Urinary excretion of pyridinolines crosslinks measured by immunoassay and HPLC techniques in normal subjects and in elderly patients with vitamin D deficiency.
Hydroxylysylpyridinoline (HP) and lysylpyridinoline (LP) are specific constituents of mature skeletal collagens excreted in urine. Their measurement represents a sensitive index of bone resorption. In this study, we have measured urinary excretion of pyridinolines crosslinks by immunoassay (ELISA) and HPLC methods in 80 patients with different bone resorption rates. We chose a sample of 44 healthy adults (30 men and 14 women) and a sample of 36 elderly patients (7 men and 29 women) presenting a secondary hyperparathyroidism due to a vitamin D deficiency. The correlation between HPLC (x) and ELISA (y) was judged satisfactory (y = 0.794x + 6.947, r = 0.92). The sensitivity of pyridinolines estimation was 50 nmol/l for immunoassay and 20 nmol/l for HPLC. The intra-assay and inter-assay coefficients of variation for the two analytical methods was < 10%. The mean excretion of crosslinks (nmol/mmol of creatinine) measured by both methods in the sample of healthy adults was higher in women than in men. The amount of pyridinolines crosslinks excreted by elderly patients with vitamin D deficiency are three time higher than those of normal adults when measured by ELISA and HPLC methods. The distribution of different molecular forms of urinary pyridinoline crosslinks was investigated. Values of pyridinolines measured by HPLC in our samples of elderly patients have shown that free and peptide-bound pyridinolines with molecular weight (mol. wt.) smaller than 1000 Da represent approximately 80% of the total pyridinolines contained in urinary samples. A study on the evaluation of the antiserum used in the immunoassay for reacting with the different molecular forms isolated from urine showed a high affinity for free and peptide-bound pyridinolines with molecular weight smaller than 10,000 Da and that do not react strongly with peptide-bound with molecular weight greater than 10,000 Da. We conclude that, although this immunoassay does not measure total pyridinolines and does not distinguish between HP and LP, it seems convenient for diagnostic of metabolic bone diseases.
Kamel
S
S
Laboratoire de Pharmacie Clinique, Faculté de Pharmacie, Amiens, France.
Brazier
M
M
Picard
C
C
Boitte
F
F
Samson
L
L
Desmet
G
G
Sebert
J L
JL
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000596:Amino Acids / Q000652:urine*; D002851:Chromatography, High Pressure Liquid; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D014808:Vitamin D Deficiency / Q000652:urine*
D000596:Amino Acids; C015484:pyridinoline; C036020:deoxypyridinoline