Application and Methodology of the Non-destructive ¹⁹F Time-domain NMR Technique to Measure the Content in Fluorine-containing Drug Products

The overall goal of this low field Fluorine 19 time domain nuclear magnetic resonance methodology, is to determine the average content of Florinated drugs in formulated drug products. This is a non destructive technique for drug development and manufacturing in the pharmaceutical industry. This method can help answer key questions within the pharmaceutics field, in the process of formulating drugs and drug products.

Suggest the average content of drugs in tablets and capsules. The main advantage of this technique is that it is non destructive and faster than conventional methods such as high performance liquid chromatography when measuring the average content of drugs in drug products. Weigh the samples for calibration in the appropriate NMR tubes until the marking line, or around 30 millimeters in height.

Also weigh the formulated drug product samples in the appropriate NMR tubes until the marking line or around 30 millimeters in height. Next, place the Teflon standard sample inside a 25 or 26 millimeter NMR tube. Place the NMR tube into the probe inside the magnet.

Allow the tube to equilibrate for five to 10 minutes, before tuning the instrument to f 19. Select auto o one, under the commands menu in the RINMR software. Repeat the measurement at least three times.

And take the last value of the o one parameter. Navigate to the commands menu in the RINMR software and select auto p 90, to calibrate the 90 degree pulse for f 19, with the standard Teflon sample, using the standard automated calibration sequence available in the instrument. For a better signal to noise ratio, place the API sample, prepared in the 25 millimeter NMR tube, inside the f 19 probe, installed in the magnet and let it equilibrate for at least 10 minutes.

Measure the t one relaxation time following the instructions of the instrument manual, for each API sample, using the standard experiment for inversion recovery, from the RINMR software. To measure the relaxation time, navigate to sequence and select load in the RINMR software. Then select the inverse recovery pulse sequence and click open.

Check that the basic parameters have the correct values by clicking the a tab. Under the tools menu, select scripts and a window will pop up. Then select the tab for t one and click the green arrow.

Another window will pop up. Select the delay list file with the appropriate delays. Or create the list after clicking open.

Once the delay list is satisfactory, click okay. A window will display a prompt for the creation of a file for all spectra that will be collected, in order to calculate the t one value. Click on save for the instrument to start acquiring the data.

And automatically save the files in the selected folder. A window will pop up, with the t one relaxation curve and the time constant or t one value. To build the calibration curve, first place one calibration sample in the magnet, for a particular API.

Equilibrate the samples for a minimum of five minutes for the smaller tubes and 10 minutes for the larger tubes. Acquire a free induction decay or solid experiment with the appropriate parameters, following the instructions in the instrument manual. In the sequence tab of the RINMR software, load the pulse sequence by selecting the sequence and clicking open.

Check that the basic parameters have the correct values. Measure all samples, prepared for each API, after they are equilibrated to the temperature of the magnet, for five to 10 minutes, to generate the calibration curves for every API sample. Save the data in a folder, giving a distinct name to each experimental run.

Calculate the weight percentage of the samples, based on the amount in each tube. Considering the purity of the API sample, in the largest tube. Place the already prepared formulated drug product samples in the magnet, to measure them one at a time.

Equilibrate the samples five minutes for small NMR tubes or 10 minutes for the samples in the larger NMR tubes. Acquire the same NMR experiment, with the same conditions, as for the calibrated samples for each API. Open the RI calibration software.

Recall all the solid data by clicking on the clip symbol to open the FID or solid files from a particular API and its'drug products samples as tablets. For all the samples measured, enter the weight percentages under the concentration column and the weights under the mass column. Select the appropriate region, using the average method to build the calibration curve.

Save the calibration curve. Next, select the region of the solid experiment from five to 300 points, to create the calibration curves. The results of the calibration, using the average method appear in table form.

As a second method of calibration, use the Fit method in the RI calibration software, to build the calibration curve with the same selected region. Save the calibration curve. The results of the calibration, using the Fit method appear in table form.

As a third method of calibration, open the Mnova software. Drag all the solid data from a particular API. And enter the weight percentages of the samples.

In the advanced menu, select time domain and then quantitation. A window will pop up. Click on the blue plus sign to select the integration area.

Then integrate the appropriate region using the magnitude mode, to build the calibration curve. Save the calibration curve. In the table, enter the percentages of drugs per sample under the concentration column only for the standards.

And the weights for all the samples under the mass column. The calibration samples should be in red. Make sure that the signal function is y equal to s over m, to normalize the signal to mass.

For reproducibility and repeatability, measure each sample three times, on different days and build a calibration curve in both software packages. Be sure to save the calibration curve files. Read the NMR data from the measured samples and enter their weights to determine the weight percentage of Flourinated API in the formulated tablets or capsules.

Use the calibration curves obtained from each of the three different calibration methods. Finally, calculate the amount of Flourinated drug per tablet or capsule as the average value, using the equation found in the text protocol. And compare with the values calculated by the software, based on the calibration curves.

Shown here are representative results of the calibration curve of Cinacalcet HCL, calculated as free base and triplicate measurements. Using RI calibration software in the average mode. Use of the RI calibration software in the Fit mode results in a similar calibration curve of Cinacalcet HCL in triplicate measurements.

The calibration curve of Cinacalcet HCL in triplicate measurements, using the Mnova software area method in the magnitude mode, yields comparable results. Shown here is a table of results for the calculated doses of Cinacalcet free base in commercial tablets. With their statistics by RI calibration software using the average and fit methods as well as Mnova software.

Once mastered, this technique can be done in 15 to 20 minutes. For example, the calibration samples can take one hour. And they only need to be done once if it is performed properly.

After watching this video, you should have a good understanding of how to quantify the average content of Flourinated drugs in formulated drug products, by the non destructive technique of low field Fluorine 19 TD NMR. Don't forget that working with drugs and NMR instruments can be extremely hazardous and precautions such as personal protective equipment, and no metal objects near the magnet, should always be taken while performing this procedure.