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Nutrition Obesity Research Center at Harvard

Core Facilities

Mass Spectrometry

Tom Jaksic M.D., Ph.D.
Principle Investigator

The NORC Mass Spectrometry Core has two major goals: (i) to provide a resource for CNRU investigators for the use of stable isotopes estimated with mass spectrometry in nutritional investigations and (ii) to act as a center for training CNRU investigators in the development and application of stable isotope methods. Dr. Tom Jaksic, Associate Professor at Children's Hospital, Boston is the Core PI. Joining Dr. Jaksic as the Core's Senior Staff are co-Directors Joanne K. Kelleher, PhD and Yong-Ming Yu, MD PhD.

A strength of the Mass Spectrometry Core is the experience of the Senior Staff at every level of stable isotope methodology from mathematical modeling to cellular, animal and clinical applications. The team is actively involved in stable isotope nutritional research developing and applying new techniques for quantifying metabolic reactions in vivo. The Senior Staff serves as an experienced consultative group to be used by any NORC participants interested in stable isotopic experiments. The core is prepared to assist investigators with tracer selection, infusion preparation, infusion protocols, data analysis modeling and institutional review board applications. Thus, the core provides the resources necessary for NORC investigators to carry our stable isotope studies.

An important objective of nutritional studies is to quantify metabolic reactions in vivo. Stable isotope metabolic tracers provide a valuable tool for accomplishing this task. They offer a noninvasive tool for exploring metabolic reactions across cells, tissues, and organs. In additional, because stable isotopes are not radioactive, they are ideal for human nutritional studies. Commonly used stable isotopes for nutritional studies include 2H, 13C, 15N and 18O. Molecules containing these stable heavy isotopes are detected in the Core by mass spectrometry. Our core has access to a variety of instruments well suited for nutritional and metabolic studies incorporating the use of stable isotopes as nutrients or metabolites. The combination of the expertise of the Senior Staff and the mass spectrometry resources available allows our Core to provide strong support for metabolite stable isotope studies by NORC investigators.

Nutritional Mass Spectrometry Methods

Isotope Dilution to estimate in vivo rate of appearance

The Core has extensive experience with amino acid metabolism and protein turnover studies. These techniques form the basis for many techniques in nutritional mass spectrometry. A second key area is glucose metabolism. A classic experimental technique in glucose and amino acid studies is to estimate the rate of appearance in plasma of specific amino acids or glucose. This is accomplished by a constant infusion of a stable isotope labeled amino acid. When the plasma enrichment reaches steady state the rate of appearance of the amino acids is calculated from the dilution of the isotope in plasma. This type of study is classified as “isotope dilution”. When isotope is infused at a constant rate, isotope dilution studies provide quantitative information on interorgan metabolite movement or flux. This data may be further analyzed to estimate rates of rates of protein synthesis and degradation.

Isotope Incorporation to estimate de novo biosynthesis.

Another classic issue in nutritional studies is to estimate the rate of biosynthesis of metabolites in vivo. A hallmark of this type of study is that the stable isotope tracer is a precursor, directly incorporated into the metabolite of interest. Cholesterol may serve as an example for this type of study. The core has experience with estimating cholesterol synthesis in vivo via the incorporation of precursors of acetyl CoA as shown here. (Only six of the 27 carbons are shown in this simplified model.) The concept is that the labeling pattern of the product molecule, cholesterol, can be used to solve for both the fraction of newly synthesized cholesterol and the precursor enrichment. More recently, the Core has been developing techniques for estimating de novo synthesis using deuterated water (2H2O) as the precursor. 2H2O may be used to estimate the biosynthesis of a large number of compounds that incorporate 2H from water during biosynthesis. These include glucose, cholesterol and fatty acids. This approach has many potential applications in clinical research and the Core is presently developing additional assays based on the use of H2O.

Isotope Detection as 13CO2 to Estimate Substrate Oxidation.

A third class of assays estimates substrate oxidation. These “breath tests” detect oxidation of a 13C labeled metabolite or drug to 13CO2. Breath tests following ingestion of 13C labeled fatty acids have been used to estimate the defects b-oxidation. Other assays detect drug oxidation. Yet another use of breath tests is to estimate organ function with assays such as methionine oxidation depicted here. The principle underlying this assay is that methionine oxidation is largely, or exclusively, confined to hepatic mitochondria. Thus, rapid production of 13CO2 following administration of13C-methionine is an indicator of hepatic function.

The brief descriptions above provide an overview of the basic assays performed at the Mass Spectrometry Core. Actual studies are often more complicated. Studies may include more than one type of assay and may utilize multiple stable isotope tracers. As the complexity of the experiment grows, the data analysis becomes a significant project. At a most basic level the data analysis must consider the natural abundance of stable isotopes, especially 13C, which complicate the mass spectrometry data. The Core Senior Staff has expertise in mass spectrometry data analysis and modeling and can offer assistance to investigators. The interpretation of the data is a joint effort between Core Senior Staff and the investigator to insure that experimental and methodological issues are each considered appropriately.

Center Access to Core

An important objective of the Core is to assist researchers in a timely and effective manner. The Senior Staff has weekly conferences to discuss the progress of ongoing research projects. These meetings also address new techniques and projects. Every investigator who wishes to use the facility presents his/her project at one of these meetings. This provides the Senior Staff an opportunity to provide input into the project design and the type of measurements that may be provided by the Core. Before projects are included in the Core the following procedures are followed to facilitate the smooth and efficient conduct of this share facility. A written protocol describing the background of the project, hypothesis to be tested and all the experimental methods is distributed to the Senior Staff. The analytical methods or requirements are fully described. It is the responsibility of the CNRU investigator to assist the Core staff in obtaining all necessary methodology. The Core personnel as a service for other CNRU investigators will carry out a limited amount of analytical effort. However, for most studies, the investigator supplies samples prepared for mass spectrometry. A section on expected results is requested as part of the protocol. The investigator will present the project for the information of the Core staff at one of the weekly meetings. Following review of the protocol, one staff member is assigned major responsibility for the analytical data to be collected in the project. All data pertaining to the project will be forwarded to this person. The Investigator is expected to present the completed study at meeting of the Senior Staff. At this final meeting the Senior Staff and NORC investigators can discuss enhancements and new techniques that will facilitate additional studies.

Enrichment Program

The Mass Spectrometry Core provides a "clearing house" for information regarding the use of new methods for use of mass spectrometry and stable isotope tracers in nutritional investigations. The core organizes seminar presentations by leaders in the field with expertise in techniques of interest to NORC investigators. In the past year our invited lectures included a presentation on doubly labeled water by Dr. William Wong of USDA Children's Nutrition Research Center, Baylor College of Medicine and a presentation on stable isotope contributions to metabolomics by Dr. Henri Brunengraber, Chair of the Department of Nutrition at Case Western Reserve University. These lectures provide an important component of the enrichment program for the Core staff. As new stable isotope methods for human nutritional research continue to appear, the Core will explore such methods and provide a resource for investigators on the practical implementation of new techniques. Since one of the mandates of the Core is to provide training in specialized techniques, part of the enrichment program offered by the facility will be individual technical training and possibly group training in the form of seminars and workshops on topics pertinent to NORC studies.

For more information about this core, please contact:

Tom Jaksic, MD, PhD
General Surgery
300 Longwood Avenue, Fegan-3
Boston MA 02115

Phone: 617-355-9600
Fax: 617-730-0477


Dr. Jaksic’s biographical sketch is linked here.

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Updated 1/25/2008