Materials.
Azithromycin Dihydrate was purchased for Sigma-Aldrich (Germany). Poly (meth) acrylates (Eudragit L100, S100) were obtained from Evonik Co., Ltd (Germany). All reagents used were AR grade and HPLC grade. Acetonitrile (HPLC grade-Fischer Scientific), Dipotassium hydrogen phosphate (Rankem), and Ortho Phosphoric acid (Rankem) were used for analysis. Other solvents were of analytical grade.
Methods.
Preparation of solid dispersion of AZI by solvent evaporation method.
Solid dispersion of Azithromycin Dihydrate was prepared by a solvent evaporation method. The drug and polymer were wholly dissolved in ethanol/dichloromethane (1:1, v/v). The organic solvent was evaporated at 60°C under stirring until complete evaporation and in a static oven for the next 24 hours. The prepared solid dispersion was crushed and passed through suitable sieves to harvest solid dispersion. 4 formulations were prepared with Poly (meth) acrylates (Eudragit L100, S100) (Table 1).
Table 1
Formulations for solid dispersions of AZI.
Trial
|
AZI (g)
|
Eu L100 (g)
|
Eu S100 (g)
|
DCM:Ethanol (1:1, ml)
|
F1
|
1
|
2
|
-
|
28
|
F2
|
1
|
4
|
-
|
28
|
F3
|
1
|
-
|
2
|
28
|
F4
|
1
|
-
|
4
|
28
|
Characterization of the solid dispersion:
1. Production yield, and drug loading.
The production yield was defined by Eq. (1). The drug loading was determined using the HPLC method and calculated using Equations 2.17
Chromatographic Conditions: 18
The HPLC instrument used was the Shimadzu LC-20AD system supplied with a photodiode array (PDA) detector.
The mobile phase was Acetonitrile, and phosphate buffer in the ratio of (35:65 v/v) and pH 6.5 adjusted with orthophosphoric acid. It was filtered through a 0.45 µ membrane filter. All determinations were conducted using C8, (250×4.6 mm, 5 µm), reverse phase column (GL Science) at ambient temperature (25°C). The column effluent was set at 200 nm. The injection volume was 20 µl with a flow rate of 1.5 ml/min.
Production yield =\(\frac{ \text{t}\text{o}\text{t}\text{a}\text{l} \text{m}\text{a}\text{s}\text{s} \text{o}\text{f} \text{s}\text{o}\text{l}\text{i}\text{d}\text{d}\text{i}\text{s}\text{p}\text{e}\text{r}\text{s}\text{i}\text{o}\text{n}}{\text{t}\text{o}\text{t}\text{a}\text{l} \text{m}\text{a}\text{s}\text{s} \text{o}\text{f} \text{r}\text{a}\text{w} \text{m}\text{a}\text{t}\text{e}\text{r}\text{i}\text{a}\text{l}\text{s}}\) × 100% (1)
Loading% = \(\frac{\text{d}\text{r}\text{u}\text{g} \text{r}\text{e}\text{t}\text{a}\text{i}\text{n}\text{e}\text{d} \text{i}\text{n}\text{t}\text{o} \text{s}\text{o}\text{l}\text{i}\text{d} \text{d}\text{i}\text{s}\text{p}\text{e}\text{r}\text{t}\text{i}\text{o}\text{n}}{\text{I}\text{n}\text{i}\text{t}\text{i}\text{a}\text{l} \text{d}\text{r}\text{u}\text{g} \text{c}\text{o}\text{n}\text{c}\text{e}\text{n}\text{t}\text{r}\text{a}\text{t}\text{i}\text{o}\text{n}}\) × 100% (2)
2. Comparison of the bitter taste threshold of AZI with the solid dispersion in vitro.
The taste-masking capacity of AZI solid dispersion was determined by in vitro drug release test because none of the polymers combined in taste-masked AZI solid dispersion had a bitter taste.
According to previous literature, the bitter taste threshold was between 25.3 and 30.4 µg/ml 19. Solid dispersion containing about 200 mg azithromycin was put into a test tube containing 10 ml distilled water. The mixture was directly shaken for 30 s and then filtered through membranes 0.45 µm. AZI concentration in the filtrate was analyzed by high-performance liquid chromatography (HPLC) and compared to the bitter taste threshold. If the drug concentration in the filtrate was lower than the bitter taste threshold, the solid dispersion was considered to have masked the bitter taste of AZI.
3. Drug release in different media.
For the in vitro dissolution test to mimic pH situations in vivo solid dispersion containing 200 mg AZI was tested using the paddle method with a rotation speed of 100 rpm.
A volume of 900 ml for hydrochloric acid solution (pH = 1.2 to study stability formulations in acid media) or phosphate buffer (pH = 6.80 corresponding to saliva pH) was used as the dissolution media at 37 ± 0.5°C. Aliquots of 5 mL of solution samples were withdrawn for 5 min for phosphate buffer pH 6.8 and 2 hours for hydrochloric acid, and an equal volume of the fresh medium with the same temperature was added immediately. The aliquot samples were immediately filtered through a 0.22 µm membrane and quantified on a HPLC-UV system20, 21.
4. Fourier transform infrared (FT‑IR) spectroscopy.
FTIR spectroscopy was conducted using FT/IR (Shimadzu IR, Germany). Azithromycin, Eudragit L100, physical mixtures, and formulation F2 were prepared by compressing each sample with pure Potassium bromide (KBr).
5. Thermal analysis.
Azithromycin, Eudragit L100, physical mixtures, and formulation F2 were tested using DSC 131, SETARAM, France. Samples were prepared in open aluminum pans (2–5 mg). The samples were heated at 10 C/min under a nitrogen atmosphere in a temperature range between 0 and 250°C.
6. Powder X‑ray diffraction (PXRD) analysis.
Azithromycin, Eudragit L100, physical mixtures, physical mixtures, and formulation F2 were tested using STOE-Stadi P diffractometer with a Cu Kα radiation source tube and 1.54 Å X-ray wavelengths. Emission filament voltage and amperage were 40 kV and 30 mA respectively. The scanning range of 5–59.98° 2θ with a step size of 0.02° and reflection mode was used.
7. Scanning electron microscopy (SEM).
The surface morphology of Formula F2 was analyzed with scanning electron microscopy (SEM). Solid dispersion was mounted on double-faced adhesive tape, coated with a thin gold-palladium layer by a sputter-coated unit, and analyzed with a scanning electron microscope (MIRA3 LMU).