The environmental destruction due to human activities in the current era has become the focus of society, governments and manufactures. Since transport has a negative impact on the environment it has become one of the areas with considerable potential within this scope. The proactive approach to address these negative environmental impacts from transport is named as Green Freight practices. The motivation to adopt green practices is the ability to use it as a differentiation strategy. This research was conducted to identify the relationship between Green freight transport practices and differentiation strategy. The primary objective of this study was to analyse the relationship between green freight transportation practices and differentiation strategies of an organization in order to gain competitive advantage. As the sample of the study initially seventy five companies were selected in Colombo district. The sample was selected based on the convenience sampling method as it was difficult to obtain data from certain organizations. This study focused on freight transport organizations where green activities already exist. Primary data was obtained through a questionnaire survey and interviews with the executives and the management staff of organizations. Twelve green freight transport practices were considered. SPSS was used for data analysis. The correlation matrix was derived during the analysis. The findings state that there is a positive relationship between six of the green practices out of the twelve with differentiation strategy. The six practices are transporting in bulk packages, direct transportation, using of trained drivers, avoiding congestion hours, using a transport time table and using multiple modes to transport freight. Transporting goods in bulk packages, direct transportation, use of trained drivers and using a transport time table have significantly strong positive correlations with differentiation (at the significance level of 90 percent).Therefore it can be concluded that the implementation of practices such as transporting in bulk packages, direct transportation, using of trained drivers and using a transport time table contributes to enhance the differentiation of freight transporters.