In this thesis I have studied the fragmentation of biological important molecules with soft ionization methods. Through this thesis I studied and analyzed the fragmentation pattern and how they defer depending on the alcohol solvent used. Then I went deeper into understanding the fragmentation pattern of different molecules and how they defer from each other using ESI as the fragmentation technique to better understand the factors that cause this differences. Finally I have analyzed and compared the fragmentation pathway in ESI and APCI and the Fragmentation pathway in Positive and Negative ion mode.