Isolation and structure determination of a new amino acid, α-amino-γ,δ-dihydroxyadipic acid, from the hydrolysate of normal human urine

Abstract Hydroxylysyl-galactosyl-glucose (HGG) and hydroxylysyl-galactose (HG) were isolated from normal human urine and shown to be identical with hydroxylysine glycosides (OHLG's) whose structure had been elucidated in other laboratories. We devised a procedure for separating the OHLG's from many other urinary constituents by preliminary fractionation on a column of Sephadex. The OHLG's in the fractions so obtained were then isolated, uncontaminated with other ninhydrin-positive components, in a single pass through an analytical cation-exchange column equipped for splitting the effluent stream. In addition, a procedure was devised for determining as little as 10-9 mol of the OHLG's in 100 µl of filtrate (corresponding to 83.3 µl, of urine) on standard ion-exchange chromatographic amino acid analyzers. In normal adult urine, 1.5, µmol of each glycoside was present per 100 mg of creatinine; the molar ratio of HHG to HG was about 1.1 to 1.2. Each was present in plasma in about 400- to 500-fold smaller concentration than in urine. After surgical fusion of the spine, excretion of OHLG's increased and the ratio of HGG to HG decreased, changes we interpreted as indicating a more rapid bone-collagen turnover consequent to surgical damage.

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IMMUNOREACTIVE GROWTH HORMONE IN HUMAN URINE

Acta Endocrinologica ◽

10.1530/acta.0.0710665 ◽

1972 ◽

Vol 71 (4) ◽

pp. 665-676 ◽

Cited By ~ 19

Author(s):

Kristian F. Hanssen

Keyword(s):

Molecular Weight ◽

Growth Hormone ◽

Linear Relationship ◽

Human Urine ◽

Gel Filtration ◽

Human Growth ◽

Normal Subjects ◽

Normal Human ◽

Radio Immunoassay

ABSTRACT By using a double antibody radio-immunoassay (pre-precipitation technique) for the determination of immunoreactive human growth hormone (IRHGH) in normal human urine concentrated by dialysis and lyophilization, a factor was revealed that displaces 125I-HGH from HGH antibodies. This displacement was neither due to salts nor to glucose; it is suggested that it is due to IRHGH in the urine. A linear relationship between dilution of urine and the measured IRHGH concentration was obtained. Recovery of exogenous HGH was between 70–105%. The recovery of IRHGH from different volumes of urine following dialysis and lyophilization was between 97–110%. Plasma IRHGH and urinary IRHGH was measured simultaneously after HGH injection in a normal subject. A correlation was shown between plasma IRHGH and urinary IRHGH. In 9 normal subjects, the urinary IRHGH ranged from 28–53 ng/24 h. The excretion of urinary IRHGH was increased in acromegaly and was diminished in some, but not in all patients with adult hypopituitarism. The urinary IRHGH was further studied by gel filtration. It was recovered in one peak corresponding to a molecular weight of approximately 20 000 – 30 000. However, in the present work it was not clarified whether the urinary IRHGH represents pituitary HGH excreted in the urine or a metabolite of high molecular weight with retained immunological properties.

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