full issue - Association of Biotechnology and Pharmacy
full issue - Association of Biotechnology and Pharmacy
full issue - Association of Biotechnology and Pharmacy
You also want an ePaper? Increase the reach of your titles
YUMPU automatically turns print PDFs into web optimized ePapers that Google loves.
Current Trends in <strong>Biotechnology</strong> <strong>and</strong> <strong>Pharmacy</strong><br />
Vol. 5 (3) 1298 -1308 July 2011, ISSN 0973-8916 (Print), 2230-7303 (Online)<br />
1303<br />
while it was 5.3×10 6 cells/ml in the control<br />
(culture medium without excess ammonium).<br />
Therefore, the threshold concentration <strong>of</strong><br />
ammonium for the rCHO cells is 6 mM. The<br />
effect <strong>of</strong> ammonium stress on the activity <strong>of</strong><br />
rhFVIII is shown in Table 2. Recombinant<br />
protein production was completely stopped at 11<br />
mM NH 4<br />
Cl.<br />
Cell density 10 6 cells/ml<br />
Ammonium concentration<br />
Fig. 4. The effect <strong>of</strong> ammonium concentration on the<br />
rCHO cell density<br />
Table 2. Activity <strong>of</strong> rhFVIII under ammonium stress<br />
Ammonium rhFVIII Ammonium rhFVIII<br />
concentration activity concentration activity<br />
(mM) (%) (mM) (%)<br />
Control 6.4 9 2.6<br />
2 4.4 10 2.1<br />
4 4.1 11 0<br />
6 3.2 12 0<br />
To mitigate the adverse effects <strong>of</strong><br />
ammonium, some amino acids were selected <strong>and</strong><br />
then added to the culture medium. According to<br />
the isoelectric point <strong>of</strong> amino acids <strong>and</strong> their role<br />
in metabolic pathways related to ammonium<br />
formation (14), four amino acids consisting <strong>of</strong><br />
proline, threonine, glycine <strong>and</strong> asparagine were<br />
selected. As it is shown in Figure 5, proline <strong>and</strong><br />
glycine increased the ammonium-stressed rCHO<br />
cell density by up to 24% <strong>and</strong> 28 % (ANOVA,<br />
p