This work presented and used for the first time a new printer, 3DForMe®, which is based on the ability to extrude powders directly, thus overcoming the filament preparation problem associated with FDM. Different blends consisting of NCS and various concentrations of HPMC, HP-β-CD and PEG 6000 were first characterized and then extruded as such through 3DForMe®, obtaining sustained release tablets. A complete solid-state characterization of the different blends and tablets achieved has been described, together with in vitro dissolution studies of the printed tablets.
NCS Phase solubility studies
Phase solubility studies of the drug in the presence of HP-β-CD were performed, allowing the evaluation of the stability constant and to foresee the stoichiometric ratio of the NCS- HP-β-CD complex. Pure NCS has an intrinsic solubility of 8 µg/mL in water. The phase solubility plot displays a significant and linear increase in NCS solubility with increasing HP-β-CD concentration (Fig. 2). The drug shows a solubility of 54.46 µg/mL in the presence of HP-β-CD at a concentration of 0.26 M. The regression analysis (R2 = 0.9846) describes an AL-type curve, according to the Higuchi-Connors classification, with a slope of less than 1, which could be associated with a 1:1 molar ratio between NCS and HP-β-CD. By Higuchi Connors equation was calculated the constant stability equal to 876.1 M−1 indicating a good interaction between NCS and HP-β-CD. Indeed, in literature stability constant values between 50–2000 M−1 are regarded favorable .
The addition of HPMC at HP-β-CD solution significantly improved the aqueous solubility of NCS, confirming that the use of hydrophilic polymers such as HPMC increased the CD complexation efficiency for drugs . In all samples containing the ternary system (HPMC/HP-β-CD/NCS), a tenfold increase in aqueous solubility of NCS was observed in comparison to solution containing the HP-β-CD (Fig. 3). Based on these results, it could be stated that HPMC and HP-β-CD showed a synergistic effect in enhancing NCS solubility.
Visual inspection and homogeneity of HME filaments
The filaments share some common characteristics, even though they are derived from different powder blends. In fact, all four filaments show a common color variation during extrusion: the filament at the beginning of the extrusion has a very dark color that tends to lighten at the end of the process (Fig. 4). These variations in terms of color are most probably due to the different time the powder mixtures were kept in the extrusion head. The four filaments had a smooth surface and were not brittle at handling.
The yield of the extrusion process was very high, considering that almost all the powder in the hopper was completely extruded and that the weight of the entire filament coincided with the weight of the blend from which it was obtained.
Each piece of filaments, cut in equal length, had a similar weight and although the change in color of the extruded filament might suggest degradation or a change in the stability of the drug, which often occurred during the HME process, the NCS present within the entire filament obtained with the DPE technique retained its stability (see Table 2). Indeed, respecting the correct extrusion order, the single pieces were analyzed to assess the concentration of NCS present in them. The study of filament homogeneity shows that in the four filaments there are fragments in which the drug is under-extruded, which are compensated by fragments in which the drug is over-extruded (Fig. 5). For all the filaments, it can be seen that the values of the percentage of drug are in a range between 6.34 and 13.57 (% w/w). These results are in agreement with the literature , as Vidik et al. justified the presence of under-extruded and over-extruded fragments as a consequence of demixing of the raw materials during the printing process. However, unlike the data reported by Vidik et al. in which the average drug concentrations in the different filament fragments were lower than the theoretical value, in the present study the average of the values obtained is very close to the value of the theoretical concentration. The results demonstrated that the adopted process was not detrimental for the drug (Table 2) despite the contact with the high temperatures of the extruder.
Direct powder extrusion 3D-printing
The single screw extruder printer model was used for the first time to enable DPE. The blend powder realized was inserted in the hopper, heated to a specific temperature, and used for direct tablet printing. This innovative printing technique overcomes the limitations associated not only with conventional tablet printing systems , but also with established printing processes such as HME. The elimination of the preliminary HME step followed by FDM printing makes the DPE printing process much simpler and faster. In fact, the overall process of printing one tablet takes 13 min. In addition, with no intermediate steps, the amount of raw material waste is also greatly reduced. The design of the extruder with a vertical orientation and its adequate distance from the hopper facilitate the flow of powder into the screw. A functional amount of powder can be placed inside the hopper to ensure a production from 1 to 15 tablets for cycle. This makes the 3D printer suitable for formulating personalized medicine often required with galenic preparations.
All powder blends were found to be suitable for DPE 3DP. Only the extrusion of powder blend 3 led to the formation of final tablets considered unsatisfying. Different temperatures and printing parameters (screw speed) were tested in order to improve the characteristics, but none of the modifications improved the printing quality. A possible cause may be attributable to the presence of cyclodextrin, which tended to moisten the powder as the residence time in the printer increased, creating vacuum zones along the screw, and leading to extruder blockage. This problem did not occur during the filament extrusion phase due to the reduced time the powder spent in contact with the extruder. This demonstrates that the DPE requires that the powders must have a certain degree of fluidity and homogeneity to ensure that the flow through the extruder is uniform . Blend 3 was then modified by adding 2% silica and 1% talc, as dehydrating and glidant agents, respectively, which allowed printing without any problems It should be noted that Blend 4 also consisted of a percentage of HP-β-CD; however, the presence of the plasticizer and glidant agent (PEG 6000) helped the printed process, favoring the powder to flow within the screw. The resulting NCS printed tablets showed a cylindrical shape and good adhesion between the printed layers (Fig. 6).
Characterization of tablets
The tablets showed good uniformity in physical dimensions and high efficiency in reproducing the CAD drawing, which was set at 12 mm in diameter and 3.7 mm in height. The printed tablets had an average diameter ranging from 11.63 mm to 12.15 mm, and the average height ranged from 3.49 mm to 3.74 mm. This accuracy in the printing of the tablets was confirmed by data reported in the literature obtained with the same printing method [41, 42]. The average mass was described as ranging from 414.00 to 506.80 mg, this variation could be due to the different composition of the individual blends, which gives the powders different flow properties in the extruder. Despite these differences, all formulations satisfied the weight uniformity assay performed, showing weight deviations from the mean value congruent with the required acceptance limits (± 10%) . All tablets had functional mechanical properties for packaging and handling. The required breaking load of the tablets has values above 200 N up to the maximum value measurable with the durometer, 484 N. The strength of the printer tablets was also confirmed by the friability test conducted, which was satisfied as each formulation had a weight loss significantly less than 1%, which is indicated by F.U.I. XII as the maximum acceptable value . From the content uniformity test, the concentration of drug present in 10 tablets of each formulation was obtained, confirming the closeness to the theoretical value of the drug, and demonstrating the absence of drug degradation during extrusion. All the data described are shown in Table 3.
Regarding the disaggregation test, as expected no formulations showed signs of disaggregation or breakage at the end of the test. In fact, the 3D printed tablets were intact after both acid and basic media exposure. Therefore, the results confirmed that these pharmaceutical dosage forms did not disaggregate, but they released the loaded drug following swelling of the matrix and solubilization of the excipients.
SEM images show the surface and transverse planes of the four different formulations obtained (Fig. 7). Although the tablets were derived from blends consisting of different excipients at various concentration, they are morphologically similar. In the cross section, the three-dimensional layer-by-layer structure, characteristic of tablets obtained by 3D printing, can be seen. Each layer has a thickness of 0.2 mm, as determined by the printing parameters. The surface plane shows the concentric geometry of the chosen infill.
The data obtained from the chemical microanalysis show a homogeneous presence of the elements N (green in Fig. 7) and Cl (red in Fig. 7), which, being atoms exclusively present in the structure of the drug, indicate an equal distribution of NCS within the printed tablets.
Solid state characterization of printed tablets
The possible amorphization process undergone by the drug during extrusion was verified by solid-state characterization studies of the printed tablets. Using FT-IR it was observed that characteristic peaks in the NCS spectrum were present at 1218, 1520, 1570 and 1650 cm−1 (Fig. 8). The presence of these peaks was confirmed in the blend with the drug spectrum for each formulation. Peaks around 3300 cm−1 are characteristic of HP-β-CD as seen in the blends with and without the drug (Fig. 8 C-D). From the analysis of the spectrum relative to the formulations obtained from the above blends, we can see a relative widening of the HP-β-CD peak in a range between 3350 and 3100 cm−1. This widening could be reasonably attributed to the interaction of HP-β-CD with HPMC and NCS during the printing step . The same observation can be made regarding the NCS peaks between 1600 cm−1 and 1800 cm−1 and between 1250 cm−1 and 1100 cm−1. The absence of important peaks characteristic of NH stretching and the stretching of the carbonyl group of the drug could suggest the possible amorphization of the drug inside the printed tablets and/or the interaction between the drug and the HP-β-CD . These results could confirm the formation of intimate complexes of HP-β-CD, NCS and HPMC.
A second test to verify this condition was performed by DSC. NCS shows a strong endothermic peak at 230 °C, descriptive of its crystalline nature (Fig. 9). A smaller endothermic peak is present in the thermogram of the physical blend with the drug. In contrast, the peak at 230 °C is completely absent in the thermogram of all formulations, probably due to amorphization of the drug during the printing phase and/or the formation of an inclusion complex between the NCS and HP-β-CD.
In Fig. 10 the powder diffraction patterns of NCS, blend with and without drug and formulations, are reported for each compound under investigation. The sharp peaks present in the pure NCS powder diffraction pattern indicate its crystallinity, while their complete disappearance in the patterns of formulations, clearly shows the NCS amorphization during the printing phase. This phenomenon, already pointed out by FT-IR and DSC results is confirmed by the PXRD analysis.
TGA analysis was conducted on both blend 4 and formulation 4. The choice of these samples was determined by the need to study the behavior of all the used excipients. The results obtained (Figure S1) for mixture and formulation 4 described a perfect sigmoidal pattern, confirming the stability of all components at the printing temperature of 180 °C. In addition, Figure S1 reported the first derivatives of the obtained values, indicating that the maximum peak of complete degradation of both the powder mixture and the formulation occurred at 300 °C, thus excluding any possible degradation during the printing phase.
To assess the stability of NCS during the printing process, HPLC analyses were carried out on a sample of NCS left for 15 min at 180 °C (printing time and extrusion temperature for one tablet) and on formulation 4 after complete solubilization. The chromatograms reported in Figure S2 show no sign of drug degradation, with only one NCS peak with a retention time of 6.1 min. We could therefore confirm that the thermal and mechanical stress induced by the printing process does not interfere with the stability of the drug.
Further confirmation of the stability of the drug was obtained by mass spectroscopy analysis performed on NCS kept at 180 °C for 15 min. Single quadrupole scanning in negative ion mode of the NCS confirmed that the main ion was the drug (m/z ion 326.97, [M]− H+) (data not shown).
Dissolution testing of printed tablets
Dissolution studies of tablets obtained by direct extrusion of the four blends were performed for 48 h (Fig. 11). NCS release was studied in simulated gastric fluid for the first two hours and in simulated enteric fluid for the remaining hours. All formulations enhanced the dissolution profile of the pure drug, offering further confirmation of the advantage of this technique, which induced the loss of the drug's crystalline network during the printing process. Although at different times, all printed tablets showed a sustained and complete drug release within 48 h. In fact, HP-β-CD-containing formulations (3 and 4) achieved 100% NCS release in 24 h. In contrast, formulations 1 and 2 released 65% and 70% of NCS at 24 h, respectively, achieving complete drug release at 48 h. The different release profile of the tablets could be related to the different concentration of HPMC that constitutes them. In fact, the HPMC interacting with the aqueous fluid led to the formation of a gel layer, which acts as a diffusion barrier that counteracted drug release . Several studies have been found in the literature concerning the behavior of HPMC when it is placed in contact with aqueous fluids . Palugan et al. demonstrated the previously described phenomenon and provided as a potential solution the use of cellulolytic products capable of degrading the HPMC matrix more rapidly. The thickness of this layer may depend on the concentration of the polymer, in formulations 1 and 2 was present in higher concentrations than tablets containing HP-β-CD, causing a NCS release slower, but within 48 h. The high hydrophilicity of HP-β-CD and the ability of HPMC to stabilize the NCS/HP-β-CD complex in solution [26, 31] justify the improved release profile obtained from formulations 3 and 4. In addition, to produce sustained release tablets, a concentric filling pattern was selected in the set printing parameters. In fact, Obein et al. examined the influence on release profiles of four different filling patterns, showing that tablets with tri-hexagonal or concentric fillings had a longer release when compared to tablets with different filling geometries but the same composition . In addition, they had shown that the selection of appropriate excipients affects the type of formulation that can be obtained, demonstrating that the use of HPMC allows the development of sustained-release tablets. Plotting the data of cumulative releases vs. time, for the first 8 h, obtained straight lines with R2 ˃ 0.96 for all tablets, suggesting that the release profiles follow kinetics of order 0.
The results obtained confirm the amorphization of the drug and the formation of a ternary inclusion complex between HP-β-CD, HPMC and NCS, inducing an improvement on drug loading and release.
The stability of NCS in printed tablets was evaluated for 3 months (storage parameters 25 °C, 60% RH). The results highlighted that the amorphous state of the NCS, achieved following the extrusion process, is maintained and no signs of drug degradation were detected (data not shown).