SEC-MALS characterization of microbial polyhydroxyalkanoates

Area of Science:

• Polymer Science
• Analytical Chemistry

Background:

• Polyhydroxyalkanoates (PHAs) like PHB and PHBV are biodegradable polymers.
• Accurate characterization of PHAs is crucial for understanding their properties and applications.
• Chloroform is a common solvent for PHA analysis, but sample preparation can impact results.

Purpose of the Study:

• To characterize poly-3-hydroxybutyric acid (PHB) and poly-3-hydroxybutyric-co-valeric acid (PHBV) in chloroform.
• To evaluate the effect of different sample preparation methods on PHA dissolution and integrity.
• To determine absolute molar mass, molar mass distribution, and radius of gyration.

Main Methods:

• Size Exclusion Chromatography coupled with Multi-Angle Light Scattering (SEC-MALS).
• Analysis of three sample preparation methods: room temperature dissolution, 60°C dissolution, and thermal pretreatment (annealing at 180°C).
• Differential Scanning Calorimetry (DSC) and Nuclear Magnetic Resonance (NMR) for degradation analysis.

Main Results:

• Dissolution at 60°C and of thermally pretreated samples yielded molecularly dissolved PHB and PHBV.
• No degradation was observed for PHB/PHBV in chloroform at 60°C for up to 120 minutes.
• Thermal annealing at 180°C caused significant degradation, with slightly higher rates for PHB than PHBV.
• Both polymers formed random coils in chloroform, as indicated by the radius of gyration-molar mass relationship.

Conclusions:

• Optimized sample preparation (dissolution at 60°C or post-thermal pretreatment) is essential for accurate PHA characterization in chloroform.
• High-temperature annealing leads to polymer degradation, affecting molar mass averages and distribution.
• SEC-MALS provides reliable absolute molar mass determination for PHB and PHBV in solution.