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Pure mucin isolated from canine submaxillary glands was treated with alkaline borohydride and products of degradation were fractionated by a procedure that included ion-exchange chromatography, gel filtration, high-voltage electrophoresis and paper chromatography.Seven acidic, reduced oligosaccharides were isolated being divided into two distinct types : one, containing sialic acid and no glucosamine or sulfate (type A) and the other containing glucosamine and sulfate but no sialic acid (type B).The most complete oligosaccharide of type A designated component (I) was a tetrasaccharideThe most complex oligosaccharide of type B designated component (V) has a hexasaccharide with the following structure : a-L-fucopyranosyl-(I -i2)-P-~-galactopyranosyl-(l+3, 4, orAll the other acidic isolated oligosaccharides were derived from these two main components.No oligosaccharide containing both sialic acid and glucosamine sulfate could be demonstrated. Based on these structural features, a possible biosynthetic pathway leading to the two different oligosaccharide chains is discussed.Variations in ratios of sugars observed in canine submaxillary mucin and especially the reciprocal relationship between sialic acid and fucose noted earlier (1962) by Dische, Pallavicini, Kavasaki, Smirnow, Cizek and Chien may reflect variations in the proportions of these two types of oligosaccharides.Mucins from the submaxillary glands of a variety of species have been shown to consist of a polypeptide chain with many relatively small oligosaccharide side chains linked to the peptide through glycosidic bonds between the anomeric carbon of N-acetylgalactosamine and the hydroxyl group of serine or threonine [I].The oligosaccharides from ovine and bovine submaxillary mucins have been shown to be mainly t Deceased (September 28, 1972).Abbreviations. Fuc, L-fucose; Gal, D-galaCtOSe; AcNeu, N-acetylneuraminic acid; GlcNAc, N-acetylglucosamine (2-deoxy-2-acetamide-~-glucose); GalNAcol, N-acetylgalactosaminitol(2-deoxy-2-acetamido-galactitol).Enzymes (CBN Recommendations 1972). a-I,2-~-Fucosidase (EC 3.2.1.63); P-N-acetyl-glucosaminidase (EC 3.2.1.30); 8-galactosidase (EC 3.2.1.23); neuraminidase (EC 3.2.1 .18).sialyl-N-acetylgalactosamine [I 1. The oligosaccharides of porcine submaxillary mucin are more complex, consisting of structures up to the size of pentasaccharides, some having blood group A activity [2-51.The structures of oligosaccharides from canine submaxillary mucin have not been investigated. However, this is a problem of considerable interest, since Dische [6,7] has found that the ratios of sialic acid to fucose in the submaxillary secretions from the dog may vary by a factor of 10 depending upon the nature of the stimulus eliciting the secretion. Although the ratio of the sugars varies widely, their sum remains constant. It was originally suggested that the two sugars, which are both terminal at the non-reducing ends of oligosaccharide chains, had a reciprocal relationship in which, under one set of secretory conditions, oligosaccharide...

The isolation and properties of mucin from submaxillary glands of a number of species of animals have been reported (reviewed by Pigman & Gottschalk, 1966).The present communication describes a method for preparation of purified canine submaxillary mucin involving gel filtration on Sephadex G-200 with an additional fractionation step using Bio-Gel CM-100. This method yields a native and highly purified material from submaxillary glands and from submaxillary secretions. The composition of the purified mucin indicates that it is a sulphated glycoprotein containing 35 % protein, 61 % carbohydrate and 3.8% sulphate. The findings extend previous work on canine submaxillary mucin reported by Hashimoto & Pigman (1962).Canine submaxillary glands either were obtained from dogs under anaesthesia and immediately frozen in liquid N2, or were obtained frozen from Pel-Freeze Inc. (Rogers, Ark., U.S.A.). The glands were decapsulated, freed from connective tissue and fat, cut into small pieces and extracted with 2vol. of ice-cold distilled water containing NaN3 (5mM) for 24h at 4°C (extract 1). Extracts 2 and 3 were obtained by repeating the extraction twice with 1 vol. of water for 48h. The extracts were combined and dialysed overnight in the cold against 0.1 M-sodium acetate buffer, pH 3.9, and a small precipitate was removed by centrifugation.Then 80ml of the combined dialysed supernatant, containing 610mg of protein, was applied to a Sephadex G-200 column (5cm x 81cm) previously equilibrated with 0.1 M-sodium acetate buffer, pH3.9. Elution was carried out with the same buffer at a flow rate of 36ml/h. Fractions of volume 20ml were collected, and samples were analysed for protein by measuring the absorbance at 280nm, for hexoses by the orcinol method (Winzler, 1955) and for sialic acid by the direct Ehrlich method (Werner & Odin, 1952). Virtually all of the sialic acid, 93 % of the hexose and 43 % of the protein were eluted in the void volume. The remaining protein was eluted in two retarded peaks, one of which contained some hexose. Recoveries of sialic acid, hexose and protein were greater than 95 %. For large-scale preparations 300ml (2.29g of protein) was applied to a column of larger size (9.7cm x 105 cm). The fractions constituting the Vol. 128

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