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- Posts: 2
- Joined: Mon Jun 16, 2014 7:58 am
Currently our HPLC column in addition to a Shodex Sugar is the Aminex HPX-87H:
The eluant is 5mM sulphuric acid
The flow rate is 0.60 ml/min
The temperature is 50 degrees celsius
The detection is UV 210nm.
This is the existing sample preparation with additional points added as a result of the advice given on this forum:
1)Firstly we are going to adjust the acidic hydrolysate(as a result of the acidic pretreatment phase) to pH 7, using NaOH
2) Then we are going to run the sample through an SPE cartridge filled with a cation exchanger in the hydrogen form to get rid of the sodium which otherwise will accumulate on the analytical column (advice from the forum)
3) We are then going to centrifuge
4) We are then going to take 1 ml of the adjusted sample and dilute with 9 ml de-ionized water for a dilution factor of 1 in 10.
5) We are then going to filter the sample using a 0.45 µm syringe
6) We are going to transfer 1 ml to an HPLC vial for analysis
Now my question is: is this the appropriate dilution factor for this column and carbohydrate analysis (since we have just appropriated the 1:10 dilution factor from our Shodex column methodology). Can we also dilute with de-ionized water or should we use an internal standard and de-ionized water?
Should we perhaps adjust the dilution factor to 1:5 or perhaps use a dilution factor of 1:20 for carbohydrate and organic acid analysis on the Aminex HPX-87H?