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[Protein analysis of enzyme tablets]
B Schulz1, S Stoeva, W Voelter
1Abteilung für Physikalische Biochemie des Physiologisch-Chemischen Instituts, Eberhard-Karls-Universität, Tübingen.
Die Pharmazie
|October 12, 2001
Summary
Enzyme tablets containing butyrylcholine esterase (CHE) and peroxidase (POD) retain some activity after high-pressure compaction. However, significant changes in circular dichroism spectra indicate structural alterations in these enzymes.
Area of Science:
- Biochemistry
- Materials Science
- Enzyme Engineering
Background:
- Enzymatic activity is crucial for many applications.
- Compaction processes can affect enzyme stability and function.
- Understanding enzyme behavior under pressure is important for formulation development.
Purpose of the Study:
- To investigate the impact of high-pressure compaction on the enzymatic activity and structural integrity of butyrylcholine esterase (CHE) and peroxidase (POD).
- To compare the effects of compaction with those of thermal treatment on enzyme properties.
Main Methods:
- Enzyme tablets were prepared using high-pressure compaction (495 MPa).
- Enzymatic activity assays were performed.
- Spectroscopic techniques including ultraviolet absorbance and fluorescence spectroscopy were used.
- Far-ultraviolet circular dichroism (CD) spectroscopy was employed to assess secondary structure.
- Polyacrylamide gel electrophoresis (PAGE) was used to evaluate protein integrity.
- Thermal treatment (150°C) was applied to solid enzyme samples for comparison.
Main Results:
- Compaction at 495 MPa led to a partial loss of enzymatic activity for both CHE and POD.
- No significant changes in ultraviolet absorbance or fluorescence intensity were observed in tablet solutions compared to original enzymes.
- Far-UV CD spectra showed only minor alterations after compaction, suggesting limited changes in secondary structure.
- PAGE analysis revealed no missing or additional protein bands, indicating no major degradation or aggregation.
- Solid enzyme samples heated to 150°C retained partial enzymatic activity.
- Thermal treatment caused an increase in ultraviolet absorbance and significant changes in CD spectra, indicating denaturation.
Conclusions:
- High-pressure compaction partially reduces enzymatic activity of CHE and POD but causes less structural damage compared to severe thermal treatment.
- Enzymes in solid-state tablet formulations may exhibit different stability profiles than in solution.
- Further studies are needed to optimize compaction parameters for preserving enzyme function in solid dosage forms.