137

Clinica Chimica Acto, 69 (1976) 137-138 @ Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands

BRIEF TECHNICAL NPTE ____-

~--

_

-

CGA 7176

QUANTITATIVE

DETERMINATION

OF DERMATAN SULFATE IN URINE

R. HUMBEL State Pediatric

Clinic, Luxembourg

(G.D.L.)

(Received December 29,1975)

Di Ferrante et al. [l] described in 1971 a specific reaction for the determination of dermatan sulfate (DS) which is an application of the periodate-acid Schiff’s reaction of Scott [2]. This reaction is specific for DS providing that glycogen and glycoproteins are removed. Glycoproteins are known to coprecipitate with glycosaminoglycans when using quaternary ammonium salts as precipitating reagent [ 31. However, few publications refer to the utilization of the Di Ferrante method. In the present note we wish to report a simple purification step for urinary glycosaminoglycans and an application of this method to the quantitative determination of DS in urine of patients with mucopolysaccharidosis. Material and method Urinary glycosaminoglycans were isolated from 10 ml urine by precipitation with cetylpyridinium chloride as previously described. The dry extract was dissolved in 1 ml 0.6 M NaCl and kept at 4°C for 30 min. After centrifugation the clear supernatant was used for microchromatography [ 41. A micro-column (height 60 mm, diameter 3 mm) was packed with DOWEX I X 2 Cl- type (200-400 mesh) suspended in 0.6 M NaCl. The urinary extract was applied to the column. Glycoproteins were eliminated by washing with 1 ml 0.6 M NaCl. Glycosaminoglycans were eluted with two portions of 1 ml 2.0 M NaCl. Determination of DS was performed on 0.2 ml of the latter eluate as indicated by Di Ferrante [ 11. The standards of DS were also prepared in 2.0 M NaCl. Results Table I indicates the different results obtained in the various types of mucopolysaccharidosis. As can be seen, increased amounts of DS are found only in

138

TABLE

I

QUANTITATIVE

DETERMINATION

OF

DS

IN

Dermatan (mg/g ___~. Normal

controls

(17)

URINE

SPECIMENS

sulfate

creatinine) ~~~~

~~~

_~ ~_

~~~ ~~~

~

..~~

0.55-5.90

Mucopolysaccharidoses

-_.

57

(39)

Type

IH (14)

57.43-280.00

Type

II (9)

32.30-350.00

Type

Is

(4)

65.70-116.70

TYPC

VI (4)

29.10-280.00

Type III (8) .___~

2.85-9.28

mucqpolysaccharidoses of Types I,, II, I, and VI which is in agreement with known findings. Differential determination of iduronic and glucuronic acids using the carbazol and naphthoresorcinol methods leads to the characterization of DS [ 51 whereas the present method gives quantitative results. References Di Ferrante, Scott,

N.,

J.E.

Di Ferrante, Humbel, Teller,

(1974)

(1967)

P.V.

Biochim.

N. (1967)

R. W.

Donnelly.

(1968)

Anal.

Clin. Z. Klin.

and

Berglund.

Biophys.

Acta

Biochem.

Chin Chem.

Acta Klin.

21. 52,

170,

R.K.

(1971)

471

98

173

Biochem.

5, 29

Biochem.

J. 124.

549

Quantitative determination of dermatan sulfate in urine.

137 Clinica Chimica Acto, 69 (1976) 137-138 @ Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands BRIEF TECHNICAL NPTE __...
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