Gemperline came to the notice of a larger scientific community in 1984 with the publication of a paper describing DISNET in the Journal of Automated Methods and Management in Chemistry. (The journal title was changed to Journal of Analytical Methods in Chemistry in 2013.) Gemperline and his colleagues provided methodologies which underlay the improvements in calibration accuracy, computer-based data acquisition and mathematical analysis in chemometrics. The qualitative advances helped open new scientific fields such as molecular modeling and QSAR, cheminformatics, the ‘-omics’ fields of genomics, proteomics, metabonomics and metabolomics, process modeling and process analytical technology.
Gemperline is a self-taught chemometrician, and he has been most influential by dispersing knowledge of his chemometric methodologies through his publications. Perhaps the best example of this is his book, Practical Guide to Chemometrics, for which he served as both editor and as a contributor. Better process methods can create inflection points that became the foundation for radical improvements and transitions in the scientific enterprise. It can be decades before the implications of particular advances are intuited, especially those related to the basic training of the next generations of chemists. His research in chemometrics has been "focused on development of new algorithms and software tools for analysis of multivariate spectroscopic measurements using pattern recognition methods, artificial neural networks, multivariate statistical methods, multivariate calibration, and non-linear model estimation." Sandia National Laboratories researcher, David Haaland, called Gemperline a "chemometrician extraordinaire," with a "deep understanding of chemometrics" and "wide-ranging" contributions to the research literature, and said after his first meeting with Gemperline he "always read his publications and attended his talks at conferences knowing that I would learn something new every time."
According to former graduate student, Patrick Cutler, another important aspect of Gemperlines research and teaching career has been his ability to collaborate with industry, which allowed "opportunities for students to gain invaluable experience." Cutler mentions pharmaceutical company Burroughs Wellcome in particular which at one time operated a production plant in Greenville, North Carolina, in close proximity to East Carolina University. Gemperline collaborated with Burroughs Wellcome in the 1980s to develop software for multivariate pattern recognition analysis of near-infrared reflectance spectra for rapid, non-destructive testing of pharmaceutical ingredients and products. His research and publications in this area furthered his international recognition. Cutler notes that Gemperline, "built his career and has produced world-renowned chemometrics research with the limited resources of mostly undergraduates and masters level students."
Gemperline also has had research collaborations with Pfizer, Inc. and GlaxoSmithKline and he has received "significant" funding for his research from the National Science Foundation (NSF) and the Measurement and Control Engineering Center (MCEC) at University of Tennessee, Knoxville, an NSF-sponsored Industry-University Cooperative Research Center (IUCRC). He collaborated with David Haaland at Sandia National Laboratories to develop chemometric tools to study kinetics in cells using hyperspectral fluorescence imaging. Their efforts were judged successful, and led to the 2010 Meggers Award for them and coauthors Patrick Cutler and Erik Andres for their two 2009 publications in Applied Spectroscopy about their research.
Gemperline served as the editor-in-chief of Journal of Chemometrics for ten years, from 2007 to 2017, and before that as the North American editor for five years, from 1996 to 2001.  The journal honored him with a special issue in July 2020. He joined the editorial advisory board of Spectroscopy Magazine in 2020.