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- Posts: 37
- Joined: Tue Feb 20, 2007 12:36 pm
I am currently trying to optimize an existing HPLC-method for the determination of Hyaluronic acid. The method uses a Shodex OH-Pak SB-806 HQ, 300 x 8.0 mm column which is based on size exclusion (SEC). The Hyaluronic acid peak comes at 10 minutes and has a width of 8 - 12.5 minutes which is a very broad peak. The problem is a very close negative peak at 13.5 minutes, followed by two more peaks. I was wondering if anyone can provide some ideas how to make the Hyaluronic acid peak come earlier, hence moving it away from the other peaks.
I have inserted a typical chromatogram below and the method details are as follows:
Column: as mentioned above
Guard column: Phenomenex Security Guard C18, 4 x 3mm
Flow rate: 0.8 ml/min
RI -Detection
Columntemp: 21C
Detectortemp: 35C
Run time: 25 min
Mode: Isocratic
Mobile phase: 11.7 g Sodium chloride + 0.1 g sodium azide, dissolved and diluted to 1000ml with water.
Sample diluent: same as mobile phase
Many thanks in advance!
Max