G. A. Levvy

Preparation and Properties of β-Glucuronidase

Publisher Summary This chapter discusses the preparation and properties of β–glucuronidase. All mammalian tissues contain a group-specific enzyme that catalyzes the hydrolysis of the biosynthetic β-D-glucopyranosiduronic acids of all types to the aglycons and D-glucuronic acid. This enzyme is commonly known as “β-glucuronidase,” a term that will be employed in the following pages. In the same way, for the sake of brevity, β-D-glucopyranosiduronic acids will often be referred to as β-glucuronides. Enzymes that decompose β-glucuronides have been obtained from diverse sources other than the mammalian cell. β-Glucuronidase finds its major application at the present time as a reagent for the hydrolysis of urinary steroid D-glucuronides, for which purpose it has certain advantages over mineral acids, and the rapid changes in tissue β-glucuronidase activity induced in vivo by certain steroid and pituitary hormones offer possibilities for the rapid measurement of the hormones. The enzyme might perhaps be employed with advantage as a hydrolytic agent in carbohydrate chemistry. Most of the assay methods depend upon colorimetric determination of the amount of aglycon liberated from a specified concentration of substrate at a fixed pH, the results being read from a standard curve. Unit β-glucuronidase activity is then almost universally expressed as that which liberates 1 μg. of aglycon in 1 hr. at 38°; a similar type of unit is employed when the liberation of D-glucuronic acid is followed.

Methyl and Phenyl Glycosides of the Common Sugars

Publisher Summary This chapter discusses the methyl and phenyl glycosides of the common sugars. Several new general methods or modified procedures for glycoside synthesis have been introduced. The O -acylglycosyl halides, where the acyl group at the potential reducing group in a fully acylated reducing sugar has been replaced by halogen, are the most generally useful intermediates in the synthesis of alkyl and aryl glycosides, and the O -acetylglycosyl bromides (“acetobromosugars”) are by far the most commonly employed. They are more reactive than the chlorides and more stable than the iodides. In some instances, however, the bromide is so unstable that the chloride is preferred. Yields of O -acetylglycosyl bromides obtained from other sugars by this method were: lactose 85%, arabinose 50%, maltose 60%, cellobiose 72%, and galactose 75%. With cellobiose, some glacial acetic acid has to be added at the start to maintain solution. In general, the products are pure enough to be used without recrystallization in the next stage of glycoside synthesis. Apart from their use as starting materials for the preparation of O -acetyl-glycosyl halides, polyacetylated sugars are employed extensively in glycoside synthesis by the Helferich reaction, in which they are condensed directly with low-melting phenols.

The determination of the individual neutral and amino sugars in carbohydrates.

Abstract An experimental study has been made of procedures for the acid hydrolysis and methanolysis of sugar polymers. Individual neutral and amino sugars in the products were examined by colorimetric methods and by gas-liquid chromatography of the trimethylsilyl derivatives. Quantitative N -acetylation of amino sugars was carried out with dilute acetic anhydride in aqueous acetone (1:1, v/v) on a column of Dowex 1 (CO 3 2− form). Methanolysis was found to be preferable for the release of neutral sugars from glyco-proteins, whereas acid hydrolysis should be employed for amino sugars.

The heterogeneity of ovalbumin glycopeptide

Abstract When passed repeatedly down a Sephadex G-25 column, pronase digests of ovalbumin gave glycopeptide fractions with hexose: hexosamine ratios varying progressively from 5:4.8 to 5:2.5. The bulk of the material approximated to a 5:3 ratio, but showed continuous dispersion on recycling on Sephadex. The preparation of pronase contained hexose and yielded a glucose-containing glycopeptide on autodigestion. Errors from this source were avoided by purifying the proteolytic enzyme before use. Pronase was examined for some of the commoner glycosidase activities, with negative results.