Plant cell walls contain a complex yet important polymer known as lignin. In addition to offering structural support, it acts as a natural barrier against microbial invasion and facilitates the transport of water and nutrients. Despite its abundance and nearly a century of efforts to transform it into valuable products, lignin remains insufficiently utilized in the majority of existing biorefineries. Lignin is primarily used to generate heat and power due to its complicated structure, which hinders its utilization. In the last decade, interest in lignin valorization has increased exponentially to decrease our dependence on nonrenewable energy resources. This chapter highlights the structural properties of lignin which make it a suitable candidate for biofuel. Also, the structural characterization of lignin using different analytical techniques like nuclear magnetic resonance (NMR) spectroscopy, ultraviolet-visible spectroscopy, Raman spectroscopy, and Fourier transform infrared spectroscopy (FTIR) will be explored in this chapter.