#METABOLOMICS WORKBENCH ryant_20201130_112026 DATATRACK_ID:2335 STUDY_ID:ST001616 ANALYSIS_ID:AN002651 PROJECT_ID:000000 VERSION 1 CREATED_ON December 3, 2020, 11:15 am #PROJECT PR:PROJECT_TITLE Unique metabolomic profile of skeletal muscle in chronic limb threatening PR:PROJECT_TITLE ischemia PR:PROJECT_TYPE NMR PR:PROJECT_SUMMARY This project is focused on a cross-sectional analysis of non-PAD controls and PR:PROJECT_SUMMARY CLTI patients undergoing either a vascular intervention or undergoing limb PR:PROJECT_SUMMARY amputation was performed and involved a detailed assessment of the limb muscle PR:PROJECT_SUMMARY metabolome using HR-MAS and solution state NMR spectroscopy. It was hypothesized PR:PROJECT_SUMMARY that patients undergoing limb amputation would present with altered muscle PR:PROJECT_SUMMARY metaolite features compared with non-PAD controls. PR:INSTITUTE University of Florida PR:DEPARTMENT Applied Physiology and Kinesiology PR:LABORATORY University of Florida PR:LAST_NAME Ryan PR:FIRST_NAME Terence PR:ADDRESS 1864 Stadium Rd, FLG 114, Gainesville, FL, 32611, USA PR:EMAIL ryant@ufl.edu PR:PHONE 352-294-1700 PR:FUNDING_SOURCE National Institutes of Health and the National Heart, Lung, and Blood, Institute PR:FUNDING_SOURCE numbers R01-HL149704 (to T.E.R.) and R01-HL148597 (to S.T.S.); as well as the PR:FUNDING_SOURCE American Heart Association grant number 18CDA34110044 (to T.E.R.). A portion of PR:FUNDING_SOURCE this work was supported by pilot funds (awarded to T.E.R.) from The University PR:FUNDING_SOURCE of Florida Claude D. Pepper Older Americans Independence Center P30AG028740. (if PR:FUNDING_SOURCE applicable) PR:PROJECT_COMMENTS PAD metabolomic study via NMR.A detailed assessment of the limb muscle PR:PROJECT_COMMENTS metabolome using semi-solid and solution state NMR spectroscopy. PR:PUBLICATIONS Journal of Clinical Medicine PR:CONTRIBUTORS Ram B. Khattri, Kyoungrae Kim, Trace Thome, Zachary R. Salyers, Kerri A. PR:CONTRIBUTORS O’Malley, Scott A. Berceli, Salvatore T. Scali, Terence E. Ryan* #STUDY ST:STUDY_TITLE Unique metabolomic profile of skeletal muscle in chronic limb threatening ST:STUDY_TITLE ischemia (aqueous phase samples ) ST:STUDY_TYPE NMR ST:STUDY_SUMMARY This project is focused on a cross-sectional analysis of non-PAD controls and ST:STUDY_SUMMARY CLTI patients undergoing either a vascular intervention or undergoing limb ST:STUDY_SUMMARY amputation was performed and involved a detailed assessment of the limb muscle ST:STUDY_SUMMARY metabolome using HR-MAS and solution state NMR spectroscopy. It was hypothesized ST:STUDY_SUMMARY that patients undergoing limb amputation would present with altered muscle ST:STUDY_SUMMARY metaolite features compared with non-PAD controls. ST:INSTITUTE University of Florida ST:DEPARTMENT Applied Physiology and Kinesiology ST:LABORATORY Rm 42 and Rm 43 ST:LAST_NAME Ryan ST:FIRST_NAME Terence ST:ADDRESS University of Florida, Applied Physiology and Kinesiology, 1864 stadium RD, ST:ADDRESS Gainesville, FL 32611 ST:EMAIL ryant@ufl.edu ST:PHONE 352-294-1700 ST:NUM_GROUPS 3 ST:TOTAL_SUBJECTS 30 ST:NUM_MALES 25 ST:NUM_FEMALES 5 ST:STUDY_COMMENTS PAD metabolomic study via NMR. A detailed assessment of the limb muscle ST:STUDY_COMMENTS metabolome using semi-solid and solution state NMR spectroscopy ST:PUBLICATIONS Journal of Clinical Medicine #SUBJECT SU:SUBJECT_TYPE Human SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 SU:AGE_OR_AGE_RANGE 55-82 years SU:GENDER Male and female SU:HUMAN_RACE unknown SU:HUMAN_ETHNICITY unknown SU:HUMAN_MEDICATIONS Asprin, ACE inhibitor, statins, cilostazol SU:HUMAN_SMOKING_STATUS Controls = 4, CLTI Pre-surgery = 7, and CLTI Amputation = 9 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - CLTI_Amputation1a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation1a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation2a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation2a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation3a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation3a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation4a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation4a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation5a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation5a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation6a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation6a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation7a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation7a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation8a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation8a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation9a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation9a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Amputation10a Group:CLTI_Amputation RAW_FILE_NAME=CLTI_Amputation10a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Pre_surgery1a Group:CLTI_Pre_surgery RAW_FILE_NAME=CLTI_Pre_surgery1a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Pre_surgery2a Group:CLTI_Pre_surgery RAW_FILE_NAME=CLTI_Pre_surgery2a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Pre_surgery5a Group:CLTI_Pre_surgery RAW_FILE_NAME=CLTI_Pre_surgery5a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Pre_surgery6a Group:CLTI_Pre_surgery RAW_FILE_NAME=CLTI_Pre_surgery6a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Pre_surgery7a Group:CLTI_Pre_surgery RAW_FILE_NAME=CLTI_Pre_surgery7a.raw SUBJECT_SAMPLE_FACTORS - CLTI_Pre_surgery9a Group:CLTI_Pre_surgery RAW_FILE_NAME=CLTI_Pre_surgery9a.raw SUBJECT_SAMPLE_FACTORS - Control2a Group:Control RAW_FILE_NAME=Control2a.raw SUBJECT_SAMPLE_FACTORS - Control3a Group:Control RAW_FILE_NAME=Control3a.raw SUBJECT_SAMPLE_FACTORS - Control4a Group:Control RAW_FILE_NAME=Control4a.raw SUBJECT_SAMPLE_FACTORS - Control5a Group:Control RAW_FILE_NAME=Control5a.raw SUBJECT_SAMPLE_FACTORS - Control6a Group:Control RAW_FILE_NAME=Control6a.raw SUBJECT_SAMPLE_FACTORS - Control7a Group:Control RAW_FILE_NAME=Control7a.raw SUBJECT_SAMPLE_FACTORS - Control8a Group:Control RAW_FILE_NAME=Control8a.raw SUBJECT_SAMPLE_FACTORS - Control10a Group:Control RAW_FILE_NAME=Control10a.raw #COLLECTION CO:COLLECTION_SUMMARY Gastrocnemius muscle specimens were obtained from ten older adult non-PAD CO:COLLECTION_SUMMARY controls (Control), ten patients with critical limb ischemia (CLI) undergoing CO:COLLECTION_SUMMARY surgical intervention (CLTI Pre-surgery), and ten CLI patients undergoing limb CO:COLLECTION_SUMMARY amputation (CLTI Amputation). Five pre-surgery patients underwent bypass CO:COLLECTION_SUMMARY interventions and five underwent endovascular procedures. Muscle specimens were CO:COLLECTION_SUMMARY collected within the confines of the operating rooms (CLI patients) or via CO:COLLECTION_SUMMARY percutaneous muscle biopsy using sterile procedures. A portion of the muscle was CO:COLLECTION_SUMMARY quickly trimmed of fat/connective tissue and snap frozen in liquid nitrogen for CO:COLLECTION_SUMMARY metabolomics analysis. CO:SAMPLE_TYPE Muscle CO:COLLECTION_METHOD Muscle specimens were collected within the confines of the operating rooms (CLI CO:COLLECTION_METHOD patients) or via percutaneous muscle biopsy using sterile procedures. A portion CO:COLLECTION_METHOD of the muscle was quickly trimmed of fat/connective tissue and snap frozen in CO:COLLECTION_METHOD liquid nitrogen for metabolomics analysis. CO:STORAGE_CONDITIONS -80℃ CO:STORAGE_VIALS cryovials #TREATMENT TR:TREATMENT_SUMMARY This was a prospective cohort study that examined the metabolomic profile of TR:TREATMENT_SUMMARY skeletal muscle from CLTI patients undergoing surgerical intervention or TR:TREATMENT_SUMMARY amputation, as well as a cohort of non-PAD controls. CLTI patients exhibited TR:TREATMENT_SUMMARY severe symptomology (Rutherford Classification 3-6), with high incidence of TR:TREATMENT_SUMMARY common PAD risk factors including hypertension, hyperlipidemia, coronary artery TR:TREATMENT_SUMMARY disease, and diabetes.Ten patients with critical limb ischemia (CLI) undergoing TR:TREATMENT_SUMMARY surgical intervention (CLTI Pre-surgery), and ten CLI patients undergoing limb TR:TREATMENT_SUMMARY amputation (CLTI Amputation). Five pre-surgery patients underwent bypass TR:TREATMENT_SUMMARY interventions and five underwent endovascular procedures. Muscle specimens were TR:TREATMENT_SUMMARY collected within the confines of the operating rooms (CLI patients) or via TR:TREATMENT_SUMMARY percutaneous muscle biopsy using sterile procedures. TR:HUMAN_FASTING non-fasted TR:HUMAN_ENDP_CLINICAL_SIGNS N/A #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Extraction. Both polar and non-polar metabolites were extracted from the SP:SAMPLEPREP_SUMMARY gastrocnemius muscle specimens using FOLCH extraction. In brief, wet weigh of SP:SAMPLEPREP_SUMMARY the frozen tissues were determined and immediately homogenized in 1 mL of SP:SAMPLEPREP_SUMMARY ice-cold methanol using a PowerLyzer 24 Homogenizer (QIAGEN Group, Hilden, SP:SAMPLEPREP_SUMMARY Germany). All enzymatic activities are halted once the sample was homogenized in SP:SAMPLEPREP_SUMMARY the methanol. Homogenization was followed by centrifugation (13.2K r.p.m., 4 oC, SP:SAMPLEPREP_SUMMARY 30 minutes) and supernatant was transferred into a new glass vial consisting a SP:SAMPLEPREP_SUMMARY mixture of 3 mL of ice-cold chloroform and methanol (2:1 v/v) ratio. The cold SP:SAMPLEPREP_SUMMARY mixture was vortexed for several minutes and left in an ice bath 15 minutes to SP:SAMPLEPREP_SUMMARY allow for phase separation. Next, 1 mL of ice-cold 0.9% saline was added to the SP:SAMPLEPREP_SUMMARY mixture followed by vigorous mixing. The mixture was again left in the ice bath SP:SAMPLEPREP_SUMMARY for 45 minutes for phase separation. The upper methanol/water layer was SP:SAMPLEPREP_SUMMARY transferred to a new falcon tube. To the lower chloroform layer, 1 mL of SP:SAMPLEPREP_SUMMARY ice-cold 0.9% saline was added and all steps were followed as mentioned above. SP:SAMPLEPREP_SUMMARY Following a 45-minute incubation, upper methanol/water layer was again SP:SAMPLEPREP_SUMMARY transferred to the previous falcon tube and dried using a Labconco freeze drier SP:SAMPLEPREP_SUMMARY (Labconco Corporation, MO, USA). The chloroform layer was dried under a stream SP:SAMPLEPREP_SUMMARY of nitrogen gas. The dried samples (both aqueous and organic phases) were stored SP:SAMPLEPREP_SUMMARY at -80 oC until resuspension for NMR experiments. Lyophilized aqueous phase SP:SAMPLEPREP_SUMMARY samples were re-suspended in 50 µL of 50 mM phosphate buffer (pH 7.2) SP:SAMPLEPREP_SUMMARY consisting 2 mM of EDTA along with 0.2% NaN3 and 0.5 mM D6-DSS in 100% SP:SAMPLEPREP_SUMMARY deuterated environment. SP:PROCESSING_METHOD Lyophilization and Homogenization SP:PROCESSING_STORAGE_CONDITIONS -80℃ SP:EXTRACTION_METHOD Modified FOLCH extraction for one set of samples SP:EXTRACT_STORAGE -80℃ SP:SAMPLE_RESUSPENSION In 50 microliter of 50 mM phosphate buffer (pH 7.2) with 2 mM EDTA, 0.5 mM DSS SP:SAMPLE_RESUSPENSION and 0.2% sodium azide for aqueous phase samples. SP:SAMPLE_SPIKING 0.5 mM of DSS for aqueous phase samples #ANALYSIS AN:DATA_FORMAT fid, 1r #NMR NM:INSTRUMENT_NAME Bruker Avance Neo 600 MHz/54mm console NM:INSTRUMENT_TYPE FT-NMR NM:NMR_EXPERIMENT_TYPE 1D-1H NM:FIELD_FREQUENCY_LOCK Deuterium NM:STANDARD_CONCENTRATION 0.5 mM DSS NM:SPECTROMETER_FREQUENCY 600.2328273 MHz NM:NMR_PROBE 1.7 mm TXI CryoProbe NM:NMR_SOLVENT Phosphate buffer (pH 7.2) + 2 mM EDTA + 0.5 mM DSS + 0.2% of sodium azide in NM:NMR_SOLVENT deuterated environment NM:NMR_TUBE_SIZE 1.7 mm O.D. NM:SHIMMING_METHOD Topshim NM:PULSE_SEQUENCE noesypr1d NM:WATER_SUPPRESSION presat NM:PULSE_WIDTH 90-degree NM:RECEIVER_GAIN 101 NM:OFFSET_FREQUENCY 2827.31 Hz NM:CHEMICAL_SHIFT_REF_CPD DSS (4,4-dimethyl-4-silapentane-1-sulfonic acid) NM:TEMPERATURE 25 o C NM:NUMBER_OF_SCANS 128 scans NM:DUMMY_SCANS 8 NM:ACQUISITION_TIME 4 s NM:RELAXATION_DELAY 1 s NM:SPECTRAL_WIDTH 7142.9 Hz NM:NUM_DATA_POINTS_ACQUIRED 28571 NM:REAL_DATA_POINTS 65536 NM:LINE_BROADENING 0.22 Hz NM:ZERO_FILLING 65,536 points NM:APODIZATION Exponential NM:BASELINE_CORRECTION_METHOD Spline NM:CHEMICAL_SHIFT_REF_STD 0 ppm for DSS #NMR_METABOLITE_DATA NMR_METABOLITE_DATA:UNITS A.U. NMR_METABOLITE_DATA_START Samples CLTI_Amputation1a CLTI_Amputation2a CLTI_Amputation3a CLTI_Amputation4a CLTI_Amputation5a CLTI_Amputation6a CLTI_Amputation7a CLTI_Amputation8a CLTI_Amputation9a CLTI_Amputation10a CLTI_Pre_surgery1a CLTI_Pre_surgery2a CLTI_Pre_surgery5a CLTI_Pre_surgery6a CLTI_Pre_surgery7a CLTI_Pre_surgery9a Control2a Control3a Control4a Control5a Control6a Control7a Control8a Control10a Factors Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Amputation Group:CLTI_Pre_surgery Group:CLTI_Pre_surgery Group:CLTI_Pre_surgery Group:CLTI_Pre_surgery Group:CLTI_Pre_surgery Group:CLTI_Pre_surgery Group:Control Group:Control Group:Control Group:Control Group:Control Group:Control Group:Control Group:Control Leucine 22.4405388 11.67555373 6.47223163 17.28769094 5.54733629 5.927198371 4.403771407 3.458077768 3.461177326 4.779684835 3.794311813 1.758059723 3.976793361 1.960784264 2.790398133 3.433114292 3.268513271 3.043341919 3.315022548 4.032985446 2.868263183 3.228257108 3.792465207 3.400749288 Valine1 14.16811618 7.254776072 4.731295338 10.45071081 2.890758578 4.532224503 3.809449642 2.616907157 2.973632004 4.036959359 2.935949391 1.482287123 3.171081418 1.473606764 2.228454418 2.775632374 2.562967015 2.97166071 2.543985849 3.353760123 2.333389386 3.081978245 3.0608464 2.61969254 Isoleucine 11.35431972 6.237536675 3.552170509 9.84590946 2.823949626 3.154013525 2.453891808 1.690319274 2.143569447 2.655380188 2.328493925 1.192072566 2.607462274 1.167672382 1.828856262 2.22339429 1.650009436 1.72696141 1.887591056 2.202583339 1.810393596 1.853526055 2.090373882 2.109132375 Valine2 13.98810923 7.118450872 5.759432068 10.16372149 3.031449055 5.609925616 4.36223311 3.548143225 3.624941146 4.6253726 5.275007747 2.169996559 4.443204244 2.575931815 3.903979162 4.319555456 3.093888203 3.544127623 3.462352953 4.867278265 2.889814238 3.931602947 4.274177189 4.063518962 Isobutyrate 5.470538864 3.061235466 5.273130896 2.207972259 1.49324143 5.561298769 3.238407544 3.655102409 3.786481669 4.572937403 8.005095991 2.659186864 4.222100589 3.574189579 5.39022179 5.192549837 2.527220624 2.378920034 3.16459986 4.777700293 2.883992585 3.418833012 5.988578124 4.662941387 3-methyl-2-oxovalerate 5.557861732 3.67576972 4.07722742 2.360242153 1.860548998 4.608683193 3.31325704 2.198233927 2.874604472 3.797658415 5.124525531 2.362500295 4.742893637 2.574843705 3.893814292 4.195836465 2.555559502 2.361976289 3.438101112 3.709143318 3.080463206 2.790787483 4.076122036 4.008350802 Lactate1 484.9445001 67.24312451 99.19112331 66.3280391 38.48554823 220.6693699 90.57461348 108.8452396 64.26845067 168.6650033 13.95111514 6.931142413 16.68340843 7.590114323 14.94564782 14.48499611 17.3152326 32.00712601 15.89926404 20.20925924 13.12226032 16.09575807 18.55047909 15.5766084 Alanine 71.6345335 38.63031967 40.54546026 24.82476061 11.54392306 30.29353637 28.4945082 27.60497917 33.46114979 31.22734728 20.67267201 9.030562157 19.548547 9.11575473 12.72058555 15.63680271 17.88077925 20.64567791 14.61421551 21.41337959 13.54460364 21.45875604 23.11636161 15.32885987 Arginine 12.46027318 11.50869455 12.54476418 14.70891693 5.490889532 14.21791076 12.4461742 0.721948674 11.01314043 13.51692386 11.90279273 8.208565077 12.34561488 2.007076656 9.374199702 12.68354011 5.823377452 13.07753523 7.642971635 8.802613374 8.732288853 10.04665953 3.682195385 11.65677912 Lysine 10.07273271 8.470938556 12.76261512 5.797706683 4.06421786 16.6408848 11.85414867 3.209258778 9.97133574 14.60298953 16.48151044 9.135587228 14.62557578 4.247901362 12.34535083 15.88368821 6.440320477 11.71234186 7.996762757 9.75650607 8.860546084 10.68457499 7.724773849 13.82490163 Acetate 15.45252225 5.357541611 22.83102017 4.746104532 3.03378546 18.67886593 6.861372786 12.35143959 11.16417075 19.18662015 32.13412405 7.482643065 9.03751167 15.53088927 20.52439533 12.1917511 4.746376693 4.947369213 6.315792455 17.31369198 6.933907379 6.882293963 27.78377336 11.09581539 Glutamate1+Proline 34.45739649 36.56928785 46.21744452 26.248961 9.831545018 42.36120236 41.24233563 5.349747688 41.57182474 46.5767615 37.847471 23.94755112 32.77938811 5.185423493 27.25144244 34.0000423 22.07818606 52.58617092 25.23670379 33.91334432 25.7100439 35.50993347 9.953541364 32.58843791 Glutamate+Glutamine+methionine+O-Acetylcholine 113.820553 66.39885716 74.82138766 51.86830808 15.97729511 51.00390925 80.30614954 25.37735057 100.2924178 82.03113193 34.37268469 18.43104249 26.77110399 7.684645947 23.18407594 25.74577976 70.5876796 75.97553216 32.5264318 69.77306052 39.05407935 70.83750086 30.48968746 26.557221 2-Aminoadipate 3.149018942 2.904248881 3.803588027 1.796283008 1.17068222 4.200135435 3.224970888 0.938286135 2.942578162 4.010058859 4.890996948 2.493900662 4.138656316 1.577763682 3.37848257 3.752929112 1.918077245 3.266236505 2.759762373 3.024921988 2.531884968 2.887806629 2.230732804 3.669394599 Glutamate2 25.21768562 26.01438321 31.05449565 14.47257052 6.875611976 24.69721059 25.11465452 8.341109744 31.06048177 27.52672667 25.08393255 12.87239826 20.38538976 7.163843046 15.87496652 16.46131159 19.37143103 30.56034425 18.79371289 23.64874909 15.70176584 24.73674271 8.964085364 18.10237081 Pyruvate 24.1203337 17.85136761 19.67056256 15.0974677 8.340768133 19.73857174 17.40531102 8.297617002 14.41076039 19.08368992 26.62788289 11.8610353 22.70077759 11.36900645 16.9486839 18.7515663 13.26589093 14.04057291 17.13704601 16.77757551 13.43415283 13.49915987 16.79243726 20.44976819 Succinate 12.25121127 5.593461967 6.920687748 6.292279285 1.975577457 4.511971669 5.491316243 3.154981312 5.581410563 7.401705178 4.268991122 2.261953706 3.736194318 1.453389503 2.877289087 3.206911464 2.686924502 4.264943172 2.94296516 3.408773247 2.689907801 3.209356533 1.97473203 3.421641386 Glutamine2 110.5606764 64.15547795 76.02240837 44.16183247 14.3814832 42.76678982 84.35307145 22.07040694 81.54569932 68.16145734 41.09376656 22.04212608 32.75765279 11.94691345 26.62532849 27.34826657 62.01677759 79.87769387 36.2714902 66.87443033 40.32126284 69.07273353 30.08315289 30.9028313 Creatine1 283.7281008 121.5047721 164.807404 54.16304388 11.50494025 107.1502489 149.4563858 118.0899475 133.2804451 128.739802 30.65933059 18.27336845 79.32698647 55.00384853 60.09950734 79.24572705 76.61531313 81.45526255 74.73303826 73.68736788 65.52656118 98.58693309 64.88704645 67.43141683 Creatinine 59.91989937 63.83163414 33.24744612 19.59821156 1.764356296 19.85184455 45.22254498 18.14254244 71.92646269 27.891426 48.56522868 32.54728385 20.83358184 10.92682216 18.21812781 20.52099535 41.71510351 74.66686754 23.26510518 62.67229662 49.27138543 59.15168616 32.91425846 20.27147458 Malonate? 7.971115152 5.221935859 9.467859888 7.688777681 4.276227078 12.33402134 12.30069414 7.630492396 5.417103532 9.398043581 12.24807325 8.263669759 14.09874646 9.180398501 10.32217411 13.284352 7.646851812 9.434743654 10.02203255 8.119726624 10.08150752 11.28660081 7.696784877 12.29790611 Glycerol 63.65419306 20.91933652 20.57604567 15.66294962 17.70675197 21.27379713 18.18901997 1.680375794 14.88485325 27.72132949 16.64043402 11.8334767 13.8557627 5.418393772 11.90144466 15.28776748 8.493104472 22.32933864 13.06575961 13.71475166 11.26055977 13.1438559 3.386464016 14.30156395 Glycine 10.86588455 6.099715805 6.628877578 4.615823331 1.580230176 4.454065262 4.406144923 3.006447314 4.560205847 6.051441489 2.648393925 1.828615656 2.189433478 1.041750369 1.903945434 2.230323518 2.122332366 4.194922078 2.06803532 3.284299299 1.978350649 2.71450813 0.994504659 2.210113972 3-methyl histidine 12.20653207 4.977362545 5.943657271 3.949390996 6.831731406 9.036442894 8.3490958 7.610001695 4.476836413 8.577836974 15.84496527 9.856675765 16.21786623 9.681515184 12.88518637 14.83456115 7.83263071 9.086400752 11.96789793 7.053304082 11.63414874 9.089492787 9.279488723 15.01600299 Betaine 18.04328495 7.989507157 11.91791472 16.10630201 13.3007412 20.30140483 17.58867484 16.60352047 10.01601779 19.03888139 27.01476026 18.61124258 32.71547264 18.41804374 22.68852754 33.6361952 16.72263729 17.67053644 23.20198343 12.70437142 22.86160592 18.93503906 18.65778129 31.32641677 Aspartate1 25.09327096 8.741797677 7.335826715 7.40634224 11.52755967 9.298459731 7.030246092 3.130812406 6.652242565 9.286110954 6.961369318 5.142476711 9.421688041 4.063916283 5.441592547 10.11476553 5.427480547 8.156418541 6.775755131 6.243650099 7.939441211 7.478600533 4.266024875 9.590522263 Aspartate2 24.20158355 8.597333411 8.750838692 6.42478104 9.245701674 10.512583 7.808448063 3.323354082 8.200647847 10.62605979 7.980562224 5.794658989 8.418769746 3.699416706 6.581952356 10.2997629 6.077651696 9.400850291 6.398660806 7.095339448 6.396339811 8.280391114 4.217112189 8.16500248 Creatine2 209.3254451 85.11568208 121.6425212 34.75177235 9.27132881 84.5976552 107.2697864 84.32409117 98.01491573 99.73517291 18.99727416 11.76933849 59.90383279 40.26365204 43.55380679 62.22377448 55.94279187 57.92535273 54.53021911 51.70795283 47.7586355 72.05627426 45.17608461 50.83749252 Myo-inositol 15.40331983 5.791723357 6.175803069 2.607753015 1.351324376 8.012289174 6.226527939 0.871325982 6.644245759 10.77341895 2.801262191 2.764585027 3.66235435 0.217581566 1.915179923 5.824666749 2.97106408 7.221831207 2.803440744 3.577272685 3.919852303 4.920408256 0.989515662 4.345757837 O-Phosphoethanolamine 82.78953137 16.26894029 24.57574305 10.95700493 8.370892244 49.45896464 21.84376481 17.64465043 19.16938036 41.317367 6.986770489 6.891384572 9.313294015 0.278331656 4.974372837 13.83329909 7.449536513 15.82737286 5.009185329 7.949139737 8.039423988 10.45508317 2.523626615 11.12134094 Lactate2 82.78953137 16.26894029 24.57574305 10.95700493 8.370892244 49.45896464 21.84376481 17.64465043 19.16938036 41.317367 6.986770489 6.891384572 9.313294015 0.278331656 4.974372837 13.83329909 7.449536513 15.82737286 5.009185329 7.949139737 8.039423988 10.45508317 -2.523626615 11.12134094 Sn-Glycero-3-phosphocholine 13.35663452 3.562964121 9.905384305 2.812189555 0.461224376 14.62635259 8.578989676 8.533431815 19.93800205 13.27113628 2.292901444 2.44663924 2.691354812 -0.125226163 0.867431563 4.722516235 6.143003273 15.97169062 2.762365071 12.71924758 3.817682216 9.223872954 1.572568345 3.64749197 Beta-Glucose 0.048536007 0.072773922 0.035003949 0.157760562 0.680073381 0.015842566 0.008197824 3.015371938 0.017374343 0.039448278 0.01480645 0.019576132 0.061040724 0.001964188 0.015432921 0.143620086 0.039798765 0.015499627 0.028939224 0.006516287 0.015247647 0.019798498 0.005461525 0.005035087 Alpha-Glucose.1 12.43153145 3.528391442 0.427125206 3.087413977 6.255047169 0.017200272 0.153415588 1.125059574 0.000286741 0.550789756 0.107880284 0.000652909 0.128948826 0.070213469 0.0651134 0.007987644 0.074260678 0.139219961 0.091215516 0.127810958 0.019781089 0.056939456 0.145665324 0.030660167 Maltose 0.384315039 0.438640369 0.996194307 1.302356141 0.805629902 0.297162298 0.358074311 0.799203399 0.09493411 0.282962001 0.522924874 0.005899847 0.626593537 0.028126059 0.455921415 1.1189754 1.034721026 0.535287906 0.921542564 0.098548634 0.187963138 0.375380358 0.917252183 1.025407179 UDP-sugar 3.822115007 1.839790175 0.754904535 1.274970267 0.458626708 1.282780846 1.129674463 0.641429481 3.138531858 1.224013073 0.266926394 0.085432234 0.015789538 0.078635962 0.069578657 0.312498978 3.03900183 0.616566329 0.054462933 1.220907208 0.221697487 0.846995987 0.614541469 0.198296949 Fumarate 0.417133294 0.377982546 0.250139079 0.238342401 0.092469546 0.253731639 0.240981606 0.283283157 0.238689097 0.242552326 0.008156352 0.029805503 0.025109849 0.009140511 0.017008348 0.038929921 0.137594197 0.130457372 0.081044778 0.088093489 0.072140667 0.145669271 0.083895322 0.02422111 Tyrosine1 2.844083458 0.957461552 0.591471953 2.021340634 0.414924109 0.298520004 0.121729669 0.31495029 0.414697868 0.475080076 0.00039102 0.013796008 0.020041103 0.041053271 0.002931594 0.010437395 0.124599248 0.440654172 0.059851529 0.364607796 0.024206886 0.531672919 0.025165733 0.010351202 Histidine1 36.92957695 12.46405551 8.771260079 5.444530584 0.3879209 4.055259503 9.777334529 4.961426028 8.25961483 6.960603963 0.236626455 0.005880357 0.258448935 0.296904705 0.192883092 0.042309217 2.568250308 11.05781018 2.544898689 7.735298128 2.433299811 4.73831444 4.538590406 0.421134677 Tyrosine2 3.141792879 1.329132528 1.014706155 2.569406045 0.614388496 0.44027959 0.108858581 0.220185018 0.69981637 0.753830538 0.392233239 0.030470941 0.228802134 0.235796088 0.087903729 0.26305768 0.414705963 0.773286016 0.412963549 0.824348909 0.137812929 0.681830203 0.30502682 0.099092854 Phenylalanine 3.981306284 2.062335641 1.239825177 3.617444129 1.217884444 0.851912941 0.232342334 1.54719829 1.460955879 1.516178431 0.594175923 0.23329593 0.305958155 0.37859393 0.397447314 0.202232736 0.484204599 0.256731829 0.070142295 0.506729975 0.688902295 1.143267487 0.954702469 0.816449897 Benzoate 3.516666554 4.992950595 2.736998347 3.996714634 1.750846149 2.343078943 3.01386257 2.007184049 2.630289021 3.201362961 2.37062498 1.186555555 3.416121357 1.208107869 1.925164976 2.45586409 3.978282143 1.887957981 3.464681904 2.503457932 2.819115308 2.428900077 3.619236902 1.99265559 Pyridoxine? 0.734830373 0.870309429 0.440672951 0.511322174 0.169605096 0.473532007 0.284275158 0.253136735 0.431704745 0.531746658 0.356917502 0.328087453 0.380492795 0.049957656 0.224724585 0.41381462 0.495020812 0.251361905 0.31343456 0.41744174 0.269669208 0.545343368 0.397561303 0.32355352 Histidine2 40.71864037 14.99085825 12.99572497 6.623366457 2.018296944 5.79003813 10.07066816 6.100116926 11.3558847 10.00400138 2.452221544 0.460712894 2.455071372 1.515740435 1.380898991 1.528730569 4.591031245 11.9594073 4.478820299 10.6509087 3.973642148 8.235332685 7.644008433 2.414156775 Inosine 3.22952703 0.69242458 1.472103228 0.71118653 0.431906118 4.275042786 1.036415284 1.951131846 1.915808476 2.804012119 0.152137168 0.055877314 0.417147578 0.051273219 0.128017955 0.310497891 0.169511062 0.050598923 0.166724296 0.262193018 0.200172902 0.624752605 0.349479122 0.390467485 Formate 6.029614035 3.645201482 4.830750266 3.080552811 1.652829597 5.29917942 2.774568303 2.634649701 2.838477577 5.574773558 5.036650614 2.430934622 5.070364377 3.635060142 3.894076477 5.270619692 1.794893506 1.436493485 3.335581968 3.831668532 2.915354333 2.65196187 5.07406804 3.827388604 NMR_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Leucine Valine1 Isoleucine Valine2 Isobutyrate 3-methyl-2-oxovalerate Lactate1 Alanine Arginine Lysine Acetate Glutamate1+Proline Glutamate+Glutamine+methionine+O-Acetylcholine 2-Aminoadipate Glutamate2 Pyruvate Succinate Glutamine2 Creatine1 Creatinine Malonate? Glycerol Glycine 3-methyl histidine Betaine Aspartate1 Aspartate2 Creatine2 Myo-inositol O-Phosphoethanolamine Lactate2 Sn-Glycero-3-phosphocholine Beta-Glucose Alpha-Glucose.1 Maltose UDP-sugar Fumarate Tyrosine1 Histidine1 Tyrosine2 Phenylalanine Benzoate Pyridoxine? Histidine2 Inosine Formate METABOLITES_END #END