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Stability study of biological products plays an important role for determination of product changes in maintenance period and ensuring of safety and efficacy of vaccines. In this research, accelerated and long-term stability study performed for six batches of full and reduced-dose cattle Iriba strain brucellosis vaccine that manufactured by Razi vaccine and serum research institute as a new vaccine. After sampling, the vaccines were tested for accelerated stability, after four days storage at 22 0C and tested intervals in three months until 24 months for long-term stability after storage at 2-8 0C. The result indicated all batches of vaccines in accelerated stability met the specification recommended by OIE 2012 and the mean loss of activity for full-dose was 16.68, 18.87 and 17.79 % and for reduced-dose was 38.85, 36.06 and 34.98 %. In long term stability, the quality control tests including colony forming unit, purity, dissociation and physicochemical tests in all samples until 24 months, met the specification recommended by OIE 2012. The full-dose vaccines showed a mean loss of activity of 30.73, 25.53 and 32.45 % and the reduced-dose vaccines showed 63.51, 58.60 and 60.83 %. The mean increasing of moisture content was, 187.85, 214.13 and 160.77 % for full-dose and 142.35, 110.23 and 164.47 % for reduced-dose. So, the results of this research indicated in spite of moisture content increasing in second year, the brucellosis vaccines with this strain are stable at least 24months if the cold chain considered properly but the best expiry date for the vaccine is one year.

Background: Thimerosal (Merthiolate) is a well-known preservative used in pharmaceutical products, the safety of which was a matter of controversy for decades. Thimerosal is a mercury compound, and there is a debate as to whether Thimerosal exposure from vaccination can contribute to the incidence of mercury-driven disorders. To date, there is no consensus on Thimerosal safety in Vaccines. In 1977, a maximum safe dose of 200 μg/ml (0.5 mM) was recommended for Thimerosal by the WHO experts committee on biological standardization. Up-to-date guidelines, however, urge national control authorities to establish their own standards for the concentration of vaccine preservatives. We believe such safety limits must be studied at the cellular level first. The present study seeks a safe yet efficient dose of Thimerosal exposure for human and animal cells and control microorganism strains., Methods: The safety of Thimerosal exposure on cells was analyzed through an MTT cell toxicity assay. The viability of four cell types, including HepG2, C2C12, Vero Cells, and Peripheral blood mononuclear cells (PBMCs), was examined in the presence of different Thimerosal concentrations and the maximum tolerable dose (MTD) and the half maximal inhibitory concentration (IC50) values for each cell line were determined. The antimicrobial effectiveness of Thimerosal was evaluated on four control strains, including Pseudomonas aeruginosa, Staphylococcus aureus, Candida albicans, and Aspergillus brasiliensis, to obtain the minimum inhibitory concentration (MIC) of Thimerosal. The MIC test was performed in culture media and under optimal growth conditions of microorganisms in the presence of different Thimerosal concentrations., Results: The viability of all examined cell lines was suppressed entirely in the presence of 4.6 μg/ml (12.5 μM) of Thimerosal. The MTD for HepG2, C2C12, PBMC, and Vero cells was 2, 1.6, 1, and 0.29 μg/ml (5.5, 4.3, 2.7 and 0.8 μM), respectively. The IC50 of Thimerosal exposure for HepG2, C2C12, PBMC, and Vero cells was 2.62, 3.17, 1.27, and 0.86 μg/ml (7.1, 8.5, 3.5 and 2.4 μM), respectively. As for antimicrobial effectiveness, the growth capability of Candida albicans and Staphylococcus aureus was suppressed entirely in the presence of 6.25 µg/ml (17 μM) Thimerosal. The complete growth inhibition of Pseudomonas aeruginosa in culture media was achieved in 100 µg/ml (250 µM) Thimerosal concentration. This value was 12.5 µg/ml (30 μM) for Aspergillus brasiliensis., Conclusion: According to our results Thimerosal should be present in culture media at 100 μg/ml (250 µM) concentration to achieve an effective antimicrobial activity. We showed that this amount of Thimerosal is toxic for human and animal cells in vitro since the viability of all examined cell lines was suppressed in the presence of less than 5 μg/ml (12.5 μM) of Thimerosal. Overall, our study revealed Thimerosal was 333-fold more cytotoxic to human and animal cells as compared to bacterial and fungal cells. Our results promote more study on Thimerosal toxicity and its antimicrobial effectiveness to obtain more safe concentrations in biopharmaceuticals., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier GmbH. All rights reserved.)