Effect of ethylenediamine on chemical degradation of insulin aspart in pharmaceutical solutions.
OBJECTIVE
To examine the effect of different amine compounds on the chemical degradation of insulin aspart at pharmaceutical formulation conditions.


METHODS
Insulin aspart preparations containing amine compounds or phosphate (reference) were prepared and the chemical degradation was assessed following storage at 37 degrees C using chromatographic techniques. Ethylenediamine was examined at multiple concentrations and the resulting insulin-ethylenediamine derivates were structurally characterized using matrix assisted laser desorption ionization time-of-flight mass spectroscopy. The effects on ethylenediamine when omitting glycerol or phenolic compounds from the formulations were investigated.


RESULTS
Ethylenediamine was superior in terms of reducing formation of high molecular weight protein and insulin aspart related impurities compared to the other amine compounds and phosphate. Monotransamidation of insulin aspart in the presence of ethylenediamine was observed at all of the six possible Asn/Gln residues with Asn(A21) having the highest propensity to react with ethylenediamine. Data from formulations studies suggests a dual mechanism of ethylenediamine and a mandatory presence of phenolic compounds to obtain the effect.


CONCLUSIONS
The formation of high molecular weight protein and insulin aspart related impurities was reduced by ethylenediamine in a concentration dependant manner.
Poulsen
Christian
C
Pharmaceutics, Diabetes Department, Novo Nordisk A/S, Novo Alle, room 6B1.58.1, 2880 Bagsvaerd, Denmark. chpo@novonordisk.com
Jacobsen
Dorte
D
Palm
Lisbeth
L
D000595:Amino Acid Sequence; D002852:Chromatography, Ion Exchange; D003408:Cresols / Q000737:chemistry; D004355:Drug Stability; D005029:Ethylenediamines / Q000737:chemistry*; D005990:Glycerol / Q000737:chemistry; D007004:Hypoglycemic Agents / Q000737:chemistry*; D007328:Insulin / Q000031:analogs & derivatives* / Q000737:chemistry; D061267:Insulin Aspart; D008969:Molecular Sequence Data; D019800:Phenol / Q000737:chemistry; D012996:Solutions
D003408:Cresols; D005029:Ethylenediamines; D007004:Hypoglycemic Agents; D007328:Insulin; D012996:Solutions; C032538:4-cresol; D019800:Phenol; C031234:ethylenediamine; D061267:Insulin Aspart; D005990:Glycerol