Fabrication of paper-based enzyme immobilized microarray by 3D-printing technique for screening α-glucosidase inhibitors in mulberry leaves and lotus leaves.

BACKGROUND

The discovery of bioactive compounds in traditional Chinese medicine (TCM) has become an important field in TCM modernization. Ligand fishing is a suitable method for discovery of bioactive compounds in complex mixtures such as TCM with high selectivity. Because of unique advantage of low cost and convenience, paper-based microdevices can be good carriers for enzyme immobilized ligand fishing.

METHODS

As an important enzyme for glucose metabolism, α-glucosidase was immobilized on polycaprolactone-chitosan-modified paper to prepare the microdevice with unique microfluid structure generated by 3D printing technology, which can be easily applied to screen active compounds in herbal extracts. The preparation conditions of the paper microarray were optimized. The activity of immobilized α-glucosidase was verified by colorimetric reactions which can be easily monitored by cellphone. The paper microarray with α-glucosidase immobilized was used to screen active compounds in the water extracts of mulberry leaves and lotus leaves.

RESULTS

Several key parameters including NaCO solution concentration, NaCO solution volume, glutaraldehyde concentration, crosslinking time of glutaraldehyde and time of α-glucosidase immobilization were optimized. The proposed paper-based microarray was successfully applied in screening active compounds in two herbal extracts. Four compounds including chlorogenic acid, quercetin-3--glucuronide, isoquercetin, and quercetin were identified as α-glucosidase inhibitors. The compounds with significant non-specific adsorption caused by chitosan, such as isoquercitrin, astragalin, quercetin, were also found to be active compounds.

CONCLUSIONS

An enzyme immobilized paper microarray was designed and fabricated in this work. Polycaprolactone and chitosan were used to modify filter paper to prepare paper microarrays. Parameters of paper device preparation were optimized. Our findings suggested that 3D-printing paper-based microarrays can be a simple and low-cost approach for discovery of active compounds of TCM.