I'm working on the liposome digestion and relevant cumulative release now and have one question. So based on several reference papers, it seems that they mixed the liposome supsension with simulated gastric fluid or simulated intestinal fluid in a large volume ratio such as (1:3 or 1:4). They did this because if large volume of liposomes is added the absorbance of the bioactive release will be easily measured. If liposome was added in a small amount, it may not be measured easily. However, if I do so, liposome suspension seems to have a large volume proportion in the final mixed aqueous phase. I'm concerned that my pepsin or pancreatin concentration and the pH will be changed if I follow the procedure and mixed my liposome suspensino with prepared SGF and SIF. But if I freeze dry my sample, it may change my particle size and morphology to some extent. I'm not sure if this protocol is accurate. Anyone have some suggestions on it?
Actually, the problems you are facing are very important to study the stability of liposome in vitro. Because in vitro experiment can just simulate the in vivo circumstances to very little extent. I think the protocol you mentioned should be fine. However, you can make different concentrations of pepsin or pancreatin to study if you worried about that. Besides, you are also able to use acids or bases to modify the final pH when you mixed liposome with SGF or SIF to avoid the change of pH. If you tend to use freeze dried liposome, I suggest you should screen the diluents to protect your liposome during freeze drying, such as mannitol, sucrose or lactose. Looking forward to continuing to discuss this with you.
Hi there, thank you so much for your kind information. Yes, you are definitely right. If I use the freeze drying, the use of cryoprotectants like trehalose is necessary to prevent liposome aggregation. But it may change the particle size and PdI after the rehydration process. Based on this aspect, I assume that mixing liposomal suspension with SGF or SIF may be a better choice. It seems that the protocols in reference papers varies from people to people which makes it hard to summarize. I guess may be I can mix relatively large volume of liposome suspension with SGF or SIF phase and adjust the final concentration for each salt and enzymes and pH to their original level. For this aspect, there is no reference supported so I'm not confident on this. What do you think? This problem will not occur in the digestion of many solid-cored nanoparticles since these particles don't have aqueous cores and easy to be concentrated based on the centrifugation.
Dear Lisha Zhao, if you want to reduce volume of liposome dispersion, you can use centrifugation, dialysis or size exclusion chromatography. Centrifugation is most commonly used and will be helpful to you if you can get dense mas of liposomes at low speed i.e. 10,000 rpm or less, as higher speed may damage liposomes. Dialysis can be performed by dialysis bag which is routinely used for in vitro drug release studies of nanoparticles.
Your idea of making SGF and SIF by adding salts and eznymes in liposome dispersion seems interesting but you should contact technical services expert of manufacturer. Alternatively, you can contact corresponding author of the papers you ar following.
Regards,
Mubashar
Hi there. Thank you so much for your information! I believe your answer make sense. I have tried to use 8000 rpm to concentrate my liposomes before placing into SGF or SIF so that the final concentration can be maintained. My previous idea of making SGF and SIF by adding salts and enzymes in liposome dispersion is not a standardized protocol in USP protocols and may not work well considering the osmotic pressure and the acidic pH of SGF medium. Thank you so much for your help!