Separation of enantiomers on HPLC chiral stationary phases based on human plasma alpha1-acid glycoprotein: effect of sugar moiety on chiral recognition ability.
HPLC chiral stationary phases based on human plasma alpha1-acid glycoprotein (AGP) and partially deglycosylated AGP (pd-AGP) were prepared to investigate the effects of sugar moiety of AGP on chiral discrimination of various solutes. Removal of a sugar moiety of AGP by treatment with N-glycosidase was confirmed by high-performance capillary electrophoresis, reversed-phase HPLC and matrix-assisted laser desorption-time of flight (MALDI-TOF) mass spectrometry. The average molecular weights of AGP and pd-AGP were estimated to be ca. 33,000 and 30,600, respectively, by MALDI-TOF mass spectrometry. Next, AGP and pd-AGP were bound to aminopropyl-silica gels activated with N,N '-disuccinimidylcarbonate. The retentivity+ and enantioselectivity of the neutral, acidic and basic solutes tested on the pd-AGP column were significantly or not significantly larger in most solutes than those on the AGP column. This is ascribable to that by cleavage of a sugar chain(s) by N-glycosidase, pd-AGP could become more hydrophobic than AGP, and/ or that a solute could be easily accessible to the specific and/or non-specific binding sites of pd-AGP. It is interesting that warfarin enantiomers are not resolved on the pd-AGP column, but resolved on the AGP column. A sugar chain(s) of AGP cleaved by N-glycosidase might be involved in the enantioselective binding of warfarin enantiomers.
Haginaka
J
J
Faculty of Pharmaceutical Sciences, Mukogawa Women's University, Hyogo, Japan. haginaka@mwu.mukogawa-u.ac.jp
Matsunaga
H
H
D000526:Alprenolol / Q000737:chemistry / Q000302:isolation & purification; D001573:Benzoin / Q000737:chemistry / Q000302:isolation & purification; D002045:Bupivacaine / Q000737:chemistry / Q000302:isolation & purification; D002241:Carbohydrates / Q000737:chemistry*; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D019075:Electrophoresis, Capillary; D006026:Glycoside Hydrolases / Q000494:pharmacology; D006031:Glycosylation; D006801:Humans; D006827:Hydantoins / Q000737:chemistry / Q000302:isolation & purification; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D009961:Orosomucoid / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D010096:Oxprenolol / Q000737:chemistry / Q000302:isolation & purification; D010654:Phenylbutyrates / Q000737:chemistry / Q000302:isolation & purification; D011422:Propionates / Q000737:chemistry / Q000302:isolation & purification; D011433:Propranolol / Q000737:chemistry / Q000302:isolation & purification; D011485:Protein Binding; D012822:Silicon Dioxide; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013237:Stereoisomerism; D013329:Structure-Activity Relationship; D014859:Warfarin / Q000737:chemistry / Q000302:isolation & purification
D002241:Carbohydrates; D006827:Hydantoins; D009961:Orosomucoid; D010654:Phenylbutyrates; D011422:Propionates; C021611:aminopropyl silica gel; C084606:ethotoin; D010096:Oxprenolol; D014859:Warfarin; D012822:Silicon Dioxide; D000526:Alprenolol; C408480:2-phenoxypropanoic acid; D011433:Propranolol; D006026:Glycoside Hydrolases; D001573:Benzoin; C039714:2-phenylbutyric acid; D002045:Bupivacaine