The exact method will depend on the drug and nanoparticle formulation but here is a method I used for polymeric nanoparticles:
To measure drug release from nanoparticles, nanoparticles should be lyophilized, weighed, and resuspended in buffer (I used two different PBS/0.1% Tween-80 buffers, one at pH 7.4 and one at pH 6.5, but you can use what ever buffer is suitable for your drug/nanoparticle preparation) and then incubate in a 37 °C water bath. At various time points (I measured between 15 min and ten days) an aliquot of eluted drug medium should be removed for quantification; this volume must replaced with fresh buffer to prevent sink conditions. Drug release can be quantified by measuring the absorbance of the release media using a plate reader or HPLC, this value, of course, will be unique to the drug you are using. As a control you should also measure blank nanoparticles (with no drug) and make suitable standard curves for your drug concentration in buffer.
Please see the two attached papers of mine for methods and if you have any other questions please let me know.
Article Pharmacokinetics and biodistribution of lonidamine/paclitaxe...
Please see above.
Article Development of EGFR-Targeted Polymer Blend Nanocarriers for ...
If the drug is unstable, I would look for degradation products. As an example Temozolomide is not stable at pH 7 and above but you can monitor the the formation of MTIC or AIC as a degradation product and can be used to quantify the original drug.
I would follow an easy and direct approximation: make the release in a buffer/media where it will not degrade, for example acetate buffer. If your release is not very pH dependent (i.e. you are not using a pH-sensitive) you will get an idea of what is going on.
If your formulation is suspension or liquid, you can use Dialysis Bag Method and Amicon Ultra Tubes.
We used keep in NPs (nanoparticles) suspension in a suitable dialysis bag, and immerse this bag in a larger volume of receptor compartment (stirred buffer container). Take samples at scheduled intervals, which again depends on type NPs. Usually it is good to have some very early points to check if there is any burst release
see the published paper ;
Aldo Laganà, Iole Venditti, Ilaria Fratoddi, Anna Laura Capriotti, Giuseppe Caruso,
Chiara Battocchio, Giovanni Polzonetti, Filippo Acconcia, Maria Marino, Maria Vittoria Russo
“Nanostructured functional co-polymers bioconjugate integrin inhibitors”
J. Colloid Interface Sci. (2011), 361 ( 2) 465-471 DOI: 10.1016/j.jcis.2011.05.
You can take different time interval and measured absorption with particular range .
I would suggest Lara Milanie's method. You might have gone through the simple interval measuring method.
You have not mentioned if the drugs will be administered orally or by injection(IV,Im).It has great influence on the conditions of release study which will simulate the conditions in the body.
As I understand, you have encapsulated drug in nanoparticles which is not stable in pH 7. So it is a drug considered to deliver the active ingredient in intestinal tract (upper or lower part). So:
1-first of all you are going to study that the encapsulating agent is resistant in low pH in stomach? This will be carried on by putting the encapsulated nanoparticles in stomach simulated release medium. If the particles remain intact it is OK to protect active ingredient therein.
2- in second steps you will prepare release medium according intestinal condition (or colon which do you prefer).The release mediums preparation are described in detail in USP or other pharmacopoeia.
When you say you want to study the encapsulated drug release in vitro, will it be in solution without cells or will it be in the presence of cells? In the presence of cells, you will have to deal with the cell medium or DPBS. Without the cells, you will be using physiological buffers to mimic the physiological conditions where you want the drugs to be released. in either case, if your drug has optical properties such as fluorescence or absorbance, establish the properties of the solution before you add your drug. After that, measure the optical properties at different time points which may or may not (better if it does) correspond to the predicted half-life of the drug in vivo. The release maybe pH controlled or temperature controlled; that you have to establish before you study the timed release.
this may sound silly but i do not know how to plot a graph of accumulated concentration vs time, what is the formula to calculate the accumulated concentration? is it from the calibration curve equation and absorbance? please help me.
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