Enough Is Enough—How Many Replicates Are Needed for Non-Targeted Analysis with LC/HR-MS?

Description:
Most analytical methods used for food safety applications target specific compounds or compound classes. While this is crucial, any analytes that are not included as part of these routine screening methods will not be detected or identified. A more global analysis approach can be taken with liquid chromatography/high-resolution mass spectrometry, where unknown compounds can be detected and identified without focusing on a particular compound class. Because food is a chemically-rich sample matrix, this technique can generate data sets with thousands of detected compounds for each sample; however, it can be challenging to determine if compounds are reliably detected and extracted for each data file. Replicates can help in this regard, but to our knowledge, no studies have focused on how many samples should be prepared and/or analyzed to obtain reliable results. We designed an experiment where we prepared five replicates and analyzed each of these five times to determine the best balance of found molecular features (including spiked compounds), time to prepare samples, and time to analyze data files. We began by analyzing a strawberry composite market basket sample from the FDA Total Diet Study and plan on analyzing other matrices to cover different regions of the AOAC food triangle. Furthermore, one of the goals of non-targeted data analysis software is to find all (or as many as possible) molecular features in a sample, which is more challenging than searching for defined molecular targets. Because of this, many software applications recursively search all data files for features found in each individual data file in the same analysis queue. As a result, more features are found as more data files are processed together. While beneficial, we want to determine how many samples should be processed together to obtain high-quality data. Tradeoffs will be discussed, as well as practical guidance for future experimental design.

Speaker: Ann Knolhoff - U.S. Food and Drug Administration
Dr. Ann Knolhoff is currently acting as a Team Lead for the Spectroscopy and Mass Spectrometry Branch at the Food and Drug Administration (FDA) in the Center for Food Safety and Applied Nutrition. She received her B.Sc. in Chemistry at Truman State University and her Ph.D. in Chemistry at the University of Illinois at Urbana-Champaign under the direction of Professor Jonathan Sweedler. Dr. Knolhoff has been at the FDA since 2011, first as a post-doctoral fellow and then transitioning to a Research Chemist. Her research has primarily focused on the development of non-targeted screening approaches using liquid chromatography and high-resolution mass spectrometry for food safety applications.