Chromatography Forum

Hey all

I am trying to investigate a possible interaction between aluminum and liposomes consisting of DOPC. Basically i extrude my liposome with and without aluminum. I extrude the liposomes with aluminum to see if aluminum get inside the liposome. I also extrude the liposome without aluminum and add aluminum after the extrusion to see if aluminum coordinates to the outisde of the liposome. (Due to attraction of aluminum toward the negative charge on the DOPC headgroup.)

I have these solutions presumably containing free aluminum ions and liposomes with aluminum ions in water. Is it possible for me by an HPLC column to separate the aluminum ions in solution from the aluminum trapped or adsorped by the lipsomes. The column should not remove the aluminum adsorped to the lipsome, if this occurs.

I guess that the free aluminum ions will make complex binding to the silica base used in RP & NP columns? So these cannot be used.

The setup i have can only run in isocratic mode.

I hope you can help me with my problem.

All best
Teis

Interesting...

I've not thought this through completely, but there are a few avenues of possible pursuit:

1. Polymeric RP columns would be less likely to bind Al than silica based columns. You'll have lower separation efficiency, but you should be able to work around that.

2. You can separate & purify the liposomes via SEC (open column) and assay fractions for Al via atomic absorption spectroscopy or ICP. You can also look at DOPC in these as well. Aluminum can also be determined spectrophotometrically, apparently:

http://dx.doi.org/10.1016/0026-265X(69)90062-9

3. Differential Scanning Calorimetry (DSC) could be instructive here, too.

Will keep thinking on this and post anything I come up with if you wish...

Cheers,

CJ

Hey

It is some good ideas that you have posted. I have also thought about using some kind of polymeric column, such as ion chromatography columns. It should be possible to retain the aluminum but i do not exactly know about the lipids and the aluminum bound to them.

I'll have a look at the article you posted.

I really appreciate your help, and if you get some new ideas i'm very interested in hearing them

What about using centrifugal filters? People use them to study drug encapsulation in liposomes.

I think also zeta potential measurements could be useful here.

Thanks for the input. I will take a look at those methods

I was thinking about your project again, it is really interesting.
If your liposomes are negatively charged on the surface, they definitely should change their charge upon interaction with Al ions, look at the Fig 1 here http://www.azonano.com/Details.asp?ArticleID=1214 and try to imagine it while using Al ions instead of PEG.
Look at two instruments which measure zeta potential: one of Zetasizers (Malvern) and Mobius (Wyatt).
If all Al ions get inside liposomes and liposomes are not destructed during centrifugation you can (1) measure Al concentration before extrusion with e.g. ICP (2) after extrusion, filter your liposomes with centrifugal filters (3) measure Al concentration in supernatant and (4) wash your liposomes, devastate them to release Al ions which you can measure again with ICP.

Sounds reasonable? I have no experience with liposomes.

It sounds very promising with the zeta potential. I will take contact to a person in another department to ask if i can borrow their zeta sizer.
It also seems possible with the centrifugation aso., but i think i will try a polymer based column before trying your suggestion. But it definitely is a good idea.
Again thanks a lot