title: Ascorbic acid glycation: the reactions of L-threose in lens tissue.
abstract: L-Threose is a significant degradation product of ascorbic acid at pH 7.0 in the presence of oxygen. When compared to several other ascorbate-derived degradation products, it had the greatest ability to glycate and crosslink lens proteins in vitro. To determine whether L-threose was formed in the lens, the sugars in a TCA-soluble extract from human lenses were reduced to polyols with NaBH4, acetylated and analysed by gas-liquid chromatography. The threitol levels measured were 3.4 +/- 0.8 micrograms per lens (n = 4). GC-MS measurements made after reduction with NaBD4 indicated that threitol, but little or no threose, was originally present in the human lens. Rat lenses were incubated with [1-13C]D-threose for 24 hr, and considerable D-threitol formation was seen by NMR spectroscopy. Analysis of the lenses after medium removal showed that only [1-13C]threitol was present within the lenses indicating a rapid reduction of threose within the lens, presumably by aldose reductase. Assays with human recombinant aldose reductase and with human lens cortical and nuclear extracts all exhibited sorbinil-inhibitable aldose reductase activity with L-threose as substrate. This was confirmed by incubating a preparation of [1-14C]L-tetrose (a mixture of 40% L-threose and 45% L-erythrose) with both the pure aldose reductase and crude lens extracts followed by the subsequent identification of the [1-14C]L-threitol formed by thin layer chromatography. L-Threose degrades very slowly in 0.1 M phosphate buffer at pH 7.0, but the addition of a four-fold excess of N alpha-acetyl-L-lysine accelerated the rate of disappearance of threose 30-fold, indicating a rapid glycation reaction. When [1-14C]L-tetrose was incubated with a complete bovine lens homogenate, a linear incorporation into protein was observed over a 24 hr period. Increasing levels of lens extract exhibited increasing incorporation into protein. These data confirm a rapid reactivity of L-threose with lens protein and argue that glycation would occur in vivo in spite of the presence of aldose reductase.
authors.affiliation: Mason Institute of Ophthalmology, University of Missouri, Columbia 65212.
authors.forename: B J
authors.initials: BJ
authors.lastname: Ortwerth
authors.forename: J A
authors.initials: JA
authors.lastname: Speaker
authors.forename: M
authors.initials: M
authors.lastname: Prabhakaram
authors.forename: M G
authors.initials: MG
authors.lastname: Lopez
authors.forename: E Y
authors.initials: EY
authors.lastname: Li
authors.forename: M S
authors.initials: MS
authors.lastname: Feather
mesh_terms: D000449:Aldehyde Reductase / Q000378:metabolism; D000818:Animals; D001205:Ascorbic Acid / Q000378:metabolism*; D002417:Cattle; D002849:Chromatography, Gas; D003459:Crystallins / Q000378:metabolism; D046508:Culture Techniques; D006031:Glycosylation; D007908:Lens, Crystalline / Q000187:drug effects / Q000378:metabolism*; D008239:Lysine / Q000031:analogs & derivatives / Q000494:pharmacology; D013402:Sugar Alcohols / Q000378:metabolism; D013780:Tetroses / Q000378:metabolism*
chemical_list: D003459:Crystallins; D013402:Sugar Alcohols; D013780:Tetroses; C002315:N(alpha)-acetyllysine; C003460:threitol; D000449:Aldehyde Reductase; D008239:Lysine; D001205:Ascorbic Acid; C073321:erythrose