I am preparing a chitosan system containing two drugs, and one of them is 5FU. However, the results of the LC% and EE% are very low.

In short, after freeze drying I prepared a solution of 1 mg/ml (3 mg of the NPs in 3 ml of the used mobile phase, filtered by a 0.22 syringe filter, and 20 ppm of the solution was injected triplicate in UPLC/UV at 266 nm. The weight is calculated automatically from the software by a calibration curve of (20, 40, 60, 80, 100 ppm). Now, I am struggling in the LC% and EE% calculation, because the obtained weights are very low, for example; from 0.56 to 3.33 ug, and reflected badly on the LC% and EE% values using the below equations:

LC%= mass of the encapsulated drug /mass of the NPs*100.

EE%= mass of the encapsulated drug/ mass of drug(initial amount).