Chromatographic analysis and cytotoxic effects of lidocaine HCl and NaOCl admixtures: An in vitro study

Aim: This study aimed to analyze the reaction mixtures of 2% lidocaine hydrochloride with adrenaline (LA) and 2.5% sodium hypochlorite (NaOCl) using high-performance liquid chromatography (HPLC)-mass spectrometry and assess their cytotoxicity on 3T3 mouse fibroblast cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Methods: The experiment was conducted with two different NaOCl fractions (1 and 5 mL of 2.5% NaOCl) mixed with 1 mL of LA solution that served as test groups, whereas LA/distilled water as well as independent LA and NaOCl solutions served as control groups. The data were statistically analyzed using GraphPad Prism 10.4.1 with a level of significance set at P < 0.05. One-way analysis of variance and Tukey’s post hoc test with Bonferroni correction was performed to compare the mean viability scores between the various groups. Results: The HPLC peak eluted at a retention time of 19.80–20.00 min and the corresponding mass was detected in mass spectroscopy. The compound mass (m/z 143) was identified as 2,6-dimethylaniline (2,6-xylidine), corresponding to (M[121] + Na [23] -1)+ values MTT results revealed that Group 1 (2% LA) exhibited the highest cell viability (98.47 ± 1.00), while the Group 2 (2.5% NaOCl) showed the lowest viability (45.87 ± 35.87). Pairwise comparisons revealed statistically significant differences between Group 2 with all the groups except Group 5 (1 mL of 2% LA + 5 mL of 2.5% NaOCl). The addition of NaOCl with 2% LA decreased cell viability; however, the differences were highly significant in Group 5 ( P < 0.001). Conclusion: The formation of an intermediary reactionary compound following NaOCl/LA admixed solutions is observed to be cytotoxic. Hence, the immediate copious use of NaOCl following intrapulpal anesthetic administration should be avoided in the clinical scenario.