Studies on labeling and application to microanalysis of biotin related compounds, 5

The isotope dilution method was established for the determination of d-dethiobiotin. This compound reacts with p-(dimethylamino) cinnamaldehyde (p-DACA) to develop an intense color having an absorption maximum at 533 mμ. By using the proposed isotope dilution analysis in conjunction with the absorbancy and radioactivity, the average recovery rate of d-dethiobiotin and d-biotin from synthetic mixed sample showed as 100.9% for d-dethiobiotin and 100.4% for d-biotin. By this method d-dethiobiotin and d-biotin were determined periodically up to 10 days in the culture filtrate of Aspergillus niger incubated in a medium containing d-dethiobiotin. As the cultivation proceeds, the amount of d-biotin produced increased gradually and reached approximately 14% of the d-dethiobiotin added, which decreased simultaneously. By means of the reverse isotope dilution method, the amount of 3H-d-biotin produced from 3H-d-dethiobiotin (4.67 μmole) was 1.48% for a 2-day incubation and 5.97% for 4 days, as a total yield in the cell and the culture filtrate.