Physicochemical characterization of complex drug substances: evaluation of structural similarities and differences of protamine sulfate from various sources.
The purpose of this study was to characterize and evaluate differences of protamine sulfate, a highly basic peptide drug, obtained from five different sources, using orthogonal thermal and spectroscopic analytical methods. Thermogravimetric analysis and modulated differential scanning calorimetry showed that all five protamine sulfate samples had different moisture contents and glass transition and melting temperatures when temperature was modulated from 25 to 270°C. Protamine sulfate from source III had the highest residual moisture content (4.7 ± 0.2%) at 105°C, resulting in the lowest glass transition (109.7°C) and melting (184.2°C) temperatures compared with the other four sources. By Fourier-transform infrared (FTIR) spectroscopy, the five sources of protamine sulfate had indistinguishable spectra, and the spectra were consistent with a predominantly random coil conformation in solution and a minor population in a β-sheet conformation (~12%). Circular dichroism spectropolarimetry confirmed the FTIR results with prominent minima at 206 nm observed for all five sources. Finally, proton ((1)H) nuclear magnetic resonance spectroscopy showed that all five protamine sulfate sources had identical spectra with backbone amide chemical shifts between 8.20 and 8.80 ppm, consistent with proteins with predominantly random coil conformation. In conclusion, thermal analyses showed differences in the thermal behavior of the five sources of protamine sulfate, while spectroscopic analyses showed the samples had a predominantly random coil conformation with a small amount of β-sheet present.
Division of Product Quality Research, OTR, OPS, CDER, FDA, Silver Spring, Maryland, USA.
David
D
Awotwe-Otoo
Cyrus
C
Agarabi
David
D
Keire
Sau
S
Lee
Andre
A
Raw
Lawrence
L
Yu
Muhammad J
MJ
Habib
Mansoor A
MA
Khan
Rakhi B
RB
Shah
D002152:Calorimetry, Differential Scanning; D002942:Circular Dichroism; D009682:Magnetic Resonance Spectroscopy; D011479:Protamines / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared; D013818:Thermogravimetry
D011479:Protamines