Electronic Thesis and Dissertation Repository

Thesis Format

Monograph

Degree

Master of Engineering Science

Program

Chemical and Biochemical Engineering

Supervisor

Rehmann Lars

Abstract

Disinfectant screening is an important step for manufacturing and approval of disinfectants. Screening helps identify the effectiveness, target microbe, minimum inhibitory concentration, and contact time. A standard screening test must be able to assess these parameters effectively over a broad range of formulations and microbes. Currently, there is no unified method to analyze disinfectants, a standard test is selected based on the application. This project aimed to identify a standard method that can be used over a broad range of microbes irrespective of disinfectant composition. Microbicidal activity of Hydrogen peroxide and Quaternary ammonium-based disinfectants was evaluated by Fluorescein diacetate (FDA), Methyl tetrazolium (MTT), and growth assays. M. terrae and C. albicans pellets were formed in 96 well plates. In comparison to FDA and MTT assay, growth assay was more accurate and had a larger range. It required no chemical reagents, thus eliminating interaction with disinfectant components. Therefore, growth assay can potentially be used as a unified method for disinfectant studies. Further tests on a broader range of microbes and field studies have to be conducted to confirm this claim.

Summary for Lay Audience

Disinfectant testing is used to estimate the potency of a formulation. Manufacturers use these methods while formulating and optimizing disinfectants. While developing a successful disinfectant formulation, the composition is varied several times. However, the current testing methods are specific to certain chemicals and microbes. Changing the method of analysis every time a new component is added is time-consuming and expensive. A method that can be used for all disinfectant samples across a broad range of microorganisms would be more advantageous. This project aims to identify this method.

Two disinfectants, E2O (hydrogen peroxide-based) and Germicide (Quaternary ammonium-based) were analyzed using three viability assays namely, Fluorescein diacetate (FDA), 3-(4,5-dimethyl-2-thiazolyl)-2-5-diphenyl-2H-tetrazolium bromide (MTT), and growth viability assays. Viability assays determine the effect of compounds by measuring cell activity or cell death. Mycobacterium terrae and Candida albicans were used as model microbes. FDA assay worked for E2O however, it couldn’t analyze Germicide due to the interaction of its components with Fluorescein diacetate dye used in FDA analysis. MTT assay worked for all samples but, it had a limited range, and the results were not reliable for E2O samples. Growth assay successfully analyzed C. albicans activity in both the disinfectants. However, it did not work for M. terrae.

A harmonized screening tool is required due to varying global regulatory requirements, aid selection and comparison of disinfectant, Growth assay is simple, reliable, and requires no chemical reagents. Therefore, it could be used to analyze disinfectants irrespective of their components by manufacturing industries and researchers. However, further tests on a wider range of disinfectants and microbes have to be performed.

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