The overall goal of this work is to show the synthesis of cyclodextrin nanosponges, convert them into drug-loaded hydrogels, and to study the drug diffusivity inside the hydrogel by high-resolution magic angle spinning, nuclear magnetic resonance spectroscopy. Cyclodextrin nanosponge are prepared by step-growth polymerization between cyclodextrins, or CDs, and the EDTA DNA dried. If we don't mention, our network of CD units is formic, capable to absorb and release organic and inorganic species.
The main advantage of high-resolution magic angle spinning NMR is to monitor the transfer property of the encapsulated drug within a polymeric host with potential interest for drug delivery. The synthetic approach is very simple, and it is performed at room temperature. The step-growth polymerization reaction takes place in about 10 minutes.
We have chosen ibuprofen as an example of a drug trapped in polymeric hydrogen metrics, representative of a really six car fold down delivery. Dry beta cyclodextrin in an oven at 100 degrees Celsius up to a constant weight. Also, dry 250 milliliters of DMSO, and 100 milliliters of triethylamine over four Angstrom molecular sieves for 24 hours before using them in the protocol.
Introduce 25 milliliters of DMSO into a 50-milliliter, one-neck round bottom flask. Under magnetic stirring, add 5.675 grams of beta-cyclodextrin. Add the beta-cyclodextrin powder in small portions to DMSO to reduce the formation of lumps.
After about 30 minutes, add six milliliters of triethylamine to the homogenous solution using a 10 milliliter graduated pipette. Plunge the flask into a water bath maintained at room temperature, and keep the mixture under stirring for 15 minutes. For preparation of cyclodextrin nanosponge one-four, add 5.124 grams of EDTA dianhydride under intense stirring.
And for preparation of cyclodextrin nanosponge one-eight, add 10.248 grams of EDTA dianhydride under intense stirring. After three hours, remove the solid material, which is the cyclodextrin nanosponge, from the flask using a spatula and crush it grossly with a mortar and pestle. Wash the solid material in an acetone bath, then wash and filter the solid material on filter paper.
After drying all the solid material in the air at room temperature for 48 hours, crush the dried material finely using a mortar and pestle. Then, keep it under vacuum for two hours at 45 degrees Celsius. For high-resolution magic angle spinning NMR sample preparation, first prepare a solution of 0.27 molar ibuprofen sodium salt in deuterated water.
Add 20 milligrams of cyclodextrin nanosponge one-four, and two milligrams of anhydrous sodium carbonate to 150 microliters of the solution in a two milliliter glass vial. Mix the contents of the vial with a small spatula to homogenize it. Repeat this process for the cyclodextrin nanosponge one-eight polymer.
Obtaining a homogenous, perfectly swollen hydrogel is the first key step for a good HR mass NMR spectrum. The hydrogel should be placed in the NMR rotor very carefully, avoiding bubbles which may cause incorrect rotor spinning. Insert the gel into a five millimeter NMR rotor suitable for HR mass experiments using a small spatula.
Then, insert the rotor into the magnet of the NMR spectrometer. Once in the magnet, set the rotor in rotation by a pneumatic unit up to 4, 000 hertz. To perform HR mass proton NMR experiments, set up the instrumental parameters with a rotor spinning speed of four kilohertz at the mass pneumatic control unit, and a sample temperature at 305 kelvin in the variable temperature unit.
Run a one-dimensional spectrum, quire the diffusion HR mass spectra of ibuprofen in cyclodextrin nanosponge one-Four and cyclodextrin nanosponge one-eight hydrogels by using the pulsed gradient spin-echo pulse sequence on the proton resonance. The diffusion experiments are performed by using the BP led pulse sequence. This is a pseudo two-dimensional experiment with a gradient ramp increasing linearly from 2%to 100%gradient strength in the indirect dimension.
The signal intensity is attenuated depending on the diffusion time and gradient pulse P30. The optimization of these parameters is required before properly running an experiment. The optimization is done by running two one-dimensional measurements, in which the diffusion time is kept constant while P30 is varied.
Variable diffusion time measurements are carried out by incrementing the diffusion time in the range of 50 to 200 milliseconds. For each experiment, an array of 32 spectra is collected. Then, process the data as described in the text protocol.
The basic pulsed field gradient spin-echo experiment allows for measurement of the molecular mean-squared displacement of ibuprofen in the hydrogels. The NMR signal intensity decays as the gradient strength increases. The decay fitting provides the molecular mean-squared displacement, indicated as Z-squared.
In turn, the mean-squared displacement is related to the type of diffusion regime. The alpha exponent is obtained experimentally by a log-log linear regression of the mean-squared displacement as a function of the diffusion time. The value of the alpha exponent is a descriptor of the diffusive regime observed for ibuprofen in the hydrogel matrix.
Isotropic unrestricted diffusion exists when alpha equals one. A subdiffusive regime is indicated for an alpha value of less than one. And a superdiffusive regime is represented by an alpha value of greater than one.
Ibuprofen entrapped in the hydrogel matrix shows two different diffusion regimes according to the polymer preparation. Hydrogels of nanosponges one-four are represented by a subdiffusive regime. Conversely, hydrogels of nanosponges one-eight are characterized by a slightly superdiffusive regime.
Once mastered, this reaction can be done in high-yield and purity in one hour if it is performed properly. The materials show interesting capability of water uptake by forming viscous hydrogels, in which, biologically active molecules can be easily incorporated. The molecular state in the transfer properties of a biologically active molecule encapsulated in such polymeric matrixes can be studied experimentally by using HMS NMR techniques.
The NMR results on the diffusion of ibuprofen in the hydrogels prepared from cyclodextrine nanosponges indicate that the diffusion can be modulated by a suitable preparation of the polymers. Ibuprofen shows both subdiffusive and superdiffusive regime in cyclodextrin nanosponges hydrogels. This finding can be exploited for original design of the release properties of cyclodextrin nanosponges hydrogen scaffold.
