Quantification of lactobionic acid and sorbitol from enzymatic reaction of fructose and lactose by high-performance liquid chromatography.
Experimental conditions for complete separation and quantification of mixtures containing lactobionic acid, sorbitol, lactose and fructose are discussed for the first time. These mixtures appear in the enzymatic bioconversion of fructose and lactose catalyzed by glucose-fructose oxidoreductase (GFOR) and glucono-delta-lactonase (GL) enzymes of Zymomonas mobilis cells. The high-performance liquid chromatography (HPLC) separation was carried out in a strong cation ion exchange resin (hydrogen form) based on a copolymer of styrene divinylbenzene (PS-DVB). A stationary phase of beta-cyclodextrin was also evaluated. An efficient separation was obtained with PS-DVB column eluted with sulfuric acid 0.450 mM solutions (pH 3.0-3.2) at 75 degrees C. The formation of lactones was observed, which is associated with the dissolution of lactobionic acid crystals; however, by dissolving the lactobionic acid crystals on alkaline calcium hydroxyde solution in equimolar ratio a single lactobionic acid chromatographic peak without lactobionolactone is obtained.
Pedruzzi
Israel
I
Laboratory of Separation and Reaction Engineering, Associate Laboratory, Faculty of Engineering University of Porto, Rua Dr Roberto Frias, 4200-465 Porto, Portugal.
Malvessi
Eloane
E
Mata
Vera G
VG
Silva
Eduardo A B
EA
Silveira
Mauricio M
MM
Rodrigues
A E
AE
D002411:Cation Exchange Resins / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004187:Disaccharides / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D005632:Fructose / Q000737:chemistry*; D007785:Lactose / Q000737:chemistry*; D015203:Reproducibility of Results; D013012:Sorbitol / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D047392:beta-Cyclodextrins / Q000737:chemistry
D002411:Cation Exchange Resins; D004187:Disaccharides; D047392:beta-Cyclodextrins; D005632:Fructose; D013012:Sorbitol; C005608:lactobionic acid; D007785:Lactose; C031215:betadex