Petroleum is a complex mixture of hydrocarbons that exist naturally in gaseous (natural gas), liquid (crude oil), and solid (asphalt) states. It is derived from a variety of organic materials that are chemically converted over long periods of time (hundreds of millions of years) under different geological and thermal conditions. Crude oil is composed mainly of carbon and hydrocarbon, but also minor amounts of sulfur, oxygen, and nitrogen as well as trace amounts of metals are present. Refined petroleum products are fractions derived by distillation from crude oil. Thus, due to variations in crude oil feed stocks and in the refining process individual oil samples have unique chemical fingerprints, which provide a basis for distinguishing oils and identifying the source of spilled oil. Biological markers or biomarkers are one of the most important hydrocarbon groups in petroleum for chemical fingerprinting. They are complex molecules derived from formerly living organisms. Biomarkers are useful for chemical fingerprinting because they retain all or most of the original carbon skeleton of the original natural product, and this structural similarity reveals more information about oil source than do other compounds in oil. This chapter focuses on biomarker chemistry, biomarker genesis, overview of analytical methodologies for biomarker separation and analysis, identification of biomarkers, biomarker distributions in crude oils and various petroleum products, and sesqueterpane and diamondoid biomarkers in oils and lighter petroleum products.