Establishing industry QA/QC best practices for global metabolomics
3 Ways Machine Learning Propels Metabolomics at Metabolon
Metabolon and DNA Genotek: Collaborating to Advance Functional Understanding of the Microbiome
For a Successful Metabolomics Study, Don’t Risk Going it Alone
New Hope for Patients with Inborn Errors of Metabolism and Rare Diseases
Sometimes, there is no playbook. That’s often the case with inborn errors of metabolism (IEM), defined as rare genetic disorders that cause alterations or deficiencies in enzymes involved in metabolism, and other rare diseases. A patient may exhibit clinical signs that don’t map to specific disorders, or a screening panel may come back with no clues. In these frustrating situations, clinical researchers – and patients – need more places to turn for answers.
Applying metabolomics to clinical samples can help screening efforts by vastly expanding the information available to clinical researchers. When used in a clinical setting, metabolomics can measure small molecules in large volumes to identify potential biological pathways that are disturbed in diseased patients. While whole-exome sequencing has provided a valuable systems-based approach in the clinical laboratory, a recently published paper describes a clinical metabolomics approach to drive precision medicine forward, as a method for the screening of metabolic diseases through the analysis of a multi-pronged mass spectrometry platform. In the paper, authored by a team of 11 Metabolon scientists featured in the March 2020 issue of The Journal of Applied Laboratory Medicine, the authors assert that by simultaneously measuring hundreds of metabolites in a single sample, clinical metabolomics offers a comprehensive approach to identify metabolic perturbations across multiple biochemical pathways.

To arrive at their conclusions, the authors performed a single- and multi-day precision study on hundreds of metabolites in human plasma on four, multi-arm, high-throughput metabolomics platforms. This resulted in the reproducibility of the method for the measurement of key IEM metabolites in patient samples across multiple analytical batches, proving the method to be robust and reproducible for the screening of patients with previously undiagnosed inborn errors of metabolism.
Clinical metabolomics has the potential to make precision medicine a reality delivering transformational medicine. Traditional clinical testing of patient samples involves analyzing and measuring individual analytes or biomarkers. In the case of IEM’s, the screening method typically includes three panels, which measure in the range of 50-60 small molecules. Metabolon’s application of clinical metabolomics extends the number of biomarkers that can be measured significantly. Instead of 50-60, we can identify more than 500 small molecules, giving researchers a broader range of biomarkers to mine for insights.
The application of metabolomics is exceptionally significant for IEM and rare disease detection. There are hundreds of IEM diseases, and often the clinical manifestations of those diseases are vague, or the metabolic pathway that is disturbed is not immediately apparent. Applying global, or untargeted, analytical approaches, where prior knowledge of the affected metabolic pathway is not required, provides advantages over targeted analytical approaches and can extend the diagnostic potential of IEM screens beyond the 34-58 disorders that are currently evaluated in newborn screening programs, especially in complex cases. That makes throwing out this wide net to measure as many biomarkers as possible invaluable in a clinical setting.
When evaluating large quantities of information, it is paramount to make data quality a top priority. A critical step in drawing insights from untargeted metabolomics is accurate metabolite identification. Metabolon’s Precision Metabolomics™ workflow delivers Tier 1-2 identifications for detected metabolites and uniquely provides high confidence in the identification of compounds. By contrast, most metabolomic practitioners operate primarily with annotations that only meet the standards of Tiers 3-5, where some unique features are identified. Still, there is low confidence in confirming the metabolite. Data quality is extremely important to Metabolon, with exceptionally high standards applied to our work in clinical metabolomics. From start to finish, we emphasize quality control measures and checks and balances.
Metabolon’s method of clinical metabolomics is validated by Clinical Laboratory Improvement Amendments (CLIA), a regulatory standard for all testing of patient samples. Metabolon is ISO 9001: 2015 certified for analytical and diagnostic testing of biological specimens and accredited by the College of American Pathologists for diagnostic testing on human specimens. Additionally, Metabolon has a New York State Department of Health Clinical Laboratory permit to perform Quantose IR and IGT testing for the identification of insulin resistance and impaired glucose tolerance under the categories of Clinical Chemistry and Endocrinology.
In addition to the validation work we do to demonstrate the properties of the method, such as precision and accuracy, we also run the method under very specific quality control guidelines, which are even stricter than our research guidelines, including running batch controls for each of the analytic batches. We start by setting up the assays following policies that dictate how to establish a reference population, how you qualify the normalizing matrix, and so on. The typical regulatory requirements for review of the results is exceeded, with an additional analytical and lab director review in addition to our typical sub-level review. All of these steps ensure data quality and accurate insights.
Applying our expertise to clinical metabolomics is just one example of how Metabolon reveals biological insights otherwise unseen through other technologies by leveraging our proprietary discovery platform and one of the world’s richest and most diverse patient data sets. We are unique in our ability to receive samples, process them under a clinically validated metabolomics platform that’s controlled for clinical testing, and provide impactful results. Expanding the capabilities of metabolomics to the clinical setting has enormous potential for researchers and patients alike.
Curious about how metabolomics could enrich your study? Contact us today at hello@metabolon.com to learn more.
Delivering Clinical Insights with Targeted Biomarker Assays
Metabolomics reveals biological insights otherwise unseen, making it a crucial component of drug development. For a successful metabolomics study, both small molecule discovery and the ability to dig deeper into specific biomarkers of interest are needed to uncover actionable insights that propel new therapeutic developments. A specific combination of technology and expertise is required to identify these biomarkers of interest and develop assays that are sensitive enough to explore them fully.
Metabolomics reveals biological insights otherwise unseen, making it a crucial component of drug development. For a successful metabolomics study, both small molecule discovery and the ability to dig deeper into specific biomarkers of interest are needed to uncover actionable insights that propel new therapeutic developments. A specific combination of technology and expertise is required to identify these biomarkers of interest and develop assays that are sensitive enough to explore them fully.
At Metabolon, we understand the crucial role targeted small molecule assays play in drug development, and we’ve established best-in-class expertise. Metabolon has developed quantitative assays for more than 900 biochemicals in over 20 different matrices, including more than 150 qualified and/or validated targeted biochemical assays for pharmaceutical and biotechnology research and development. These assays focus on specific metabolites or metabolic pathways and can be used to track biomarkers and enhance biological understanding across preclinical and clinical research.
Our approach speaks directly to the questions biopharma research directors are seeking to answer, such as understanding their therapeutic’s mechanism of action, establishing pharmacodynamics, gauging safety and efficacy, defining patient segmentation, and conducting post-market surveillance. Once the critical biomarkers that are most important to researchers are identified through discovery, a more precise and accurate quantitation of the biomarkers is sought. Metabolon’s strategic services offer biopharma clients global discovery followed with a direct targeted approach to dig deeper into promising and relvant biomarkers.
Metabolon’s targeted assay program is uniquely situated to fit into this workflow.
Targeted biomarker assay development is a natural progression following discovery using Metabolon’s first-in-class metabolomics platform. Metabolon’s discovery platform uses statistical approaches to assess the relative quantity of thousands of biomarkers. Our targeted assay program picks up from there, taking the discovery work and developing a more precise, accurate measurement of those few molecules that are of most interest. We apply LC-MS/MS based methods to determine absolute quantities of important biomarkers, using labeled internal standards as well as calibrators.
Our expertise in the measurement of endogenous biomarkers is unique and second to none. Most bioanalytical services offer expertise in exogenous biomarker measurements, such as the drug of interest or compounds that are exogenously introduced into a system. Metabolon targeted assays measure endogenous biomarkers naturally found at various levels within biological systems. Since endogenous biomarkers are sometimes present at the lower limits of analytical detection, the sensitivity of the assay becomes paramount. Furthermore, modest changes in biomarker levels may have biological relevance; Metabolon’s highly accurate targeted assays are required to tease out even modest changes in analyte levels. Our team of Ph.D. level method development scientists have a degree of experience in endogenous biomarker assay development that few places, if any, in the world can match. Our scientists are highly efficient and experienced in endogenous small molecule analysis, bringing additional data and actionable insights to drug development and clinical trials. No other bioanalytical company can translate metabolomic data and biomarker discovery into valuable biological insight using targeted LC-MS/MS analysis in the way that we can.
Finally, Metabolon’s capability is unique because we can operate under a research or clinical setting. In addition to our targeted work meeting specific client study demands, we also develop assays specifically used in clinical testing. Some of the pre-developed targeted assays offered by Metabolon include:
- Fatty acid metabolism panel
- Short-chain fatty acid panel
- Bile acid panel
- Cholesterol metabolism panel
- Stratum corneum panel
- Sebum lipid panel
- accuGFR™ panel (to estimate glomerular filtration rate)
- 7 alpha-hydroxy-4-cholesten-3-one (C4) panel
- Metal ion panel
LC-MS/MS-based assays can be customized based on client needs and developed under the quality system for research use only (RUO) or validated under Good Clinical Practice (GCP) and Clinical Laboratory Improvement Amendments (CLIA). Metabolon is ISO 9001: 2015 certified for analytical and diagnostic testing of biological specimens and accredited by the College of American Pathologists for diagnostic testing on human specimens.
Nowhere else will you find a laboratory like Metabolon’s that combines a world-class metabolomics discovery platform with a highly sophisticated analytical chemistry laboratory doing targeted analysis the way we do it. We have spent nearly 20 years building the best metabolomics competency in world to enable, accelerate and support drug development through biomarker discovery, understanding mechanism of action, patient stratification and more.
We proudly cover the whole breadth of the biopharma pipeline, from discovery to targeted analysis to clinical trial support, because your success is our goal.
Is your study yielding all the insights it could be? Download our free infographic to learn more about the five risks of drug development and how you can avoid them with metabolomics, and contact us today at hello@metabolon.com to get started.
Front lines of COVID-19
Metabolon scientists are collaborating with researchers on world-leading programs to use multi-omic deep phenotyping technologies, including metabolomics, to advance human health. A primary focus right now is on improving our collective understanding of the SARS-CoV-2 virus and the COVID-19 disease during this unprecedented pandemic.
Metabolon scientists are collaborating with researchers on world-leading programs to use multi-omic deep phenotyping technologies, including metabolomics, to advance human health. A primary focus right now is improving our collective understanding of the SARS-CoV-2 virus and the COVID-19 disease during this unprecedented pandemic. Metabolon has a strong history of working closely with scientists at the Institute for Systems Biology (ISB) like Dr. James R. Heath, Dr. Leroy Hood, Dr. Andrew Magis, Dr. Nathan Price, Dr. John Earls and Dr. Sean Gibbons.
Now, we are teaming up with ISB again to understand and treat coronavirus. Longitudinal blood samples are being collected from patients with varying COVID-19 disease severity to track their health over time. One goal for the project is to understand better which patients are at the highest risk of severe infection so that we can most effectively deploy medical resources. The work will also help to develop a deeper understanding of effective immune and organ system responses during the onset and recovery of affected patients.
The findings will be vital in helping to treat patients effectively and design optimal vaccines and therapies in response to viral outbreaks in the future. Similar to the SARs outbreak in 2003, this important work will also help us understand if there are any long-term health implications for those that recover from COVID-19.
Previous studies of viral infections suggest that patients’ metabolic profiles become altered during disease progression. These metabolic changes influence how a patient responds both through the degree of immune response and clearance of the virus from the lungs and other organs. Even in the early stages of the most recent COVID-19 pandemic, it was clear that pre-existing conditions such as coronary heart disease, diabetes, asthma, and hypertension have an impact on outcomes. As the virus has spread globally, no clear patterns of effects with age, gender and these pre-existing conditions have emerged. Given that therapeutic options are limited, and vaccine development remains in progress, there remain gaps in our understanding of the mechanisms of the virus to host interactions and the trajectory of the infection cycle that metabolomics studies looking across bodily systems will help reveal.
Metabolon is the industry leader in the ability to reveal metabolic perturbations across all biochemical pathways including amino acids, carbohydrates, lipids, nucleotides, microbiota metabolism, energy, cofactors and vitamins, xenobiotics, and novel metabolites. Given that metabolism represents the integration of genetic and non-genetic factors such as microbiome and lifestyle, it is a uniquely poised modality to assess health outcomes and severity of the disease, particularly relevant to understanding the trajectory of COVID-19 infection. Within this context, Metabolon has been given special dispensation by the state of North Carolina to continue all laboratory activity during this crisis due to our capacity to analyze human samples under both clinical and research environments. We are focused on maintaining our high levels of service while protecting our staff and maintaining the physical distancing protocols. We are committed to fast track mission-critical projects linked to COVID-19 research to support this vital work.
Our dedicated team of Study Directors, including experts with years of deep, infectious disease and drug development experience, are available now to address your questions, lead metabolomics study design and draft final biological interpretation reports. Please contact us today via metabolon.com/contact-us to learn how our experts may be able to support expediting your COVID-19 research.
References:
Wang, T et al. Comorbidities and multi-organ injuries in the treatment of COVID-19. The Lancet. 2020;395(10228): e52.
Wu et al. Altered lipid metabolism in recovered SARS patients twelve years after infection. Scientific Reports. 2017:7(9110).
Metabolon: A Rich Past and A Promising Future
When Annie Evans, Ph.D., first joined Metabolon, she had no idea of the breakthroughs she would be a part of, and how metabolomics would grow into an important ‘omics technology. “We’re impacting areas of the life sciences that I never thought possible,” she noted. After completing her Doctorate in Biological Mass Spectrometry at the University of Virginia, she joined Metabolon in 2004 as a research scientist. Early on, she was praised for her straightforward nature and presentation style, two things that played a crucial role in the development of the Metabolon we know now. Today, she is the Director of Research and Development.
Dr. Evans’ background in chemistry led to a fascinating discovery early in Metabolon’s history that may have otherwise gone unnoticed. The study involved a pharmaceutical company that wanted to pinpoint diagnostic markers of a drug that caused gut toxicity. The researchers ran plasma samples of drugs that were known to cause gut issues and some that did not.

Dr. Evans discovered a molecule that biologists deemed impossible to occur in mammals, but the presence of the molecule could not be denied. That is when metabolomics revealed that gut toxicity could be recognized through metabolites from the gut that passed into circulating plasma. What they were seeing for the first time were microbial derived metabolites in the blood, a discovery made 13 years ago. Even today, metabolomics is the only ‘omic able to identify gut microbiome activity in the blood.
Clinical metabolomics has the potential to make precision medicine a reality delivering transformational medicine. Traditional clinical testing of patient samples involves analyzing and measuring individual analytes or biomarkers. In the case of IEM’s, the screening method typically includes three panels, which measure in the range of 50-60 small molecules. Metabolon’s application of clinical metabolomics extends the number of biomarkers that can be measured significantly. Instead of 50-60, we can identify more than 500 small molecules, giving researchers a broader range of biomarkers to mine for insights.
The application of metabolomics is exceptionally significant for IEM and rare disease detection. There are hundreds of IEM diseases, and often the clinical manifestations of those diseases are vague, or the metabolic pathway that is disturbed is not immediately apparent. Applying global, or untargeted, analytical approaches, where prior knowledge of the affected metabolic pathway is not required, provides advantages over targeted analytical approaches and can extend the diagnostic potential of IEM screens beyond the 34-58 disorders that are currently evaluated in newborn screening programs, especially in complex cases. That makes throwing out this wide net to measure as many biomarkers as possible invaluable in a clinical setting.
When evaluating large quantities of information, it is paramount to make data quality a top priority. A critical step in drawing insights from untargeted metabolomics is accurate metabolite identification. Metabolon’s Precision Metabolomics™ workflow delivers Tier 1-2 identifications for detected metabolites and uniquely provides high confidence in the identification of compounds. By contrast, most metabolomic practitioners operate primarily with annotations that only meet the standards of Tiers 3-5, where some unique features are identified. Still, there is low confidence in confirming the metabolite. Data quality is extremely important to Metabolon, with exceptionally high standards applied to our work in clinical metabolomics. From start to finish, we emphasize quality control measures and checks and balances.
Metabolon’s method of clinical metabolomics is validated by Clinical Laboratory Improvement Amendments (CLIA), a regulatory standard for all testing of patient samples. Metabolon is ISO 9001: 2015 certified for analytical and diagnostic testing of biological specimens and accredited by the College of American Pathologists for diagnostic testing on human specimens. Additionally, Metabolon has a New York State Department of Health Clinical Laboratory permit to perform Quantose IR and IGT testing for the identification of insulin resistance and impaired glucose tolerance under the categories of Clinical Chemistry and Endocrinology.
In addition to the validation work we do to demonstrate the properties of the method, such as precision and accuracy, we also run the method under very specific quality control guidelines, which are even stricter than our research guidelines, including running batch controls for each of the analytic batches. We start by setting up the assays following policies that dictate how to establish a reference population, how you qualify the normalizing matrix, and so on. The typical regulatory requirements for review of the results is exceeded, with an additional analytical and lab director review in addition to our typical sub-level review. All of these steps ensure data quality and accurate insights.
In the early 2000s when Metabolon was formed, there were very few metabolomics practitioners. In fact, the term “metabolomics” was not yet part of the common scientific language. The practitioners mainly composed of academic universities referred to the science as “metabonomics,” and primarily used Nuclear Magnetic Resonance (NMR) technology. Metabolon, on the other hand, focused on a mass spectrometry-based platform that delivered on a global scale. Metabolon ultimately decided to standardize its methodology on the use of LCMS because the company quickly noticed the short-comings of both GCMS and NMR technology in terms of specificity and sensitivity. By choosing LCMS, Metabolon was able to identify more compounds than NMR technology allowed, while also enabling greater processing speed and reducing process variability beyond what is possible with GCMS. While using LCMS in the study of metabolomics was less common at the time, this methodology has allowed Metabolon to maintain its focus on the biology and the meaning of its findings—to interpret and make sense of the data that is revealed.
In its early years, Metabolon employed fewer than 15 scientists who studied just 50 to 100 small molecules. Today, the company has more than 200 employees, has run over 10,000 projects and has identified more than 2,000 named molecules in human plasma from a database of over 5,300 biologically relevant compounds. This comprises the world’s largest metabolomics knowledgebase.
In addition to traditional metabolites, Metabolon’s library features more than 1,000 complex lipids, analyzed on the Lipidyzer™, which the company co-developed with Sciex in 2012.
Metabolon’s compound database is remarkable not only for its size, but for its quality. From the beginning, Metabolon adopted a validated identification methodology, which required the identification of a compound based on multiple orthogonal criteria to an authentic standard analyzed using the same methodology. Compound identification confidence levels were later described and formalized in Sumner et. al., as Tiers ranging from 1 to 5 for each metabolite, with Tiers 1 and 2 being of the highest level of confidence. In untargeted metabolomics, accurate metabolite information is the key step in separating noise from insight. Today, more than 85% of compounds in Metabolon’s library meet the Tier 1 criteria. Though tier rating is a critical part of confidence in compound identification, it is a difficult and expensive process that takes tremendous resources and experience to be successful. As a result, few other metabolomics service providers are able to deliver the same level of confidence in metabolite identification offered from Metabolon’s Precision Metabolomics™ solution.
Today, Metabolon remains committed to investing in and advancing leading-edge metabolomics technologies. This emphasis is improving the understanding of systems biology, enhancing identification of clinically relevant biomarkers, accelerating drug development, improving patient stratification and revealing new insights in the rapidly advancing field of microbiome research.

For Dr. Evans, each success is a motivator to make the science even better. Once the power of the science has been revealed, the reward motivates both her and other scientists to continue making it better for everyone who is involved. She remarks that each scientist has been heavily involved in the trajectory of the company, each playing pivotal roles to move the science and technology from the small lab they began in to where they are today. “How could you not just fall in love with any technology where you can take something that you love and really feel like you’re impacting people and improving lives,” Dr. Evans said.

More about Dr. Annie Evans:
Dr. Annie Evans leads the discovery metabolomics and lipidomics profiling research and development team at Metabolon, Inc. The metabolomics and lipidomics platforms developed under Dr. Evans have been the analytical basis for thousands of commercial studies from over 700 institutions since 2004. She has over 30 publications covering the analytical methodology as well as informatics, data processing requirements and approaches for global profiling metabolomics and lipidomics workflows. These publications have been cumulatively cited over 2000 times, with a foundational metabolomics methodology paper being cited over 650 times. These publications have spanned various technology journals such of the Analytical Chemistry and the Journal of Chromatography, biological journals such as Blood and PNAS as well as high impact journals such as Nature Genetics. Dr. Evans currently holds an h-index of 35, with a JCR of 13. She is involved in industry establishment of quality control methods through the National Institutes of Health Metabolomics Quality Assurance & Quality Control Consortium (mQACC).
Tier 1 Metabolite Identifications: A Compass to Rich Research Insights
Research is like a treasure hunt. Without the right navigation tools, precious time is spent on trial and error exploration to find the right path, increasing the cost as well as the risk of program failure. Metabolomics provides uniquely valuable guidance for navigating a biological system, but the signal-to-noise ratio from vast amounts of data can cloud your ability to see the actionable insight.

In untargeted metabolomics, the critical step to separating noise from insight is accurate metabolite identification (often also called annotation). As pointed out by Schrimpe-Rutledge et al, “metabolite annotation is the crucial link between acquired data and meaningful biological information.”
Phrased another way, the biological insight from a study is only as good as the metabolite annotation – if you want the highest quality metabolomic insight you need the highest quality annotation. In recognition of this fact, Sumner et al proposed a schema for stratifying the quality of metabolomic annotations. Tiers, or levels, ranging from 1 to 5 are commonly used to convey metabolite identification confidence. Tiers refer to the level of detail for each metabolite, and thus the assigned confidence for accurate identification. Tier 5, the lowest level of identification, offers a unique feature in the metabolite, but lacks the information required for confirmation. Through increasing precision measurement, additional unique characteristics are discoverable enabling a definitive identification of the molecule to be made. It is not until the appropriate level of detail is reached in Tier 1 that definitive compound identification based on multiple orthogonal measurements and comparison to data from an authentic standard can be achieved. Therefore, Tier 1 identifications represent the highest level of confidence in the annotation. Tiers 2 through 5, on the other hand, represent decreasing levels of confidence based on less rigorous or more ambiguous criteria.
While most metabolomic practitioners operate primarily with annotations that only meet the standards of Tiers 3-5, Metabolon’s Precision Metabolomics™ workflow is uniquely designed to deliver Tier 1-2 identifications for detected metabolites. This unique level of confidence in the annotations is made possible by Metabolon’s use of a chemocentric approach to metabolomics that uniquely detects metabolite features and matches them against Metabolon’s vast in-house library. Metabolon built this library through the analysis of >5,000 authentic standards run in-house using our methods and instruments. This approach to untargeted metabolomics means that all metabolite annotations meet the stringent criteria required for a Tier 1 or 2 identification.
Our method stands in stark contrast to the more traditional metabolomics workflow in which the individual ion features, the instrument signal from a mass spectrometer, undergo statistical analysis. Only the most significant are subjected to an attempt at a high Tier identification. Metabolon’s approach leverages this vast library to ensure accurate annotation of not only the metabolites, which show statistically significant changes in a study, but also those which remain unchanged, and therefore add crucial insight into the underlying biology.
The field of untargeted metabolomics continues to expand due to its growing track record of providing a crucial understanding of biological processes including aging, disease, and the role of the microbiome in health. Metabolon sets the standard in delivering high-quality metabolomic data. By harnessing the power of our extensive library and delivering metabolite measurements with Tier 1 identifications, Metabolon leads the way to unlocking the information stored in the metabolome and revealing the contained biological story.
To learn more about how metabolomics can help you uncover actionable insights with our Precision Metabolomics platform, contact us at hello@metabolon.com.
References: