S35-sulfate was injected intraperitoneally- into 7-day-old rats and their long bones were removed after intervals of time. The epiphyses were separated from the diaphyses for analysis.
From the diaphyses freed of bone marrow about 82 per cent of the S35 which they contained was extracted with a 2.5 N solution of sodium hydroxide. More, about 91 per cent of the S35, was thus extracted from the epiphyses.
Dialysis of the extracts against water showed that the fraction of S35 which was dialyzable decreased rapidly with time. After 1 hour about 80 per cent and 50 per cent of the S35 in the extracts of diaphyses and epiphyses, respectively were found in the dialysates, after 24 hours about 20 per cent and 4 per cent, and after 120 hours 12 per cent and 1 per cent. Similar values for the S35 in inorganic sulfate were found when the extracts were chromatographed on an anion exchange resin, dowex-2.
The S35, other than inorganic sulfate, was in the form of bound sulfate, which was released by acid hydrolysis. Uronic acid and hexosamines, primarily galactosamine, were associated with the S35. Indeed, on paper electrophoretograms and paper chromatograms the major S35-labelled component which was seen resembled chondroitin sulfate in its mobility. On the paper chromatograms, also a second S35-labelled component with a mobility lower than that of chondroitin sulfate was found. It is unlikely that the latter is a breakdown product of chondroitin sulfate, produced in the course of extraction with the sodium hydroxide solution. In fact, both components were also found in sodium versenate homogenates which had been dialyzed extensively against water.
On the basis of these results it is suggested that the greatest part of the S35-labelled materials previously demonstrated by autoradiography to be progressively deposited in the metaphyses after 24 hours,—as the concentration of S35-sulfate concurrently decreased in the epiphyseal cartilage plates,—are akin to the chondroitin sulfate of the epiphyseal cartilage plates and are derived from the latter.