TY - JOUR
T1 - Secondary structure characterization of microparticulate insulin powders
AU - Yeo, Sang‐Do ‐D
AU - Debenedetti, Pablo G.
AU - Patro, Sugunakar Y.
AU - Przybycien, Todd M.
PY - 1994/12
Y1 - 1994/12
N2 - The secondary structure content of microparticulate insulin powders produced by the supercritical antisolvent (SAS) precipitation technique was investigated via Raman spectroscopy. Precipitate samples were generated at 25 and 35 °C processing temperatures. Both precipitate samples gave amide I band spectra that were shifted roughly +10 cm−1 relative to the commercial powder. The corresponding secondary structure estimates had significantly increased β‐sheet contents with concomitant decreases in α‐helix contents relative to the commercial protein; the sum of β‐turn and random coil content remained essentially unchanged. The magnitude of the perturbation was slightly greater for the 35 °C sample. Dissolution of the commercial powder and precipitates in 0.01 M HCl yielded solution structure estimates similar to that of the commercial powder. An analysis of insulin in dimethyl sulfoxide, the suspending solvent in the SAS process, indicated that some, but not all, of the structural change observed for the precipitate samples may be attributed to solvent exposure. These results are suggestive of extensive β‐sheet‐mediated intermolecular interactions in precipitate states, consistent with analyses of irreversible protein aggregate/fibril states. Interestingly, unlike irreversible protein aggregates, the insulin powders recover essentially full biological activity on reconstitution.
AB - The secondary structure content of microparticulate insulin powders produced by the supercritical antisolvent (SAS) precipitation technique was investigated via Raman spectroscopy. Precipitate samples were generated at 25 and 35 °C processing temperatures. Both precipitate samples gave amide I band spectra that were shifted roughly +10 cm−1 relative to the commercial powder. The corresponding secondary structure estimates had significantly increased β‐sheet contents with concomitant decreases in α‐helix contents relative to the commercial protein; the sum of β‐turn and random coil content remained essentially unchanged. The magnitude of the perturbation was slightly greater for the 35 °C sample. Dissolution of the commercial powder and precipitates in 0.01 M HCl yielded solution structure estimates similar to that of the commercial powder. An analysis of insulin in dimethyl sulfoxide, the suspending solvent in the SAS process, indicated that some, but not all, of the structural change observed for the precipitate samples may be attributed to solvent exposure. These results are suggestive of extensive β‐sheet‐mediated intermolecular interactions in precipitate states, consistent with analyses of irreversible protein aggregate/fibril states. Interestingly, unlike irreversible protein aggregates, the insulin powders recover essentially full biological activity on reconstitution.
UR - http://www.scopus.com/inward/record.url?scp=0028597537&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0028597537&partnerID=8YFLogxK
U2 - 10.1002/jps.2600831203
DO - 10.1002/jps.2600831203
M3 - Article
C2 - 7891290
AN - SCOPUS:0028597537
SN - 0022-3549
VL - 83
SP - 1651
EP - 1656
JO - Journal of Pharmaceutical Sciences
JF - Journal of Pharmaceutical Sciences
IS - 12
ER -