Determination of Dextran with Anthrone

Scott, Thomas Allan; Melvin, Eugene H.

doi:10.1021/ac60083a023

Cited by 1,141 publications

(417 citation statements)

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“…This was determined with the anthrone reaction (Scott & Melvin, 1953) with galactose as standard. Glycosaminoglycuronans interfere with the anthrone reaction.…”

Section: Concentration Of Fractions Polysaccharide Fractionsmentioning

confidence: 99%

The nature of the non-ultrafilterable glycosaminoglycans of normal human urine

Wessler

1971

Biochemical Journal

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1. The non-ultrafilterable acidic glycosaminoglyeans from pooled urine ofnormal men, aged about 20, were isolated and characterized. The isolation procedure included digestion with sialidase and pronase, and fractionation by stepwise elution from an ECTEOLA-cellulose column. The glycosaminoglycans in each fraction were separated from each other by preparative electrophoresis in sodium barbital buffer and in barium acetate. 2. Approximate relative amounts of the different glycosaminoglycans were: chondroitin sulphate 60%, chondroitin 2%, hyaluronic acid 4%, dermatan sulphate 1%, heparan sulphate 15% and keratan sulphate 18%. Chondroitin sulphate-dermatan sulphate hybrids seemed to occur in trace amounts. 3. Chondroitin sulphate, heparan sulphate and keratan sulphate were heterogeneous with respect to degree of sulphation. Two distinct groups of chondroitin sulphate fractions were found, with sulphate/hexosamine molar ratios of about 0.5 and 1 respectively. The sulphate/hexosamine molar ratios in the heparan sulphate fractions varied from 0.5 to 0.9;theN-sulphate/hexosamineratiowas about 0.5 in all fractions. The sulphate/hexosamine molar ratios in the keratan sulphate fractions varied from 0.2 to 0.7.All of the types of glycosaminoglycan that occur in human tissues, with the exception of heparin, have also been found in normal urine (

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“…This was determined with the anthrone reaction (Scott & Melvin, 1953) with galactose as standard. Glycosaminoglycuronans interfere with the anthrone reaction.…”

Section: Concentration Of Fractions Polysaccharide Fractionsmentioning

confidence: 99%

The nature of the non-ultrafilterable glycosaminoglycans of normal human urine

Wessler

1971

Biochemical Journal

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“…Colorimetric methods were used for determination of hexose [19], 6-deoxyhexose [20], sialic acid [21], and protein Sugar analyses were performed by gas-liquid chromatography [23] and mass spectrometry [24]. Optical rotations were recorded using a Perkin-Elmer 241 polarimeter.…”

Section: Methodsmentioning

confidence: 99%

Isolation and Characteristics of Human Urinary Sialoglycoproteins

Berggård¹,

Ekström²,

Lisowska

et al. 2005

European Journal of Biochemistry

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Three sialoglycoproteins (Sl, S, , and S, ) have been isolated from normal human urine by ultrafiltration, zone electrophoresis, gel chromatography, and ion-exchange chromatography. The average yields per litre of urine were 0.39 mg (S I)r 0.40 mg (S2), and 1.9 mg (S3). The isolated substances were homogeneous on ultracentrifugation both at neutral and acid pH with sedimentation coefficients of 3.6s (S,), 2.4s (S2), and 0.93s ( S 3 ) . Equilibrium ultracentrifugation gave molecular weights of 77 900 (Si), 37 000 (S, ), and 5300 (S3). All three components were rich in carbohydrate (64 to 76 x) and contained 28 to 35 '%, ofsialic acid. Alkaline borohydride degradation of component S, yielded two sialylated oligosaccharides which were partially characterized.The origin of the isolated substances is unknown. The molecular size of S, (Stokes' radius of 2.0 nm) is compatible with passage from the blood by glomerular filtration whereas the size of S, (Stokes' radius of 6.5 nm) would suggest a renal origin.Normal human urine is known to contain macromolecules characterized by a particularly high content of sialic acid [I]. Only limited information is available concerning the chemical nature and origin of these substances. One of us [2] has previously reported the preparation by electrophoresis of a urinary glycoprotein fraction rich in sialic acid. The present paper describes the isolation from this material of three sialoglycoproteins (designated S SZ, and S,, respectively). Some of the physical and chemical properties of the isolated substances are also given. EXPERIMENTAL PROCEDURE MaterialsUrine from healthy male individuals was collected and pooled, and bacterial growth was prevented by addition of phenylmercuric nitrate to a final concentration of 1 : 50 000. The urine was filtered at 4 "C, and immediately ultrafiltered.Pevikon C-870 (Kema-Nord AB, Stockholm, Sweden), DEAE-cellulose (Serva Entwicklungslabor, Heidelberg, FRG), and Sephadex (3-25, G-100, G-200 and Blue Dextran 2000 (AB Pharmacia, Uppsala, Sweden) were used according to the instructions furnished by the manufacturer. Rabbit anti-M and anti-N sera were produced in the Laboratory of Blood Groups (Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wroclaw, Poland). Extract of Vicia grarninea seeds was prepared [3] at pH 6.5. Neuraininidase from Vibrio cholerae was obtained from Behringwerke AG (Marburg/Lahn, FRG). General MethodsConcentration by Ulfrqfiltration. The urinary macromolecules were concentrated by ultrafiltration with 23/32-inch * Deceased in June 1978.AhbrPriarions. NeuAc, N-acetylneuraminic acid; II-G~I, n-galactose: Galg, galactopyrenose; D-G~INAcOH, 2-acetamido-2-deoxy-o-gaIactitol. Electrophore Tis. Preparative zone electrophoresis in blocks of Pevikon C-870 were carried out as previously described [5-71. All experiments were done in 0.1 M sodium acetate buffer pH 4.5.Determinations of' Stokes' Radii, Dff"Usion Coqfficien f s , Sedimentation Coefficients, Molecular Weights, and Frictionul Ratios. Stokes' molec...

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“…Total lipid determinations were performed according to Frings and Dunn (1970) and carbohydrates, according to Scott and Melvin (1953).…”

Section: Methodsmentioning

confidence: 99%

“…For determination of glycogen, the fat bodies were homogenized in a KOH 30%, boiled for 30 min and precipitated with ethanol (Hassid & Abraham 1957). Then, the carbohydrates were determined by the anthrone method (Scott & Melvin 1953).…”

Section: Determination Of Lipophorin Hdlpmentioning

confidence: 99%