I have prepared the drug loaded PLGA nanoparticle. I want to calculate the exact concentration, so that I can carry out further experiments. Please suggest how.
What kind of drug? Lipophile? Hydrophile? The main problem I guess is not the determination, but the isolation. You can try with dialysis with membranes in the "Dalton" scale or isolating the particles via ultracentrifugation, I normally use 20, 000 g for 1 hr and 15°C. For that purpose you need to know the sink conditions of you drug...
Depends how much drug you added and how much drug is retained. Depending on the electrical nature of your NP formulation and your drug you may be able to carry out a serial gel-shift assay that will tell you how much is retained. Multiply the retained percentage by your original amount of drug added. Best of luck.
Dear Rivera, I have isolated the particle but need to know the exact concentration, so that I can make some toxicity tests and other biochemical assays.
Dear Felix, I didn't get you. Could you please suggest me a paper related to your answer. That will be much helpful.
Thanks.
If your drug molecule has absorbion in the UV range, for example, you can estimate the drug uptake by measuring the absorbance at a certain wavelength of the solution collected after the loading experiments. Hence, the difference between the initial amount of drug used in the loading experiment and the amount of drug in the resulted supernatant solution will direct you towards the conecntration of the drug in the obtained hybrid material. Prior to this experiment you have to perform a calibration line, using drug solutions with well-known concentrations. In order to obtain reliable interpolation results, your calibration line must have high correlation coefficients.
first of all isolate your drug form you NP
Second there is two methods to quantify your drug concentration.
1-using Spectrophotometry, but as Br. Silviu Said you must do calibration curve for your drug and you must get high correlation coefficient 99.99.
2- Using HPLC method and I prefer this method because give u full detailed status of your Drug, for example ,if there is any degradation of drug or interaction with polymers but please validate your HPLC method before run it which means select column, select mobile phase, do calibration curve ....etc
Thanks Silviu & Ahmad for the valuable suggestions. I will try it.
Regards
It seemed to me that you wanted to determine the particle concentration not the amount of drug on particle. For this you may do this based on mass after drying or by particle counting methods. Concentration of drug loaded particles should be equal to initial particle concentration.
You should have a good account on your PLGA weigh ratio to your drug. I think you are asking to answer dosage amount of your PLGA containing your drug.
Depending on your drug property, you may use HPLC of your filtrate (as suggested by reader). Then back calculate how much in and out. I normally used vivaspin for a quick analysis or size exclusion column attached with light or uv detector.
The simple method by weighing the final product w.r.t polymer and drud wt in percentage.
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