Nitrile gloves are commonly used in Central Sterile Services Department in order to be often used when manually cleaning medical equipment such as personal protective equipment. As a result they are resistant to a wide range of chemicals and germicides, including alcohols, peroxides, and other sterilizing agents. In addition to providing chemical protection, nitrile gloves are resistant to punctures and tears, which is important when working with sharp medical instruments. This research focuses on the efficacy of gloves in relation to the multi-chemicals or compounds of nitrile gloves and observation at the point of use to remove soil and contact with chemicals on medical devices at the Central Sterile Supply Department. The powder-free nitrile exam gloves and extended cuff methods of chemical analysis of permeation testing of standard practice for assessment of resistance of medical gloves to permeation by chemotherapy drugs (ASTM D 6978-05), EN455 Part 1: 2000 were used to test the test method for single-use medical gloves. The standard of protective gloves against dangerous chemicals and microorganisms was tested by EN 420:2003 + A1:2009, EN388: 2016, ISO 374-1:2016, and ISO 374-5:2016. The testing of chemotherapy drugs and UV/NIS absorption spectrometry were used to measure the absorbance of test chemicals, which permeated through the specimens into the collection medium. The collection medium was circulated in a closed loop at an 11 ml/minute flow rate through the testing period. As a result, a glove user must protect against the specific chemicals or chemical mixtures to which they were exposed while manually cleaning medical devices. The physical properties of the gloves were evaluated by visual observations at the point of use for soil removal and chemical exposure to medical devices at the Central Sterile Supply Department of five hospitals. The results showed that nitrile gloves protected chemical exposure during manual cleaning of medical equipment at the point of use.