Cross-linked glucose isomerase crystals as a liquid chromatographic separation material

Area of Science:

• Biochemistry
• Analytical Chemistry
• Materials Science

Background:

• Enzyme immobilization is crucial for developing reusable biocatalysts.
• Glucose isomerase is a well-studied enzyme with potential applications beyond its native function.
• Developing novel stationary phases for liquid chromatography is essential for advancing separation science.

Purpose of the Study:

• To characterize cross-linked glucose isomerase crystals (CLGI) as a stationary phase for liquid chromatography.
• To evaluate the separation capabilities of CLGI for various molecules, including polyethylene glycols and n-alcohols.
• To assess the potential of CLGI for chiral separations.

Main Methods:

• Size exclusion chromatography was employed to determine porosity and pore size distribution using D(2)O and polyethylene glycols.
• The separation of n-alcohols (C1-C8) was investigated, analyzing hydrophobic interactions.
• The chiral separation of D- and L-arabitol was performed to evaluate enantioselective properties.

Main Results:

• CLGI crystals exhibited a porosity of 0.47, with an average apparent pore diameter of 29 Å, indicating a microporous structure.
• Effective separation of polyethylene glycols <1000 g/mol was achieved.
• CLGI demonstrated separation of n-alcohols C1-C8, with Height Equivalent to a Theoretical Plate (HETP) values ranging from 1.6 to 0.89 mm.
• Chiral separation of D- and L-arabitol with a resolution (Rs) of 0.58 was accomplished.

Conclusions:

• Cross-linked glucose isomerase crystals are a viable material for liquid chromatographic separations.
• CLGI shows potential for both normal-phase and chiral chromatography due to its unique pore structure and hydrophobic/chiral properties.
• Further investigation into CLGI for chiral separation of amino acids is warranted.