Poor oral cavity hygiene is the major cause of the many dental related health issues. In order to develop efficient oral cavity cleaning products, there should be enough scientific information about the oral microbial diversity in bio-film. Such scientific data is not available here for Nepalese population. Due to various geographic variations and multiple ethnic populations with multiple cultures and living customs, different oral microbial composition must be obtained. In the present research project, we isolated 49 different species belonging to 19 different genera of oral microbial diversity in Nepalese population from various altitudinal locations, ethnic groups, and various age groups. In addition, genetic variation of microbial species with respect to geographical location and their biochemical and physiological variation were also been investigated. We also investigated 140 different compounds secreted by different microbial flora in the oral bio-film. Similarly, microbial enzyme production were also be explored. The co-relation between the oral disease and microbial diversity along with chemical composition of bio-film and extracellular enzymes secreted by microbial species will be discussed. The overall scientific data has paved the way to describe the distribution, pathogenic potential, physiology and genetic variation of pathogenic oral micro-organisms in Nepalese communities. Such information will be useful to design and produce proper and effective commercial oral cleaning products.