocin and citrate. The site of reaction was shown to be the N-terminal amine of cysteine. The adducts have been found to form for oxytocin formulated in citrate buffer over the pH range of 3-6; the extent of formation is greatest at a pH of 4-4.5. We have additionally identified these same adducts in samples of oxytocin formulated in citrate buffer that had been stored in the dark for 3 months at room temperature. Altogether, these results demonstrate that reaction between citrate and oxytocin leads to the formation of covalent amide- and imide-linked adducts.
authors.affiliation: Division of Drug Delivery Technology, Leiden/Amsterdam Center for Drug Research (LACDR), 2333 CC Leiden, The Netherlands.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Poole
authors.forename: Piotr Tomasz
authors.initials: PT
authors.lastname: Kasper
authors.forename: Wim
authors.initials: W
authors.lastname: Jiskoot
mesh_terms: D000577:Amides / Q000737:chemistry*; D002021:Buffers; D002626:Chemistry, Pharmaceutical; D002851:Chromatography, High Pressure Liquid; D056148:Chromatography, Reverse-Phase; D019343:Citric Acid / Q000737:chemistry*; D003545:Cysteine; D004355:Drug Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007094:Imides / Q000737:chemistry*; D015394:Molecular Structure; D010121:Oxytocin / Q000737:chemistry*; D053719:Tandem Mass Spectrometry; D013678:Technology, Pharmaceutical / Q000379:methods
chemical_list: D000577:Amides; D002021:Buffers; D007094:Imides; D019343:Citric Acid; D010121:Oxytocin; D003545:Cysteine