1. Introduction

2. Materials and Methods

2.2. Methods

2.2.1. Thermal Analysis

Differential Scanning Calorimetry (DSC)

Pure substances were analyzed via DSC (Mettler DSC 820, Mettler-Toledo GmbH, Germany) regarding their melting point, glass transition temperature, and possible recrystallization. Printed tablets were measured after 2-4 weeks of storage. Approximately 10-15 mg samples were accurately weighed into sealed aluminum pans with punctured lids. Measurements used a heat-cool-heat cycle to determine the melting point in the first heating and glass transition temperature during the second heating. Information about the miscibility of loperamide and PVA was expected to be found during DSC trials. One of the approaches for estimation of glass transition temperature is described by the Gordon-Taylor equation, which can be applied to miscible blends.

$$T_{g,mix}\approx \frac{\left[{\omega }_1\ast T_{g,1}+K\ast {\omega }_2\ast T_{g,2}\right]}{{\omega }_1+K\ast {\omega }_2}$$

Equation 1

with T_(g,mix) and T_(g,i) representing the glass transition temperature of the mixture and the components, ω_i is the mass fraction component I and K is an adjustable fitting parameter.

It is expected for two miscible substances to show one glass transition temperature instead of two individual glass transition temperatures. The detected temperature should be partially composed of the individual glass transition temperatures [18].

Simultaneous Thermal Analyzer (STA)

Thermal degradation as well as loss of water taken up from air humidity during printing and storage was tested using a Netzsch STA 409 PG/1/G Luxx (Erich NETZSCH GmbH & Co. Holding KG, Germany). All samples were treated the same way, measuring the mass of a sample using Al₂O₃ as a reference during heating of the samples up to 250°C. Pure substances and printed tablets were measured at a mass of approximately 22 mg.

2.2.2. XRD-Measurements

Wide angle X-ray (powder) diffraction patterns were obtained in an angular range of 10-50° 2θ with a stepwise size of 0.02° on a Bruker D8 Advance diffractometer (Bruker Corp., USA) using monochromatic CuKα radiation (λ = 0.15406 nm). Pure substances were measured in powdered form, while printed tablets were measured intact.

2.2.3. Meltrheology

To measure the viscosity of the molten mixtures and thus link the rheological properties to the printing process, the shear rate occurring during printing was determined. Two different methods related to different extrusion processes were compared.

The first method is usually used in FDM 3D printers that run on filament. Here, only the shear rate in the nozzle is considered [19].

Volume flow (Q) was determined empirically by accurately weighing printed tablets with 100% infill and measuring their volume (V) using a gas displacement pycnometer (Accupyc 1330, Micromeritics Instrument Corporation, USA). Temperature was set to 25°C while the remaining volume was flushed with Helium. During printing, the time (t) for each individual tablet was recorded. The following equation was employed afterwards:

$$Q=\frac{V\left[m{m}^3\right]}{t\left[s\right]}$$

Equation 2

The apparent shear rate at the nozzle wall (${\dot{\gamma }}_{wa}$) can be calculated as:

$${\dot{\gamma }}_{wa}=\frac{4Q}{\pi r_n^3}$$

Equation 3

with a radius (r) of the equipped nozzle of 0.04 cm.

The second method is applied to extrusion processes and used to calculate shear rates inside single-screw extruders. Approximation of the shear rate γ ̇ in the screw channel can be achieved from the Couette shear rate:

$$\dot{\gamma }=\frac{v_b}{H}=\frac{\pi DN}{H}$$

Equation 4

with v (velocity), H (channel depth = 0.55 mm), D (diameter = 8.0 mm) and N (screw speed) [20].

Melt rheology was conducted using compact rheometer (Anton Paar Physica MCR 501, Anton Paar GmbH, Germany) equipped with single-use stainless steel plates with a radius of 10 mm in a plate-plate configuration. Sample specimens used for the melt rheology were produced using the MeltPrep device (MeltPrep GmbH, Austria) with a 20 mm disc geometry at temperatures 10°C above printing temperature yielding ASDs with consistent geometric shape [21]. Since the viscosity of non-newtonian fluids is a product of temperature and shear rate [22], samples were measured at a range of temperatures after determination of the linear viscoelastic region (LVR). Determination of LVR was conducted for every measured temperature individually by performing amplitude sweeps from 0.01% to 100% deformation (0.0103 to 103 mrad) at 10 rad/s. Frequency sweeps from 100 Hz to 0.1 Hz were performed afterwards at 1% amplitude.

2.2.4. HME of PVA/Sorbitol Mixtures

Apart from PVA/loperamide mixtures, also plasticized mixtures were used. Sorbitol was used as the plasticizer. Direct powder extrusion of PVA/sorbitol mixtures was not possible due to the different melting points. To be able to still print this mixture a single screw extruder (Noztek Pro, Noztek, England) was used to extrude these two excipients before adding the API and printing the mixture. Afterward, it was milled using an ethanol–cooled mill and sieved while the fraction < 0.400 mm was used in further printing steps.

2.2.5. Preparation of Powder Mixtures

All powder mixtures for printing were produced by sieving (0.400 mm mesh size) and accurately weighing the compounds needed and subsequent mixing in a tumbler mixer (Turbula Type T2C, Willy A. Bachofen AG, Switzerland) for 20 min.

Table 1. Physical mixtures for 3D printing.

Table 1. Physical mixtures for 3D printing.

Batch	Parteck MXP	Parteck MXP/Sorbitol 15% Extrudate	Loperamide	Aerosil
PAR-LOP5%-AER1%	94.0%		5.0%	1.0%
PAR_LOP10%_AER0.5%	89.0%		10.0%	1.0%
PAR-SOR15%E_LOP5%_AER1.5%		93.5%	5.0%	1.5%

2.2.6. 3D Printing Using Direct Powder Extrusion Tool of M3dimaker

A computer-aided design (CAD) tablet-shaped geometry was created using Autodesk^® Fusion 360 (Autodesk GmbH, München). The .stl file was sliced by Repetier-Host (Hot-World GmbH & Co. KG, Germany). Objects were printed with 2 outer perimeters, 3 bottom and top layers using varying infills to change the dosage of printed tablets on the M3dimaker 3D printer (FabRx Ltd., London) equipped with the direct powder extrusion (DPE) tool. Print speed and temperature were adjusted according to the properties of the mixtures in molten state.

Figure 1. stl file of tablet shaped geometry (d = 5 mm; h = 5mm).

Figure 1. stl file of tablet shaped geometry (d = 5 mm; h = 5mm).

Figure 2. stl file of improved geometry (d = 5 mm; h = 5mm; edges rounded with r = 2mm).

Figure 2. stl file of improved geometry (d = 5 mm; h = 5mm; edges rounded with r = 2mm).

2.2.7. Confocal Raman Microspectroscopy

Witec’s alpha 300R system (WITec GmbH, Ulm, Germany) was employed for the measurements. Pure substances were measured with 30 mW power of the laser (λ = 532 nm) and the evaluated spectra were used for the true component analysis to show the distribution of loperamide in the tablet.

Printed tablets were measured on the outer surface to find information on potential demixing processes, varying concentrations, and recrystallization on the outer surface.

One printed tablet of each mixture was measured using 30 mW laser (λ = 532 nm). The surface of the measurement area was evaluated and corrected by the TrueSurface Mk II module. An area of 20 x 2000 µm with 4 x 400 pixels was measured using the autofocus function.

For the evaluation of drug content, the true component analysis was employed using the previously obtained spectra of individual substances. The software calculates based on individual spectra the amount of drug found in each pixel measured and reports a colored picture of the distribution and intensity of the signal found.

2.2.8. Drug Content Analysis by HPLC

Evaluation of the drug content was performed on the powdered mixtures as well as on the printed tablets. From powdered samples, an equivalent of 5 mg loperamide was accurately weighed and dissolved in 100 ml of the mobile phase used for the subsequent HPLC analysis (purified water/acetonitrile/0.5% ammonium acetate solution 29/36/35 (v/v)). Tablets were also dissolved in the mobile phase using stirring and an ultrasonic bath. Drug contents were determined by HPLC-UV/VIS (System 20A, Shimadzu Ltd., Japan) at a wavelength of 254 nm. The method showed a linearity of R2=0.99680 between 0.0023 mg/ml and 0.07 mg/ml with a tested LOQ of 0.00166 mg/ml and an estimated LOD of 0.0005488 mg/ml calculated according to ICH guidelines [23]. All experiments were performed in triplicate.

2.2.9. In Vitro Dissolution

Dissolution testing was adapted from standard procedures of the FDA [24] used for loperamide capsules using a Pharma Test PT-DT7 (Pharma Test Apparatebau AG, Germany) manual dissolution tester. 0.1M HCl was used as dissolution medium (500 or 900 ml, depending on the tablet strength) at 37°C with 100 RPM paddle speed. Sampling of 1 ml took place every 30 min for 5.5 hours and a last sample was drawn after 24 hours. Samples were measured using UV-HPLC as described in 2.2.8. For each formulation and infill setting printed, 4 tablets were measured and the mean value including the standard deviation is shown. For each formulation, a linear regression of the correlation between mass and infill is calculated and the best-fit values are given. Slopes are compared using two-tailed testing with the null hypothesis that the slopes are identical.

3. Results and Discussion

3.5. Confocal Raman Microspectroscopy

Visualization of API content, distribution, and potential recrystallization can be achieved via confocal Raman microspectroscopy (CRM). The overlay of the single spectra of loperamide, sorbitol, and PVA showed peaks that can be used for the evaluation of contained API in the printed tablets. The peak in the high wavenumber range (3040 – 3090 1/cm) as well as the peak in the fingerprint region (1575 – 1615 1/cm) have no significant overlapping with peaks from other substances and can be used to identify loperamide. The low drug load of 5 and 10%, respectively lead to a decreased signal intensity of these peaks in the resulting spectra of printed tablets relative to the single spectra obtained from pure substances.

Figure 15. Obtained single spectra (0.5 s; 10 Acc.) red = Parteck MXP, blue = loperamide, green = sorbitol.

Figure 15. Obtained single spectra (0.5 s; 10 Acc.) red = Parteck MXP, blue = loperamide, green = sorbitol.

Figure 16 shows an example of the measured area on the side of the printed tablet. The true surface module was able to cover most of the differences in the height of the surface area. Fine-tuning to achieve good spectra was done via auto-focus.

Obtained signals were color-coded (Figure 18) and exemplary spectra were given for the individual colors used (Figure 17). Red shows signal where loperamide was found, and spectrum intensity was sufficient. With the loss in signal intensity (as seen in the lighter and blue-colored regions) the signal-to-noise ratio of the loperamide peaks gets worse, although loperamide concentration remains constant.

Raman imaging was able to show consistent spectra containing polymer and API over a range of points distributed over roughly 40% of the printing time. No “hotspots” or crystalline API were found during these measurements, which supports findings from the XRPD measurements of a uniform ASD.

4. Conclusion

References

1. West, T.G.; Bradbury, T.J. 3D Printing: A Case of ZipDose^® Technology - World’s First 3D Printing Platform to Obtain FDA Approval for a Pharmaceutical Product. In 3D and 4D Printing in Biomedical Applications; Maniruzzaman, M., Ed.; Wiley-VCH Verlag GmbH & Co. KGaA: Weinheim, Germany, 2019; pp. 53–79. ISBN 9783527813704. [Google Scholar]
2. Li, Q.; Guan, X.; Cui, M.; Zhu, Z.; Chen, K.; Wen, H.; Jia, D.; Hou, J.; Xu, W.; Yang, X.; et al. Preparation and investigation of novel gastro-floating tablets with 3D extrusion-based printing. International Journal of Pharmaceutics 2018, 535, 325–332. [Google Scholar] [CrossRef] [PubMed]
3. Khaled, S.A.; Burley, J.C.; Alexander, M.R.; Yang, J.; Roberts, C.J. 3D printing of tablets containing multiple drugs with defined release profiles. International Journal of Pharmaceutics 2015, 494, 643–650. [Google Scholar] [CrossRef] [PubMed]
4. Zheng, Y.; Deng, F.; Wang, B.; Wu, Y.; Luo, Q.; Zuo, X.; Liu, X.; Cao, L.; Li, M.; Lu, H.; et al. Melt extrusion deposition (MED™) 3D printing technology - A paradigm shift in design and development of modified release drug products. International Journal of Pharmaceutics 2021, 602, 120639. [Google Scholar] [CrossRef] [PubMed]
5. Kempin, W. Entwicklung und Charakterisierung 3D-gedruckter, wirkstoffhaltiger Darreichungsformen am Beispiel von Tabletten und Implantaten; Universität Greifswald, 2018.
6. Goyanes, A.; Fina, F.; Martorana, A.; Sedough, D.; Gaisford, S.; Basit, A.W. Development of modified release 3D printed tablets (printlets) with pharmaceutical excipients using additive manufacturing. International Journal of Pharmaceutics 2017, 527, 21–30. [Google Scholar] [CrossRef] [PubMed]
7. Goyanes, A.; Scarpa, M.; Kamlow, M.; Gaisford, S.; Basit, A.W.; Orlu, M. Patient acceptability of 3D printed medicines. International Journal of Pharmaceutics 2017, 530, 71–78. [Google Scholar] [CrossRef] [PubMed]
8. Goyanes, A.; Wang, J.; Buanz, A.; Martínez-Pacheco, R.; Telford, R.; Gaisford, S.; Basit, A.W. 3D Printing of Medicines: Engineering Novel Oral Devices with Unique Design and Drug Release Characteristics. Molecular pharmaceutics 2015, 12, 4077–4084. [Google Scholar] [CrossRef] [PubMed]
9. Fu, J.; Yin, H.; Yu, X.; Xie, C.; Jiang, H.; Jin, Y.; Sheng, F. Combination of 3D printing technologies and compressed tablets for preparation of riboflavin floating tablet-in-device (TiD) systems. International Journal of Pharmaceutics 2018, 549, 370–379. [Google Scholar] [CrossRef] [PubMed]
10. Chai, X.; Chai, H.; Wang, X.; Yang, J.; Li, J.; Zhao, Y.; Cai, W.; Tao, T.; Xiang, X. Fused Deposition Modeling (FDM) 3D Printed Tablets for Intragastric Floating Delivery of Domperidone. Sci. Rep. 2017, 7, 2829. [Google Scholar] [CrossRef] [PubMed]
11. Prasad, L.K.; Smyth, H. 3D Printing technologies for drug delivery: a review. Drug Dev. Ind. Pharm. 2016, 42, 1019–1031. [Google Scholar] [CrossRef] [PubMed]
12. Goyanes, A.; Buanz, A.B.M.; Basit, A.W.; Gaisford, S. Fused-filament 3D printing (3DP) for fabrication of tablets. International Journal of Pharmaceutics 2014, 476, 88–92. [Google Scholar] [CrossRef] [PubMed]
13. Zhang, J.; Feng, X.; Patil, H.; Tiwari, R.V.; Repka, M.A. Coupling 3D printing with hot-melt extrusion to produce controlled-release tablets. International Journal of Pharmaceutics 2017, 519, 186–197. [Google Scholar] [CrossRef] [PubMed]
14. Goyanes, A.; Allahham, N.; Trenfield, S.J.; Stoyanov, E.; Gaisford, S.; Basit, A.W. Direct powder extrusion 3D printing: Fabrication of drug products using a novel single-step process. International Journal of Pharmaceutics 2019, 567, 118471. [Google Scholar] [CrossRef] [PubMed]
15. Chavda, H.V.; Patel, C.N.; Anand, I.S. Biopharmaceutics classification system. Syst Rev Pharm 2010, 1, 62. [Google Scholar] [CrossRef]
16. Wei, C.; Solanki, N.G.; Vasoya, J.M.; Shah, A.V.; Serajuddin, A.T.M. Development of 3D Printed Tablets by Fused Deposition Modeling Using Polyvinyl Alcohol as Polymeric Matrix for Rapid Drug Release. J. Pharm. Sci. 2020, 109, 1558–1572. [Google Scholar] [CrossRef] [PubMed]
17. Zaki, N.M.; Artursson, P.; Bergström, C.A.S. A modified physiological BCS for prediction of intestinal absorption in drug discovery. Molecular pharmaceutics 2010, 7, 1478–1487. [Google Scholar] [CrossRef] [PubMed]
18. Bochmann, E.S.; Neumann, D.; Gryczke, A.; Wagner, K.G. Micro-scale prediction method for API-solubility in polymeric matrices and process model for forming amorphous solid dispersion by hot-melt extrusion. Eur. J. Pharm. Biopharm. 2016, 107, 40–48. [Google Scholar] [CrossRef] [PubMed]
19. Boetker, J.; Water, J.J.; Aho, J.; Arnfast, L.; Bohr, A.; Rantanen, J. Modifying release characteristics from 3D printed drug-eluting products. European journal of pharmaceutical sciences: official journal of the European Federation for Pharmaceutical Sciences 2016, 90, 47–52. [Google Scholar] [CrossRef] [PubMed]
20. Rauwendaal, C. Polymer extrusion, 5th edition; Hanser Publications; Hanser Publication: Munich, Cincinnati, 2014; ISBN 9781569905166. [Google Scholar]
21. Treffer, D.; Troiss, A.; Khinast, J. A novel tool to standardize rheology testing of molten polymers for pharmaceutical applications. International Journal of Pharmaceutics 2015, 495, 474–481. [Google Scholar] [CrossRef] [PubMed]
22. Jaluria, Y. Heat and Mass Transfer in the Extrusion of Non-Newtonian Materials. In Transport Phenomena in Materials Processing; Elsevier: Amsterdam, The Netherlands, 1996; pp. 145–230. ISBN 9780120200283. [Google Scholar]
23. CHMP/ICH. Q 2 (R1) Validation of Analytical Procedures: Text and Methodology.
24. FDA/CDER/”Yeaton, A. Guidance for Industry. In Managing the Documentation Maze; Gough, J., Nettleton, D., Eds.; John Wiley & Sons, Inc: Hoboken, NJ, USA, 2010; pp. 431–450. ISBN 9780470597507. [Google Scholar]
25. LaFountaine, J.S.; McGinity, J.W.; Williams, R.O. Challenges and Strategies in Thermal Processing of Amorphous Solid Dispersions: A Review. AAPS PharmSciTech 2016, 17, 43–55. [Google Scholar] [CrossRef] [PubMed]
26. Goyanes, A.; Buanz, A.B.M.; Hatton, G.B.; Gaisford, S.; Basit, A.W. 3D printing of modified-release aminosalicylate (4-ASA and 5-ASA) tablets. Eur. J. Pharm. Biopharm. 2015, 89, 157–162. [Google Scholar] [CrossRef] [PubMed]