7 research outputs found
Nanoparticles in Endodontics Disinfection: State of the Art
Endodontic-related diseases constitute the fourth most expensive pathologies in industrialized countries. Specifically, endodontics is the part of dentistry focused on treating disorders of the dental pulp and its consequences. In order to treat these problems, especially endodontic infections, dental barriers and complex root canal anatomy should be overcome. This constitutes an unmet medical need since the rate of successful disinfection with the currently marketed drugs is around 85%. Therefore, nanoparticles constitute a suitable alternative in order to deliver active compounds effectively to the target site, increasing their therapeutic efficacy. Therefore, in the present review, an overview of dental anatomy and the barriers that should be overcome for effective disinfection will be summarized. In addition, the versatility of nanoparticles for drug delivery and their specific uses in dentistry are comprehensively discussed. Finally, the latest findings, potential applications and state of the art nanoparticles with special emphasis on biodegradable nanoparticles used for endodontic disinfection are also reviewed. Keywords: PLGA; dentistry; endodontics; metal nanoparticles; nanoparticles
Novel strategies enhancing endodontic disinfection: antibacterial biodegradable calcium hydroxide nanoparticles in an ex vivo model.
Due to the high failure rates associated to endodontic disinfection, this study aimed to investigate the antibacterial properties of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) loaded with Ca(OH)2 for endodontic disinfection procedures. Ca(OH)2 NPs production and physicochemical characterization were carried out as well as multiple antibacterial tests using three bacterial strains and an ex vivo model of endodontic infection with extracted human teeth. Agar diffusion test and broth dilution determined the inhibition growth zones (n=5) and the minimal inhibitory concentration (MIC, n=5), respectively. Cell viability was assessed using Live/Dead staining with confocal microscopy (n=5). Data was analysed using ANOVA followed by post-hoc analysis. After 24 hours of incubation, Ca(OH)₂ NPs demonstrated a MIC of 10 µg/mL for Porphyromonas gingivalis (p<0.001) and Enterococcus faecalis and 5 µg/mL for Fusobacterium nucleatum (p<0.001). Although the agar diffusion test did not exhibit any inhibition for Ca(OH)2 nor for Ca(OH)₂ NPs area probably due to the buffering effect of the agar medium. However, the antibacterial capacity was confirmed in an ex vivo model, where instrumentalized teeth were infected with Enterococcus Faecalis and treated after 28 days of culture. A significant reduction in bacterial metabolic activity was confirmed for Ca(OH)2 NPs (40% reduction with a single dose) and confirmed by Live/Dead staining. In conclusion, Ca(OH)₂-loaded PLGA NPs present promising antibacterial efficacy for endodontic disinfection procedures
Nanoparticles: The next step in endodontic medicaments.
Aim: The aim of the current study was to develop two formulations loading calcium hydroxide (Ca(OH)â‚‚)
and Clobetasol Propionate into poly lactic-co-glycolic acid (PLGA) biodegradable nanoparticles (NPs) to
be used in the field of endodontics as an intracanal medications, including NPs optimization and
characterization, plus drug release profile of the NPs compared to free drug counterpart. Additionally,
comparison of the depth and area of penetration of the NPs inside the dentinal tubules against the free
drug was carried out. Furthermore, the therapeutic efficacy for both formulations were examined using
antibacterial and anti-inflammatory assays.
Methodology: NPs were prepared using the solvent displacement method. Optimization of the NPs was
carried out with a central composite design to obtain a final optimized formulation. The NPs morphology
was examined under transmission electron microscopy (TEM), plus characterization was done by x-ray
diffraction (XRD), fourier transform infrared spectroscopy (FTIR) and differential scanning calorimetry
(DSC). Moreover, the optimal conditions and cryoprotectants for freeze drying of the NPs were examined
and tested. The drug release profile of the NPs and free drug was evaluated up to 48 hours. Additionally,
the depth and area of penetration inside the dentinal tubules was examined for both the NPs and free
drug. Antibacterial testing was performed on the antibacterial NPs to evaluate their efficacy in
eliminating different bacterial strains. Agar disk-diffusion and broth dilution were used to determine the
inhibition growth zones and the minimal inhibitory concentration (MIC). Likewise, anti-inflammatory
testing was performed on the Clobetasol Propionate anti-inflammatory NPs, in which their response and
reaction against inflammatory cells, in particular against macrophages was tested using the enzyme-
linked immunosorbent assay (ELISA) to examine the cytokine release of IL-1β and TNF-α.
Results: Using the solvent displacement method, both formulations of NPs were successfully optimized.
The characteristics of the NPs utilizing the optimized formula showed a polidpsersity index (PI) value
lower than 0.2, characteristic of monodisperse systems and an average size below 200 nm along with a
highly negative surface charge (measured as zeta potential, ZP) and maximum entrapment efficiency (EE)
percentage. For both the Ca(OH)â‚‚/PLGA and the clobetasol propionate/PLGA NPs, the characterization of
the NPs was performed, and the spherical morphology was confirmed using TEM. Moreover, interaction
studies were carried out including XRD, FTIR and DSC to examine the profile of the NPs. These studies
confirmed that no covalent bonds were formed during the preparation process and that the drug
appeared to be encapsulated inside the NPs. Examining the drug release profile for the Ca(OH)â‚‚/PLGA
NPs, the Ca(OH)â‚‚ NPs exhibited a prolonged and steady release with higher concentrations than those
obtained with the free Ca(OH)â‚‚, that remained stable up to 48 hours. Moreover, clobetasol
propionate/PLGA NPs, exhibited a prolonged and steady release with only around 21 % of the
encapsuled drug released after 48 hours, in comparison the free drug that was completely released after
just 6 hours. For the freeze drying of the Ca(OH)â‚‚/PLGA NPs, the combination of 5% of (2-
Hydroxypropyl)-β-cyclodextrin and 15% D-mannitol gave rise to the most stable outcome and the best
appearance after lyophilization being able to recover the initial physicochemical characteristics of the
NPs after their resuspension. Unfortunately, for the clobetasol propionate/PLGA NPs, we were not able
to achieve a suitable combination. Using the confocal laser scanning microscopy, it was shown that both
the Ca(OH)â‚‚/PLGA and the clobetasol propionate/PLGA NPs had a better depth and area of penetration
inside the dentinal tubules when compared to the free drug. Plus, the NPs displayed higher values mean
fluorescence intensity (MFI) and integrated density compared to the free drug. Assessing the
antibacterial efficiency of the Ca(OH)â‚‚/PLGA NPs using the agar diffusion test, it was noted that the NPs
showed clear maximal zones of growth inhibition around the filter papers comparable only to the
positive group of antibiotics. Additionally, the MIC of the NPs was measured in regard to inhibiting
bacterial growth for 3 bacterial species. It was noted that after 24 hours of incubation the NPs were able
to inhibit bacterial growth at all tested concentrations. The anti-inflammatory capacity of the clobetasol
propionate/PLGA NPs was also examined, in which ELISA assays were used to evaluate the inflammatory
cytokines release of the macrophages in response to the NPs. The releases of TNF-α was reduced
considerably to almost undetectable amounts with NPs compared to the LPS negative control, displaying
results similar and even less than the TCP positive control after 48 hours. However, resulted in the
release of IL-1β in higher amounts which were similar to the TCP positive control and lesser than the LPS
negative control group. Additionally, the inflammatory cytokines release of IL-1β and TNF-α from the LPS
pretreated macrophages, was not reduced with the NPs at all tested concertation to limits below the
amounts obtained by the LPS pretreated macrophages or the TCP positive control group after 48 hours.
Conclusions: In the present study, Ca(OH)â‚‚ loaded PLGA NPs and clobetasol propionate loaded PLGA NPs
were successfully optimized and characterized, and their therapeutic efficacy was tested with aims to
increase the antibacterial and anti-inflammatory effect through controlled drug release and reach areas
of complicated root canal anatomy due to the smaller mean nanoscopic size. However, further future
studies are still required in order for the developed NPs to take the next step to be utilized in a clinical
condition and to achieve its goal as an intracanal medicament.El objetivo del presente estudio ha sido el desarrollo de dos formulaciones encapsulando
hidróxido de calcio (Ca(OH)â‚‚) y clobetasol en nanopratÃculas (NPs) biodegradables de ácido poli(láctico-
co-glicólico) (PLGA) para ser usadas en el campo de la endodoncia como medicación intra-conducto,
incluyendo la optimización y caracterización de las NPs y el perfil de liberación del principio activo en
comparación con el fármaco libre. Adicionalmente se comparó la cantidad y el área de penetración de las
NPs en el interior de los túbulos dentinarios. Las eficacias terapéuticas de ambas formulaciones fueron
examinadas mediante test antibacterianos y antiinflamatorios.
Resultados: Las formulaciones de NPs con hidrócido de calcio y clobetasol se prepararon utilizando el
método de desplazamiento de solvente. Las caracterÃsticas de las formulaciones optimizadas
presentaron una población homogénea con un valor de Ãndice de polidispersión (PI inferior a 0,2) y
tamaño promedio adecuado, inferior a 200 nm. Además, su carga superficial medida en base al potencial
zeta fue altamente negativa y se obtuvo una eficiencia de asociación elevada en ambos casos, superior
para el clobetasol. Tanto para las NP de Ca (OH) â‚‚ / PLGA como para las NP de propionato de clobetasol /
PLGA, se confirmó su morfologÃa esférica y su superficie lisa mediante MET. Se realizaron diferentes
estudios de interacción fisicoquÃmica confirmando que ambos fármacos se encontraban en el interior de
la matriz polimérica y que no se producÃan nuevos enlaces en la formación de los sistemas
nanoestructurados. Se examinó el perfil de liberación para ambas formulaciones. En el caso de las NP de
Ca (OH) ₂ / PLGA, las NP de Ca (OH) ₂ mostraron una liberación prolongada y constante del fármaco con
concentraciones más altas que en el caso dl fármaco libre, que permanecieron estables hasta 48 horas
después. Las NP de propionato de clobetasol / PLGA exhibieron una liberación prolongada y constante
con una liberación del 21 % del fármaco encapsulado liberado después de 48 horas, en comparación con
el fármaco libre que se liberó por completo después de solo 6 horas. Para la liofilización de las NP de Ca
(OH) ₂ / PLGA, se obtuvo un resultado el mejor resultado con la combinación de 5% de (2-hidroxipropil) -
β-ciclodextrina y 15% de D-manitol. Estas formulaciones pudieron ser resuspendidas de forma
instantánea consiguiendo los parámetros originales. Desafortunadamente, para los NP de propionato de
clobetasol / PLGA, no pudimos lograr una combinación adecuada de crioprotectores y condiciones de
liofilización óptimas. Usando la microscopÃa confocal, se demostró que tanto el Ca (OH) â‚‚ / PLGA como el
propionato de clobetasol / PLGA NP tenÃan una mejor profundidad y área de penetración dentro de los
túbulos dentinarios en comparación con el fármaco libre. Además, los NP mostraron valores más altos de
intensidad de fluorescencia media (mean fluorescence intensity, MFI) y densidad integrada en
comparación con el fármaco libre. Al evaluar la eficacia antibacteriana de las NP de Ca (OH) ₂ / PLGA
utilizando la prueba de difusión en agar, se observó que las NP mostraban zonas claras máximas de
inhibición del crecimiento alrededor de los papeles de filtro comparables solo al grupo positivo de
antibióticos. Además, se midió la concentración mÃnima inhibitoria (CMI) de las NP con respecto a la
inhibición del crecimiento bacteriano para 3 especies bacterianas. Se observó que después de 24 horas
de incubación, las NP inhibieron el crecimiento bacteriano en todas las concentraciones probadas. Se
realizaron estudios para medir la capacidad antiinflamatoria de las NPs de propionato de clobetasol
/PLGA, para lo cual se utilizaron test ELISA para evaluar la respuesta de los macrófagos en la liberación
de citoquinas inflamatorias en presencia de las NP. Se observó que las liberaciones de TNF-α se
redujeron considerablemente a cantidades casi indetectables con NPs en comparación con el control
negativo (LPS), mostrando resultados similares e incluso menores que el control positivo (TCP) después
de 48 horas. Sin embargo, se obtuvo la liberación de IL-1β en cantidades mayores que eran similares al
control positivo de TCP y menores que al grupo de control negativo de LPS. La liberación de citoquinas
inflamatorias de IL-1β y TNF-α de los macrófagos pretratados con LPS no se redujo con las NP en ninguna
concertación probada a lÃmites por debajo de las cantidades obtenidas por los macrófagos pretratados
con LPS o el grupo de control positivo de TCP después de 48 horas.
__Conclusiones: En el presente estudio, las NP de PLGA encapsulando Ca(OH)â‚‚ y las NP de PLGA
encapsulando propionato de clobetasol han sido optimizadas y caracterizadas con éxito, y se ha probado
su eficacia terapéutica con el objetivo de aumentar el efecto antibacteriano y antiinflamatorio. La
penetración de las mismas en áreas de anatomÃa complicada del conducto radicular (túmulos
dentinarios) ha sido demostrada, debido al tamaño nanoscópico de las mismas. Sin embargo, aún se
requieren más estudios futuros para que las NP formuladas puedan ser utilizados en una condición
clÃnica
Nanoparticles: The next step in endodontic medicaments.
Aim: The aim of the current study was to develop two formulations loading calcium hydroxide (Ca(OH)â‚‚) and Clobetasol Propionate into poly lactic-co-glycolic acid (PLGA) biodegradable nanoparticles (NPs) to be used in the field of endodontics as an intracanal medications, including NPs optimization and characterization, plus drug release profile of the NPs compared to free drug counterpart. Additionally, comparison of the depth and area of penetration of the NPs inside the dentinal tubules against the free drug was carried out. Furthermore, the therapeutic efficacy for both formulations were examined using antibacterial and anti-inflammatory assays.
Methodology: NPs were prepared using the solvent displacement method. Optimization of the NPs was carried out with a central composite design to obtain a final optimized formulation. The NPs morphology was examined under transmission electron microscopy (TEM), plus characterization was done by x-ray diffraction (XRD), fourier transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC). Moreover, the optimal conditions and cryoprotectants for freeze drying of the NPs were examined and tested. The drug release profile of the NPs and free drug was evaluated up to 48 hours. Additionally, the depth and area of penetration inside the dentinal tubules was examined for both the NPs and free drug. Antibacterial testing was performed on the antibacterial NPs to evaluate their efficacy in eliminating different bacterial strains. Agar disk-diffusion and broth dilution were used to determine the inhibition growth zones and the minimal inhibitory concentration (MIC). Likewise, anti-inflammatory testing was performed on the Clobetasol Propionate anti-inflammatory NPs, in which their response and reaction against inflammatory cells, in particular against macrophages was tested using the enzyme-linked immunosorbent assay (ELISA) to examine the cytokine release of IL-1β and TNF-α.
Results: Using the solvent displacement method, both formulations of NPs were successfully optimized. The characteristics of the NPs utilizing the optimized formula showed a polidpsersity index (PI) value lower than 0.2, characteristic of monodisperse systems and an average size below 200 nm along with a highly negative surface charge (measured as zeta potential, ZP) and maximum entrapment efficiency (EE) percentage. For both the Ca(OH)₂/PLGA and the clobetasol propionate/PLGA NPs, the characterization of the NPs was performed, and the spherical morphology was confirmed using TEM. Moreover, interaction studies were carried out including XRD, FTIR and DSC to examine the profile of the NPs. These studies confirmed that no covalent bonds were formed during the preparation process and that the drug appeared to be encapsulated inside the NPs. Examining the drug release profile for the Ca(OH)₂/PLGA NPs, the Ca(OH)₂ NPs exhibited a prolonged and steady release with higher concentrations than those obtained with the free Ca(OH)₂, that remained stable up to 48 hours. Moreover, clobetasol propionate/PLGA NPs, exhibited a prolonged and steady release with only around 21 % of the encapsuled drug released after 48 hours, in comparison the free drug that was completely released after just 6 hours. For the freeze drying of the Ca(OH)₂/PLGA NPs, the combination of 5% of (2-Hydroxypropyl)-β-cyclodextrin and 15% D-mannitol gave rise to the most stable outcome and the best appearance after lyophilization being able to recover the initial physicochemical characteristics of the NPs after their resuspension. Unfortunately, for the clobetasol propionate/PLGA NPs, we were not able to achieve a suitable combination. Using the confocal laser scanning microscopy, it was shown that both the Ca(OH)₂/PLGA and the clobetasol propionate/PLGA NPs had a better depth and area of penetration inside the dentinal tubules when compared to the free drug. Plus, the NPs displayed higher values mean fluorescence intensity (MFI) and integrated density compared to the free drug. Assessing the antibacterial efficiency of the Ca(OH)₂/PLGA NPs using the agar diffusion test, it was noted that the NPs showed clear maximal zones of growth inhibition around the filter papers comparable only to the positive group of antibiotics. Additionally, the MIC of the NPs was measured in regard to inhibiting bacterial growth for 3 bacterial species. It was noted that after 24 hours of incubation the NPs were able to inhibit bacterial growth at all tested concentrations. The anti-inflammatory capacity of the clobetasol propionate/PLGA NPs was also examined, in which ELISA assays were used to evaluate the inflammatory cytokines release of the macrophages in response to the NPs. The releases of TNF-α was reduced considerably to almost undetectable amounts with NPs compared to the LPS negative control, displaying results similar and even less than the TCP positive control after 48 hours. However, resulted in the release of IL-1β in higher amounts which were similar to the TCP positive control and lesser than the LPS negative control group. Additionally, the inflammatory cytokines release of IL-1β and TNF-α from the LPS pretreated macrophages, was not reduced with the NPs at all tested concertation to limits below the amounts obtained by the LPS pretreated macrophages or the TCP positive control group after 48 hours.
Conclusions: In the present study, Ca(OH)â‚‚ loaded PLGA NPs and clobetasol propionate loaded PLGA NPs were successfully optimized and characterized, and their therapeutic efficacy was tested with aims to increase the antibacterial and anti-inflammatory effect through controlled drug release and reach areas of complicated root canal anatomy due to the smaller mean nanoscopic size. However, further future studies are still required in order for the developed NPs to take the next step to be utilized in a clinical condition and to achieve its goal as an intracanal medicament.Objetivo: El objetivo del presente estudio ha sido el desarrollo de dos formulaciones encapsulando hidróxido de calcio (Ca(OH)â‚‚) y clobetasol en nanopratÃculas (NPs) biodegradables de ácido poli(láctico-co-glicólico) (PLGA) para ser usadas en el campo de la endodoncia como medicación intra-conducto, incluyendo la optimización y caracterización de las NPs y el perfil de liberación del principio activo en comparación con el fármaco libre. Adicionalmente se comparó la cantidad y el área de penetración de las NPs en el interior de los túbulos dentinarios. Las eficacias terapéuticas de ambas formulaciones fueron examinadas mediante test antibacterianos y antiinflamatorios.
MetodologÃa: Las NPs fueron preparadas por el método de desplazamiento de solvente. La optimización fue realizada mediante un diseño factorial central compuesto obtenido la formulación final. Se realizó un estudio de la morfologÃa de las NP mediante microscopÃa electrónica de transmisión (MET). Además, las NPs se caracterizaron mediante cristalografÃa de Rayos X, espectroscopÃa infrarroja de Fourier y calorimetrÃa diferencial de barrido. También se examinaron y llevaron a cabo el estudio de las condiciones óptimas para el proceso de liofilización de las NP. Se evaluó el perfil de liberación del fármaco desde el interior de las NP comparándolo con el del fármaco libre, hasta las 48 horas. El examen de penetración dentro de los túbulos dentinarios de las NP en comparación con el fármaco libre se realizó mediante microscopÃa confocal. Para evaluar la capacidad antibacteriana de las NP de hidróxido de calcio se realizaron los test de método Kirby-Bauer (Test de difusión en agar) y la concentración mÃnima inhibitoria (CMI). En las NP de propinado de clobetasol se evaluó su respuesta y reacción frente a células inflamatorias, en particular frente a macrófagos, utilizando el ensayo inmunoabsorbente ligado a enzimas (ELISA) para examinar la liberación de citocinas de IL -1β y TNF-α.
Resultados: Las formulaciones de NPs con hidrócido de calcio y clobetasol se prepararon utilizando el método de desplazamiento de solvente. Las caracterÃsticas de las formulaciones optimizadas presentaron una población homogénea con un valor de Ãndice de polidispersión (PI inferior a 0,2) y tamaño promedio adecuado, inferior a 200 nm. Además, su carga superficial medida en base al potencial zeta fue altamente negativa y se obtuvo una eficiencia de asociación elevada en ambos casos, superior para el clobetasol. Tanto para las NP de Ca (OH) â‚‚ / PLGA como para las NP de propionato de clobetasol / PLGA, se confirmó su morfologÃa esférica y su superficie lisa mediante MET. Se realizaron diferentes estudios de interacción fisicoquÃmica confirmando que ambos fármacos se encontraban en el interior de la matriz polimérica y que no se producÃan nuevos enlaces en la formación de los sistemas nanoestructurados. Se examinó el perfil de liberación para ambas formulaciones. En el caso de las NP de Ca (OH) â‚‚ / PLGA, las NP de Ca (OH) â‚‚ mostraron una liberación prolongada y constante del fármaco con concentraciones más altas que en el caso dl fármaco libre, que permanecieron estables hasta 48 horas después. Las NP de propionato de clobetasol / PLGA exhibieron una liberación prolongada y constante con una liberación del 21 % del fármaco encapsulado liberado después de 48 horas, en comparación con el fármaco libre que se liberó por completo después de solo 6 horas. Para la liofilización de las NP de Ca (OH) â‚‚ / PLGA, se obtuvo un resultado el mejor resultado con la combinación de 5% de (2-hidroxipropil) -β-ciclodextrina y 15% de D-manitol. Estas formulaciones pudieron ser resuspendidas de forma instantánea consiguiendo los parámetros originales. Desafortunadamente, para los NP de propionato de clobetasol / PLGA, no pudimos lograr una combinación adecuada de crioprotectores y condiciones de liofilización óptimas. Usando la microscopÃa confocal, se demostró que tanto el Ca (OH) â‚‚ / PLGA como el propionato de clobetasol / PLGA NP tenÃan una mejor profundidad y área de penetración dentro de los túbulos dentinarios en comparación con el fármaco libre. Además, los NP mostraron valores más altos de intensidad de fluorescencia media (mean fluorescence intensity, MFI) y densidad integrada en comparación con el fármaco libre. Al evaluar la eficacia antibacteriana de las NP de Ca (OH) â‚‚ / PLGA utilizando la prueba de difusión en agar, se observó que las NP mostraban zonas claras máximas de inhibición del crecimiento alrededor de los papeles de filtro comparables solo al grupo positivo de antibióticos. Además, se midió la concentración mÃnima inhibitoria (CMI) de las NP con respecto a la inhibición del crecimiento bacteriano para 3 especies bacterianas. Se observó que después de 24 horas de incubación, las NP inhibieron el crecimiento bacteriano en todas las concentraciones probadas. Se realizaron estudios para medir la capacidad antiinflamatoria de las NPs de propionato de clobetasol /PLGA, para lo cual se utilizaron test ELISA para evaluar la respuesta de los macrófagos en la liberación de citoquinas inflamatorias en presencia de las NP. Se observó que las liberaciones de TNF-α se redujeron considerablemente a cantidades casi indetectables con NPs en comparación con el control negativo (LPS), mostrando resultados similares e incluso menores que el control positivo (TCP) después de 48 horas. Sin embargo, se obtuvo la liberación de IL-1β en cantidades mayores que eran similares al control positivo de TCP y menores que al grupo de control negativo de LPS. La liberación de citoquinas inflamatorias de IL-1β y TNF-α de los macrófagos pretratados con LPS no se redujo con las NP en ninguna concertación probada a lÃmites por debajo de las cantidades obtenidas por los macrófagos pretratados con LPS o el grupo de control positivo de TCP después de 48 horas.
Conclusiones: En el presente estudio, las NP de PLGA encapsulando Ca(OH)â‚‚ y las NP de PLGA encapsulando propionato de clobetasol han sido optimizadas y caracterizadas con éxito, y se ha probado su eficacia terapéutica con el objetivo de aumentar el efecto antibacteriano y antiinflamatorio. La penetración de las mismas en áreas de anatomÃa complicada del conducto radicular (túmulos dentinarios) ha sido demostrada, debido al tamaño nanoscópico de las mismas. Sin embargo, aún se requieren más estudios futuros para que las NP formuladas puedan ser utilizados en una condición clÃnica
Post-Instrumentation Dentinal Microcracks Induced by Two NiTi Rotary Systems with Increased Super Elasticity and Shape Memory: A MicroCT Comparative and Methodological Ex Vivo Study
This study aimed to determine the incidence of dentinal microcracks after instrumenting the root canals of maxillary premolars using two novel rotary instrumentation systems. Micro-computed tomography (microCT) scans and images generated by sectioning and observation with a stereomicroscope were used to assess the specimens. Twenty-two freshly extracted maxillary premolars were collected and the specimens were divided into two groups of eleven. The tested radicular shaping systems were XP-Endo shaper and TRUShape (i.e., single file). The specimens were scanned with microCT pre- and post-instrumentation, and the newly formed microcracks were detected. The post-instrumentation scans were also compared with images obtained by sectioning method and stereomicroscope inspection, comparing the incidence of microcracks in either microCT scans or images. The results identified an overall incidence of 0.49% of newly formed microcracks, with no statistically significant differences (p = 0.689) between the shaping systems (0.11% for TRUShape and 0.87% for XP-Endo shaper). There were statistically significant differences (p < 0.001) between the microcracks incidence in microCT scans and the sectioning method (16.6% more for the latter). In conclusion, the results show that neither TRUShape nor XP-Endo shaper created dentinal microcracks during root canal instrumentation. The sectioning method with stereomicroscope evaluation overestimates the presence of microcracks with a statistically significant difference compared to microCT scans