https://doi.org/10.47108/jidhealth.Vol5.Iss3.233 Mohammad FK., Journal of Ideas in Health 2022;5(3):725-726 © The Author(s). 2022 This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (https://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article unless otherwise stated. e ISSN: 2645-9248 Journal homepage: www.jidhealth.com Open Access Calculation of 95% confidence interval of the median lethal dose determined by the up-and-down procedure: a letter to editor Fouad Kasim Mohammad 1* Dear Editor; Acute toxicity of chemicals can be detected by the widely advocated up-and-down procedure (UDP) used to measure the median lethal dose (LD50) in laboratory animals [1,2]. A recent article by Zhang et al. [3] reliably introduced an improved UPD (iUDP) to measure the LD50 values of substances in animals. In an innovative way, the iUDP procedure was presented in a suitable manner to include the 95% confidence intervals (CI) for the LD50 values using the equation: 95% confidence interval = LD50 + the standard error (SE) SE = SD x √2/N SD is the standard deviation of all dosages in N trials in animals. However, not all the reports using the UDP report 95% CI; some studies use elaborate software programming to calculate the CI [1,3,4]. Others have tried calculating the CI with the arithmetic means (as an LD50) + standard error of the doses used in the UDP [5]. To further benefit from the iUDP and its simple way of calculating the CI [3], here we suggest an additional input for researchers in toxicology by using the above-mentioned equations to calculate the 95% CI for UDP using the table of the maximum likelihood estimation of LD50-the section with a standard error of 0.61 of Dixon [2]. This is because this version of UDP is still being used, and reports are still being published without mentioning the CI. In this context and for demonstration purposes, we recalculated selected LD50 examples from published literature [6-9] and included their 95% CI (Table 1) according to Zhang et al. [3]. Table 1: Acute (24 h) median lethal doses (LD50) and their 95% confidence intervals (C.I.) of drugs or toxicants administered in laboratory animals Reference Animal Drug/ Toxicant doses Dose interval Response X: dead O: alive LD50 Mean + SD C.I. = LD50 + SE Naser and Mohammad [6] Chicks Propofol 100, 80, 60, 80, 60, 40, i.p. 20 XXOXXO 57.22 70.0 + 20.98 45.11, 69.33 Mohammad et al. [7] Chicks Diazinon 15, 10, 5, 10, 5, 10, orally 5 XXOXOX 6.32 9.17 + 3.76 4.15, 8.49 Al-Baggou and Mohammad [8] Mice Cadmium chloride 10, 8, 10, 8, 10, i.p. 2 XOXOX 8.6 9.2 + 1.1 7.9, 9.3 Mohammad et al. [9] Mice Tetramisole 50, 60, 50, 60, 50, s.c. 10 OXOXO 57.0 54.0 + 5.48 53.5, 60.5 All doses are in mg/kg of body weight; i.p.: intraperitoneal injection; s.c.: subcutaneous injection; SD: standard deviation; SE: standard error (= SD x √2/number of animals) calculated for the 95% C.I. (= LD50 + SE) [3]. LD50 was determined by the up- and-down procedure, and the table of the maximum likelihood estimation of LD50-the section with a standard error of 0.61 of Dixon [2] was used for this purpose. Abbreviations UDP: Up-and-Down Procedure; LD50: Median Lethal Dose; iUDP: improved UPD; CI: Confidence Intervals ___________________________________________________ fouadmohammad@yahoo.com 1Department of Physiology, Biochemistry and Pharmacology, College of Veterinary Medicine, University of Mosul, Mosul, Iraq. Keywords: Confidence Interval, LD50, Median Lethal Dose, Up-and- Down Procedure https://doi.org/10.47108/jidhealth.Vol5.Iss3.233 http://www.jidhealth.com/ Mohammad FK., Journal of Ideas in Health 2022;5(3):725-726 726 Declarations This is a correspondence, and no experimental work has been done for this letter. Acknowledgment None Funding The author received no financial support for the research, authorship, and/or publication of this article. Availability of data and materials Data will be available by emailing fouadmohammad@yahoo.com. Authors’ contributions Fouad Kasim Mohammad (FKM) is the principal investigator of this manuscript (Letter to Editor). FKM is responsible for the study concept, design, writing, reviewing, editing, and approving the manuscript in its final form. FKM read and approved the final manuscript. Ethics approval and consent to participate The corresponding Author conducted the research following the Declaration of Helsinki; however, Letter to Editor Article needs no ethics committee approval. Consent for publication Not applicable Competing interest The author declares that he has no competing interests. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article unless otherwise stated. Author Details 1Department of Physiology, Biochemistry and Pharmacology, College of Veterinary Medicine, University of Mosul, Mosul, Iraq. Article Info Received: 19 June 2022 Accepted: 27 July 2020 Published: 20 August 2022 References 1. Rispin A, Farrar D, Margosches E, Gupta K, Stitzel K, Carr G, Greene M, Meyer W, McCall D. Alternative methods for the median lethal dose (LD (50)) test: the up-and-down procedure for acute oral toxicity. ILAR J. 2002;43(4):233- 43. doi: 10.1093/ilar.43.4.233. 2. Dixon WJ. Efficient analysis of experimental observations. Annu Rev Pharmacol Toxicol. 1980; 20:441-62. doi: 10.1146/annurev.pa.20.040180.002301. 3. Zhang YY, Huang YF, Liang J, Zhou H. Improved up-and- down procedure for acute toxicity measurement with reliable LD50 verified by typical toxic alkaloids and modified Karber method. BMC Pharmacol Toxicol. 2022 Jan 4;23(1):3. doi: 10.1186/s40360-021-00541-7. PMID: 34983670; PMCID: PMC8725450. 4. Manage A, Petrikovics I. Confidence limit calculation for antidotal potency ratio derived from lethal dose 50. World J Methodol. 2013 Mar 26;3(1):7-10. doi: 10.5662/wjm.v3.i1.7. PMID: 25237618; PMCID: PMC4145567. 5. Ahur VM, Anika SM, Onyeyili PA. Age-sex dimorphisms in the estimation of median lethal dose (LD50) of lead diacetate in rabbits using up-and-down procedure (Arithmetic method). Sokoto J Vet Sci.2018; 16(4):64-72. https://www.ajol.info/index.php/sokjvs/article/view/18303 3 6. Naser AS, Mohammad FK. Central depressant effects and toxicity of propofol in chicks. Toxicol Rep. 2014 Aug 13; 1:562-568. doi: 10.1016/j.toxrep.2014.08.003. 7. Mohammad FK, Al-Badrany YM, Al-Jobory MM. Acute toxicity and cholinesterase inhibition in chicks dosed orally with organophosphate insecticides. Arh Hig Rada Toksikol. 2008 Sep;59(3):145-51. doi: 10.2478/10004-1254-59- 2008-1873. 8. Al-Baggou BK, Mohammad FK. Effects of cadmium on the acute toxicity of cholinesterase inhibiting insecticides in mice. Arabian J Sci Res. 2022. (In press). 9. Mohammad FK, Faris GA, Rhayma MS, Ahmed K. Neurobehavioral effects of tetramisole in mice. Neurotoxicology. 2006 Mar;27(2):153-7. doi: 10.1016/j.neuro.2005.08.003.