I have digested my plant sample (50 mg) in 1 mL of 6N HCl for amino acid analysis. How should I proceed to dry the sample? I have a freeze dryer but no acid trap so I cant lyophilize. Please advise!
Dear Anna,
you have to adjest the pH of your sample to set with pH of mobile phase prior to HPLC injection.
Piyal Aravinna
The simple route is to neutralise as Aravinna suggests. If that is not possible then you are correct to NOT use a basic freeze dryer, as you say, the HCl will damage the metal components and destroy the vacuum pump, perhaps not on the first time you try it, but it will kill the system quite quickly.
If you have one, or a few samples, a glass rotary evaporator will be OK for evaporation to remove the HCl, however, take care with the vacuum pump, use a dry diaphragm type pump, not a mechanical oil based pump (as is used with a freeze dryer). Take care where the vapours from the pump are exhausted to ... I have seen these destroy laboratory hardware and instruments if you are not careful. For similar reasons I would not recommend a nitrogen blowing system as the HCl will be in the atmosphere!
The company Genevac, produce a truly acid resistant evaporation system for multiple samples which withstand routine use with 6M HCl. The downside is that these systems are very expensive.
Best regards,
Rob
Thanks Aravinna and Rob.
I have a BUCHI rotavap with a trap attached, so I may be able to attempt drying that way but I'm not sure that I'll be able to pull off the HCl at 37-40 deg C. Our (central) vacuum is likely not strong enough. I will try this. If its not successful- then neutralizing it is! Thanks again
PS: A concern with neutralizing with NaOH would be formation of excess salts? Could this have negative impacts on my HPLC analysis? How could I de-salt the samples?
Anna,
To get water to boil at 37°C then a vacuum level of 50mbar is required.
The lower the pressure, the lower the boiling point ... however, if you boil it, you have to catch it in the cold trap (or the pressure will rise). The condensation point = the boiling point, so if you have a cold trap at 10°C and then you can't boil water colder than this.
Kind regards,
Rob
you should neutralize the excess HCl with alkali such as NaOH till pH =7
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