Fluorometric determination of mucin-type glycoproteins by the galactose oxidase-peroxidase method.
We developed a convenient and specific method for the determination of mucin-type glycoproteins using galactose oxidase and horseradish peroxidase on the basis of the contents of galactosyl and N-acetylgalactosaminyl residues in glycoproteins. Galactose and galactosamine residues released from glycoproteins after hydrolysis were oxidized with galactose oxidase and subsequently the resultant hydrogen peroxide was determined by a combination of horseradish peroxidase and 3-(p-hydroxyphenyl) propionic acid as a fluorogenic substrate. The contents of galactose/galactosamine residues in N- and O-glycans, as determined by the galactose oxidase-peroxidase method, were in good agreement with those described in the previous reports. We applied the present method to determine mucin-type glycoproteins secreted from rat gastric mucosa by stimulation with misoprostol, a prostaglandin E(1) analogue in vivo. Thus, the galactose oxidase-peroxidase method is useful for the determination of mucin-type glycoproteins in biological materials.
Faculty of Pharmaceutical Sciences, Kinki University, Kowakae 3-4-1, Higashi-osaka 577-8502, Japan.
M
M
Kinoshita
K
K
Inagake
A
A
Kawabata
R
R
Kuroda
Y
Y
Oda
K
K
Kakehi
D000818:Animals; D002138:Calibration; D002417:Cattle; D004305:Dose-Response Relationship, Drug; D005688:Galactosamine / Q000737:chemistry; D005690:Galactose / Q000737:chemistry; D005692:Galactose Oxidase / Q000494:pharmacology*; D005753:Gastric Mucosa / Q000378:metabolism; D006023:Glycoproteins / Q000032:analysis*; D006735:Horseradish Peroxidase / Q000494:pharmacology*; D008297:Male; D016595:Misoprostol / Q000494:pharmacology; D008956:Models, Chemical; D009077:Mucins / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence; D013552:Swine; D013997:Time Factors
D006023:Glycoproteins; D009077:Mucins; D016595:Misoprostol; D005688:Galactosamine; D005692:Galactose Oxidase; D006735:Horseradish Peroxidase; D005690:Galactose