Limited enzymatic digestion for the determination of the quantities of minor diastereomeric impurities in preparations of RMP-7, a peptide containing a reduced peptide bond.
RMP-7 is a bradykinin analogue containing all "L" amino acids and a reduced dipeptide bond between amino acids eight and nine. This reduced dipeptide bond [4-Me-Tyr-psi(CH2NH)-Arg] is created under synthetic conditions which could result in inversions of the chiral centers of either 4-Me-Tyr or Arg. Stereoisomers of RMP-7 would be expected to have altered biological specificity. Current chromatographic methods are not sufficiently sensitive to distinguish the anticipated stereoisomeric variants of the intact molecule. Therefore we have devised an analytical method based on limited enzymatic digestion of the compound followed by reversed-phase HPLC analysis of the peptide fragments. Using this method we have been able to carry out precise and reliable quantitative analysis of the stereoisomeric content of different batches of peptide prepared for biological testing.
Alkermes, Inc., Cambridge, MA 02139.
J A
JA
Straub
A
A
Akiyama
P
P
Parmar
G F
GF
Musso
D000595:Amino Acid Sequence; D001920:Bradykinin / Q000031:analogs & derivatives* / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D004151:Dipeptides / Q000032:analysis*; D004798:Enzymes / Q000737:chemistry*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D013056:Spectrophotometry, Ultraviolet; D013237:Stereoisomerism
D004151:Dipeptides; D004798:Enzymes; C089876:RMP 7; D001920:Bradykinin