Degradation of peptide drugs by immobilized digestive proteases.
An in vitro assay using immobilized digestive enzymes and LC/MS has been developed to measure the susceptibility of orally administered peptide drugs to hydrolysis by the digestive system. Six different bioactive peptides and peptide drugs were digested using an established in vitro system incorporating four sets of immobilized digestive enzymes, which simulated gastric digestion in the stomach, digestion by pancreatic enzymes in the intestinal lumen, and digestion by mucosal peptidases at the intestinal brush border. Hydrolysis products were rapidly identified using LC/MS and coincided with those expected based on the substrate specificities of the immobilized enzymes. The LC/MS system consisted of a reversed phase HPLC connected to a mass spectrometer through a continuous-flow fast atom bombardment ionization system. Data acquired on the susceptibility of peptide drugs to hydrolysis by digestive enzymes could be used during the formulation of orally administered drugs to protect them from degradation prior to absorption in the gut. Also, this assay could be used to quickly identify those compounds in a structurally related series of new drugs that are most resistant to digestive proteases.
Van Breemen
R B
RB
Department of Chemistry, North Carolina State University, Raleigh 27695.
Bartlett
M G
MG
Tsou
Y H
YH
Culver
C
C
Swaisgood
H
H
Unger
S E
SE
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D002918:Chymotrypsin / Q000378:metabolism; D004064:Digestive System / Q000201:enzymology; D010450:Endopeptidases / Q000378:metabolism*; D004800:Enzymes, Immobilized; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism*; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D016339:Spectrometry, Mass, Fast Atom Bombardment; D014357:Trypsin / Q000378:metabolism
D004800:Enzymes, Immobilized; D010455:Peptides; D004364:Pharmaceutical Preparations; D010450:Endopeptidases; D002918:Chymotrypsin; D014357:Trypsin