Summary of Study ST000149

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench,, where it has been assigned Project ID PR000130. The data can be accessed directly via it's Project DOI: 10.21228/M81P4M This work is supported by NIH grant, U2C- DK119886.


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Study IDST000149
Study TitleHigh Insulin Combined With Essential Amino Acids Stimulates Skeletal Muscle Mitochondrial Protein Synthesis While Decreasing Insulin Sensitivity in Healthy Humans
Study TypeHigh and low insulin with and without essential amino acids
Study SummaryThirty participants were randomized to 3 groups of 10 each with each participant studied twice. Study groups comprised (1) low and high insulin, (2) low insulin with and without EAAs, and (3) high insulin with and without EAAs.
Mayo Clinic
Last NameNair
First NameSreekumaran
Submit Date2015-03-05
Num Groups3
Raw Data AvailableNo
Analysis Type DetailLC-MS
Release Date2015-03-05
Release Version1
Sreekumaran Nair Sreekumaran Nair application/zip

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Project ID:PR000130
Project DOI:doi: 10.21228/M81P4M
Project Title:High insulin combined with essential amino acids stimulates skeletal muscle mitochondrial protein synthesis while decreasing insulin sensitivity in healthy humans
Project Type:Targeted metabolomics
Project Summary:CONTEXT:Insulin and essential amino acids (EAAs) regulate skeletal muscle protein synthesis, yet their independent effects on mitochondrial protein synthesis (MiPS) and oxidative function remain to be clearly defined. OBJECTIVE:The purpose of this study was to determine the effects of high or low insulin with or without EAAs on MiPS. DESIGN:Thirty participants were randomized to 3 groups of 10 each with each participant studied twice. Study groups comprised (1) low and high insulin, (2) low insulin with and without EAAs, and (3) high insulin with and without EAAs. SETTING:The study was conducted in an in-patient clinical research unit. PARTICIPANTS:Eligible participants were 18 to 45 years old, had a body mass index of <25 kg/m(2), and were free of diseases and medications that might impair mitochondrial function. INTERVENTION:Low (? 6 ?U/mL) and high (? 40 ?U/mL) insulin levels were maintained by iv insulin infusion during a somatostatin clamp while maintaining euglycemia (4.7-5.2 mM) and replacing GH and glucagon. The EAA infusion was 5.4% NephrAmine. l-[ring-(13)C6]Phenylalanine was infused, and muscle needle biopsies were performed. MAIN OUTCOMES:Muscle MiPS, oxidative enzymes, and plasma amino acid metabolites were measured. RESULTS:MiPS and oxidative enzyme activities did not differ between low and high insulin (MiPS: 0.07 ± 0.009 vs 0.07 ± 0.006%/h, P = .86) or between EAAs and saline during low insulin (MiPS: 0.05 ± 0.01 vs 0.07 ± 0.01, P = .5). During high insulin, EAAs in comparison with saline increased MiPS (0.1 ± 0.01 vs 0.06 ± 0.01, P < .05) and cytochrome c oxidase activity (P < .05) but not citrate synthase (P = .27). EAA infusion decreased (P < .05) the glucose infusion rates needed to maintain euglycemia during low (? 40%) and high insulin (? 24%). CONCLUSION:EAAs increased MiPS and oxidative enzyme activity only with high insulin concentrations.
Institute:Mayo Clinic
Laboratory:Dr. Sreekumaran Nair's lab
Last Name:Nair
First Name:Sreekumaran
Address:Mayo Clinic, 200 First Street SW, Rochester, MN
Funding Source:5T32DK007352, 5R01DK041973, UL1 TR000135, U24DK100469
Project Comments:25222757


Subject ID:SU000168
Subject Type:Human
Subject Species:Homo sapiens
Taxonomy ID:9606
Species Group:Human


Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Sample Group
SA0080903DHigh Insulin
SA0080914DHigh Insulin
SA0080925CHigh Insulin
SA0080935DHigh Insulin
SA0080943CHigh Insulin
SA0080952DHigh Insulin
SA0080961DHigh Insulin
SA0080971CHigh Insulin
SA0080982CHigh Insulin
SA0080996CHigh Insulin
SA0081004CHigh Insulin
SA0081019CHigh Insulin
SA00810210DHigh Insulin
SA0081036DHigh Insulin
SA0081049DHigh Insulin
SA0081058DHigh Insulin
SA00810610CHigh Insulin
SA0081078CHigh Insulin
SA0081087DHigh Insulin
SA0081097CHigh Insulin
SA00811022DHigh Insulin + EAA
SA00811124CHigh Insulin + EAA
SA00811223CHigh Insulin + EAA
SA00811322CHigh Insulin + EAA
SA00811420DHigh Insulin + EAA
SA00811524DHigh Insulin + EAA
SA00811620CHigh Insulin + EAA
SA00811721CHigh Insulin + EAA
SA00811821DHigh Insulin + EAA
SA00811929DHigh Insulin + EAA
SA00812028CHigh Insulin + EAA
SA00812129AHigh Insulin + EAA
SA00812228DHigh Insulin + EAA
SA00812327DHigh Insulin + EAA
SA00812427CHigh Insulin + EAA
SA00812525DHigh Insulin + EAA
SA00812626CHigh Insulin + EAA
SA00812726DHigh Insulin + EAA
SA00812825CHigh Insulin + EAA
SA00812923DHigh Insulin + EAA
SA00813025BHigh Insulin+Saline
SA00813126AHigh Insulin+Saline
SA00813224BHigh Insulin+Saline
SA00813324AHigh Insulin+Saline
SA00813423BHigh Insulin+Saline
SA00813526BHigh Insulin+Saline
SA00813627AHigh Insulin+Saline
SA00813729BHigh Insulin+Saline
SA00813829CHigh Insulin+Saline
SA00813928BHigh Insulin+Saline
SA00814028AHigh Insulin+Saline
SA00814127BHigh Insulin+Saline
SA00814223AHigh Insulin+Saline
SA00814325AHigh Insulin+Saline
SA00814420BHigh Insulin+Saline
SA00814521BHigh Insulin+Saline
SA00814620AHigh Insulin+Saline
SA00814722AHigh Insulin+Saline
SA00814821AHigh Insulin+Saline
SA00814922BHigh Insulin+Saline
SA00815014DLow Ins +EAA
SA00815130CLow Ins +EAA
SA00815215DLow Ins +EAA
SA00815319CLow Ins +EAA
SA00815430DLow Ins +EAA
SA00815519DLow Ins +EAA
SA00815612CLow Ins +EAA
SA00815715CLow Ins +EAA
SA00815816CLow Ins +EAA
SA00815916DLow Ins +EAA
SA00816018CLow Ins +EAA
SA00816118DLow Ins +EAA
SA00816213CLow Ins +EAA
SA00816317DLow Ins +EAA
SA00816413DLow Ins +EAA
SA00816517CLow Ins +EAA
SA00816611CLow Ins +EAA
SA00816712DLow Ins +EAA
SA00816811DLow Ins +EAA
SA00816914CLow Ins +EAA
SA00817013ALow Ins+ Saline
SA00817113BLow Ins+ Saline
SA00817212ALow Ins+ Saline
SA00817311BLow Ins+ Saline
SA00817411ALow Ins+ Saline
SA00817514ALow Ins+ Saline
SA00817630BLow Ins+ Saline
SA00817730ALow Ins+ Saline
SA00817812BLow Ins+ Saline
SA00817915BLow Ins+ Saline
SA00818018ALow Ins+ Saline
SA00818117ALow Ins+ Saline
SA00818215ALow Ins+ Saline
SA00818316ALow Ins+ Saline
SA00818417BLow Ins+ Saline
SA00818518BLow Ins+ Saline
SA00818616BLow Ins+ Saline
SA00818719BLow Ins+ Saline
SA00818819ALow Ins+ Saline
SA00818914BLow Ins+ Saline
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Collection ID:CO000154
Collection Summary:Percutaneous muscle biopsy specimens of the vastus lateralis (?300 mg) were performed at 180 and 480 minutes under local anesthesia (2% lidocaine). Muscle samples were blotted, were dissected free of fat and connective tissue, and then were frozen. Biopsy specimens were collected from the opposite legs on a single visit. Indirect calorimetry was performed at 11:00 am for 20 minutes using a ventilated hood (Parvo Medics). The final 10 minutes of steady-state oxygen consumption (Vo2) and carbon dioxide production (Vco2) determined the respiratory exchange ratio (RER = Vco2/Vo2). Urine was collected throughout the clamp and analyzed for urea content at a core laboratory.
Sample Type:Muscle


Treatment ID:TR000173
Treatment Summary:Low Insulin | High Insulin | Low Ins+ Saline | Low Ins +EAA | High Insulin+Saline | High Insulin + EAA

Sample Preparation:

Sampleprep ID:SP000168
Sampleprep Summary:MMP and muscle fractions were isolated from frozen samples using differential centrifugation (18, 19). Biopsy samples were homogenized with protease and phosphatase inhibitors (Halt; Thermo Fisher Scientific) and centrifuged to pellet myofibrillar (MYO) proteins. The supernatant was centrifuged to pellet mitochondrial (MITO) proteins, and the final supernatant was deproteinated with cold ethanol (1:9, v/v) and then centrifuged to pellet sarcoplasmic (SARC) proteins. Aliquots from MMP, MYO, SARC, and MITO were acid hydrolyzed, and free amino acids were purified using cation exchange columns and then were dried.
Plasma phenylalanine enrichment was determined using gas chromatography (GC) and mass spectrometry (MS) as described previously (19). Samples were derivatized to a heptafluorobutyryl isobutyl ester and identified with a Micromass Quattro Micro triple quadrupole GC-MS system (Waters) operating under negative ion chemical ionization using isobutane as the reactant gas. Selected ion monitoring of m/z 399.2 and 403.2 M + 2 and M + 6 fragments of phenylalanine and the l-[ring-13C6]phenylalanine, respectively, was performed.

Combined analysis:

Analysis ID AN000236
Analysis type MS
Chromatography type
Chromatography system
MS instrument type Triple quadrupole
MS instrument name ABI Sciex API 5000 QQQ
Units uM


Chromatography ID:CH000165


MS ID:MS000187
Analysis ID:AN000236
Instrument Name:ABI Sciex API 5000 QQQ
Instrument Type:Triple quadrupole
MS Comments:Dried amino acids were prepared as isobutyl ester derivatives, and the mass was determined by an API 5000 triple quadrupole mass spectrometer with a TurboIonSpray source (Applied Biosystems).