#METABOLOMICS WORKBENCH ryant_20201211_122551 DATATRACK_ID:2352 STUDY_ID:ST001626 ANALYSIS_ID:AN002661 PROJECT_ID:000000 VERSION 1 CREATED_ON December 15, 2020, 11:26 am #PROJECT PR:PROJECT_TITLE Tissue-specific 1H-NMR metabolomic profiling in mice with adenine-induced PR:PROJECT_TITLE chronic kidney disease PR:PROJECT_TYPE Metabolomic profiling of the heart, liver, kidney, and skeletal muscles obtained PR:PROJECT_TYPE from mice with and without CKD via 1H NMR PR:PROJECT_SUMMARY This project is focused on a metabolomic analyses of the heart, liver, kidney, PR:PROJECT_SUMMARY and skeletal muscles obtained from mice with and without CKD. To accomplish this PR:PROJECT_SUMMARY objective, we extracted tissues from mice with CKD induced by long-term (24 PR:PROJECT_SUMMARY week) adenine-supplemented diet as well as their control-diet fed counterparts PR:PROJECT_SUMMARY with normal kidney function. Metabolites were extracted from tissues and 1H PR:PROJECT_SUMMARY nuclear magnetic resonance (NMR) was performed and coupled with multivariate PR:PROJECT_SUMMARY statistical analysis. PR:INSTITUTE University of Florida PR:DEPARTMENT Applied Physiology and Kinesiology PR:LABORATORY Rm 42 and Rm 43 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 This research was funded by grants from the National Institutes of Health and PR:FUNDING_SOURCE the National Heart, Lung, and Blood, Institute numbers R01-HL149704 (to T.E.R.) PR:FUNDING_SOURCE and the American Heart Association grant number 18CDA34110044 (to T.E.R.). PR:PROJECT_COMMENTS CKD metabolomic study via NMR using mice model PR:PUBLICATIONS MDPI PR:CONTRIBUTORS Ram B. Khattri, Trace Thome, and Terence E. Ryan #STUDY ST:STUDY_TITLE Tissue-specific 1H-NMR metabolomic profiling in mice with adenine-induced ST:STUDY_TITLE chronic kidney disease - aqueous phase Heart (part-III) ST:STUDY_TYPE Metabolomic profiling of the heart, liver, kidney, and skeletal muscles obtained ST:STUDY_TYPE from mice with and without CKD via 1H NMR ST:STUDY_SUMMARY This project is focused on a metabolomic analyses of the heart, liver, kidney, ST:STUDY_SUMMARY and skeletal muscles obtained from mice with and without CKD. To accomplish this ST:STUDY_SUMMARY objective, we extracted tissues from mice with CKD induced by long-term (24 ST:STUDY_SUMMARY week) adenine-supplemented diet as well as their control-diet fed counterparts ST:STUDY_SUMMARY with normal kidney function. Metabolites were extracted from tissues and 1H ST:STUDY_SUMMARY nuclear magnetic resonance (NMR) was performed and coupled with multivariate ST:STUDY_SUMMARY statistical analysis. 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 1864 Stadium Rd, FLG 114, Gainesville, FL, 32611, USA ST:EMAIL ryant@ufl.edu ST:PHONE 352-294-1700 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 17 ST:NUM_MALES All ST:STUDY_COMMENTS CKD metabolomic study via NMR using mice model ST:PUBLICATIONS MDPI #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN C57BL/6J SU:AGE_OR_AGE_RANGE 13 months SU:WEIGHT_OR_WEIGHT_RANGE (32.86±1.21 g (control mice) vs 23.57±1.27 g (CKD mice), P < 0.0001, SU:WEIGHT_OR_WEIGHT_RANGE N=7/group). SU:GENDER Male SU:ANIMAL_ANIMAL_SUPPLIER Jackson Labs (Stock # 000664) SU:ANIMAL_HOUSING Housed in a temperature of 22 oC SU:ANIMAL_LIGHT_CYCLE 12-hour light/12-hour dark SU:ANIMAL_FEED Ad libitum (Casein control diet vs. adenine-supplemented diet to induce CKD) SU:ANIMAL_WATER free access to food and water (3-5 animals per cage). SU:ANIMAL_INCLUSION_CRITERIA (3-5 animals per cage). #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 - CKD1Heart_Aq Group:CKD RAW_FILE_NAME=CKD1Heart_Aq SUBJECT_SAMPLE_FACTORS - CKD2Heart_Aq Group:CKD RAW_FILE_NAME=CKD2Heart_Aq SUBJECT_SAMPLE_FACTORS - CKD3Heart_Aq Group:CKD RAW_FILE_NAME=CKD3Heart_Aq SUBJECT_SAMPLE_FACTORS - CKD4Heart_Aq Group:CKD RAW_FILE_NAME=CKD4Heart_Aq SUBJECT_SAMPLE_FACTORS - CKD5Heart_Aq Group:CKD RAW_FILE_NAME=CKD5Heart_Aq SUBJECT_SAMPLE_FACTORS - CKD6Heart_Aq Group:CKD RAW_FILE_NAME=CKD6Heart_Aq SUBJECT_SAMPLE_FACTORS - CKD7Heart_Aq Group:CKD RAW_FILE_NAME=CKD7Heart_Aq SUBJECT_SAMPLE_FACTORS - Con1Heart_Aq Group:Control RAW_FILE_NAME=Con1Heart_Aq SUBJECT_SAMPLE_FACTORS - Con2Heart_Aq Group:Control RAW_FILE_NAME=Con2Heart_Aq SUBJECT_SAMPLE_FACTORS - Con3Heart_Aq Group:Control RAW_FILE_NAME=Con3Heart_Aq SUBJECT_SAMPLE_FACTORS - Con4Heart_Aq Group:Control RAW_FILE_NAME=Con4Heart_Aq SUBJECT_SAMPLE_FACTORS - Con5Heart_Aq Group:Control RAW_FILE_NAME=Con5Heart_Aq SUBJECT_SAMPLE_FACTORS - Con6Heart_Aq Group:Control RAW_FILE_NAME=Con6Heart_Aq SUBJECT_SAMPLE_FACTORS - Con7Heart_Aq Group:Control RAW_FILE_NAME=Con7Heart_Aq #COLLECTION CO:COLLECTION_SUMMARY While under isoflurane anesthesia, tissues were rapidly dissected and snap CO:COLLECTION_SUMMARY frozen in liquid nitrogen and stored at -80°C until metabolite extraction. The CO:COLLECTION_SUMMARY following tissues were used in this study: kidney, liver, heart (left CO:COLLECTION_SUMMARY ventricle), skeletal muscle (quadriceps). Euthanasia was carried out by CO:COLLECTION_SUMMARY thoracotomy followed by cervical dislocation. CO:SAMPLE_TYPE Heart CO:COLLECTION_METHOD While under isoflurane anesthesia, tissues were rapidly dissected and snap CO:COLLECTION_METHOD frozen in liquid nitrogen and stored at -80°C until metabolite extraction CO:COLLECTION_LOCATION University of Florida, Applied Physiology and Kinesiology, 1864 stadium RD, CO:COLLECTION_LOCATION Gainesville, FL 32611 CO:STORAGE_CONDITIONS -80℃ CO:STORAGE_VIALS cryovials #TREATMENT TR:TREATMENT_SUMMARY To induce CKD, we utilized an established adenine-diet model. Briefly, mice were TR:TREATMENT_SUMMARY assigned to a casein-base chow for 7-days, followed by induction of renal TR:TREATMENT_SUMMARY tubular injury by supplementing the diet with 0.2% adenine for 7-days, and TR:TREATMENT_SUMMARY subsequently maintained on a 0.15% adenine diet for 5 months and two weeks. CKD TR:TREATMENT_SUMMARY mice were then placed back on casein control diet for 2-weeks prior to TR:TREATMENT_SUMMARY euthanasia and terminal experiments, allowing for a washout period of adenine TR:TREATMENT_SUMMARY based chow. Control mice received casein diet for the duration of the study. TR:TREATMENT_SUMMARY Total duration of CKD encompassed 6-months. TR:ANIMAL_VET_TREATMENTS none TR:ANIMAL_ANESTHESIA isoflurane TR:ANIMAL_FASTING non-fasted TR:ANIMAL_ENDP_EUTHANASIA Euthanasia was carried out by thoracotomy followed by cervical dislocation. TR:ANIMAL_ENDP_TISSUE_COLL_LIST kidney, liver, heart (left ventricle), skeletal muscle (quadriceps) #SAMPLEPREP SP:SAMPLEPREP_SUMMARY A modified form of FOLCH extraction protocol was used to extract metabolites SP:SAMPLEPREP_SUMMARY from the tissues. Wet weights of all tissue samples were recorded prior to SP:SAMPLEPREP_SUMMARY extraction. Tissue samples were immediately homogenized to prevent any possible SP:SAMPLEPREP_SUMMARY enzymatic action using 1 mL of ice-cold methanol in a PowerLyzer 24 Homogenizer SP:SAMPLEPREP_SUMMARY (QIAGEN Group, Hilden, Germany). The mixture was centrifuged using 13,200 rpm at SP:SAMPLEPREP_SUMMARY 4oC for 30 minutes and the resulting supernatant was transferred to a new glass SP:SAMPLEPREP_SUMMARY vial consisting 3 mL of ice cold chloroform:methanol (2:1, v/v) mixture. The SP:SAMPLEPREP_SUMMARY homogenate was vortexed and left in an ice bath for 15 minutes to allow for SP:SAMPLEPREP_SUMMARY phase separation. Next, 1 mL of 0.9% of saline was added, vortexed it for couple SP:SAMPLEPREP_SUMMARY of minutes followed by a second incubation in an ice bath for 30-45 min for SP:SAMPLEPREP_SUMMARY complete phase separation. The upper aqueous layer was transferred to a new SP:SAMPLEPREP_SUMMARY falcon tube. To the remaining organic phase sample, 1 mL of 0.9% of saline was SP:SAMPLEPREP_SUMMARY added again followed by vigorous mixing and letting it stand in ice bath (15 SP:SAMPLEPREP_SUMMARY minutes) for a second phase separation. This second aqueous phase was combined SP:SAMPLEPREP_SUMMARY with the first. The resulting aqueous and organic layers were dried separately. SP:SAMPLEPREP_SUMMARY The aqueous layer was dried overnight with a Labconco freezer dryer (Labconco SP:SAMPLEPREP_SUMMARY Corporation, MO, USA) and the organic layer was dried via inert nitrogen gas. SP:SAMPLEPREP_SUMMARY These two dried powders (aqueous and organic phases) were stored at -80oC until SP:SAMPLEPREP_SUMMARY performing NMR experiments. SP:PROCESSING_METHOD Lyophilization and Homogenization SP:PROCESSING_STORAGE_CONDITIONS -80℃ SP:EXTRACTION_METHOD Modified FOLCH extraction 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 CKD1Heart_Aq CKD2Heart_Aq CKD3Heart_Aq CKD4Heart_Aq CKD5Heart_Aq CKD6Heart_Aq CKD7Heart_Aq Con1Heart_Aq Con2Heart_Aq Con3Heart_Aq Con4Heart_Aq Con5Heart_Aq Con6Heart_Aq Con7Heart_Aq Factors Group:CKD Group:CKD Group:CKD Group:CKD Group:CKD Group:CKD Group:CKD Group:Control Group:Control Group:Control Group:Control Group:Control Group:Control Group:Control Incaproate 2.080664953 3.551155646 4.969544894 2.950483712 2.874338663 2.177291613 3.727701594 4.061899861 7.37794019 4.506655851 6.514673126 4.062614403 4.580415839 3.273879633 Pantothenate? 0.722743189 1.276199179 1.813956279 1.547175701 1.303068691 1.310750064 1.644195821 1.131344653 1.754845164 1.123765648 1.782972608 1.048732534 1.094878437 0.990962933 Pantothenate2? 0.706362087 1.132654655 1.581986193 1.473542085 1.160751109 1.310940428 1.616940067 0.986778996 1.589142528 1.110042644 1.58379154 0.999315045 1.087373976 0.989988213 Leucine 3.338618563 3.968360564 7.358110155 5.327400312 5.341515903 4.940472008 6.744774535 6.557007365 11.69948318 7.592544022 10.77548365 6.651396006 7.448569437 6.988445736 Valine 1.261103481 1.465941038 2.703001293 1.821445456 2.213186673 1.9812008 2.44221607 2.427306153 4.474738834 2.864431042 4.023986089 2.484562944 2.727661997 2.915440612 Isoleucine 0.718463077 0.875987556 1.652422665 1.165559672 1.282199088 1.112769557 1.400042091 1.423519478 2.784395074 1.755530767 2.588356111 1.513591135 1.538443123 1.595223015 Valine2 1.258281331 1.471361263 2.876486957 2.100787384 2.290614357 1.97144545 2.410024532 2.563252516 4.783442617 3.023887578 4.589234881 2.623038488 2.93633112 2.859807525 Isobutyrate1 0.16590764 0.19489834 0.407568102 0.301116422 0.377052588 0.282263477 0.312148559 0.30516825 0.675598139 0.411405685 0.68298395 0.385480938 0.328517104 0.305978926 Isobutyrate2 0.267378175 0.343153912 0.673876136 0.521610789 0.554056629 0.418569235 0.493908052 0.496040697 1.099120401 0.644632943 1.099403499 0.60212844 0.552432311 0.509889311 Lactate1 64.29879974 108.5561389 133.3998442 176.8619844 180.9227125 174.5036859 158.1212432 80.21643986 151.3863259 114.0422651 176.9455899 121.4300691 130.5703568 156.0379144 Alanine 11.9598142 12.4126909 22.38056787 21.46303133 23.59460396 22.24438694 20.03277349 18.24113989 35.40126551 21.43503136 33.64837249 22.4201277 22.11246652 24.87995603 Lysine 3.042118155 6.618868038 12.25834965 8.880229219 10.35054879 9.023418514 10.84673959 9.175811503 17.58150099 11.698628 17.92786591 10.27626327 10.20247058 11.27541834 Acetate 2.007502777 4.587554424 9.442309673 8.299475475 6.024607088 3.594621065 5.468851919 5.866707822 13.78480549 8.138504806 12.80792538 6.816839181 8.086344877 3.645929624 Glutamate 7.594810298 11.49580315 22.33335719 15.68619329 21.14025796 20.27199377 18.9201745 23.38703505 31.4885809 23.10143735 28.10260111 21.16363937 18.35898344 20.12221115 Glutamate2+Glutamine1+Methionine+O-Acetylcholine 20.05971571 36.39174164 54.0896106 40.98225892 49.15678893 53.27081868 53.3679273 59.9577767 64.8559261 64.044615 70.11818952 45.46161043 43.53003773 65.34031282 Acetone 0.81254657 1.992305344 3.353320058 2.11661615 2.151601121 1.824918279 2.252587814 2.143812403 4.589663071 2.445346967 4.232205241 2.402811519 2.220295627 1.998778526 Glutamate2 10.67262978 11.65781676 26.99165899 17.32564013 26.69947359 25.29607509 22.27138496 29.42219481 38.34450172 28.48043667 31.24172895 26.09975076 22.70663817 24.20392693 Pyruvate 3.236169801 4.995096948 9.091876106 6.94722751 5.592942537 4.538044352 6.151980403 7.389106298 15.64384958 8.175957704 15.5705239 7.376357682 7.087945563 5.916524547 Succinate 4.106005313 7.735423782 11.77624156 8.880275926 9.189824709 14.01998645 13.09124775 10.7464876 12.17741425 11.36454778 13.99849556 10.02176011 10.15685938 12.56059486 Glutamine2 13.5358203 30.95516715 42.6565516 32.28584991 35.39905425 37.92041557 40.72095043 46.54892514 50.5842311 48.99838622 59.44344515 33.21167487 32.24122229 52.15427889 Unknown2.54to2.56ppm 3.698366114 14.75028472 13.64686951 16.98966861 13.67390392 9.69564571 15.52450846 15.15663442 19.76365018 12.04277892 25.06614058 11.02425304 11.1477807 13.00938136 Aspartate 0.739751599 2.123800887 4.24570276 2.76000833 3.323733544 2.824806733 3.091897422 3.798568508 6.665718806 3.200670182 6.046830837 3.003654132 2.956297137 3.147628567 Aspartate2 0.673420723 2.101770674 3.835737852 2.510912612 2.871860645 2.513514342 2.745310104 3.25797716 5.733960347 2.905344778 5.510907524 2.761997665 2.56819855 2.725082424 N,N-Dimethylformamide 0.722267895 2.071654668 2.924859012 4.301579141 2.070808275 1.84939486 2.458813937 2.290886389 3.964369244 2.058476213 3.770493995 2.219500937 2.03015473 2.01290777 Creatine1 39.89257167 54.15434204 100.6071995 63.64361581 86.79838669 88.3805875 93.62240281 76.41516609 118.0788689 103.7860259 109.6632095 84.12059819 80.80498851 100.3227974 Creatinine 2.800847576 5.255764745 8.225041942 4.708941057 6.417966134 8.592494472 5.442425779 5.205348285 10.41126758 8.637366837 8.752323818 7.206143816 5.091189285 5.723446761 Unknown3.06to3.17ppm 15.47974427 49.31301114 53.22349683 53.12005041 44.99247925 38.15719428 54.6274839 49.35218262 72.20127096 45.01730722 86.73197539 41.71061138 40.12816533 45.01520264 O-phosphocholine 6.591159101 7.815508071 22.66676906 18.03719485 11.77400745 12.62734117 17.3124775 13.15612038 20.86345436 17.44752847 26.23575381 10.0263387 15.8612705 11.10452403 Sn-Glycero-3-Phosphocholine 10.38572557 12.36196303 34.04926286 21.94928839 20.06544591 16.99729714 23.19417553 21.17763153 35.43785229 28.11806073 44.00551607 19.94162417 28.14585319 27.63375797 Trimethyl-N-oxide 78.71447325 198.3499073 247.1110552 184.8177526 159.5294882 111.2489115 210.1108534 129.9119933 280.2938562 143.5058112 303.1617906 159.5105985 145.2544327 119.4846207 Taurine1 91.91554606 138.7497211 221.9868935 153.5078794 174.0821763 210.60056 221.4287663 143.6383926 273.149001 213.9099767 245.9118279 179.733623 171.1107328 197.4673835 Taurine2 85.18247795 123.9065142 207.7177381 132.2090247 171.6861771 181.1039786 207.5677932 143.0504033 241.2186382 206.8121337 231.5709804 174.4606064 168.4621821 199.3787944 Glycine 1.82275761 3.579339876 4.671925216 4.210757194 3.097270504 3.218173212 4.251181056 3.072129867 5.581869487 3.108299737 5.698286005 3.317269653 2.896971873 2.6596197 Creatine2 32.57494303 46.56698041 84.39368405 54.03619877 71.23141412 74.12416285 76.22843738 64.33879277 98.74321198 85.69833007 92.24145378 68.99477846 66.05434333 81.22469345 Lactate2 11.64304496 26.74707535 32.90834788 40.12857731 39.52374845 39.27916287 36.54244859 20.69341427 38.06595901 26.78725294 41.9119946 27.96483721 27.87665288 33.51035258 Adenosine1 0.902598703 2.973922812 8.613146095 3.517254264 5.494011989 3.5666769 5.05039824 7.397410506 9.503913674 6.130623739 8.642388907 5.68881876 3.207662705 4.293825415 ATP/AMP1 2.705367322 7.241175245 10.96521729 8.327444383 9.103798775 10.90485612 11.0292449 9.05909927 12.72565977 11.72538931 13.97801499 8.393952333 9.962741795 11.94886905 Adenosine2 0.18350733 1.312368395 4.749530225 1.617548634 2.783785303 2.118381248 3.406980356 5.188511277 6.08557815 3.846171069 5.121343407 3.708136201 1.342584946 2.162515481 ATP/AMP2 0.839161048 2.470908609 3.514803177 2.906497947 2.907220909 4.462208797 4.282383761 3.067684687 4.633970178 4.133373169 4.957290646 2.900082248 3.239038934 4.089479955 Glucose1 0.18023259 0.151848346 0.020542849 0.178727835 0.064732548 0.101454322 0.004951148 0.069946715 0.005322542 0.040335714 0.020474946 0.017310408 0.094938633 0.007684031 Glucose2 0.339015759 0.772352719 0.216951335 1.162483294 0.392639871 0.657277034 0.319719835 0.494856037 0.267255787 0.135237967 0.193713955 0.248510353 0.519485948 0.219408206 Glucose-1-Phosphate 1.576664996 0.770887476 0.401421237 2.47032134 0.236860676 0.766083357 0.16416581 0.415247465 0.110989312 0.069506672 0.277721812 0.04336097 0.120966027 0.190406275 NAD+1 0.295395338 0.473914489 0.88991857 0.766541973 0.87742108 0.96701533 0.964538028 1.081766531 1.062934774 1.230451706 1.050876546 0.981279669 1.081967969 0.900431507 NAD+2 0.565597143 0.471295808 1.674801831 0.778822237 1.392031177 1.255943199 1.285573513 2.219203613 2.065211899 1.931739907 2.323483419 1.613909836 1.06004643 1.362008436 ATP/AMP3 2.365120143 3.521505 4.957338688 4.249924522 4.758652677 6.121711171 5.508347971 3.95933405 5.817485416 6.283955431 6.804469474 3.882027033 5.837538016 6.530898973 Fumarate 0.439671928 0.403599264 0.854710433 0.617302931 0.778290708 0.512854537 0.682157808 0.605555384 1.053604371 0.647582313 1.233172394 0.569280946 0.597433668 0.67953336 Tyrosine1 0.239394575 0.150179142 0.3991574 0.200134981 0.519248537 0.620920736 0.57964042 1.206815407 1.228414954 0.803971868 0.980312347 0.745429699 0.959567327 1.672484662 Histidine 0.27891207 0.208733745 0.480196036 0.373211218 0.518025456 0.609365952 0.448089336 0.61194977 0.780852818 0.68642374 0.782423505 0.571319736 0.536678025 0.694525121 Tyrosine2 0.390559223 0.298049061 0.678742123 0.348823633 0.673315226 0.81734961 0.866175694 1.473663673 1.806278168 1.077057106 1.331632933 0.962165047 1.218072086 1.972724417 Trytophan1 1.64485855 2.029055968 3.932136917 2.684857354 1.898739998 1.473140653 2.491755479 3.063417901 7.074553479 3.476336461 7.083205937 2.966953016 3.097038605 2.219576682 Phyenylalanine 0.383917449 0.484581453 0.824198285 0.362673014 0.597441733 0.68773368 0.775881845 0.886279005 1.53518045 0.869954795 1.164129234 0.562995642 0.63267012 0.895695298 Benzoate1 1.053573332 1.22480082 2.545221105 1.48123871 1.436669287 1.212298903 2.933899248 1.504525856 3.512425447 1.963761771 3.31110463 1.374609755 1.61850412 2.001751537 Nicotinurate1 0.511869203 0.575295197 0.837436262 0.565245618 0.904107799 0.863105145 0.810926753 1.039010633 1.642510173 1.023212639 1.147714768 0.553405027 0.577887977 0.945522045 Tryptophan2 1.088587628 1.321136406 2.533459251 2.046674945 1.32068318 0.992511838 1.551600282 2.210995167 5.460847758 2.529942379 5.031791541 2.057400721 2.131779406 1.466789707 Xanthine? 0.040881189 0.031178014 0.074536626 0.048681755 0.111738628 0.057516274 0.057230797 0.212414889 0.347691995 0.207296505 0.242177141 0.042921742 0.056117234 0.129169119 ATP/AMP4 0.569593954 0.364881698 2.129237252 0.321288978 1.172048439 0.642059378 0.952350803 2.365769447 2.163001527 1.592298855 1.870793377 1.314173919 0.706598803 0.910178708 Inosine1 1.567780013 2.384183841 3.73137917 3.039532039 3.393110324 4.080879577 3.768696258 2.754137319 4.649564004 4.56822156 4.718078687 2.774526764 4.067615934 4.111723725 Inosine2 1.109352489 0.711897989 4.539153274 0.76626804 2.668360056 1.35655237 2.144852517 5.245984304 5.234195726 3.490934704 4.398024732 3.160474924 1.253700278 2.024637233 NAD+3 0.378936982 0.57717188 1.012702698 0.925469649 0.869676188 1.051662619 1.198605104 1.248151874 1.177543561 1.282421706 1.148900091 1.016684338 1.123367438 0.987879246 Formate 0.675720691 1.053382696 1.6646659 1.432726112 1.82675681 0.811700221 1.103587762 1.433918079 3.348099487 1.560268786 3.705081822 1.29215267 1.812758283 1.256725685 ATP2 0.121399313 0.205397681 0.48417332 0.543457238 0.399672345 0.464234511 0.414274883 0.2198557 0.427161794 0.769699913 0.440007842 0.275836124 1.080277136 0.665846824 Unknown 0.090743604 0.129741941 0.04679326 0.33276133 0.172453447 0.323618479 0.283691681 0.509998686 0.343410622 0.244453099 0.352667203 0.222648997 0.18661524 1.190085206 ATP/AMP5 2.216025959 3.262058629 4.056245684 3.407349181 3.832009105 5.650405283 4.873197434 3.23819163 5.350553165 5.29874545 5.728029723 3.170058782 4.591960512 4.928710945 Nicotinurate2 0.358230361 0.399631387 0.551186264 0.605545285 0.706925335 0.527830135 0.671335116 0.709459536 0.872120097 0.654719151 0.791922965 0.453206028 0.485305075 0.745663199 NAD+4 0.315037853 0.424089214 0.824837882 0.791617545 0.711704715 0.913954566 0.98466385 1.059340613 0.932520646 1.144025587 0.906109658 0.887544561 0.907998508 0.834515389 Nicotinurate3 0.360904938 0.339931551 0.493878364 0.500193544 0.602286755 0.532271695 0.552076006 0.593833886 0.764567918 0.622577853 0.861568164 0.465209983 0.502417745 0.639766 NAD+5 0.386779083 0.507713528 1.015323363 0.881352597 0.763716063 1.066378928 1.138023727 1.108432978 1.102920393 1.161316099 1.068795171 1.004191923 1.084389343 0.872542451 NAD+6 0.382990059 0.579975768 1.027420164 0.914131107 0.79909853 1.010470083 1.152966556 1.194261403 1.170472366 1.183929153 1.259934051 1.011152962 1.074981239 0.934203232 NMR_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Incaproate Pantothenate? Pantothenate2? Leucine Valine Isoleucine Valine2 Isobutyrate1 Isobutyrate2 Lactate1 Alanine Lysine Acetate Glutamate Glutamate2+Glutamine1+Methionine+O-Acetylcholine Acetone Glutamate2 Pyruvate Succinate Glutamine2 Unknown2.54to2.56ppm Aspartate Aspartate2 N,N-Dimethylformamide Creatine1 Creatinine Unknown3.06to3.17ppm O-phosphocholine Sn-Glycero-3-Phosphocholine Trimethyl-N-oxide Taurine1 Taurine2 Glycine Creatine2 Lactate2 Adenosine1 ATP/AMP1 Adenosine2 ATP/AMP2 Glucose1 Glucose2 Glucose-1-Phosphate NAD+1 NAD+2 ATP/AMP3 Fumarate Tyrosine1 Histidine Tyrosine2 Trytophan1 Phyenylalanine Benzoate1 Nicotinurate1 Tryptophan2 Xanthine? ATP/AMP4 Inosine1 Inosine2 NAD+3 Formate ATP2 Unknown ATP/AMP5 Nicotinurate2 NAD+4 Nicotinurate3 NAD+5 NAD+6 METABOLITES_END #END