Hyaluronidase reaction kinetics evaluated by capillary electrophoresis with UV and high-resolution mass spectrometry (HRMS) detection
| Authors | |
|---|---|
| Year of publication | 2017 |
| Type | Article in Periodical |
| Magazine / Source | Analytica Chimica Acta |
| MU Faculty or unit | |
| Citation | |
| Doi | http://dx.doi.org/10.1016/j.aca.2016.11.036 |
| Field | Biochemistry |
| Keywords | capillary electrophoresis; mass spectrometry; enzymatic reaction; hyaluronidase |
| Attached files | |
| Description | A capillary electrophoresis (CE) method with enzymatic reaction inside the capillary was developed for the study of hyaluronidase activity. Capillary electrophoresis was used for the first time for the determination of the kinetic constants (Km, Vmax and IC50) of the enzymatic degradation of hyaluronic acid. Km and Vmax were determined using CE with UV and MS detection. Reaction products were identified using CE-MS. IC50 of a standard natural inhibitor, epigallocatechin gallate, was also determined by CE-UV and CE-MS. Kinetic constant values obtained by CE compared well with literature which validated the developed CE-based assay. Homemade oligosaccharides were tested to search for new hyaluronidase inhibitors (or activators). Sulfatation in 6- and 4- position on chondroitin tetrasacharides were found to inhibit hydrolysis of hyaluronic acid. Small modification of one hydroxyl group in a trisaccharide compound was shown to be enough for changing effect on hyaluronidase activity. CE-based assay can be used for the screening of natural and synthetic inhibitors of hyaluronidase activity for cosmetic and therapeutic applications. |
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