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Author ORCID Identifier



Open Access Dissertation

Document Type


Degree Name

Doctor of Philosophy (PhD)

Degree Program


Year Degree Awarded


Month Degree Awarded


First Advisor

Igor Kaltashov

Second Advisor

Richard Vachet

Third Advisor

Anne Mason

Fourth Advisor

Daniel Hebert

Subject Categories

Analytical Chemistry


Biotherapeutics consist of biopolymers (proteins, polysaccharides, DNA, and RNA) that are used to treat a wide range of conditions from cancer to autoimmune disease to enzyme replacement. In recent years biotherapeutics have experience tremendous growth due to advances in technology and our understanding of human biology. They are very important to modern medicine due to their ability to treat diseases which are unable to be treated with small molecule-based drugs. Unlike small molecule drugs which are synthetically produced, biotherapeutics are expressed inside cells. Produced biotherapeutics are not made up of a single homogenous population but instead a population of highly similar variants. The source of these variations are enzymatic and non-enzymatic post-translational modifications. By characterizing these modification, a profile is built that links the in vivo response of a drug to its modifications. The complexity and size of these biopolymers makes their characterization very challenging and demands the development of robust analytical techniques. Mass spectrometry- and liquid chromatography-base methods are an integral part of protein characterization. Mass spectrometry provides accurate mass measurements that are invaluable for confirming the identity of a protein and any modifications. Additionally, mass spectrometry is used to assess a protein’s higher order structure. Liquid chromatography is a very powerful tool that allows for different post-translational modified populations of a biotherapeutic sample to separate by their chemical or physical properties. Separated populations can further be characterized to identify and analyze their chemical or structural composition. The presented work utilized a blend of mass spectrometry and liquid chromatography methods to characterize proteins with biotherapeutic potential.