Preparation and disinfection of the root canal aims to remove organic and inorganic tissue from within the root canal, reduce the number of microorganisms, neutralize endotoxins inside the dentin, and prepare the root canal for proper obturation. These aims can be achieved in many cases to a degree that promises a high chance of success through a combination of mechanical preparation and chemical disinfection—in other words, irrigation and medication. Preparation and disinfection both physically and chemically act on the lumen of the root canal itself, trying to remove as much infected material as possible, but also act on the surrounding dentin and all of its components including the dentinal tubules. The chemomechanical process attempts to remove the adhering biofilm as well as microorganisms and their by-products. It is important to investigate the effect of various treatment options on root canal dentin, the periodontal ligament, and the surrounding alveolar bone. When mechanical processes and chemical agents are applied to a substrate such as dentin, this implies that the dentin will be altered or modified in structure, composition, and/or physical properties. This review aims to summarize the present knowledge on the effects of mechanical instrumentation and chemical irrigation on the root canal dentin and surrounding tissues.