#METABOLOMICS WORKBENCH cclish_20250908_095052 DATATRACK_ID:6396 STUDY_ID:ST004222 ANALYSIS_ID:AN007025 PROJECT_ID:PR002663 VERSION 1 CREATED_ON September 25, 2025, 3:37 pm #PROJECT PR:PROJECT_TITLE Loss of P. falciparum Amino Acid Transporter (ApiAT2) Function Increases PR:PROJECT_TITLE Intracellular Proline and Confers Resistance to Prolyl-tRNA Synthetase PR:PROJECT_TITLE Inhibitors PR:PROJECT_SUMMARY Plasmodium falciparum evades the antimalarial activity of proline-competitive PR:PROJECT_SUMMARY prolyl-tRNA synthetase (PfProRS) inhibitors, such as halofuginone (HFG), by a PR:PROJECT_SUMMARY unique resistance mechanism termed the Adaptive Proline Response (APR). The APR PR:PROJECT_SUMMARY is characterized by a marked elevation of intracellular proline following drug PR:PROJECT_SUMMARY exposure. Contrary to initial expectations, the APR is not mediated by PR:PROJECT_SUMMARY alterations in canonical proline metabolic pathways involving arginase (PfARG) PR:PROJECT_SUMMARY and ornithine aminotransferase (PfOAT). Instead, we identified loss-of-function PR:PROJECT_SUMMARY mutations in the Apicomplexan Amino acid Transporter 2 (PfApiAT2) as the primary PR:PROJECT_SUMMARY genetic driver of this resistance phenotype. Importantly, reversion of these PR:PROJECT_SUMMARY mutations to wildtype effectively suppresses the APR, establishing PfApiAT2 as PR:PROJECT_SUMMARY the molecular determinant of this novel resistance mechanism. The elucidation of PR:PROJECT_SUMMARY the APR significantly advances our understanding of antimalarial drug PR:PROJECT_SUMMARY resistance. By delineating the role of PfApiAT2 in this process, we establish PR:PROJECT_SUMMARY critical insights for the development of strategies to circumvent PfProRS PR:PROJECT_SUMMARY inhibitor resistance for future antimalarial therapies. PR:INSTITUTE Broad Institute of MIT and Harvard PR:DEPARTMENT Metabolomics Platform PR:LAST_NAME Clish PR:FIRST_NAME Clary PR:ADDRESS 300 Binney Street, Cambridge, MA, 02142, USA PR:EMAIL clary@broadinstitute.org PR:PHONE 617-714-7654 PR:FUNDING_SOURCE NIH-NIAID R01AI143723 and R21AI132981; NIH-NIGMS T32 GM008666 and F31AI129412; PR:FUNDING_SOURCE Bill and Melinda Gates Foundation OPP1132451 and OPP1086203; and the Harvard PR:FUNDING_SOURCE Defeating Malaria Initiative PR:PROJECT_COMMENTS 5 experiements PR:CONTRIBUTORS Selina Bopp, Lọla Fagbami, Amy Deik, Claudia Taccheri, Akansha Pant, Madeline PR:CONTRIBUTORS Luth, Daisy Chen, Mark A. Tye, Imran Ullah, Robert Morris, Wilhelm Haas, PR:CONTRIBUTORS Elizabeth A. Winzeler, Clary B. Clish, Amanda K. Lukens, Ralph Mazitschek, Dyann PR:CONTRIBUTORS F. Wirth #STUDY ST:STUDY_TITLE Elucidating the source of elevated proline in APR parasites ST:STUDY_SUMMARY In this study, we use a custom Triplex labeled media, containing stable isotope ST:STUDY_SUMMARY labeled 15-N proline (Pro+1), 13C,15N-arginine (Arg+2), and 13C5-glutamine ST:STUDY_SUMMARY (Gln+5) to identify the source of elevated proline levels in APR parasites. ST:STUDY_SUMMARY Labeled proline levels were assessed in Dd2 (WT) parasites and Dd2-APR parasites ST:STUDY_SUMMARY (parasites previously selected with HFG that demonstrate increased intracellular ST:STUDY_SUMMARY proline or the APR). High levels of labeled proline (+1) implicate import from ST:STUDY_SUMMARY the extracellular environment as a primary driver of the APR; high levels of ST:STUDY_SUMMARY labeled proline (+2) implicate the arginine to proline biosynthetic pathway as ST:STUDY_SUMMARY the primary source; and high levels of labeled proline (+5) implicate the ST:STUDY_SUMMARY glutamine to proline biosynthetic pathway as the primary source. Unlabeled ST:STUDY_SUMMARY proline (+0) indicates proline derived from hemoglobin digestion (either ST:STUDY_SUMMARY directly as proline, or via the arginine and/or glutamine conversion pathways). ST:INSTITUTE Broad Institute of MIT and Harvard ST:DEPARTMENT Metabolomics Platform ST:LAST_NAME Clish ST:FIRST_NAME Clary ST:ADDRESS 300 Binney Street, Cambridge, MA 02142 ST:EMAIL clary@broadinstitute.org ST:PHONE 617-714-7654 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 7 ST:STUDY_COMMENTS Study 1 of 5 #SUBJECT SU:SUBJECT_TYPE Cultured cells SU:SUBJECT_SPECIES Plasmodium falciparum SU:TAXONOMY_ID 5833 SU:SPECIES_GROUP Unicellular parasites #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 - Study1_Dd2_iRBC_10h_R1 Sample source:infected RBC (iRBC) | Parasite_type:Dd2 WT | Parasite_line:Dd2 | Timepoint:10 h Replicate=1; Tracer=15N Pro 15N,13C Arg 5C13 Gln; Date Generated=5/13/2017; RAW_FILE_NAME(Raw_file_name)=0011_MAZ_FAG10_HIL-11.raw SUBJECT_SAMPLE_FACTORS - Study1_Dd2_iRBC_10h_R2 Sample source:infected RBC (iRBC) | Parasite_type:Dd2 WT | Parasite_line:Dd2 | Timepoint:10 h Replicate=2; Tracer=15N Pro 15N,13C Arg 5C13 Gln; Date Generated=5/15/2017; RAW_FILE_NAME(Raw_file_name)=0026_MAZ_FAG10_HIL-26.raw SUBJECT_SAMPLE_FACTORS - Study1_Dd2_iRBC_10h_R3 Sample source:infected RBC (iRBC) | Parasite_type:Dd2 WT | Parasite_line:Dd2 | Timepoint:10 h Replicate=3; Tracer=15N Pro 15N,13C Arg 5C13 Gln; Date Generated=5/18/2017; RAW_FILE_NAME(Raw_file_name)=0041_MAZ_FAG10_HIL-41.raw SUBJECT_SAMPLE_FACTORS - Study1_Dd2APR_iRBC_10h_R1 Sample source:infected RBC (iRBC) | Parasite_type:Dd2 IND | Parasite_line:Dd2APR | Timepoint:10 h Replicate=1; Tracer=15N Pro 15N,13C Arg 5C13 Gln; Date Generated=5/13/2017; RAW_FILE_NAME(Raw_file_name)=0013_MAZ_FAG10_HIL-13.raw SUBJECT_SAMPLE_FACTORS - Study1_Dd2APR_iRBC_10h_R2 Sample source:infected RBC (iRBC) | Parasite_type:Dd2 IND | Parasite_line:Dd2APR | Timepoint:10 h Replicate=2; Tracer=15N Pro 15N,13C Arg 5C13 Gln; Date Generated=5/15/2017; RAW_FILE_NAME(Raw_file_name)=0028_MAZ_FAG10_HIL-28.raw SUBJECT_SAMPLE_FACTORS - Study1_Dd2APR_iRBC_10h_R3 Sample source:infected RBC (iRBC) | Parasite_type:Dd2 IND | Parasite_line:Dd2APR | Timepoint:10 h Replicate=3; Tracer=15N Pro 15N,13C Arg 5C13 Gln; Date Generated=5/18/2017; RAW_FILE_NAME(Raw_file_name)=0043_MAZ_FAG10_HIL-43.raw SUBJECT_SAMPLE_FACTORS - Study1_Dd2APR_iRBC_10h_R4 Sample source:infected RBC (iRBC) | Parasite_type:Dd2 IND | Parasite_line:Dd2APR | Timepoint:10 h Replicate=4; Tracer=15N Pro 15N,13C Arg 5C13 Gln; Date Generated=5/30/2017; RAW_FILE_NAME(Raw_file_name)=0055_MAZ_FAG10_HIL-55.raw #COLLECTION CO:COLLECTION_SUMMARY Highly synchronous (within 4 h) late trophozoite stage parasites were CO:COLLECTION_SUMMARY magnetically purified with MACS LD columns (Miltenyi Biotec Inc., San Diego, CA, CO:COLLECTION_SUMMARY USA). Each sample was incubated for 10hr with either labeled or unlabeled media, CO:COLLECTION_SUMMARY harvested by centrifugation, washed twice in PBS, and then suspended in 10 μl CO:COLLECTION_SUMMARY PBS (Life Technologies, Carlsbad, CA, USA). Polar metabolites were extracted CO:COLLECTION_SUMMARY using nine volumes of 74.9:24.9:0.2 (v/v/v) acetonitrile/methanol/formic acid CO:COLLECTION_SUMMARY containing stable isotope-labeled internal standards (0.2 ng/μl valine-d8 CO:COLLECTION_SUMMARY (Sigma Aldrich, St. Louis, MO, USA); and 0.2 ng/μl phenylalanine-d8 (Cambridge CO:COLLECTION_SUMMARY Isotope Laboratories, Tewksbury, MA, USA) and stored at -80°C prior to the CO:COLLECTION_SUMMARY metabolite profiling assays. CO:SAMPLE_TYPE Cultured cells CO:COLLECTION_LOCATION Harvard Chan School of Public Health, 665 Huntington Ave. Boston, MA 02115 CO:VOLUMEORAMOUNT_COLLECTED 100 uL CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY Cultures were cultured in 5% human O+ hematocrit and maintained under standard TR:TREATMENT_SUMMARY conditions [RPMI 1640 (Life Technologies) supplemented with 28 mM NaHCO3 TR:TREATMENT_SUMMARY (Sigma), 25 mM HEPES (Sigma), 50 mg/ml hypoxanthine (Sigma), 25 μg/ml TR:TREATMENT_SUMMARY gentamycin (Sigma), and 0.5% AlbuMAX II (Life Technologies)] until preparation TR:TREATMENT_SUMMARY for treatment. In preparation for treatment, 25mL synchronized parasites culture TR:TREATMENT_SUMMARY at ~5% parasitemia and 32-36hr staging at beginning of experiment were pelleted TR:TREATMENT_SUMMARY and resuspended in 4mL serum-free RPMI (Incomplete Media). Parasites were TR:TREATMENT_SUMMARY magnetically purified with an LD column (Miltenyi Biotec Inc., San Diego, CA, TR:TREATMENT_SUMMARY USA) and washed until the flow thru ran clear. The column was removed from the TR:TREATMENT_SUMMARY magnet and each sample eluted in 2mL incomplete RPMI. Samples were divided into TR:TREATMENT_SUMMARY two microfuge tubes, each receiving half (1mL) of the cell suspension. Cells TR:TREATMENT_SUMMARY were pelleted and resuspended in either incomplete RPMI (unlabeled) or our TR:TREATMENT_SUMMARY Triplex labeled RPMI media (RPMI media first prepared without proline, TR:TREATMENT_SUMMARY glutamine, or arginine and then supplemented with isotope labeled amino acids at TR:TREATMENT_SUMMARY regular RPMI concentrations: 15N-proline (Pro+1; 0.17 mM), 13C,15N-arginine TR:TREATMENT_SUMMARY (Arg+2; 0.38 mM), and 13C5-gluatmine (Gln+5; 2.05 mM)). Parasites were incubated TR:TREATMENT_SUMMARY in labeled or unlabeled media for 10h at 37ºC under standard malaria gas TR:TREATMENT_SUMMARY mixture. Samples were harvested by centrifugation, washed twice, and then polar TR:TREATMENT_SUMMARY metabolites extracted. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Each sample was washed twice in PBS and then suspended in 10 μl PBS (Life SP:SAMPLEPREP_SUMMARY Technologies, Carlsbad, CA, USA). Polar metabolites were extracted using nine SP:SAMPLEPREP_SUMMARY volumes of 74.9:24.9:0.2 (v/v/v) acetonitrile/methanol/formic acid containing SP:SAMPLEPREP_SUMMARY stable isotope-labeled internal standards (0.2 ng/μl valine-d8 (Sigma Aldrich, SP:SAMPLEPREP_SUMMARY St. Louis, MO, USA); and 0.2 ng/μl phenylalanine-d8 (Cambridge Isotope SP:SAMPLEPREP_SUMMARY Laboratories, Tewksbury, MA, USA) and stored at −80°C prior to the metabolite SP:SAMPLEPREP_SUMMARY profiling assays. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Chromatography method 1 (HILIC-POS) CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Shimadzu Nexera X2 CH:COLUMN_NAME Water Atlantis HILIC (150 x 2.1 mm, 3 μm) CH:SOLVENT_A 100% Water; 10 mM Ammonium formate; 0.1% Formic acid CH:SOLVENT_B 100% Acetonitrile; 0.1% Formic acid CH:FLOW_GRADIENT Chromatographic separation was performed using an isocratic elution at a flow CH:FLOW_GRADIENT rate of 250 μL/min with 5% mobile phase A (10 mM ammonium formate and 0.1% CH:FLOW_GRADIENT formic acid in water) for 1 minute. This was followed by a linear gradient to CH:FLOW_GRADIENT 40% mobile phase B (acetonitrile with 0.1% formic acid) over 10 minutes. At 10 CH:FLOW_GRADIENT minutes, the gradient was returned to initial isocratic conditions (5% mobile CH:FLOW_GRADIENT phase A) and held until 18 minutes, at which point MS acquisition was stopped. CH:FLOW_GRADIENT The column was equilibrated with 5% mobile phase A at a flow rate of CH:FLOW_GRADIENT 400 μL/min for 12 minutes, followed by a reduction to the initial flow rate CH:FLOW_GRADIENT of 250 μL/min for 2 minutes before the next injection. CH:FLOW_RATE 250 μL/min CH:COLUMN_TEMPERATURE 30 CH:INTERNAL_STANDARD L-Phenylalanine-d8 (CIL, DLM-372-1), L-Valine-d8 (Sigma, 486027) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS Mass spectroscopic (MS) analyses were performed by using electrospray ionization MS:MS_COMMENTS in the positive ion mode using full scan analysis over 70 to 800 m/z at 70 000 MS:MS_COMMENTS resolution and 3 Hz data acquisition rate. Other MS settings were as follows: MS:MS_COMMENTS sheath gas 40, sweep gas 2, spray voltage 3.5 kV, capillary temperature 350 °C, MS:MS_COMMENTS S-lens RF 40, heater temperature 300 °C, microscans 1, automatic gain control MS:MS_COMMENTS target 1e6, and maximum ion time 250 ms. Raw data were processed using MS:MS_COMMENTS TraceFinder 3.3 software (Thermo Fisher Scientific; Waltham, MA) and Progenesis MS:MS_COMMENTS QI (Nonlinear Dynamics; Newcastle upon Tyne, UK). Metabolite identities were MS:MS_COMMENTS confirmed using authentic reference standards or reference samples. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS peak areas MS_METABOLITE_DATA_START Samples Study1_Dd2_iRBC_10h_R1 Study1_Dd2_iRBC_10h_R2 Study1_Dd2_iRBC_10h_R3 Study1_Dd2APR_iRBC_10h_R1 Study1_Dd2APR_iRBC_10h_R2 Study1_Dd2APR_iRBC_10h_R3 Study1_Dd2APR_iRBC_10h_R4 Factors Sample source:infected RBC (iRBC) | Parasite_type:Dd2 WT | Parasite_line:Dd2 | Timepoint:10 h Sample source:infected RBC (iRBC) | Parasite_type:Dd2 WT | Parasite_line:Dd2 | Timepoint:10 h Sample source:infected RBC (iRBC) | Parasite_type:Dd2 WT | Parasite_line:Dd2 | Timepoint:10 h Sample source:infected RBC (iRBC) | Parasite_type:Dd2 IND | Parasite_line:Dd2APR | Timepoint:10 h Sample source:infected RBC (iRBC) | Parasite_type:Dd2 IND | Parasite_line:Dd2APR | Timepoint:10 h Sample source:infected RBC (iRBC) | Parasite_type:Dd2 IND | Parasite_line:Dd2APR | Timepoint:10 h Sample source:infected RBC (iRBC) | Parasite_type:Dd2 IND | Parasite_line:Dd2APR | Timepoint:10 h phenylalanine-d8 85857758 76214478 68386723 84959768 87625417 65353895 73920449 valine-d8 25865658 22228385 23636231 24560828 24074827 28845316 31383117 1-methyladenosine 1-methylhistamine 1-methylnicotinamide 1-pyrroline-5-carboxylate 1-pyrroline-5-carboxylase[15]N 1-pyrroline-5-carboxylate[13]C1[15]N1 1-pyrroline-5-carboxylate[13]C5 2-deoxycytidine 271862 6440257 1468650 464179 847258 1180553 962604 5-adenosylhomocysteine 1704512 3313055 581247 3645746 4043444 399988 75462 5-aminolevulinic acid 11835132 20353681 14189097 15363479 24529372 22759227 15106871 5-hydroxytryptophan acetylcholine 325913 334292 624295 adenosine 2472579 8305005 5137255 3079375 7603648 8269387 7240576 ADMA 7433653 53117249 28072305 9540948 25579548 29414681 18600197 alanine 1854324 6100943 5295679 1956525 5101637 5701311 3173680 allantoin alpha-glycerophosphocholine 16512030 128802832 89398180 23134398 63616076 74464657 33956717 anserine anthranilic acid arginine 73620487 169060746 118505001 56340445 150138469 140770643 122315651 arginine[15]N 3098184 10535494 6518446 2973090 5921520 6699854 5334962 arginine[13]C1[15]N1 343669064 1527525704 994070834 423456712 731571479 927022911 681548609 arginine [13]C5 argininosuccinate asparagine 4732364 13041633 20369509 6208942 12815841 19718173 8566401 aspartate 6614801 16097744 19517117 4894619 7409519 10766608 5367731 beta-alanine betaine 167865242 576853334 284710908 236426852 559231471 598418112 296801578 butyrobetaine carnitine 38558937 157489343 56931320 57349480 102668907 262835710 39083633 C2 carnitine 67440422 198592453 156196111 98474529 152997745 322853091 79938761 C3 carnitine 3407782 12319490 9337554 5029772 10872847 20704774 3608751 C3-DC-CH3 carnitine 4057690 7198328 8148337 5281807 7447452 10415832 4927826 C4 carnitine 343969 2520407 2543638 526126 1867520 4905018 708974 C4-OH carnitine 634811 2300728 2617980 1076626 1759041 3590965 1262627 C5 carnitine 350079 1438388 1148272 495956 1216310 2668806 642728 C5:1 carnitine C6 carnitine C7 carnitine C8 carnitine C10 carnitine C10:2 carnitine C12 carnitine C12:1 carnitine C14 carnitine 134547 219200 243317 425005 309394 C14:1 carnitine C14:2 carnitine C16 carnitine 2886244 3067931 2523662 4215568 4639912 3358197 661101 C18 carnitine 8896768 15526101 14392715 12696301 16880138 12489462 7689038 C18:1 carnitine 3705540 5982215 5341342 5716130 7775596 7538618 1741190 C18:2 carnitine 622270 834668 413665 1012196 1302899 860105 C20 carnitine 101041 424884 762275 154219 322049 718656 520051 C20:4 carnitine C26 carnitine cAMP carnosine 117071 135739 104728 452271 661449 255858 choline 370737265 2515400426 3028125324 596621806 1214426130 3695353652 1319133978 citrulline 528294 2186657 376235 218704 1147369 899882 158781 citrulline[15]N citrulline[13]C1[15]N1 citrulline[13]C5 cotinine creatine 248953995 504715354 266457852 355334211 521415269 564045204 212120501 creatinine 1707744 4971511 3134983 2327846 7340322 5967136 3425531 cytosine 8759737 96105479 28078945 13162991 19935752 17274818 17729185 dimethylglycine GABA 23148389 28017953 74452336 26905357 21802481 64137440 9775332 glucose glutamate 15917411 20383118 9726206 16608930 19302519 12329720 13188134 glutamate[15]N glutamate[13]C3[15]N1 glutamate[13]C5 14946672 28526550 26835062 19509127 22837738 62771234 50161956 glutamine 14202678 35410894 14427246 17969513 29852638 13858514 14619246 glutamine[15]N glutamine[13]C1[15]N1 glutamine[13]C5 41788329 60472944 27517852 59274834 61868098 33088514 15983580 glycine histamine 865742 3073850 1665583 738042 2350167 1696654 960583 histidine 53523296 250715322 473792959 100441121 256545268 414842435 152870935 homocysteine isoleucine 6663037 18195512 9013455 6589721 15764515 20258687 14787557 kynurenic acid leucine 7705810 19563942 7964907 9063332 24032434 17609780 15397916 lysine 57348815 501074697 190611683 21346382 85012262 68477835 39480163 methionine 718600 1885592 1054409 858628 2604065 2159936 1323558 methionine sulfoxide 157890 830375 449126 206001 882501 806851 544074 methylthioadenosine 1333345 6874040 4801649 3563414 8693434 5765450 3438055 N-carbamoyl-beta-alanine niacinamide 10237106 81452532 151996362 11446030 41830776 225129212 94146337 NMMA 980316 7948706 3971041 903659 3352691 3101409 1714545 ornithine 2740829 8524920 13152961 1028300 3276906 4922324 2444286 ornithine[15]N 229739 340249 23225 86932 202756 40932 ornithine[13]C1[15]N1 13037383 55358804 78205626 13396487 25846280 60638595 20847806 ornithine [13]C5 phenylalanine 2898309 10888750 3431625 4035726 14276149 5821179 5390245 phosphocholine 369685069 2515182581 2987891403 758964232 1048271528 3171514427 2705423025 phosphoethanolamine 4407373 57883827 54331040 6516410 14274854 50473943 32125421 pipecolic acid 10845306 185827214 35534235 58811305 134022346 67607548 73054978 pipecolic acid[15]N pipecholic acid[13]C1[15]N1 pipecolic acid [13]C5 proline 13197590 70074107 31677408 235339591 488658898 121196122 128038563 proline[15]N 30958292 75260639 46804520 36693160 64327132 122187017 66368687 proline[13]C1[15]N1 7357150 72027399 76295028 79322520 131281016 329085016 201606005 proline [13]C5 putrescine 8758321 43535464 42637302 7423134 15468364 45128077 51455427 pyroglutamic acid 20398630 50816521 20445972 26562173 42857996 18620455 20688642 s-adenosylmethionine 14016148 38780049 25646606 39484364 38946381 21307121 7446663 sarcosine SDMA 763931 5338314 2608999 1025124 2188474 1885744 1641606 serine 1044434 2578934 2659092 1143570 2784390 2640860 1677616 spermidine 1165725411 3462537081 3995203995 1670014593 2038794003 4933225606 4609725588 taurine 1792109 2453724 452070 757646 3925688 1763874 2397063 thiamine 30976714 140142001 45129328 45370249 115155855 81752283 54053417 threonine 764318 2033824 1528003 910021 2190659 1931532 1956553 thymidine trans-hydroxyproline 11904893 20353681 14189097 15363479 24529372 22759227 15106871 trans-hydroxyproline[15]N 4706715 12999754 20365809 6176404 12795089 19714492 8517447 trans-hydroxyproline[13]C1[15]N1 trans-hydroxyproline [13]C5 trimethylamine-N-oxide 2311362 1744066 1525297 2195512 2096395 1785881 2085989 tryptophan 1377021 6770777 2922267 1891432 5470270 4452394 3886666 tyrosine 854113 4611418 4516251 1267667 4018874 6385493 3701704 valine 1957451 7123382 3352676 2441503 7737686 6306975 4800787 xanthosine 5642637 9777212 12752090 5897105 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Formula m/z RT Super class Main class Sub class Pubchem CID CHEBI_ID HMDB_ID KEGG_ID INCHI_KEY phenylalanine-d8 C9H3NO2D8 174.1365 6.69 - - - - - valine-d8 C5H3NO2D8 126.1365 7.39 - - - - - 1-methyladenosine C11H15N5O4 282.1197 NA Nucleic acids Purines Purine ribonucleosides 27476 16020 HMDB0003331 C02494 GFYLSDSUCHVORB-IOSLPCCCSA-N 1-methylhistamine C6H11N3 126.1026 NA Alkaloids Histidine alkaloids Imidazole alkaloids 3614 29009 HMDB0000898 C05127 FHQDWPCFSJMNCT-UHFFFAOYSA-N 1-methylnicotinamide C7H9N2O+ 137.0715 NA Alkaloids Pyridine alkaloids Nicotinic acid alkaloids 457 16797 HMDB0000699 C02918 LDHMAVIPBRSVRG-UHFFFAOYSA-O 1-pyrroline-5-carboxylate C5H7NO2 114.0549 NA - - - - - 1-pyrroline-5-carboxylase[15]N C5H7[15]NO2 115.052 NA - - - - - 1-pyrroline-5-carboxylate[13]C1[15]N1 [13]CC4H7[15]NO2 116.0553 NA - - - - - 1-pyrroline-5-carboxylate[13]C5 [13]C5H7NO2 119.0717 NA - - - - - 2-deoxycytidine C9H13N3O4 228.0979 5.92 Nucleic acids Pyrimidines Pyrimidine deoxyribonucleosides 13711 15698 HMDB0000014 C00881 CKTSBUTUHBMZGZ-SHYZEUOFSA-N 5-adenosylhomocysteine C14H20N6O5S 385.1289 8.12 - - - - - 5-aminolevulinic acid C5H9NO3 132.0655 7.84 Fatty Acyls Fatty acids Amino FA 137 17549 HMDB0001149 C00430 ZGXJTSGNIOSYLO-UHFFFAOYSA-N 5-hydroxytryptophan C11H12N2O3 221.0921 NA Organic acids Amino acids and peptides Amino acids 439280 17780 HMDB0000472 C00643 LDCYZAJDBXYCGN-VIFPVBQESA-N acetylcholine C7H16NO2 146.1181 7.71 Organic nitrogen compounds Cholines Acyl cholines 187 15355 HMDB0000895 C01996 OIPILFWXSMYKGL-UHFFFAOYSA-N adenosine C10H13N5O4 268.104 4.91 Nucleic acids Purines Purine ribonucleosides 60961 16335 HMDB0000050 C00212 OIRDTQYFTABQOQ-KQYNXXCUSA-N ADMA C8H18N4O2 203.1503 9.79 Organic acids Amino acids and peptides Amino acids 123831 17929 HMDB0001539 C03626 YDGMGEXADBMOMJ-LURJTMIESA-N alanine C3H7NO2 90.055 7.78 Organic acids Amino acids and peptides Amino acids 5950 16977 HMDB0000161 C00041 QNAYBMKLOCPYGJ-REOHCLBHSA-N allantoin C4H6N4O3 159.0513 NA Organoheterocyclic compounds Cyclic ureas Ureides 439714 15678 HMDB0000462 POJWUDADGALRAB-SFOWXEAESA-N alpha-glycerophosphocholine C8H20NO6P 258.1101 10.5 Organic acids Organic phosphoric acids Organic phosphoric acids 657272 16870 HMDB0000086 SUHOQUVVVLNYQR-MRVPVSSYSA-N anserine C10H16N4O3 241.1295 NA Organic acids Amino acids and peptides Dipeptides 112072 18323 HMDB0000194 C01262 MYYIAHXIVFADCU-QMMMGPOBSA-N anthranilic acid C7H7NO2 138.055 NA Benzenoids Benzenes Aminobenzoic acids 227 30754 HMDB0001123 C00108 RWZYAGGXGHYGMB-UHFFFAOYSA-N arginine C6H14N4O2 175.119 9.21 Organic acids Amino acids and peptides Amino acids 6322 16467 HMDB0000517 C00062 ODKSFYDXXFIFQN-BYPYZUCNSA-N arginine[15]N C6H14[15]N1N3O2 176.1159 9.21 - - - - - arginine[13]C1[15]N1 [13]CC5H14[15]N1N3O2 177.1193 9.21 - - - - - arginine [13]C5 [13]C5CH14N4O2 180.1357 9.21 - - - - - argininosuccinate C10H18N4O6 291.1299 NA Organic acids Amino acids and peptides Amino acids 16950 184023 HMDB0000052 C03406 KDZOASGQNOPSCU-WDSKDSINSA-N asparagine C4H8N2O3 133.0608 7.89 Organic acids Amino acids and peptides Amino acids 6267 17196 HMDB0000168 C00152 DCXYFEDJOCDNAF-REOHCLBHSA-N aspartate C4H7NO4 134.0448 7.95 Organic acids Amino acids and peptides Amino acids 5960 17053 HMDB0000191 C00049 CKLJMWTZIZZHCS-REOHCLBHSA-N beta-alanine C3H7NO2 90.055 NA Organic acids Amino acids and peptides Amino acids 239 16958 HMDB0000056 C00099 UCMIRNVEIXFBKS-UHFFFAOYSA-N betaine C5H11NO2 118.0863 8.92 Organic acids Amino acids and peptides Amino acids 247 17750 HMDB0000043 KWIUHFFTVRNATP-UHFFFAOYSA-N butyrobetaine C7H16NO2 146.1176 NA Fatty Acyls Fatty esters Acyl carnitines 725 16244 HMDB0001161 C01181 JHPNVNIEXXLNTR-UHFFFAOYSA-N carnitine C7H15NO3 162.1125 8.81 Organic nitrogen compounds Carnitines Carnitines 10917 16347 HMDB0000062 C00487 PHIQHXFUZVPYII-ZCFIWIBFSA-N C2 carnitine C9H17NO4 204.123 8.62 Fatty Acyls Fatty esters Acyl carnitines 18230 86045 HMDB0240771 RDHQFKQIGNGIED-QMMMGPOBSA-N C3 carnitine C10H19NO4 218.1387 8.28 Fatty Acyls Fatty esters Acyl carnitines 188824 53210 HMDB0062514 C03017 UFAHZIUFPNSHSL-MRVPVSSYSA-N C3-DC-CH3 carnitine C11H19NO6 262.1285 8.54 Fatty Acyls Fatty esters Acyl carnitines 10177002 73017 HMDB0000736 LRCNOZRCYBNMEP-UHFFFAOYSA-N C4 carnitine C11H21NO4 232.1543 7.98 Fatty Acyls Fatty esters Acyl carnitines 213144 21949 HMDB0002013 C02862 QWYFHHGCZUCMBN-SECBINFHSA-N C4-OH carnitine C11H21NO5 248.1493 8.81 Fatty Acyls Fatty esters Acyl carnitines C5 carnitine C12H23NO4 246.17 7.71 Fatty Acyls Fatty esters Acyl carnitines 53481619 165624 HMDB0013128 VSNFQQXVMPSASB-JTQLQIEISA-N C5:1 carnitine C12H21NO4 244.1543 NA Fatty Acyls Fatty esters Acyl carnitines C6 carnitine C13H25NO4 260.1856 NA Fatty Acyls Fatty esters Acyl carnitines 3246938 84834 HMDB0000756 VVPRQWTYSNDTEA-LLVKDONJSA-N C7 carnitine C14H27NO4 274.2013 NA Fatty Acyls Fatty esters Acyl carnitines 6426896 74303 HMDB0013238 VDPCTFWULDLKHT-UHFFFAOYSA-N C8 carnitine C15H29NO4 288.2169 NA Fatty Acyls Fatty esters Acyl carnitines 11953814 18102 HMDB0000791 C02838 CXTATJFJDMJMIY-CYBMUJFWSA-N C10 carnitine C17H33NO4 316.2482 NA Fatty Acyls Fatty esters Acyl carnitines 11953821 28717 HMDB0000651 C03299 LZOSYCMHQXPBFU-OAHLLOKOSA-N C10:2 carnitine C17H29NO4 312.2169 NA Fatty Acyls Fatty esters Acyl carnitines C12 carnitine C19H37NO4 344.2795 NA Fatty Acyls Fatty esters Acyl carnitines 168381 77086 HMDB0002250 FUJLYHJROOYKRA-QGZVFWFLSA-N C12:1 carnitine C19H35NO4 342.2639 NA Fatty Acyls Fatty esters Acyl carnitines C14 carnitine C21H41NO4 372.3108 6.78 Fatty Acyls Fatty esters Acyl carnitines 53477791 84634 HMDB0005066 PSHXNVGSVNEJBD-LJQANCHMSA-N C14:1 carnitine C21H39NO4 370.2952 NA Fatty Acyls Fatty esters Acyl carnitines C14:2 carnitine C21H37NO4 368.2795 NA Fatty Acyls Fatty esters Acyl carnitines C16 carnitine C23H45NO4 400.3421 6.63 Fatty Acyls Fatty esters Acyl carnitines 11953816 17490 HMDB0000222 C02990 XOMRRQXKHMYMOC-OAQYLSRUSA-N C18 carnitine C25H49NO4 428.3734 6.53 Fatty Acyls Fatty esters Acyl carnitines 52922056 84644 HMDB0000848 FNPHNLNTJNMAEE-HSZRJFAPSA-N C18:1 carnitine C25H47NO4 426.3578 6.57 Fatty Acyls Fatty esters Acyl carnitines C18:2 carnitine C25H45NO4 424.3421 6.62 Fatty Acyls Fatty esters Acyl carnitines C20 carnitine C27H53NO4 456.4047 6.44 Fatty Acyls Fatty esters Acyl carnitines 53477833 73100 HMDB0006460 SVJLJQBGUITFLI-UHFFFAOYSA-N C20:4 carnitine C27H45NO4 448.3421 NA Fatty Acyls Fatty esters Acyl carnitines C26 carnitine C33H65NO4 540.4986 NA Fatty Acyls Fatty esters Acyl carnitines 145720497 165602 HMDB0006347 KOCKWDDTAHPJSX-WJOKGBTCSA-N cAMP C10H12N5O6P 330.0598 NA Nucleic acids Purines Purine rNMP 6076 17489 HMDB0000058 C00575 IVOMOUWHDPKRLL-KQYNXXCUSA-N carnosine C9H14N4O3 227.1139 9.67 Organic acids Amino acids and peptides Dipeptides 439224 15727 HMDB0000033 C00386 CQOVPNPJLQNMDC-ZETCQYMHSA-N choline C5H14NO 104.107 8.22 Organic nitrogen compounds Cholines Cholines 305 15354 HMDB0000097 C00114 OEYIOHPDSNJKLS-UHFFFAOYSA-N citrulline C6H13N3O3 176.103 8.34 Organic acids Amino acids and peptides Amino acids 9750 16349 HMDB0000904 C00327 RHGKLRLOHDJJDR-BYPYZUCNSA-N citrulline[15]N C6H14[15]N1N3O2 177.1 8.34 - - - - - citrulline[13]C1[15]N1 [13]C1C5H14[15]N1N3O2 178.1033 8.34 - - - - - citrulline[13]C5 [13]C5C1H14N4O2 177.1063 8.34 - - - - - cotinine C10H12N2O 177.1022 8.34 Alkaloids Pyridine alkaloids Nicotinic acid alkaloids 854019 68641 HMDB0001046 UIKROCXWUNQSPJ-VIFPVBQESA-N creatine C4H9N3O2 132.0768 8.29 Organic acids Amino acids and peptides Amino acids 586 16919 HMDB0000064 C00300 CVSVTCORWBXHQV-UHFFFAOYSA-N creatinine C4H7N3O 114.0662 6.41 Organoheterocyclic compounds Azoles Imidazolines 137319715 16737 HMDB0000562 C00791 DDRJAANPRJIHGJ-UHFFFAOYSA-N cytosine C4H5N3O 112.0505 5.94 Nucleic acids Pyrimidines Pyrimidones 597 16040 HMDB0000630 C00380 OPTASPLRGRRNAP-UHFFFAOYSA-N dimethylglycine C4H9NO2 104.0706 NA Organic acids Amino acids and peptides Amino acids 673 17724 HMDB0000092 C01026 FFDGPVCHZBVARC-UHFFFAOYSA-N GABA C4H9NO2 104.0706 7.5 Fatty Acyls Fatty acids Amino FA 119 16865 HMDB0000112 C00334 BTCSSZJGUNDROE-UHFFFAOYSA-N glucose C6H12O6 181.0707 NA Carbohydrates Monosaccharides Hexoses 5793 4167 HMDB0304632 C00031 WQZGKKKJIJFFOK-GASJEMHNSA-N glutamate C5H9NO4 148.0604 7.59 Organic acids Amino acids and peptides Amino acids 33032 16015 HMDB0000148 C00025 WHUUTDBJXJRKMK-VKHMYHEASA-N glutamate[15]N C5H9[15]NO4 149.0574 7.59 - - - - - glutamate[13]C3[15]N1 [13]C3C2H9[15]NO4 150.0608 7.59 - - - - - glutamate[13]C5 [13]C5H9NO4 153.0772 7.59 Organic acids Amino acids and peptides Amino acids 33032 16015 HMDB0000148 C00025 WHUUTDBJXJRKMK-VKHMYHEASA-N glutamine C5H10N2O3 147.0764 8.01 Organic acids Amino acids and peptides Amino acids 5961 18050 HMDB0000641 C00064 ZDXPYRJPNDTMRX-VKHMYHEASA-N glutamine[15]N C5H10[15]N1N1O3 148.0734 8.01 - - - - - glutamine[13]C1[15]N1 [13]C1C4H10[15]N1N1O3 149.0768 8.01 - - - - - glutamine[13]C5 [13]C5H10N2O3 152.0932 8.01 Organic acids Amino acids and peptides Amino acids 5961 18050 HMDB0000641 C00064 ZDXPYRJPNDTMRX-VKHMYHEASA-N glycine C2H5NO2 76.0393 NA Organic acids Amino acids and peptides Amino acids 750 15428 HMDB0000123 C00037 DHMQDGOQFOQNFH-UHFFFAOYSA-N histamine C5H9N3 112.0869 9.22 Alkaloids Histidine alkaloids Imidazole alkaloids 774 18295 HMDB0000870 C00388 NTYJJOPFIAHURM-UHFFFAOYSA-N histidine C6H9N3O2 156.0768 9.28 Organic acids Amino acids and peptides Amino acids 6274 15971 HMDB0000177 C00135 HNDVDQJCIGZPNO-YFKPBYRVSA-N homocysteine C4H9NO2S 136.0427 NA Organic acids Amino acids and peptides Amino acids 91552 17588 HMDB0000742 C00155 FFFHZYDWPBMWHY-VKHMYHEASA-N isoleucine C6H13NO2 132.1019 7.06 Organic acids Amino acids and peptides Amino acids 6306 17191 HMDB0000172 C00407 AGPKZVBTJJNPAG-WHFBIAKZSA-N kynurenic acid C10H7NO3 190.0499 NA Alkaloids Anthranilic acid alkaloids Acridone alkaloids 3845 18344 HMDB0000715 C01717 HCZHHEIFKROPDY-UHFFFAOYSA-N leucine C6H13NO2 132.1019 6.92 Organic acids Amino acids and peptides Amino acids 6106 15603 HMDB0000687 C00123 ROHFNLRQFUQHCH-YFKPBYRVSA-N lysine C6H14N2O2 147.1128 9.44 Organic acids Amino acids and peptides Amino acids 5962 18019 HMDB0000182 C00047 KDXKERNSBIXSRK-YFKPBYRVSA-N methionine C5H11NO2S 150.0583 7.01 Organic acids Amino acids and peptides Amino acids 6137 16643 HMDB0000696 C00073 FFEARJCKVFRZRR-BYPYZUCNSA-N methionine sulfoxide C5H11NO3S 166.0532 8.62 Organic acids Amino acids and peptides Amino acids 10062737 49032 HMDB0002005 QEFRNWWLZKMPFJ-ZXPFJRLXSA-N methylthioadenosine C11H15N5O3S 298.0968 4.17 Nucleic acids Purines Purine ribonucleosides N-carbamoyl-beta-alanine C4H8N2O3 133.0608 NA Organic nitrogen compounds Organonitrogen compounds Ureas 111 18261 HMDB0000026 C02642 JSJWCHRYRHKBBW-UHFFFAOYSA-N niacinamide C6H6N2O 123.0553 3.67 Alkaloids Pyridine alkaloids Nicotinic acid alkaloids 936 17154 HMDB0001406 C00153 DFPAKSUCGFBDDF-UHFFFAOYSA-N NMMA C7H16N4O2 189.1346 9.41 Organic acids Amino acids and peptides Amino acids 132862 28229 HMDB0029416 C03884 NTNWOCRCBQPEKQ-YFKPBYRVSA-N ornithine C5H12N2O2 133.0972 9.33 Organic acids Amino acids and peptides Amino acids 6262 15729 HMDB0000214 C00077 AHLPHDHHMVZTML-BYPYZUCNSA-N ornithine[15]N C5H12[15]N1N1O2 134.0941 9.33 - - - - - ornithine[13]C1[15]N1 [13]CC4H12[15]N1N1O2 135.0975 9.33 - - - - - ornithine [13]C5 [13]C5H12N2O2 138.1139 9.33 Organic acids Amino acids and peptides Amino acids 6262 15729 HMDB0000214 C00077 AHLPHDHHMVZTML-BYPYZUCNSA-N phenylalanine C9H11NO2 166.0863 6.68 Organic acids Amino acids and peptides Amino acids 6140 17295 HMDB0000159 C00079 COLNVLDHVKWLRT-QMMMGPOBSA-N phosphocholine C5H15NO4P 184.0733 11.62 Organic nitrogen compounds Cholines Cholines 135437 18132 HMDB0001565 YHHSONZFOIEMCP-UHFFFAOYSA-N phosphoethanolamine C2H8NO4P 142.0264 9.55 Organic acids Phosphate esters Phosphoethanolamines 1015 17553 HMDB0000224 C00346 SUHOOTKUPISOBE-UHFFFAOYSA-N pipecolic acid C6H11NO2 130.0863 7.95 Organic acids Amino acids and peptides Amino acids 439227 30913 HMDB0000716 C00408 HXEACLLIILLPRG-YFKPBYRVSA-N pipecolic acid[15]N C6H11[15]NO2 131.0832 7.95 - - - - - pipecholic acid[13]C1[15]N1 [13]CC5H11[15]NO2 132.0866 7.95 - - - - - pipecolic acid [13]C5 [13]C5CH11NO2 135.103 7.95 - - - - - proline C5H9NO2 116.0706 8.06 Organic acids Amino acids and peptides Amino acids 145742 17203 HMDB0000162 C00148 ONIBWKKTOPOVIA-BYPYZUCNSA-N proline[15]N C5H9[15]NO2 117.0676 8.06 - - - - - proline[13]C1[15]N1 [13]CC4H9[15]NO2 118.071 8.06 - - - - - proline [13]C5 [13]C5H9NO2 121.0874 8.06 Organic acids Amino acids and peptides Amino acids 145742 17203 HMDB0000162 C00148 ONIBWKKTOPOVIA-BYPYZUCNSA-N putrescine C4H12N2 89.1073 9.43 Organic nitrogen compounds Amines Amines 1045 17148 HMDB0001414 C00134 KIDHWZJUCRJVML-UHFFFAOYSA-N pyroglutamic acid C5H7NO3 130.0499 8.01 Organic acids Amino acids and peptides Amino acids 7405 18183 HMDB0000267 C01879 ODHCTXKNWHHXJC-VKHMYHEASA-N s-adenosylmethionine C15H23N6O5S 399.1451 10.22 Nucleic acids Purines Other purines 34756 15414 HMDB0001185 C00019 MEFKEPWMEQBLKI-AIRLBKTGSA-O sarcosine C3H7NO2 90.055 NA Organic acids Amino acids and peptides Amino acids 1088 15611 HMDB0000271 C00213 FSYKKLYZXJSNPZ-UHFFFAOYSA-N SDMA C8H18N4O2 203.1503 9.69 Organic acids Amino acids and peptides Amino acids 169148 61914 HMDB0003334 HVPFXCBJHIIJGS-LURJTMIESA-N serine C3H7NO3 106.0499 7.64 Organic acids Amino acids and peptides Amino acids 5951 17115 HMDB0000187 C00065 MTCFGRXMJLQNBG-REOHCLBHSA-N spermidine C7H19N3 146.1652 10.65 Alkaloids Ornithine alkaloids Polyamines 1102 16610 HMDB0001257 C00315 ATHGHQPFGPMSJY-UHFFFAOYSA-N taurine C2H7NO3S 126.0219 6.04 Organic acids Sulfonic acids Sulfonic acids 1123 15891 HMDB0000251 C00245 XOAAWQZATWQOTB-UHFFFAOYSA-N thiamine C12H17N4OS 265.1118 9.67 Nucleic acids Pyrimidines Thiamines 1130 18385 HMDB0000235 C00378 JZRWCGZRTZMZEH-UHFFFAOYSA-N threonine C4H9NO3 120.0655 7.58 Organic acids Amino acids and peptides Amino acids 6288 16857 HMDB0000167 C00188 AYFVYJQAPQTCCC-GBXIJSLDSA-N thymidine C10H14N2O5 243.0976 NA Nucleic acids Pyrimidines Pyrimidine deoxyribonucleosides 5789 17748 HMDB0000273 C00214 IQFYYKKMVGJFEH-XLPZGREQSA-N trans-hydroxyproline C5H9NO3 132.0655 7.85 Organic acids Amino acids and peptides Amino acids 5810 18095 HMDB0000725 C01157 PMMYEEVYMWASQN-DMTCNVIQSA-N trans-hydroxyproline[15]N C5H9[15]NO3 133.0625 7.85 - - - - - trans-hydroxyproline[13]C1[15]N1 [13]CC4H9[15]NO3 134.0659 7.85 - - - - - trans-hydroxyproline [13]C5 [13]C5H9NO3 137.0823 7.85 - - - - - trimethylamine-N-oxide C3H9NO 76.0757 8.04 Organic nitrogen compounds Amine oxides Trisubstituted amine oxides 1145 15724 HMDB0000925 C01104 UYPYRKYUKCHHIB-UHFFFAOYSA-N tryptophan C11H12N2O2 205.0972 6.48 Organic acids Amino acids and peptides Amino acids 6305 16828 HMDB0000929 C00078 QIVBCDIJIAJPQS-VIFPVBQESA-N tyrosine C9H11NO3 182.0812 6.78 Organic acids Amino acids and peptides Amino acids 6057 17895 HMDB0000158 C00082 OUYCCCASQSFEME-QMMMGPOBSA-N valine C5H11NO2 118.0863 7.38 Organic acids Amino acids and peptides Amino acids 6287 16414 HMDB0000883 C00183 KZSNJWFQEVHDMF-BYPYZUCNSA-N xanthosine C10H12N4O6 285.083 4.31 Nucleic acids Purines Purine ribonucleosides 64959 18107 HMDB0000299 UBORTCNDUKBEOP-UUOKFMHZSA-N METABOLITES_END #END