Short oligonucleotides are an emerging platform for the development of biopharmaceuticals to treat various diseases, including those that are inaccessible to small molecule- and protein-based approaches. Despite the increasing focus on short oligonucleotide therapeutics, standardized methods to rigorously characterize the structure of these molecules, which is essential for drug development and quality control purposes, are not yet fully established. High-resolution NMR spectroscopy is increasingly utilized to characterize the structure of protein-based therapeutics, including monoclonal antibodies, yet this technology has only sparingly been applied to short oligonucleotides. Recent advances in NMR hardware and methodologies allow for the rapid acquisition of NMR spectra under full formulation conditions and at natural isotopic abundance. Furthermore, the non-native chemistries added to enhance the pharmacological stability of short oligonucleotide therapeutics afford additional NMR-active handles of 19F and 31P, which, at 100 % natural isotopic abundance, are more sensitive than 13C (1.1 %) and 15N (0.36 %). Here we demonstrate the applicability of NMR methods to structurally “fingerprint” short oligonucleotide therapeutics to aid the biopharmaceutical industry and regulators during multiple phases of the clinical pipeline, including discovery, manufacturing, and quality control. The demonstration of NMR fingerprinting methods for short oligonucleotide therapeutics allows for the assessment of quality attributes at the atomic level, providing greater confidence in the overall structural integrity of the drug. This benchmarking study provides a pathway for the implementation of NMR fingerprinting as a high resolution structural technique to complement other analytical approaches used during drug development and quality. 

Learning Objectives:

1. Describe how NMR spectra act as structural “fingerprints” of short oligonucleotide therapeutics.

2. Break down how NMR methods can be applied to a variety of structural features, including both native and non-native chemistries. 

Recently, the use of the Analytical Quality by Design (AQbD) approach for method development has been increasingly accepted in the industrial, academic and regulatory circles. This is because it is a comprehensive, systematic, and risk-based approach that provides a broad knowledge about the impact of the critical chromatographic parameters on the method performance. The present work aims at employing the AQbD principles in developing an Ultra High Performance Liquid Chromatography-Mass Spectrometry method (UHPLC-MS) for the analysis of Budesonide, Formoterol, and related compounds in Symbicort®. The different AQbD method development stages including, screening, optimization, and verification will all be demonstrated. The advantages of using mass spectrometry for detecting and tracking the analytes of interest will also be presented and discussed.

Learning Objectives: 

1.    Explain the Analytical Quality by Design Principles.

2.    Discuss the benefits of using specialized software for rapid and robust method development.
 

Upon completion of the human genome project, it was determined that greater than 90% of human diseases are due not solely to a person’s genetics but a combination of genetic factors and non-genetic environmental influences. While genetic factors can be readily assessed using rapid genome sequencing technologies, measuring environmental factors is much more challenging. To date either direct or indirect measurements of exposure are often employed for their analysis. In direct measurements, specific xenobiotic compounds are analyzed in environmental samples or in biofluids and tissues, however, many xenobiotics are often excreted before responses even occur. Indirect analyses evaluate how biological processes change due to chemical exposure using one or more complementary omic techniques such as transcriptomics, proteomics, metabolomics or lipidomics. This presentation will demonstrate how combining liquid chromatography, ion mobility spectrometry and mass spectrometry (LC-IMS-MS) separations for direct xenobiotic measurements and indirect multi-omic evaluations enables an in-depth understanding of molecular responses occurring due to chemical exposures.

Learning Objectives:

1. Explain the power of multidimensional LC-IMS-MS platforms for untargeted analyses.

2. Explain the utility of multidimensional libraries for narrowing down potential feature identifications.

3. Describe LC-IMS-MS capabilities for improving the detection, identification and validation of endogenous and xenobiotic features.

Per- and polyfluoroalkyl substances (PFAS) are abundant in the environment as a result of the use and disposal PFAS containing products including aqueous film-forming foams, textiles, carpets, etc. Subsequently, PFAS are increasingly observed as environmental contaminants in groundwater, soil, sediment, and biota including humans. Routine monitoring methods, method development, and commercial analysis have therefore become more widely available for PFAS. The National Institute of Standards and Technology (NIST) provides a wide range of reference materials (RMs) that are homogeneous, well-characterized materials that are used to validate measurements and improve the quality of analytical data and methods. Currently NIST has ten different reference materials with concentration values of PFAS on the Certificate of Analysis; however, more PFAS reference materials are needed to support the needs of the analytical community. To support the creation of several new reference materials and to provide new quantitative measurements for PFAS on existing reference materials, new analytical methods were developed by liquid and gas chromatography – mass spectrometry (LC-MS/MS, GC-MS, and GC-MS/MS) for use on environmental samples. The current NIST efforts to fill this gap will be discussed. 

Learning Objectives:

1. Explain why per- and polyfluoroalkyl substances are environmentally emerging contaminants.

2. Discuss how the development of analytical methods by LC-MS/MS and GC-MS for PFAS can be used for analysis of environmental samples. 

Aroma is an important sensory factor in the perception of food.  But what exactly makes a food smell pungent, or savory, or foul?  The chemistry of aroma, or smell, can be very complex.  There are an estimated 10,000 volatile compounds across all foods that can contribute to the specific smell of a given food.  Yet only a few hundred determine the aroma of the food we eat.

In this study, we developed a novel approach to identifying aroma compounds in foods.  Using a combination of GCMS and GC-SCD (sulfur chemiluminescence detector) along with employing metabolomic techniques, we were able to identify and quantitate key aroma compounds in various foods and beverages.  Samples include several aromatic beverages (beer, wine, spirits, and coffee) and foods (kimchi, soy sauce, and cheese).  

Learning Objectives:

1. Identify the individual compounds that comprise the smell of foods.

2. Understand the analytical tools used to identify those compounds.

3. Learn about different approaches to measuring food aroma.

Aflatoxins are carcinogenic mycotoxins that have adverse health effects on both humans and animals consuming contaminated food and feed, respectively. A method has been developed for the highly sensitive and selective determination of regulated aflatoxins in a wide range of commodities. The extraction of aflatoxins from representative commodities of interest (corn, wheat, oats, red chili, black pepper, cocoa and a traditional Chinese medicine) was performed using liquid-liquid extraction and then immunoaffinity column clean-up on the new AflaTest WB SR+ column. Chromatographic separation was demonstrated using both HPLC (Alliance) and UPLC (ACQUITY UPLC H-Class PLUS) platforms, using fluorescence detection, supported with post-column derivatization and large flow cell, respectively. The performance of the method was evaluated through replicate analysis of spiked test portions of seven different matrices. Overall recovery was shown to be excellent, between 93% and 116%, with relative standard deviations lower than 8%. The method was found to be specific as no interference peaks were observed for blank samples. The method has been demonstrated as suitable for monitoring compliance with regulatory limits set for aflatoxins in food commodities globally.

Learning Objectives:

1.    Describe the importance to test for mycotoxins.

2.    Explain the difficulties encountered when analyzing complex matrices.

3.    Break down what solution(s) does Waters-VICAM offer for Aflatoxin testing in complex matrices.

Recently, a novel hybrid surface technology (HST) has been developed to mitigate metal analyte interactions in liquid chromatography. We have evaluated the performance of instruments and columns that incorporate the HST for simultaneous analysis of multiple B vitamins by ultra-high performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) and compared the results with those obtained using the conventional stainless-steel components. We demonstrate significantly improved performance in analyte peak area, limit of quantification, and carry-over using the HST in the analysis of multiple B vitamins. Not only for the phosphorylated B vitamins, but also for the non-phosphorylated B vitamins, significantly improved performance was observed. Our results suggest that this novel HST brings a clear advantage over the conventional stainless-steel in the UPLC-MS/MS analysis of B vitamins. This novel HST has a high potential to improve the LC based analysis for a wide range of compounds. 

Learning objectives:

1. Understand the main issues with the stainless-steel surface in liquid chromatography.

2. What is the new hybrid surface technology?

3. What are the main benefits of using this new HST for B group vitamins analysis?

Cannabis & Hemp pesticide and mycotoxin testing is difficult and time-consuming for a compliance testing laboratory. With dedicated fully automated cannabis & hemp flower workflows designed specifically for pesticide & mycotoxin residue testing, automation handling solutions enable a cannabis lab to minimize errors, reduce hands-on time, and increase throughput and reproducibility. This poster will present new data and material to show the efficiency gains (ie. more samples per hour), the sensitivity of the analysis to low ppb, the benefit of adding sample preparation automation to reduce labor requirements, and software integration solutions to integrate data into the laboratory LIMs efficiently. References will show that having a verifiable method, and protocol, for the cannabis and hemp testing industry is key in ensuring the highest quality of cannabis and hemp product reaches the patient or consumer. In addition, the influence of the cannabis sample type tested will be discussed as the sample type matrix (flower, extracts, and edibles) has considerable effects on the data integrity.

Learning Objectives:

1. Explain what preparation steps in pesticide testing are prone to experimental error and can cause data quality issues.

2. Discuss how quality control standards and internal standards can be used to improve data quality for challenging matrices such as food and cannabis.