Comparative Spectrophotometric Assessment of Crown Discoloration from Diverse Endodontic Sealers

Introduction

The color of a person’s teeth plays a major role in their overall appearance, and any changes in tooth color can lead to unwanted aesthetic issues [1]. The use of endodontic sealers is an indispensable part of the obturation systems used for endodontic treatments. There is a lack of long-term follow-up reports related to different endodontic sealers used in root canal treatment [2]. However, sealers transported into the coronal chamber during the root canal filling procedure can cause discoloration. This creates concerns among clinicians about the aesthetic appearance of a treated tooth and can have a significant impact on an individual’s quality of life[2]. An in vivo study evaluated the outcomes of quality of life and patient satisfaction after endodontic management. It was found that nearly 10% of the subjects were disappointed with the reduced aesthetic appearance of the treated tooth in terms of color, as well as the amount of time required and the experience of pain or discomfort during or after endodontic procedures [3,4].

The primary cause of local intrinsic discoloration in the cervical and central part of the tooth is the interaction between the materials used to fill the root canal and the coronal dentine layers of the tooth’s pulp chamber [2,5,6]. Even with improvements in the properties of endodontic sealers, discoloration of the crown remains a common occurrence. Studies have demonstrated that various root canal filling materials can cause discoloration, and the severity of this effect is influenced by the chemical consistency of the sealers[2,7,8].

Various techniques have been suggested for assessing or determining tooth discoloration related to endodontic treatment. These include visual examination and matching colors using a wide range of color tabs (based on the Munsell system) by senior evaluators, computerized evaluation of digital photos in Adobe Photoshop [9], colorimetry [10], and spectrophotometry [5,6,8]. Research has shown that there is no statistically significant reliability among various color measuring devices, and the correlations between the colorimeters and visual methods were not significant [11,12]. In contrast, spectrophotometers are commonly accepted as the primary tools in the study of color, and have been effectively utilized in dentistry for assessing tooth color [13].

The process of measuring color depends on complete reflection of the surface of the crown within the range of colors that can be seen by the human eye. Spectral reflectance values are used to gauge reflection and are visualized in the form of reflection curves, with each individual sample having its own distinct curve [14–16]. Several studies have demonstrated that spectrophotometry is a reliable, consistent, precise, and measurable technique [13,14]. A few studies have measured the changes in CIE L*, a*, b* values due to endodontic-related causes [17]. The CIE L*, a*, b* color system (CIELAB), developed by the International Commission on Illumination (CIE), is a widely used model for measuring and describing color in a three-dimensional space. It includes L* for lightness, ranging from 0 (black) to 100 (white), a* for the red-green axis, and b* for the yellow-blue axis. This system closely aligns with human vision and is ideal for detecting subtle color differences. It is commonly used for evaluating tooth color and monitoring discoloration following procedures like root canal treatment, offering an objective and standardized method for color assessment. Although previous studies have reported measurements of chromatic changes, a thorough review of the literature found few experimental reports specifically examining changes in CIE L*, a*, b* values resulting from discoloration caused by sealants [6].

To avoid discoloration of the crown, it is important to place root filling materials below the cementoenamel junction, remove any remaining sealer, and use sealers that do not cause staining. Endodontic sealers are specialized dental materials used in root canal therapy to fill the space between the root canal walls and the core filling material, usually gutta-percha. Their primary role is to create a hermetic seal that prevents ingress of bacteria and fluids and help in filling irregularities, accessory canals, and any voids, thereby enhancing the overall sealing ability and long-term success of the root canal treatment [18,19]. However, it can be difficult to completely remove sealer remnants from the pulp chamber, especially if visibility is limited or the material closely matches the color of the root dentin [2,5]. Additionally, using materials that do not contain discoloration-causing components can lead to a better aesthetic result [20]. Considering the growing desire for pleasing dental aesthetics, biomaterials should be chromatically stable, exhibit optical properties that are comparable to those of dental structures, and do not stain hard dental tissues.

Addressing and controlling coronal discoloration continues to present a significant challenge for dental practitioners, particularly when it occurs in the anterior teeth during endodontic treatment, which are decisive for an attractive smile. A recent study evaluated the influence of different sealers and the cervical limit of root fillings on tooth discoloration over a 1-year period. The findings indicated that all tested sealers caused perceptible discoloration within the first week, with the severity varying among sealers. Notably, positioning the root filling material 2 mm below the dental cervix significantly reduced crown discoloration compared to fillings placed at the dental cervix [21]. Another systematic review, published in 2024, analyzed the extent of tooth discoloration induced by endodontic sealers of different chemical compositions, showing that certain sealers, particularly those with specific chemical constituents, have a higher propensity to cause progressive pigmentation of the tooth. The discoloration process can start within a week of sealer application and can continue for up to 3 years [22]. Despite the well-documented impact of tooth discoloration on aesthetics and patient satisfaction following endodontic treatment, there is a notable lack of experimental studies specifically assessing the discoloration potential of different endodontic sealers using objective and standardized color measurement methods. While spectrophotometry has been recognized as a reliable tool for evaluating tooth color changes, very few studies have focused on quantifying changes in CIE L*, a*, b* values resulting from coronal discoloration caused by sealers. Additionally, there is limited evidence regarding the progression of discoloration over time in relation to different types of sealers. This gap highlights the need for systematic evaluation of commonly used sealers to better understand their aesthetic impact and support clinical decisions aimed at preserving tooth appearance after endodontic treatment. Thus, the present study was conducted to evaluate the crown discoloration induced by 4 different commonly used endodontic sealers: mineral trioxide aggregate MTA-based sealer, zinc oxide eugenol (ZOE)-based sealer, calcium hydroxide (Ca(OH)2)-based sealer, and resin-based sealer. A secondary objective was to evaluate the possible chronological relationship between when the teeth were exposed to the dyes and the consequent increase in the index of discoloration over time, as well as the relationship with the endodontic sealer used. Our null hypothesis was that there is no difference in discoloration of crowns associated with these 4 sealers.

Material and Methods

SAMPLE SIZE ESTIMATION:

Based on the findings of a pilot study, sample size was determined following power analysis using the formula n=2(z1−α/2 + zβ)2 σ2 {1+(m − 1)ρ}/mΔ2 with ρ=0.5, m=4 (repeated measurements), σ2=4, Δ=2.2, alpha=0.05, power=0.8 (the selected value of Δ was greater than the observed one in order to be close to the 3.7 threshold value) [23]. As a result, the sample size for each group was estimated to be close to n=12 specimens. Three specimens were added to each group (n=15) to allow for participants dropping out during the experiment.

SAMPLE SELECTION AND GROUP DIVISION:

We selected 75 human maxillary central incisors from the teeth collected for the study from the Oral Surgery Department of the institute. All enrolled patients signed a general consent form prior to any investigation or treatment, which includes consent for the use of their extracted teeth in future studies, without any personal identifying information. Teeth with cavities, cracks, restorations, or stains were not included. The teeth were first soaked in 2.5% sodium hypochlorite for 10 minutes to clean them, then kept in saline. Gross calculus and stains were removed using hand scalers and pumice with polishing cups [18,20]. Maxillary central incisors were specifically chosen due to their relatively flat labial surface, standardized anatomy, and central role in aesthetics, making them ideal for spectrophotometric evaluation, without causing anatomical variation, such as differences in enamel thickness or curvature, which could introduce inconsistencies in color measurements. Other anterior teeth, like lateral incisors and canines, present greater anatomical variability, which may have affected the reliability of the comparative results. Donor age was not individually recorded. Only fully matured, non-carious, unrestored maxillary central incisors extracted for orthodontic or periodontal reasons were included.

The 75 selected teeth were randomly divided into 5 groups: 4 experimental groups and 1 Negative Control group. Group 1 teeth received endodontic treatment with MTA-based sealer (MTA-FILLAPEX, Angelus Indústria de Produtos Odontológicos S/A, Londrina, Brazil) after performing access cavity and chemo-mechanical preparation. Group 2 teeth received endodontic treatment zinc oxide eugenol (ZOE)-based sealer (Tubliseal, manufactured by Kerr Endodontics, Kerr Dental, Brea, CA 92821). Group 3 teeth received endodontic treatment using calcium hydroxide (Ca(OH)2)-based sealer (Sealapex, Kerr Dental, Brea, CA 92821). Group 4 teeth received endodontic treatment with resin-based sealer (AH26, 13320-B, Ballantyne Corporate Pl, Charlotte, NC); Group 5 was the Negative Control group, which had access cavity, chemo-mechanical cleaning similar irrigation method and shaping, without any obturation material (Table 1).

SAMPLE PREPARATION:

After assignment to groups, the selected teeth were prepared for subsequent steps. The access cavities were prepared in all samples using a size 4 diamond round bur and high-speed handpiece. Then, Endo-access or Endo-Z burs were used to remove the pulp chamber roof. Gates Glidden drills #1, #2, and #3 were used for coronal enlargement. Pulp chambers were mechanically instrumented manually with hand K-files. All samples were irrigated with 3% sodium hypochlorite solution and final irrigation done with 17% ethylenediaminetetraacetic acid (EDTA) (Pulp dent, Watertown, MA). Then, the canals were washed with saline and dried with paper points. Later, the obturation was carried out with gutta-percha using the single-cone technique. The sealers were mixed and placed according to the manufacturer’s instructions into the pulp chamber through coronal access (0.05 ml per canal), as per group division. The exposure time matched the clinical setting, allowing the sealers to set fully in moist conditions for 24 hours. Internal axial walls were coated with lentulo-spiral filler. To simulate the forces used in the pulp chamber during the lateral condensation procedure, mild lateral pressure was applied. A dry, sterile cotton pellet was used to remove the excess sealant from the apical aspect. Self-curing glass ionomer cement (XTRACEM Medicept Dental, India) was used to seal the coronal orifices. All the teeth were kept in saline in an incubator at 37°C. In the Negative Control group, tooth canals were dried, and the access cavity was sealed using resin composite. All teeth were stored in an incubator at 100% humidity and 37°C during the study period to simulate the oral cavity environment [20].

ASSESSMENT OF CORONAL DISCOLORATION:

Crown discoloration with various endodontic sealers was evaluated by spectrophotometry, a technique used to measure the intensity of light that a sample absorbs. According to the concepts of physics and the Beer-Lambert law, which is used in spectrophotometry, the concentration of an absorber in a solution is proportional to the amount of light absorbed. Based on this principle, the positive values of L*, a*, and b*, which characterize the amount of light absorbed, can be acquired from objects with color, and the color changes before and after treatment can be accurately calculated [20,23]. Spectrophotometry offers several advantages in the assessment of tooth discoloration, making it a valuable tool. Unlike subjective methods such as visual shade matching or photographic analysis, spectrophotometry provides objective, accurate, and reproducible measurements of tooth color. This allows for consistent comparisons over time, can detecting even minor color changes, is non-invasive and safe, and allows repeated assessments without affecting the tooth structure. It also provides both numerical data and spectral reflection curves, offering a comprehensive understanding of color changes.

The spectrophotometer L* represents lightness (0, black; 100, white), a* represents chromaticity (green [−], red [+]), and b* represents chromaticity (blue [−] and yellow [+]). To prevent lighting interference, all measurements were made in the same room using uniform illumination that was set to a temperature of 6500 K. The same operator performed the measurements under the same environmental conditions, as lighting conditions (eg, natural daylight vs artificial light) can affect the perception of color differences and evaluators were blinded to the group assignments throughout the study to minimize the risk of observational bias.

Spectrophotometer equipment (Color i7 Benchtop Spectrophotometer/(CM-5, Konica Minolta, Tokyo, Japan) was used to measure the coronal discoloration of samples. The instrument was carefully positioned (the square marked with a star) (Figure 2) to make sure that the Commission Internationale de l’eclairage L*a*b* (CIELAB) reading was captured consistently from the tooth. The color shades were then directly obtained from the digital screen of the spectrophotometer device, and the CIELAB readings were recorded. The color variations and measurements of the specified buccal surfaces were recorded before treatment and considered as baseline data to which the further readings (immediately after treatment, 1 month, and 6 months after treatment) were compared. The spectrophotometer underwent calibration following the guidelines provided by the manufacturer prior to every measurement. The color differences at each time interval were calculated according to formula:

Where ΔL represents the change in brightness determined by variations in the ΔL* readings over 2 time periods. Δa and Δb indicate changes in color intensity. A ΔE value of 3.5 or higher was regarded as a clinically noticeable change in color perception [24]. The differences between various time intervals were designated as shown in Table 2.

A pilot study was conducted before the experiment to validate the proposed method. The pilot study assessed the repeatability and precision of the system used, including reproducibility and measurement error. The accuracy and margin of error of the operator’s measurements were also documented in the first 24 hours of the experimental period. The experimental model showed high positive correlation values and a ΔE of 1 unit after repeated measurements [17].

The reliability of the quantitative measurements made by 1 observer for each shade measurement was tested through intra-observer reliability tests. The same blinded observer captured the shades twice, with a 1-week gap, for 25 out of the 75 teeth in the study. Color measurements for these 25 teeth were also taken twice, with a 1-week gap.

Subsequently, the analysis of the average (with standard deviation) L0*, a0*, b0* values for all 5 groups revealed no statistically significant variances (P>0.05). This suggests that the teeth collected for each group effectively represented the original color properties of the entire sample. Each measurement was conducted twice, and the average value was recorded (Figure 3).

STATISTICAL ANALYSIS:

The quantitative tooth color measurements using spectrophotometer were obtained for samples in the 4 experiment groups and the Negative Control group. The data were obtained at baseline, immediately after treatment, at 1 month, and at 6 months for each sample in each group. The spectrophotometer parameters L, a, and b at each time point and ΔE values were obtained as a measure of discoloration.

The results are presented as mean±standard deviation for each chromatic parameter (L*, a*, b*, and ΔE) across all groups and time intervals. Additionally, ANOVA F values and P values are provided to indicate statistical significance. Where relevant, effect sizes and confidence intervals are also included to enhance the interpretation of findings. Mauchly’s test of sphericity was performed to assess whether the assumption was met. In cases where the assumption of sphericity was violated, appropriate corrections, such as the Greenhouse-Geisser adjustment, were applied to adjust the degrees of freedom and ensure the validity of the statistical inferences.

The difference in the discoloration at different times was tested for statistical significance. The values of the chromatic parameters CIE L*, a*, and b* as well as ΔE were analyzed using one-way analysis of variance (ANOVA) with repeated measures. The Shapiro-Wilk test was used to assess the univariate normality assumptions. For paired comparisons, the Tukey’s post hoc test was employed. The significant main effects of time, material, and their interactions were investigated with 2-way ANOVA with repeated measures followed by Bonferroni post hoc test for paired comparisons. All analyses were performed using SPSS version 20.0 (IBM, Inc.) software, and P<0.05 was regarded as indicating a statistically significant difference.

Results

COMPARISON OF L* CHROMATIC PARAMETER AMONG STUDIED GROUPS:

The comparison of ΔL (lightness change) across different materials – MTA, Ca(OH)₂, ZnOE, Resin, and the Negative Control – was analyzed at various time intervals using ANOVA. From baseline to immediate ΔL1, the lightness change ranged from 1.53±1.67 in MTA to 4.06±1.96 in Resin, with no statistically significant difference (F=1.86, p=0.135). However, significant differences emerged at other time intervals [ΔL2–ΔL5].

For ΔL2, the lightness change varied significantly (F=4.62, P=0.003), with Resin showing the greatest negative change (−2.30±2.97) and the Negative group having the least change (−3.98±3.34). Similarly, for ΔL3, the most substantial negative change observed in the Negative Control (−7.43±3.74) and Resin (−6.36±4.08).

For ΔL4, MTA had greater negative change (−6.98±2.93) compared to Resin (−1.69±2.69). ΔL5 had the most pronounced differences (F=19.32, P<0.001), with Resin showing a positive lightness change (0.61±1.87), while others, especially MTA and Ca(OH)2, had significant reductions. For the immediate to sixth-month interval (ie, ΔL6), no statistically significant differences were found (F=1.11, P=0.363), with all groups showing comparable lightness reductions. These results suggest that material type significantly influences lightness change over time, particularly in mid-interval assessments (Table 3, Figure 3).

Post hoc paired comparisons of ΔL (lightness change) among study materials revealed varying degrees of differences across time intervals. Overall, the Resin and the Negative groups had the most distinct patterns of ΔL over time (Table 4).

COMPARISON OF A*, B*, AND ΔE CHROMATIC PARAMETER AMONG STUDIED GROUPS:

Similar to L*, the comparison of Δa (change along the red-green axis); Δb (change along the yellow-blue axis), and total color difference (ΔE) revealed significant and non-significant differences across different time intervals.

In Δa from baseline to immediate measurement Δa1, the ANOVA test indicated a statistically significant difference (F=3.13, P=0.024), suggesting variability in red-green axis changes among the materials. MTA, Ca(OH)₂, Resin, and ZnOE all demonstrated positive mean Δa values, whereas the Negative group exhibited a slight decrease.

In Δb, from baseline to immediate measurement Δb1, significant differences were observed (P=0.014), with Resin showing the highest mean change (2.18±2.40), while MTA exhibited a negative mean change (−1.12±1.25). These differences highlight variability in color shifts among the materials during the initial phase.

In ΔE, from baseline to immediate measurement ΔE1, there was a statistically significant difference in ΔE among the materials (F=2.88, P=0.033). The highest mean change was observed in Ca(OH)₂ (5.04±2.59), followed by Resin (4.88±2.77), while the lowest was seen in MTA (2.56±1.13). However, no significant differences were found in the baseline to second-month interval (F=1.20, P=0.322) or from immediate to second-month measurements (F=2.36, P=0.068), suggesting comparable changes during these periods.

Likewise, comparisons were done for the baseline to the second month and baseline to the sixth month, immediate to the second month, immediate to the sixth-month interval, and second to sixth month and are reported in Table 3 and Figure 4A–4D.

Post hoc paired comparisons of changes in Δa (red-green axis), Δb (change along the yellow-blue axis), and total color difference (ΔE) among the study materials revealed notable differences across time intervals and are presented in Tables 4–7.

Overall, the data show that changes in Δa, Δb, and ΔE were most pronounced in specific intervals, particularly for Δa immediately after treatment to the second month and immediately after treatment to the sixth month, emphasizing the variable impact of different materials on red-green axis color changes. For Δb immediate to sixth-month interval, the differences were not statistically significant (P=0.187), although MTA displayed the largest mean increase (7.59±2.90), followed by the Negative group (6.58±2.45). These results suggest that while early-phase differences in Δb were prominent, changes tended to converge over longer periods across the study materials and for ΔE values varied significantly in some intervals, particularly during the initial and mid-stages of observation, reflecting differential color stability among the materials over time.

The 2-way ANOVA analysis demonstrated the effects of time, material, and their interaction on total color difference (ΔE), providing key insights into the factors influencing color stability. The corrected model showed statistical significance with an F value of 12.51 and P<0.001, explaining 57.3% of the variance in ΔE, indicating a strong overall model fit (Table 8).

Time emerged as a significant factor, with an F value of 59.62 and P<0.001, and an effect size of 0.525, highlighting that changes in color were heavily influenced by the duration of the study. Conversely, material did not significantly affect ΔE (F=1.53, P=0.194), with a minimal effect size (0.022), suggesting that the type of material had a limited independent impact on color change.

The interaction between time and material was significant (F=2.93, P<0.001), with an effect size of 0.178, indicating that the influence of time on ΔE varied depending on the material used. This interaction underscores the dynamic relationship between these factors, suggesting that the rate or extent of color change over time differed across materials.

Overall, time played a dominant role in influencing ΔE, while material effects were only significant when interacting with time, pointing to progressive variations in material behavior regarding color stability. MTA caused the least discoloration, while ZnOE caused the most within the first month. After 6 months, all materials had clinically perceptible crown discoloration.

Discussion

CLINICAL IMPLICATIONS:

The discoloration of the crown, which can occur after root canal treatment and reflects through the restoration, may be unsatisfactory for patients regarding the aesthetic result. The results of this study showed that certain sealers lead to significant discoloration that was perceptible by the human eye immediately after placement and even after 6 months. This clinically relevant discoloration can affect patient satisfaction with the aesthetic outcome or the longevity of the restoration. These factors should be considered and evaluated while making clinical decisions [29].

Each patient should also be informed about this aspect during the patient education and informed consent procedure. Endodontic treatment is often perceived as lacking aesthetic appeal; however, incorporating aesthetic materials into endodontic procedures is changing the trend and it should be regarded as a noteworthy development in this field. In addition, the preference for the use of endodontic sealers, which cause less discoloration, should be reinforced. Use of the extrusion and irrigation protocols to remove sealers’ residues from coronal dentin should be recommended. Consensus is clearly lacking regarding the most suitable sealer for single-visit endodontic treatment techniques. Additional researches focused on identifying the material used as sealer in single-visit endodontic treatment techniques are needed.

LIMITATIONS:

This was an in vitro study and has some limitations; therefore, caution should be exercised while applying these findings clinically as the time frame assessed in the present study was only till 6 months. Additionally, in vivo conditions involve dynamic factors such as saliva, temperature, oral hygiene, and dietary habits, which were not replicated in this in vitro study. In the future, it may be of interest to conduct in vitro and in vivo experiments with the materials for longer time periods to better simulate the clinical situation. The absence of detailed demographic data such as donor age, sex, and medical history may limit the generalizability of the findings.

RECOMMENDATIONS FOR FUTURE RESEARCH:

Crown discoloration following endodontic treatment is a complex issue that requires further investigation. Future studies should include pre- and post-obturation color measurements and explore a wider range of sealers in both in vitro and in vivo settings. Long-term studies spanning 2–5 years are needed to assess discoloration after treatment. Additionally, clinical research should evaluate patient satisfaction with tooth color and investigate operator-related variations.

Conclusions

In conclusion, endodontic sealers can cause discoloration of the coronal tooth structure, with all tested sealers inducing crown discoloration after 1 and 6 months compared to baseline. MTA-based sealer caused the least discoloration, while ZnOE-based sealer caused the most within the first month. After 6 months, all materials resulted in clinically perceptible crown discoloration. Notably, significant discoloration was observed in the Negative Control group. Root canal treatment significantly alters the spectrophotometric properties of natural teeth. Clinicians must be aware of the potential aesthetic impact of endodontic sealers when selecting materials, while also considering other contributing factors.