#METABOLOMICS WORKBENCH ryant_20201211_130105 DATATRACK_ID:2356 STUDY_ID:ST001630 ANALYSIS_ID:AN002665 PROJECT_ID:000000 VERSION 1 CREATED_ON December 15, 2020, 11:29 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 Quadricep (part-VII) 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 - CKD1Quad_Aq Group:CKD RAW_FILE_NAME=CKD1Quad_Aq SUBJECT_SAMPLE_FACTORS - CKD2Quad_Aq Group:CKD RAW_FILE_NAME=CKD2Quad_Aq SUBJECT_SAMPLE_FACTORS - CKD3Quad_Aq Group:CKD RAW_FILE_NAME=CKD3Quad_Aq SUBJECT_SAMPLE_FACTORS - CKD4Quad_Aq Group:CKD RAW_FILE_NAME=CKD4Quad_Aq SUBJECT_SAMPLE_FACTORS - CKD5Quad_Aq Group:CKD RAW_FILE_NAME=CKD5Quad_Aq SUBJECT_SAMPLE_FACTORS - CKD6Quad_Aq Group:CKD RAW_FILE_NAME=CKD6Quad_Aq SUBJECT_SAMPLE_FACTORS - CKD7Quad_Aq Group:CKD RAW_FILE_NAME=CKD7Quad_Aq SUBJECT_SAMPLE_FACTORS - Con1Quad_Aq Group:Control RAW_FILE_NAME=Con1Quad_Aq SUBJECT_SAMPLE_FACTORS - Con2Quad_Aq Group:Control RAW_FILE_NAME=Con2Quad_Aq SUBJECT_SAMPLE_FACTORS - Con3Quad_Aq Group:Control RAW_FILE_NAME=Con3Quad_Aq SUBJECT_SAMPLE_FACTORS - Con4Quad_Aq Group:Control RAW_FILE_NAME=Con4Quad_Aq SUBJECT_SAMPLE_FACTORS - Con5Quad_Aq Group:Control RAW_FILE_NAME=Con5Quad_Aq SUBJECT_SAMPLE_FACTORS - Con6Quad_Aq Group:Control RAW_FILE_NAME=Con6Quad_Aq SUBJECT_SAMPLE_FACTORS - Con7Quad_Aq Group:Control RAW_FILE_NAME=Con7Quad_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 Muscle 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 CKD1Quad_Aq CKD2Quad_Aq CKD3Quad_Aq CKD4Quad_Aq CKD5Quad_Aq CKD6Quad_Aq CKD7Quad_Aq Con1Quad_Aq Con2Quad_Aq Con3Quad_Aq Con4Quad_Aq Con5Quad_Aq Con6Quad_Aq Con7Quad_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 Isocaproate 1.004487345 3.197001653 4.503860186 3.232234434 3.007079553 7.146134247 3.110768999 2.467471529 5.097792078 4.872746872 2.550667391 5.304493421 5.095283935 6.117385873 Pantothenate1? 0.342135406 1.304993617 1.390652942 0.933342742 1.459397569 2.763270038 1.126172034 0.94807308 1.62614418 1.542018052 0.962919487 1.419550305 1.592694077 1.784600263 Pantothenate2? 0.20470385 0.985488281 1.029617847 0.769883741 0.823117455 1.461083 0.699631178 0.542414139 0.997900799 0.998981249 0.583925832 1.139633546 1.166736659 1.510225502 Leucine 2.791002092 10.34524765 10.00938251 9.516170244 12.66970422 13.02932589 10.55344608 6.869674479 16.19686982 12.30505435 10.64188782 14.27962741 14.45555115 17.74361657 Valine1 1.941652353 6.223906515 6.015594528 5.715287241 8.022706742 8.27388633 6.092667707 5.383328874 11.42483762 8.301990815 7.789033166 8.891824646 9.698399745 10.52134311 Isoleucine 0.747012769 2.759892269 2.610835916 2.473454107 3.258346136 3.5503301 2.712728107 1.839391753 4.315846836 3.438629981 3.044979349 3.969660956 4.036662538 4.767950163 Valine2 1.74380038 5.647260188 5.676646273 5.474860336 7.227016802 7.567824143 5.558977796 5.026528542 11.20599646 7.407470696 7.096242186 8.117766522 9.186816873 9.757669444 Isobutyrate 0.175191656 0.550919559 0.672358027 0.532662771 0.744993364 0.794848869 0.55625924 0.310914891 0.667232216 0.626618132 0.483834813 0.82652132 0.803231538 0.944719214 Isobutyrate2 0.223924423 0.656915146 0.735834438 0.611299144 0.858224296 0.944873173 0.644152048 0.333097535 0.788753205 0.752012312 0.593636256 0.981982769 0.870782315 1.142871829 Lactate1 219.0423378 553.7283946 573.1060124 436.4203988 640.3664335 509.2228164 504.0089764 516.7449413 772.9567075 617.1745367 550.0764956 665.9650305 586.6026019 524.4087055 Alanine 17.58701314 42.83758246 44.83143911 45.58434903 54.17232624 52.12947527 45.22528094 42.50376132 83.19675135 69.06963704 64.0841815 63.71345547 73.85582292 71.62280617 Lysine 11.06440248 7.905858835 13.77992142 7.45896818 29.30702768 19.20968521 28.0259363 11.7444976 22.80776913 16.86247983 18.68895255 21.12248014 11.88497564 19.23309912 Acetate 2.113286924 5.29852269 8.520017829 8.202248304 5.118291302 5.881374968 5.65672937 5.725040401 17.29020162 4.610571244 3.753694633 5.741550889 11.46641227 10.70727006 Glutamate1 2.924101757 6.651412232 6.629703773 7.521170045 10.90233637 12.48634803 7.46347935 12.24511433 20.66525074 18.63046388 13.95240262 12.18277197 12.79092666 20.42352473 Glutamine1+Glutamate 18.43537985 41.08067549 44.98420734 39.25002983 71.21138842 64.64045566 48.15033348 48.69353041 61.52516329 70.24925154 62.45701764 63.66650593 46.76664214 58.45813122 Methionine+O-Acetylcholine 3.6061051 9.519588224 8.904067231 10.92649948 13.72498711 13.25800213 9.888443708 7.545119197 11.25469614 10.28481873 10.38476445 9.737373019 8.629713031 9.696210969 Glutamate 4.759240556 7.252674216 10.70350887 10.96141633 20.63116122 21.0636059 13.76446888 18.88583218 33.00012785 29.72114955 24.21303199 23.86936367 17.56110307 33.71624499 Pyruvate 0.885023559 3.831424067 4.349766159 4.172945186 6.925651309 5.113671773 4.590848725 1.915061833 8.689908379 4.406310387 3.597923166 9.359420576 6.112163478 8.171453989 Succinate 2.008924072 2.431076025 4.398989106 4.15306457 10.36028687 5.249733442 5.002561191 4.988537667 9.290629043 6.682878794 7.152432394 12.95850799 3.600570285 10.00621181 Glutamate2 12.64137416 28.59830843 32.54659242 24.45974953 55.3762515 43.63586589 32.29539072 37.55313569 46.23268989 52.76704695 42.73748057 51.20380216 29.99765809 37.77104276 Citrate1 0.355026967 0.534016626 1.105564129 0.745366165 1.474840252 1.675778008 1.281759616 1.216560445 2.806211268 1.998476118 1.462213577 1.930971089 1.997559442 1.881302304 Citrate2 0.905114997 1.222763661 2.144283401 1.479348714 2.589251109 2.085658097 2.071015824 1.942766425 3.438402921 2.93474031 2.56264781 2.473965515 2.700114143 2.954492293 Unknown2.57-2.54 3.122259741 5.259150993 6.748305129 8.722304765 8.551588682 11.25705393 6.883630909 5.475160736 8.992070777 6.788248465 5.525028219 7.097412392 6.582545848 32.72635565 Anserine1 81.41607082 162.2687019 149.0959159 162.9148778 136.9943032 120.2616383 131.2448546 129.0177975 167.0821423 130.8146304 128.3028962 113.9902323 175.6316879 182.1772581 Aspartate1 0.151256679 0.344041383 0.190679786 0.991874703 0.026423352 0.358871626 0.254226889 0.423704899 1.260439354 0.177108907 0.340315291 0.288240582 0.176447007 0.395781791 Aspartate2 0.208694067 0.510066095 0.306126729 0.370696658 0.235023511 0.078619035 0.350157348 0.090179282 0.602864711 0.203134888 0.14801478 0.496230867 0.347011608 0.065153419 Trimethylamine 1.096628179 3.287727588 3.307335312 3.029561867 3.177658067 2.422193226 2.707177071 2.555011486 3.399925259 2.838182044 2.717241626 2.916569463 2.756351237 2.794981572 Creatine1 233.1064491 685.2629124 667.9036326 646.1020824 657.4451958 593.0632502 553.520706 529.5781464 699.801549 614.4657557 577.7678307 608.3003388 551.8948813 600.5385099 Creatinine 42.89994549 71.87322712 83.37761108 75.38829523 89.24347959 68.79480384 123.4939262 84.25642262 67.8377304 78.3256979 75.33360644 80.71037424 107.9971214 109.8568484 Trimethyl-N-oxide 49.51276717 147.8087986 153.939222 148.5500761 52.40428719 244.6083966 178.3121125 114.5449078 254.498817 192.980916 179.560229 220.7058986 232.8518859 164.6695677 Taurine1 340.5601182 804.8283479 739.9992431 692.4341083 888.2537437 760.4497238 742.0690206 523.8712856 762.7048194 735.6471077 707.1283318 719.09816 694.574262 797.6495414 Taurine-2 307.1566113 796.051376 741.4357871 698.7316894 745.8950064 777.3997251 715.3533848 534.9909261 751.5139897 729.4566947 696.0782296 712.4611424 697.3389317 806.3875364 Glycerol1 2.5224887 5.417408197 7.014001179 5.696191734 4.584795044 6.833298836 6.839398004 2.536859585 4.314978561 4.672409713 5.06237865 4.876316339 3.877435285 4.269546099 Glycerol2 2.23980621 2.949456624 5.281234528 2.948170791 3.845170171 6.117958742 6.137119242 1.923795318 3.502069183 3.985873362 5.720112649 3.786702311 4.348279137 3.152201998 Glycine 6.792101331 12.4937735 21.65880771 12.00244809 17.34000438 22.69489258 22.14511132 11.9743609 24.42086039 22.78979979 23.00321141 15.8636686 29.08587495 15.31081065 Glycerol3 2.056367223 3.564478243 5.96826304 3.272881756 5.027476523 5.378436868 7.018897926 3.025425961 5.192419596 4.347424953 4.425486587 5.205147696 6.03076383 4.419357089 Glycerol4 2.703424382 3.393864418 8.172831068 3.692518044 6.906065929 8.819008439 11.07640391 3.59351577 7.663363162 5.24995776 6.280954638 6.492167748 7.81758467 4.152259475 Anserine2 70.60855735 192.0745891 174.1637501 179.2305907 169.2812182 167.6605667 168.3575693 146.1862855 155.8467613 154.1475047 160.5334392 144.6794492 178.4053121 172.839807 Creatine2 200.9442409 521.5889712 511.9625302 472.1397512 555.0005619 509.3512237 468.4442007 400.9519428 507.1253592 480.1936822 473.9323777 511.733973 460.5182668 511.6070611 Creatine Phosphate? 9.486214247 20.83252155 37.21982439 24.16272114 24.08746402 32.42497978 41.6028592 33.14036189 18.69518923 31.3942471 26.24594675 21.23853231 42.6771622 35.05447507 Lactate2 43.2535517 105.0584958 103.920864 67.38554813 133.9520128 119.6806905 100.8498883 92.92380231 115.858039 127.7602355 102.8615986 130.6369529 112.8096978 128.714663 ATP+AMP1 7.511980966 19.28853281 10.84363226 12.26983705 20.25865036 19.46916049 10.80857151 16.03352497 29.64639393 31.49182706 15.10548475 22.57049581 11.65448209 40.87787735 Anserine3 7.999273546 34.84596003 27.36398979 32.13785026 29.1139994 26.66157842 28.61159405 30.67470863 56.38330521 43.36430831 21.32865748 35.07474349 39.82855468 50.83896869 Glucose1 0.539639379 3.536895628 3.463113561 1.160831281 1.361434956 0.373343502 3.809751495 2.093602127 8.491263595 1.720464811 2.117355078 4.790224186 5.050984706 0.182390384 Mannose 2.037550445 0.463320658 1.312127244 0.546338029 4.727353629 4.624029719 2.357941091 0.754998332 3.659061272 0.908286899 3.943304856 1.529012614 2.167149313 2.012523148 Glucose2 with Hexose 4.066694227 3.276499199 6.209773556 3.752752134 11.22195431 12.78777988 8.527546088 3.532726306 11.84990622 5.454081061 9.307132135 5.791681173 7.296949668 5.4842405 Allantoin 0.692487083 0.533297571 0.283540484 0.633686102 1.444155509 2.15498492 0.203720988 0.034968432 1.194745678 0.113759531 1.220441296 0.036534842 -0.028105975 1.18077554 Glucose-1-Phosphate 1.937371656 0.289125253 0.434282203 0.696036255 4.203500907 5.57929512 0.57861114 0.519494824 4.502615123 0.484549619 4.173938342 0.487625344 0.778020174 3.771457441 NAD+1 1.144722465 1.553038561 1.807393073 1.775923144 2.782957934 3.47518134 2.215106117 2.065034596 3.444081983 2.51399822 2.905495261 2.705933114 2.644131455 5.25809865 NAD+2 1.261807766 1.945931185 1.875363292 1.973107231 2.997299468 3.25559848 2.43054903 1.937866582 3.21776041 2.859574355 3.038438805 2.962660224 2.37134435 5.639703291 ATP+AMP2 13.93081549 21.51810388 11.34945782 12.55146622 20.42690333 14.95915062 11.04832078 12.89073499 22.190737 18.68329127 18.05586913 20.59321653 11.05756959 29.8219981 Fumarate 0.206990302 0.361367189 0.513260594 0.699920858 0.776654959 0.734758471 0.485848411 0.497563485 1.027907386 0.705833139 0.981619387 1.419535263 0.487201287 1.209105681 Tyrosine1 0.633064076 1.451022602 1.436183 1.202755882 2.061891169 2.38518602 1.951796479 4.149192438 4.020801985 2.959323054 2.790980091 3.362382877 3.680462723 6.870192136 Anserine4 12.31761779 47.11933198 33.50504232 41.59682951 30.1550654 30.18422214 30.39467194 31.07454873 33.55173557 28.04766374 30.08074849 25.27594894 34.22493895 33.79398511 Histidine1 3.882156017 7.914583372 11.06909638 7.424049511 12.66026341 8.615924405 10.00360519 12.50756482 14.98221287 13.13291929 12.81564845 10.19854865 18.18088699 16.38202768 Tyrosine2 0.723988345 1.539894403 1.789814481 1.582996655 2.717352267 3.143504024 2.388132434 5.250257643 5.178121146 3.788632342 3.343073959 4.298035576 4.558960055 7.790897569 Tryptophan1 0.553459324 2.605411817 2.669227385 2.523649183 3.670015153 3.208559519 2.822977621 1.071456728 4.404998322 2.461960678 1.999498373 5.220504445 3.912803671 5.712356655 Phenylalanine 0.465779803 1.492129448 1.609367428 1.655582328 1.858852772 2.101583312 1.653748718 1.537640892 1.979812676 1.545879629 1.439839238 2.245035078 1.908929924 3.444993767 Benzoate 0.315499252 1.296600359 2.075186121 1.3246842 2.258486538 2.083236343 1.864241225 2.65877174 2.087360638 2.357293212 1.258970893 2.421005925 2.191638873 3.386173962 Nicotinurate2 0.272431133 0.831812486 0.768467413 0.757714108 0.791334016 0.885731163 0.58049472 0.592800292 0.822749822 0.91053662 0.857913957 1.130245827 0.599348592 1.101905328 Tyramine? 0.077292991 0.106676977 0.189075401 0.143120784 0.197809608 0.179751943 0.169647164 0.257846914 0.224429513 0.149474397 0.203770131 0.276444739 0.252771841 0.423800553 Tryptophan2 0.323190933 1.582511264 1.647707055 1.510789436 2.56474365 2.056834503 1.844607783 0.479490426 3.038711557 1.503520988 1.299224313 3.543071513 2.60723959 3.520575706 Histidine2 2.753604372 5.433166634 7.411734145 5.216079363 7.448692087 5.646523471 7.281116818 10.20742215 12.01493165 9.251362845 10.34171359 6.925579526 14.03314358 12.88031592 Anserine5 12.10190274 45.02702512 31.66887732 37.81761204 29.86067442 27.87977079 28.7214027 29.08835449 32.92496523 26.28354681 30.00769456 24.79127735 30.97313449 31.35081375 ATP+AMP3 5.136995138 14.04353522 6.277147545 6.033458062 12.69834177 7.308886707 6.104731991 0.996654514 13.93568582 12.14990351 11.30018359 12.02149328 2.1687926 13.75853553 NAD+3 0.929843186 1.497393959 1.924463936 1.583675323 2.395423491 2.228980737 2.344234681 2.118360654 2.523580006 2.672958494 2.290391832 2.544595623 2.314877995 3.088574605 Formate 0.515408744 1.310378827 1.38641818 1.531913671 1.131525744 2.557584013 0.789274227 1.534999104 3.037062844 1.174655793 0.831914074 1.714808772 2.490714131 3.246614328 ATP+AMP4 7.358681148 18.99913877 8.344863804 9.704917373 17.57820318 10.57496181 8.329418181 9.990612575 18.47684772 16.52330231 15.14751921 16.61958853 6.619030959 18.55695388 Unknown8.59to8.60ppm 0.178473218 0.480583975 0.305042983 0.454402121 0.897066288 0.97158113 0.388368323 0.671978795 0.792033353 1.257632537 0.677835221 1.009467658 0.321902303 1.330617399 Nicotinurate3 0.343829394 0.608603196 0.478363304 0.562791642 0.791440104 0.710210496 0.475696538 0.561919887 0.865983556 0.818659724 0.83243944 0.716352026 0.457274532 0.558963297 NAD+4 0.967514129 1.360864207 1.731338078 1.394090321 2.304308303 1.97295713 2.184588031 1.919573631 2.225722983 2.508157599 2.162151612 2.257023736 2.114954616 2.704571539 Nicotinurate 4 0.264656048 0.562302888 0.439253335 0.395385267 0.618590669 0.561712484 0.440762609 0.498016821 0.678669537 0.651793258 0.815799389 0.664011902 0.462315326 0.62110627 NAD+5 1.107823451 1.687129518 2.019271299 1.676276254 2.720894148 2.339920819 2.627595801 2.162326811 2.661135994 2.966566976 2.56492925 2.785543433 2.547218943 3.231673493 NAD+6 1.100247962 1.586152765 2.09621979 1.726405828 2.73462156 2.462636376 2.607265626 2.35134741 2.961918944 3.127112482 2.615503767 2.810740536 2.648187164 3.486615238 NMR_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Isocaproate Pantothenate1? Pantothenate2? Leucine Valine1 Isoleucine Valine2 Isobutyrate Isobutyrate2 Lactate1 Alanine Lysine Acetate Glutamate1 Glutamine1+Glutamate Methionine+O-Acetylcholine Glutamate Pyruvate Succinate Glutamate2 Citrate1 Citrate2 Unknown2.57-2.54 Anserine1 Aspartate1 Aspartate2 Trimethylamine Creatine1 Creatinine Trimethyl-N-oxide Taurine1 Taurine-2 Glycerol1 Glycerol2 Glycine Glycerol3 Glycerol4 Anserine2 Creatine2 Creatine Phosphate? Lactate2 ATP+AMP1 Anserine3 Glucose1 Mannose Glucose2 with Hexose Allantoin Glucose-1-Phosphate NAD+1 NAD+2 ATP+AMP2 Fumarate Tyrosine1 Anserine4 Histidine1 Tyrosine2 Tryptophan1 Phenylalanine Benzoate Nicotinurate2 Tyramine? Tryptophan2 Histidine2 Anserine5 ATP+AMP3 NAD+3 Formate ATP+AMP4 Unknown8.59to8.60ppm Nicotinurate3 NAD+4 Nicotinurate 4 NAD+5 NAD+6 METABOLITES_END #END