Hi, I am using microfluidic device to generate PLGA nanoparticle. Briefly PLGA was dissolved in water miscible organic solvent (ACN), 1% PVA in water was used for anti-solvent. For hydrophobic drug, I've dissolved the drug in PLGA solution, and for hydrophilic drug, I've dissolved the drug in 1% PVA. After running microfluidics, I've centrifuged to remove solvent and washing procedure. Lyophilized to calculate drug loading, but it showed very poor loading. 20% drug was mixed in solution, but the result showed only 1% loading (w/w). Wondering what I did wrong. Should the solvent be completely evaporated before centrifugation?
You can use FTIR to confirm loading (measure the drug by itself, the drug-loaded NP, and empty-NP that has been treated the exact same way as the drug-loaded one), but to quantify the loading efficiency, you will have to use another method. If your drug has a strong absorbance peak, then make a standard curve of OD vs. concentration, and use that to measure the OD of the drug loaded/washed out of the NP to extrapolate the drug concentration. You can use a UV-Vis or the NanoDrop for that. If there is a peak shift and change in peak shape after loading, then you will have to use the wash-outs, rather than what is loaded. You can also use HPLC if it is available.
Take the particles, dissolved them in a naoh solution to free the drug and quantify it.
I would expect the hydrophilic drug in that setup to show fairly low loading efficiency - I'd imagine not all that much PVA is actually winding up in the final product at all. Your hydrophobic drug should (hopefully) have a higher loading efficiency. If possible, I think it's best to get all of the solvent off before centrifugation. Can you use a rotovap? Will it evaporate if you just leave it stirring for a while?
I agree that you can use FTIR to confirm loading. I've also used HPLC - run the drug alone first (I've usually done this with hydrophobic drugs that can just be dissolved in chloroform), then take your washed particles, dissolve them in chloroform, and run the whole solution to confirm that you get a peak where the drug should be. But using absorbance would be, by far, the easiest method if your drug has a strong absorbance peak.
Dear Nae Gyune,
It is so necessary to qualify and quantify drug loading in to nanocarrier for determine the loaded drug and its amount in nano-carrier. I suggest that use FT-IR and UV-Vis spectroscopy to qualify evaluation of loaded drug, initially. Then, you can utilize NMR, HPLC and eventually LC-MS characteristic analytical devices in order to quantify evaluations.
Best regards,
Drug loading confirms with DSC or STA ( thermal analysis), DLS ( Particle size and zeta potential) and HPLC.