#METABOLOMICS WORKBENCH cclish_20250922_120134 DATATRACK_ID:6470 STUDY_ID:ST004240 ANALYSIS_ID:AN007057 PROJECT_ID:PR002663
VERSION             	1
CREATED_ON             	October 1, 2025, 5:31 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 experiments
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                   	PfApiAT2 loss of function leads to Adaptive Proline Response (APR)
ST:STUDY_SUMMARY                 	Plasmodium falciparum evades the antimalarial activity of proline-competitive
ST:STUDY_SUMMARY                 	prolyl-tRNA synthetase (PfProRS) inhibitors, such as halofuginone (HFG), by a
ST:STUDY_SUMMARY                 	unique resistance mechanism termed the Adaptive Proline Response (APR). The APR
ST:STUDY_SUMMARY                 	is characterized by a marked elevation of intracellular proline following drug
ST:STUDY_SUMMARY                 	exposure. Contrary to initial expectations, the APR is not mediated by
ST:STUDY_SUMMARY                 	alterations in canonical proline metabolic pathways involving arginase (PfARG)
ST:STUDY_SUMMARY                 	and ornithine aminotransferase (PfOAT). Instead, we identified loss-of-function
ST:STUDY_SUMMARY                 	mutations in the Apicomplexan Amino acid Transporter 2 (PfApiAT2) as the primary
ST:STUDY_SUMMARY                 	genetic driver of this resistance phenotype. Importantly, reversion of these
ST:STUDY_SUMMARY                 	mutations to wildtype effectively suppresses the APR, establishing PfApiAT2 as
ST:STUDY_SUMMARY                 	the molecular determinant of this novel resistance mechanism. The elucidation of
ST:STUDY_SUMMARY                 	the APR significantly advances our understanding of antimalarial drug
ST:STUDY_SUMMARY                 	resistance. By delineating the role of PfApiAT2 in this process, we establish
ST:STUDY_SUMMARY                 	critical insights for the development of strategies to circumvent PfProRS
ST:STUDY_SUMMARY                 	inhibitor resistance for future antimalarial therapies. In this study, wild-type
ST:STUDY_SUMMARY                 	Dd2, ApiAT2 (MFR4) knockout (three independent clones: clones 1, 2, 3) parasites
ST:STUDY_SUMMARY                 	were assessed for the presence of the Adaptive Proline Response despite not
ST:STUDY_SUMMARY                 	having been exposed to HFG. These experiments were run in unlabeled media and
ST:STUDY_SUMMARY                 	total intracellular proline levels analyzed to indicate presence or absence of
ST:STUDY_SUMMARY                 	the APR. Uninfected RBCs (uRBCs) were run as a control.
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                    	5
ST:TOTAL_SUBJECTS                	30
ST:STUDY_COMMENTS                	Study 4 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           	-	Study4_uRBC_RBC_10h_R1	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Timepoint=NA; Replicate=1; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0010_MAZ_Akansha_HP-uRBC_iRBCAP_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_uRBC_RBC_10h_R2	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Timepoint=NA; Replicate=2; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0020_MAZ_Akansha_HP-uRBC_iRBCSB_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_uRBC_RBC_10h_R3	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Timepoint=NA; Replicate=3; Tracer=none; Date Generated=7/28/2020; RAW_FILE_NAME(Raw_file_name)=0030_MAZ_Akansha_HP-uRBC_iRBCSB_072820.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_uRBC_RBC_10h_R4	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Timepoint=NA; Replicate=4; Tracer=none; Date Generated=7/30/2020; RAW_FILE_NAME(Raw_file_name)=0040_MAZ_Akansha_HP-uRBC_iRBCAP_073020.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_uRBC_RBC_10h_R5	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Timepoint=NA; Replicate=5; Tracer=none; Date Generated=8/6/2020; RAW_FILE_NAME(Raw_file_name)=0050_MAZ_Akansha_HP-uRBC_iRBCAP_080620.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_uRBC_RBC_10h_R6	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Timepoint=NA; Replicate=6; Tracer=none; Date Generated=8/7/2020; RAW_FILE_NAME(Raw_file_name)=0060_MAZ_Akansha_HP-uRBC_iRBC_SB_080720.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2_iRBC_10h_R1	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Timepoint=NA; Replicate=1; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0002_MAZ_Akansha_HP-PF_Dd2_iRBC_AP_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2_iRBC_10h_R2	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Timepoint=NA; Replicate=2; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0012_MAZ_Akansha_HP-PF_Dd2_iRBC_SB_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2_iRBC_10h_R3	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Timepoint=NA; Replicate=3; Tracer=none; Date Generated=7/28/2020; RAW_FILE_NAME(Raw_file_name)=0022_MAZ_Akansha_HP-PF_Dd2_iRBC_SB_072820.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2_iRBC_10h_R4	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Timepoint=NA; Replicate=4; Tracer=none; Date Generated=7/30/2020; RAW_FILE_NAME(Raw_file_name)=0032_MAZ_Akansha_HP-PF_Dd2_iRBC_AP_073020.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2_iRBC_10h_R5	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Timepoint=NA; Replicate=5; Tracer=none; Date Generated=8/6/2020; RAW_FILE_NAME(Raw_file_name)=0042_MAZ_Akansha_HP-PF_Dd2_iRBC_AP_080620.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2_iRBC_10h_R6	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Timepoint=NA; Replicate=6; Tracer=none; Date Generated=8/7/2020; RAW_FILE_NAME(Raw_file_name)=0052_MAZ_Akansha_HP-PF_Dd2_iRBC_SB_080720.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Timepoint=NA; Replicate=1; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0004_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_1_AP_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Timepoint=NA; Replicate=2; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0014_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_1_SB_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R3	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Timepoint=NA; Replicate=3; Tracer=none; Date Generated=7/28/2020; RAW_FILE_NAME(Raw_file_name)=0024_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_1_SB_072820.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R4	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Timepoint=NA; Replicate=4; Tracer=none; Date Generated=7/30/2020; RAW_FILE_NAME(Raw_file_name)=0034_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_1_AP_073020.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R5	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Timepoint=NA; Replicate=5; Tracer=none; Date Generated=8/6/2020; RAW_FILE_NAME(Raw_file_name)=0044_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_1_AP_080620.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R6	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Timepoint=NA; Replicate=6; Tracer=none; Date Generated=8/7/2020; RAW_FILE_NAME(Raw_file_name)=0054_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_1_SB_080720.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Timepoint=NA; Replicate=1; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0006_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_2_AP_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Timepoint=NA; Replicate=2; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0016_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_2_SB_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R3	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Timepoint=NA; Replicate=3; Tracer=none; Date Generated=7/28/2020; RAW_FILE_NAME(Raw_file_name)=0026_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_2_SB_072820.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R4	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Timepoint=NA; Replicate=4; Tracer=none; Date Generated=7/30/2020; RAW_FILE_NAME(Raw_file_name)=0036_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_2_AP_073020.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R5	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Timepoint=NA; Replicate=5; Tracer=none; Date Generated=8/6/2020; RAW_FILE_NAME(Raw_file_name)=0046_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_2_AP_080620.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R6	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Timepoint=NA; Replicate=6; Tracer=none; Date Generated=8/7/2020; RAW_FILE_NAME(Raw_file_name)=0056_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_2_SB_080720.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Timepoint=NA; Replicate=1; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0008_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_3_AP_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Timepoint=NA; Replicate=2; Tracer=none; Date Generated=7/25/2020; RAW_FILE_NAME(Raw_file_name)=0018_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_3_SB_072520.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R3	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Timepoint=NA; Replicate=3; Tracer=none; Date Generated=7/28/2020; RAW_FILE_NAME(Raw_file_name)=0028_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_3_SB_072820.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R4	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Timepoint=NA; Replicate=4; Tracer=none; Date Generated=7/30/2020; RAW_FILE_NAME(Raw_file_name)=0038_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_3_AP_073020.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R5	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Timepoint=NA; Replicate=5; Tracer=none; Date Generated=8/6/2020; RAW_FILE_NAME(Raw_file_name)=0048_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_3_AP_080620.raw
SUBJECT_SAMPLE_FACTORS           	-	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R6	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Timepoint=NA; Replicate=6; Tracer=none; Date Generated=8/7/2020; RAW_FILE_NAME(Raw_file_name)=0058_MAZ_Akansha_HP-PF_MFR-4_KO_iRBC_Clone_3_SB_080720.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 10 hours 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 incomplete RPMI (unlabeled). Parasites were
TR:TREATMENT_SUMMARY             	incubated in 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 HF hybrid 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
MS:MS_COMMENTS                   	120 000 resolution and 3 Hz data acquisition rate. Other MS settings were as
MS:MS_COMMENTS                   	follows: sheath gas 40, sweep gas 2, spray voltage 3.5 kV, capillary temperature
MS:MS_COMMENTS                   	350°C, S-lens RF 40, heater temperature 300°C, microscans 1, automatic gain
MS:MS_COMMENTS                   	control 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	Study4_uRBC_RBC_10h_R1	Study4_uRBC_RBC_10h_R2	Study4_uRBC_RBC_10h_R3	Study4_uRBC_RBC_10h_R4	Study4_uRBC_RBC_10h_R5	Study4_uRBC_RBC_10h_R6	Study4_Dd2_iRBC_10h_R1	Study4_Dd2_iRBC_10h_R2	Study4_Dd2_iRBC_10h_R3	Study4_Dd2_iRBC_10h_R4	Study4_Dd2_iRBC_10h_R5	Study4_Dd2_iRBC_10h_R6	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R1	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R2	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R3	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R4	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R5	Study4_Dd2ΔApiAT2-Clone1_iRBC_10h_R6	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R1	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R2	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R3	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R4	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R5	Study4_Dd2ΔApiAT2-Clone2_iRBC_10h_R6	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R1	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R2	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R3	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R4	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R5	Study4_Dd2ΔApiAT2-Clone3_iRBC_10h_R6
Factors	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Sample source:uninfected RBC (uRBC) | Parasite_type:uRBC | Parasite_line:Control RBCs	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Sample source:infected RBC (iRBC) | Parasite_type:PF Dd2 iRBC | Parasite_line:Dd2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 1 | Parasite_line:Dd2ΔApiAT2 Clone 1	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 2 | Parasite_line:Dd2ΔApiAT2 Clone 2	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3	Sample source:infected RBC (iRBC) | Parasite_type:PF MFR-4 KO iRBC Clone 3 | Parasite_line:Dd2ΔApiAT2 Clone 3
phenylalanine-d8	102887736	95060163	88941467	97870482	95224542	103009296	106112008	106070435	95210140	108762364	121371658	112764473	102757610	105691708	100916396	111623352	109269503	114474477	110192255	98628432	98569426	103535488	127692264	129324020	110103809	103245345	100535006	104386283	121793567	119815826
valine-d8	43379436	37161148	37383547	35329970	36245807	39664596	40685875	39894824	40331867	37898255	40102785	41739693	44300278	40977836	40212859	39439027	38193815	38979072	44002725	38960644	39430459	38618831	42013454	44770284	40046253	39711508	39162752	38491821	44174239	44630277
1-methyladenosine																														
1-methylhistamine	342864	460675	285997	342568	277582	287264	522607						311186																	
1-methylnicotinamide	7748822	8516975	6045706	13636057	14561294	14878390	249794		296702		431160		571139	951969	420403		812353	213226	745355	743012	510711	291573	662324		321910	814851	608397	190636	330947	
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							3643228	5255271	1610233	9201685	4364560	5688117	2351289	4229171	2855194	7215310	4834225	6507673	3397074	3359532	2831230	8534245	4702412	4645841	1413031	3084811	2014532	6894666	3710866	4474605
5-adenosylhomocysteine							1669460	3379108	1564361	5262017	4928147	4873988	993823	2997714	4737302	3995122	5455089	6074888	2136224	2232546	3030097	5923940	4801042	3710811	977095	2601644	4156677	4192482	4785273	3460638
5-aminolevulinic acid																										20537874				
5-hydroxytryptophan	506177	2214507	940549	196447	278939	3100122		347374	251062			487907		519669	1627178	180128	199305	950519	130303	531618	975715	195771	255994	1557873		331939	1134072	119816	288520	1470798
acetylcholine	770377	1330492	949289	248615	362443	603939	253939	359335	393679	226423	203643	303031	390394	393946	317524	278640	188114	314341	366907	416493	412072	212710	248050	348601	304506	373844	395111	261043	327425	372211
adenosine	1006777	1006936	564188	1013006	1065152	1413372	4016519	8689552	2844578	5900141	8457296	4251693	2446229	4596740	7968216	4548274	5900474	4702035	4331935	4608428	6821464	6604184	7853462	4590519	4051057	4615212	4957481	4881646	4310286	4942060
ADMA	7721481	7027767	3615870	3703547	2771973	3576471	32467864	65394598	18205171	106161975	63886018	39174634	13823604	35696937	50230872	53448870	56469551	47831965	28633060	35048569	32919894	81848872	45655059	30954929	15601788	35622835	45465676	66595780	41207126	29007432
alanine	24251647	27744148	19119884	17301304	14637587	30188712	4095926	9373177	4775994	9067321	10878172	6123866	4318020	6394430	11534456	9440488	12215223	9261311	7797715	6638530	8338732	11429744	9465281	9043915	3260463	6309247	10364022	10756725	10371582	9239284
allantoin	2798701	8299337	3930818	3460703	3574595	8303334																								
alpha-glycerophosphocholine	79162081	143672510	84500168	189733407	128796219	164289323	142851945	257580371	98036879	316263465	263350565	144032266	110870890	188041582	216022334	247963377	324832520	161052499	188959062	191148943	172687784	301316893	210328662	138728743	85209822	175865718	193696648	258335113	202956911	134423190
anserine																														
anthranilic acid	304904	300314	139289		144138	246684	99582	261028	277272	248024	204801	182107	118318	206304	410321	306418	394299	411367	256673	128844	410660	422939	337101	265788	118555	179163	304691	392600	397551	275227
arginine	129492137	357306884	183649970	114081265	85327162	168001862	578140843	1606038410	487598806	2169420474	1559064145	919801013	264821636	895085437	940660741	1118528042	1233011939	1054932059	554674485	964400794	736516166	1357982172	1017092801	813540281	331767570	839982613	872829929	1246393221	856672783	763869094
arginine[15]N	1768018	4783622	2387094	1337185	1322722	1977136	7780185	22355580	6704298	29633824	20890751	13195573	3504042	12764425	12831325	15314595	16546357	15140921	7619316	14258530	10592122	19129973	14693604	11200159	4414007	11903370	11721220	16506807	12010711	10955357
arginine[13]C1[15]N1	190341	990828				295043	1447985	5795237	1376125	7191256	5026783	3259903	516620	2690645	3045423	3746576	4055794	3647219	1566007	3180993	2658817	4152871	3529187	2316069	822785	2323385	2356807	3821806	3044877	2049228
arginine [13]C5																														
argininosuccinate																														
asparagine	15927481	28457965	16194049	13444745	18575989	27376213	11135697	32512635	18700768	25697723	23348658	26379673	12071925	23906738	41559151	27073829	29239075	34808357	20207321	26175643	33304841	28360980	24449367	36534809	8779241	23236252	36373635	29077082	25321282	37272807
aspartate	2077142	14321923	11497626	1065747	956572	6566327	15987440	40565846	17618746	71475256	65966657	33825449	9803797	24147612	30199180	60464834	67610463	46050455	17078222	22757809	21521204	80293978	57965817	26836097	9051674	24027056	29140870	65575697	57364403	28344215
beta-alanine	2651500	3804290	2224884	2811580	3115128	4965673		650394		732184				651538				655402		777749	580732		968214	723862						684763
betaine	7038595672	8217402347	5693865036	3682623448	3873045882	6540613414	689957335	750892382	1686706038	1095014262	1196069225	1470383460	850626823	1147745143	2547144193	1781976790	1359935594	1569865684	1007247037	1299240226	2536656128	1683120786	1183539847	1992764732	702584146	1118369951	2415314339	1453530500	867381268	1959682478
butyrobetaine																														
carnitine	3794008201	5624832668	3252766206	3811750389	4225230466	4478901212	256463751	227368755	223035038	177783882	869119945	74082086	371752569	596084258	393631052	400824927	862635597	91333852	565420931	676746079	373724770	409278698	797439421	198323799	172951664	485140870	363297909	351035872	477452948	195048441
C2 carnitine	3375051632	4760042248	2631538545	2532106819	2839153582	3659629129	304229200	292147028	352859515	215698458	632770598	124514956	449954385	734636530	717606409	426129296	689670076	155543658	736290654	809816800	643837811	472718210	598390942	257416525	229390868	672318724	637812958	390425004	354199140	255592280
C3 carnitine	212988989	227063121	137934804	369309792	372368343	385159476	8154481	3993384	7028072	3793658	43042898	2291605	12646719	36756928	15595439	13396227	53474284	3082568	20623170	39906955	13413252	14262913	50040718	4486131	7114666	35448261	15007893	11084487	25642531	4346499
C3-DC-CH3 carnitine	45084974	55131016	26579081	24878687	26788696	40846697	7240700	8542857	7333421	6443806	8693215	6783602	9999938	11881393	14107144	8840145	8601447	8061614	12922426	13680311	12299527	8069810	6902055	11590404	6061633	11082433	12262084	8276251	4898765	11612456
C4 carnitine	20638071	41402096	21056090	62704029	69636890	65736958	4841673	4554289	2140935	858331	10139053	1006840	6286935	11434971	3575118	2202121	9835049	1193360	8858120	12723739	3069091	1848223	9388725	2176384	3773971	11047234	3395627	1753622	5867549	2249015
C4-OH carnitine	7119839	10469335	6140420	12856958	14009156	15131527	4003786	4578670	1992497	1595620	5324599	1333699	5474978	8098436	4512076	2085519	5612477	1873360	8319998	8806259	3551687	2338540	4969726	3022153	2868174	7283329	3814932	2078096	2954310	2935258
C5 carnitine	10197422	19346032	10404293	29823392	30836650	28261002	3251985	3307399	1722939	735784	6138753	673586	4533105	10532437	2839763	2112808	6804554	1496445	6925339	12030190	2709924	1891396	6570059	1586291	2348582	9826734	2646369	1739656	3881857	1563145
C5:1 carnitine	1274953	2352591	1248754	1439995	1666444	2347176	359606	245890	162778		343778		456638	667407	286167		379708		640438	789998	261959	116459	471906	191088	295511	697911	273053	166095	199135	167981
C6 carnitine	1061840	797669	834921	399575	556477			55712	156204		1149208		219353	725124	190310	274335	1105633		286407	894349			1108751		89070	712787	162341	263690	631807	
C7 carnitine																														
C8 carnitine																														
C10 carnitine	288876	519720	285655	237503	383992	404564					60340						63838						71421							
C10:2 carnitine																														
C12 carnitine	437893	694725	288833	365728	418288	764398					97771			81211			89439		68068	110532			107833			82710				
C12:1 carnitine																														
C14 carnitine	6891755	13766774	4996502	7483488	6293643	11395947	875737	1177979	665687	756510	2019998	398513	1194538	1841934	1115715	1019362	2216608	481285	1960323	1979419	1062933	772857	1885254	544183	857095	1706719	1154732	761035	1601713	574832
C14:1 carnitine	840196	1560444	612875	548906	461212	1083428		73357			148474			135664		91802	181144			171123	87597		158549			183777	88490	105042	124025	
C14:2 carnitine	794939	1155787	505002	206765	159916	572526																								
C16 carnitine	45634809	127326318	60580107	100509621	98767852	146835805	9711945	10834964	10491952	6171907	20103455	2600277	14568458	22311699	12147699	8057948	21246539	3141738	19867170	23968213	13474671	5763444	19076766	6326017	10921822	20077739	14216253	8462217	15051555	6520395
C18 carnitine	23412185	44419130	30643422	36809092	44070489	49848908	16715488	22849682	20817588	16662441	35369642	12130438	18593923	30186462	27761651	18264701	40227764	13190378	30593789	31129414	28110803	20535822	32201820	17900616	19184490	28297887	26707477	17703976	24134772	16940497
C18:1 carnitine	59234730	116178228	75140621	69702102	85757567	108979157	7071905	8025736	8551634	5619534	14702168	2636114	12450812	14132511	9682254	6803360	14875542	2997590	13789549	14818192	10085356	6759309	14675669	4503173	7662790	13327851	10155081	6853562	9637149	4527961
C18:2 carnitine	38720231	79151083	40192416	39337714	37458098	48970735	876960	635745	963737	417532	1291879		1853279	1897955	772220	512729	1222320	56432	1909894	2115882	856115	411232	1533739	353677	1117352	1864886	1090902	585348	908006	329101
C20 carnitine	1796009	2393320	1422170	809931	1151711	1640378	201364	397753	241179	336761	487937	336068	136573	356518	500728	528552	752172	326783	408326	403444	416064	508718	556024	405732	179533	250251	369025	342275	394445	369701
C20:4 carnitine	6130001	10455134	6907475	6767265	11326566	7322514	163806	224931	171599		675089		327972	549413		236929	792190		389413	396538	298527	239404	784061		129954	465170	204715		396510	104477
C26 carnitine	721429	1093533	575464	482565	905920	1031103		107017						90329	227264		232507		139519	129768	147707	102279		103254			128314	117652	90303	
cAMP							71658	297957	77765	446322	326230	247511	41122	174796	171017	287129	261677	272270	116663	96067	82194	368010	237277	150073		119916	120714	242666	207492	121829
carnosine		80100		209060		252761	1188319	4945012	2883413	5395956	9362820	6872867	715347	4959881	8449396	5097704	9138887	15051969	1209045	5636924	6399131	4277200	9705494	10583529	1356368	5679742	8042444	6196623	8138912	10979547
choline	2629636188	6780946932	4606535711	1288152293	1760399570	2493049558	3578330479	6390255350	5791939860	5802617129	4612329105	3553506930	3054675209	3151176344	5572285599	4769763933	3492338988	3566565186	3785233273	3684979597	5891696509	4719731127	3491517856	6040392384	2283211187	2943628621	5287707702	5371619089	4189740099	6166500142
citrulline	11418389	18664220	11634240	11920898	12939445	19020845	166656	221822	733293	177507	449517	378080	401949	916022	1322342	486685	746133	477330	514844	952577	1258991	496165	614117	534647	210663	829528	1159672	418306	621990	601283
citrulline[15]N																														
citrulline[13]C1[15]N1																														
citrulline[13]C5	555214	1006880	574511	438594	689332	1079624																								
cotinine	374450	348329	149271	92109	86810	184928	223278	218738	301994	76801	80671	189973	131257	220613	167124	135154	145940	184933	236893	152663	267545	179639	313782	1046625	155954	174508	315863	171848	181789	1049442
creatine	1665602634	1960391684	1386613151	1880698523	1987690457	2331233939	93737624	70356967	152051077	82291804	395642957	96162174	142421095	275430467	331956377	212286859	481652089	122147891	217377241	300787334	303216733	225195756	431453795	145385732	122280778	260856087	312495611	201380322	272340088	147013588
creatinine	328399964	427892238	304342720	399066708	285204758	375950206	621273	1515580	1429927	564532	2917986	1148932	1001869	1303731	1709543	1267433	2848009	1580901	1055504	1434170	1822830	1419687	4812074	1991692	710043	1425861	2428042	1281673	2244499	1955482
cytosine	132654	192807	162576	163835	225031	264257	22347853	28925661	6561648	61633549	31941896	29335311	13518194	25186553	11928104	39584572	28003730	31762283	20452374	16509418	11872191	48526577	28176519	22406313	11359467	15535641	9524982	45570886	20704740	22313530
dimethylglycine	8288553	11675260	7633217	5142243	4786240	8797909	3869680	4386527	2847318	2823304	2717093	2912423	3847292	3319838	2915486	2665768	2433791	2477571	3892886	3279640	2626835	2735716	2455468	2679246	3495619	3134554	2592191	2555086	2595059	2691864
GABA	556993		409933		596340		139165537	169711103	50135821	70345643	41680708	37000221	116692606	62175448	35906409	41424580	16254314	27495369	111460777	62696350	33128046	34472374	17854528	42779318	62603681	56381977	31870212	44376588	21500460	42843550
glucose																														
glutamate	3908953	5505864	3567858	4159682	3592698	2515136	56022643	154604822	69722881	150058096	111692549	112024662	34412845	85159892	56283238	86783022	67964838	100246417	38498969	89718125	56209948	95738329	63515275	85871542	30236590	80231770	52247793	83675885	60179912	84617233
glutamate[15]N								411541	143354	410054	306454	282683		195430		168600		154607		143811		182600	172753	218758		166532		168353		162334
glutamate[13]C3[15]N1	520128	1024534	531885	459872	598888	906520		1135877	532821	638350	657373	820723		523333	1313214	651791	1064244	1085897	521086	691523	931356	666031	715711	1246861	179394	561345	1075947	780021	706053	1116130
glutamate[13]C5							500704	1397737	477534	979886	769339	807761	273323	740154	893884	688724	752588	841870	579644	675001	589975	959468	497814	825052		607353	842740	890596	453976	864481
glutamine	112473766	195622987	107297025	125805992	153630617	191971968	38217381	73590642	69269626	38596999	69099771	78744789	46875669	100395223	162406104	64917169	90835667	110967731	72893400	118183229	129567420	52509538	73051947	175653295	37478566	100347283	151530586	54996377	53578196	173049677
glutamine[15]N	695764	1191338	716051	811387	946680	1203506	175966	373162	365968	186552	422080	430859	230973	588141	1068495	268514	448670	570277	367956	654365	712700	188314	367942	1020892	201886	509290	881645	201446	193485	980406
glutamine[13]C1[15]N1																														
glutamine[13]C5																														
glycine	7723465	14965853	8355552	9124763	10806231	14366308	2059261	4361886	3288564	3447192	3748948	2608988	2232124	3461360	5062774	4080621	4862178	4480510	3311623	3821997	3994687	4092091	3968664	4387103	1706596	3308351	4786938	4496156	4535586	4429191
histamine	1109405	913496	406418	354229	337097		1355600	2272719	662832	1461491	1401579	710989	1018662	844254	957609	946270	865209	935390	1073759	830023	1023703	1302493	999693	845357	804373	787098	1352630	1001254	861774	878092
histidine	83858581	145451765	68632712	66532342	77387520	100914021	157524257	296267119	110410797	351013600	407986996	103946667	103268163	147416724	286050488	219790000	326428106	202818861	217316873	168776397	197489407	290541779	329978864	185793807	165661948	157175407	261638041	299376907	312217922	181795464
homocysteine							274193	260100		844441	639800			92125		642106	756275		263746	105542		1172253	551003			135692		597905	441850	
isoleucine	45892425	200046928	123174326	38916511	44643929	205935894	5337506	31798915	33274905	14427092	13489855	40546152	5404802	33774618	82029735	17984675	18982978	81144293	7720656	38347847	65192424	21145690	14287647	98770170	3983676	33146612	71027631	17365724	17532757	98257560
kynurenic acid																														
leucine	44056855	205975000	134020281	32674509	37113874	200024374	16824188	62540852	43765170	79085632	69391108	58296834	12216413	75668467	161952332	90119711	110636684	187600738	25510696	79998767	117407492	112488541	88097414	196236532	12176671	71989840	140100636	94284407	99229892	186311598
lysine	42659240	57846886	29218483	22197136	28120139	34916723	53849539	256823872	54297920	599181412	264238990	161034553	33811067	154030609	182462272	443228399	278155043	226028623	72353677	172637820	134988074	540885208	204237330	149287008	24902110	129999251	158256254	383142515	195303484	142362671
methionine	13709587	41393041	28322741	11003764	13696661	46965585	1416721	6452061	6044378	6094978	4920303	7159205	1015151	5675445	14498500	5887601	5459300	9228516	1960514	6904826	11771736	5888699	3992497	13842736	829499	5733381	12332973	5760027	4531190	13310873
methionine sulfoxide	772915	2664832	1299907	279628	223812	2648186	232601	870056	640822	223050	441593	965257	317572	631410	1486966	257455	637129	1958994	471290	777046	1242506	436871	511679	2063540	130384	652618	1492335	352286	676126	2028866
methylthioadenosine	2982513	3970343	4179086	4954032	6014349	20975939	6077337	22763741	12651252	12606806	21196124	41906703	5810753	12242111	26762203	16050846	15412940	53487340	6882113	10698723	22445101	18518316	19881401	41774930	3328583	4993833	44040580	19527213	27311796	42853891
N-carbamoyl-beta-alanine		20456		42414	47536																									24770
niacinamide	16872939	17795118	11536707	11940296	17148707	39207697	21143527	49596832	33261880	29772705	33746630	45602780	30778658	57755779	91274401	31022595	43243622	68955363	51622698	59175329	79243212	40714078	56281018	78083998	10774301	25609728	45512164	23825837	47453916	82863364
NMMA	2278002	2020807	1041798	707990	698716	991593	9250488	26105079	5953826	37555806	19618289	16952176	3620766	14700954	19006779	24290789	19070822	20910801	10280401	14536424	14250084	35876704	14610226	12810913	3368833	13652576	17391889	20509325	14143996	11638866
ornithine	56179015	86406811	39033589	20543623	47334315	58182976	12157885	44862917	20062729	32449147	25090044	24180333	11332869	26984160	41055523	27125551	29190884	38887605	21465147	36925815	35092949	29896061	22247585	41597160	12033559	26784030	36179983	25233599	16337897	39949771
ornithine[15]N																														
ornithine[13]C1[15]N1																														
ornithine [13]C5																														
phenylalanine	22598339	113536000	72739050	20785721	24038523	124456599	9585763	46286530	22792691	51378947	44573132	35997680	6361799	29823931	61884932	47964774	51925559	67757965	13069664	31316701	48320426	59046892	41119753	69702816	5474757	27740882	55001263	48225987	46634864	65433840
phosphocholine	21264646	60733453	38790879	23874280	25539894	40252087	4495510267	10450457301	5341260923	6908567934	5882007537	6262062820	3514919473	4149258849	7028902394	5855631380	3339280902	5931547539	4022973216	4462003665	6564386018	4885537168	3147372980	7455391780	2430496152	3640835001	6429438146	6537488759	4203676148	7012234599
phosphoethanolamine							22180068	56008417	16298763	105169224	79110505	72030550	13137703	43294877	58459649	50697919	70733218	79303203	24072899	35847108	34599461	87053479	56479911	38594794	7014272	37052442	54203794	52211795	47240070	36550546
pipecolic acid	3730978	4130304	2775847	3909619	3602272	64592502	75802169	126785339	35889538	316049131	174489669	126206247	193988100	535313413	549030903	1029000609	847747988	784754856	416674459	492216076	402319124	1192023711	748144444	639099117	141419211	512006194	477491844	1020949014	750632366	633231420
pipecolic acid[15]N						191497		264775		935179	474561	292572	500298	1584908	1575710	3141254	2442519	2329823	1275718	1476046	1150471	3721067	2296271	1899791	364489	1577975	1370939	3005474	2236175	1740091
pipecholic acid[13]C1[15]N1																98679		35539										71275	66952	62679
pipecolic acid [13]C5																														
proline	495488353	794636994	487256896	262841564	305256954	596583366	67164637	177170269	80972138	256587319	166818647	95156420	1590556495	2835114699	3124018066	4128478432	3749957187	3682894271	2669532416	2753962848	2441484631	4364904402	3450773286	3361092721	1202561117	2693137610	2724055170	4441835690	3672557740	3340427603
proline[15]N	1656830	2732774	1593420	839396	948253	1846590	146581	557256	192665	724967	467092	263195	5469717	9711643	10406316	14074342	12481032	12302110	9057126	9223910	8405464	14757007	11770391	11277547	4117240	9243017	9597177	14919516	12743123	11724402
proline[13]C1[15]N1	72513			56878		93876	19346	39436	44101	41739	26934		250152	42416	271831			625158		412575	116581					16705		559633	631348	
proline [13]C5																														
putrescine	2463920	2709380	1972039	1575318	1531082	1753479	114892378	130082974	86955725	147858255	123123237	100637112	79532295	108166795	134114529	125271915	126545118	102084472	123332225	99967050	110702652	146137312	112053211	91489539	72100141	99379156	121201718	126431604	116218541	89256805
pyroglutamic acid	84644791	149142219	84042730	95203632	119670853	153853956	25741378	52553526	49893115	28708340	48970176	57092601	34107203	75299566	123480647	50989202	69028044	86147922	55112435	89142159	99232717	42884277	55879728	138387350	25747799	74931553	114693207	41646709	40640758	137265642
s-adenosylmethionine	1938226	3835450	2701752	2736109	3806538	6290754	12169629	45074461	12864802	20658545	51781499	28386892	4967838	17426169	33947129	17264762	31907425	41727768	8343513	16835577	19042243	17873793	26533250	20500683	8616790	16225703	27340904	29355316	25077739	23779810
sarcosine	20648392	23025440	16835862	23244121	24705899	29993612	1121781	617627	1661824	971707	4563018	1083554	1708524	3182017	3345489	2345257	5279639	1680600	2757309	3172445	3570141	2436963	4680946	1781017	1340950	2924648	3579978	2418072	3158575	1623609
SDMA	5803549	5626405	3007813	3700085	2646306	3411931	8580857	16013660	3874667	19252468	11784524	7829161	3186376	7866585	11042789	8554680	9280289	9671900	7188426	7911964	7028568	16170443	7418692	6691326	4147869	7019555	9765773	13370425	6914867	6187858
serine	4255804	6576583	4212259	3030455	3833078	6213121	1690094	6248213	3423592	2513460	3322823	3929937	2600944	3802335	6389778	3415796	4139900	6925380	3426125	4121672	5620896	3262826	3490467	6358571	1824438	3669489	5886645	3512997	3733673	6182166
spermidine	937510840	2386085096	1574258201	1180122891	1591705599	2143183630	8203124535	8976544894	6640000133	9433846339	8244838538	6681052330	6577160542	7338853749	7975956910	7207318814	8585925882	7118945290	8335119129	7409667912	7099140663	8174517387	8378728549	6581001159	6424750846	7719868090	7981274958	8820772935	8003978083	5426287470
taurine	34140270	56444812	35824125	26149694	31938812	71510599	360448	424905	1861928	468623	1829538	400032	627968	1458357	1862171	1156377	2424734	685979	955342	1553557	1677323	1419791	2298925	834948	322906	1423474	2023035	1083882	1622502	831917
thiamine	1477493	2120232	914706	438232	2866283	3204260	266238121	498520326	215043888	690716078	465798983	416501082	171679472	331138913	412360122	448985973	443294183	421732981	275426934	335946332	336641462	636950710	364656276	303560816	121222010	292749219	379164743	510862614	353500934	292232208
threonine	5371236	7829873	6215819	3301522	3595963	7063205	2059446	5244992	3721302	4675140	4664092	4059761	2179231	4650757	7867064	6275607	6688146	7169886	3406709	5093043	6336272	6756371	5653567	6724573	1692019	4454533	6986211	6745695	6136683	6787754
thymidine																														
trans-hydroxyproline																														
trans-hydroxyproline[15]N	15927481	28874801	15965001	13703310	18497791	27335821	11096709	32512635	18681773	25621579	23990436	26348461	12591652	23896532	41559151	27086273	29812742	34896060	20139560	26341407	33503889	29351581	24418186	36485606	8985329	23167381	36780548	29012744	25466881	37261678
trans-hydroxyproline[13]C1[15]N1	520128	1024534	531885	459872	598888	906520	307483	1135877	532821	638350	657373	820723		523333	1313214	651791	1064244	1085897	521086	691523	931356	666031	715711	1246861		561345	1075947	780021	706053	1116130
trans-hydroxyproline [13]C5																														
trimethylamine-N-oxide	48178018	73134796	46672872	40593344	43830832	61787133	2067609	1082821	1156351	1170123	1710186	1070253	1528614	1334925	495853	391429	701614		1275381	1303630		363555	825469		1566340	1421172		329144	637139	376049
tryptophan	18931565	36194593	26967175	20299611	20968084	34084264	2728959	9727699	7039436	5663042	6345049	7366571	3144597	9841085	13077020	5920350	8454746	12386180	5106138	11151873	11315628	7964645	8145100	12498743	2246165	9668369	13261539	7120291	8169846	12269291
tyrosine	23487877	46518639	27474324	20974730	24501927	45474559	5088173	19107634	10007566	12519014	13285041	14471873	4977785	16016697	26971347	14227895	18505883	25779251	9427199	17422412	22293507	16880485	15006258	27676771	3634826	14397024	24272572	14475226	16169875	25969486
valine	19797012	49455953	31238106	15036642	18047274	50466147	8289929	22101447	13888677	25409268	22084476	16231576	7175334	17442982	34128099	27463605	28784654	31275444	10679867	18812447	25615859	33925410	23248632	30890367	5888179	16728014	29532168	29294675	28315275	31187406
xanthosine							1503311	760435		1950256		401590	777339	240034		1051823		475561	1371214			2378971		448098				1533822		381386
MS_METABOLITE_DATA_END
#METABOLITES
METABOLITES_START
metabolite_name	Formula	m/z	RT
phenylalanine-d8	C9H3NO2D8	174.1365	6.87
valine-d8	C5H3NO2D8	126.1365	7.57
1-methyladenosine	C11H15N5O4	282.1197	NA
1-methylhistamine	C6H11N3	126.1026	9.84
1-methylnicotinamide	C7H9N2O+	137.0715	8.13
1-pyrroline-5-carboxylate	C5H7NO2	114.0549	NA
1-pyrroline-5-carboxylase[15]N	C5H7[15]NO2	115.0520	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	6.12
5-adenosylhomocysteine	C14H20N6O5S	385.1289	8.31
5-aminolevulinic acid	C5H9NO3	132.0655	8.02
5-hydroxytryptophan	C11H12N2O3	221.0921	6.85
acetylcholine	C7H16NO2	146.1181	7.95
adenosine	C10H13N5O4	268.1040	5.13
ADMA	C8H18N4O2	203.1503	9.95
alanine	C3H7NO2	90.0550	7.94
allantoin	C4H6N4O3	159.0513	3.52
alpha-glycerophosphocholine	C8H20NO6P	258.1101	10.61
anserine	C10H16N4O3	241.1295	NA
anthranilic acid	C7H7NO2	138.0550	1.85
arginine	C6H14N4O2	175.1190	9.33
arginine[15]N	C6H14[15]N1N3O2	176.1159	9.33
arginine[13]C1[15]N1	[13]CC5H14[15]N1N3O2	177.1193	9.33
arginine [13]C5	[13]C5CH14N4O2	180.1357	9.33
argininosuccinate	C10H18N4O6	291.1299	NA
asparagine	C4H8N2O3	133.0608	8.08
aspartate	C4H7NO4	134.0448	7.93
beta-alanine	C3H7NO2	90.0550	7.94
betaine	C5H11NO2	118.0863	8.76
butyrobetaine	C7H16NO2	146.1176	NA
carnitine	C7H15NO3	162.1125	8.62
C2 carnitine	C9H17NO4	204.1230	8.52
C3 carnitine	C10H19NO4	218.1387	8.22
C3-DC-CH3 carnitine	C11H19NO6	262.1285	8.68
C4 carnitine	C11H21NO4	232.1543	8.13
C4-OH carnitine	C11H21NO5	248.1493	8.97
C5 carnitine	C12H23NO4	246.1700	7.88
C5:1 carnitine	C12H21NO4	244.1543	7.84
C6 carnitine	C13H25NO4	260.1856	7.65
C7 carnitine	C14H27NO4	274.2013	NA
C8 carnitine	C15H29NO4	288.2169	NA
C10 carnitine	C17H33NO4	316.2482	7.14
C10:2 carnitine	C17H29NO4	312.2169	NA
C12 carnitine	C19H37NO4	344.2795	6.86
C12:1 carnitine	C19H35NO4	342.2639	NA
C14 carnitine	C21H41NO4	372.3108	6.89
C14:1 carnitine	C21H39NO4	370.2952	6.89
C14:2 carnitine	C21H37NO4	368.2795	6.96
C16 carnitine	C23H45NO4	400.3421	6.78
C18 carnitine	C25H49NO4	428.3734	6.65
C18:1 carnitine	C25H47NO4	426.3578	6.68
C18:2 carnitine	C25H45NO4	424.3421	6.74
C20 carnitine	C27H53NO4	456.4047	6.56
C20:4 carnitine	C27H45NO4	448.3421	6.63
C26 carnitine	C33H65NO4	540.4986	6.35
cAMP	C10H12N5O6P	330.0598	7.11
carnosine	C9H14N4O3	227.1139	9.49
choline	C5H14NO	104.1070	8.11
citrulline	C6H13N3O3	176.1030	8.48
citrulline[15]N	C6H14[15]N1N3O2	177.1000	8.48
citrulline[13]C1[15]N1	[13]C1C5H14[15]N1N3O2	178.1033	8.48
citrulline[13]C5	[13]C5C1H14N4O2	177.1063	8.48
cotinine	C10H12N2O	177.1022	5.18
creatine	C4H9N3O2	132.0768	8.45
creatinine	C4H7N3O	114.0662	6.29
cytosine	C4H5N3O	112.0505	6.00
dimethylglycine	C4H9NO2	104.0706	8.32
GABA	C4H9NO2	104.0706	7.67
glucose	C6H12O6	181.0707	NA
glutamate	C5H9NO4	148.0604	7.73
glutamate[15]N	C5H9[15]NO4	149.0574	7.73
glutamate[13]C3[15]N1	[13]C3C2H9[15]NO4	150.0608	7.73
glutamate[13]C5	[13]C5H9NO4	153.0772	7.73
glutamine	C5H10N2O3	147.0764	7.90
glutamine[15]N	C5H10[15]N1N1O3	148.0734	7.90
glutamine[13]C1[15]N1	[13]C1C4H10[15]N1N1O3	149.0768	7.90
glutamine[13]C5	[13]C5H10N2O3	152.0932	7.90
glycine	C2H5NO2	76.0393	8.06
histamine	C5H9N3	112.0869	9.34
histidine	C6H9N3O2	156.0768	9.45
homocysteine	C4H9NO2S	136.0427	7.19
isoleucine	C6H13NO2	132.1019	7.24
kynurenic acid	C10H7NO3	190.0499	NA
leucine	C6H13NO2	132.1019	7.11
lysine	C6H14N2O2	147.1128	9.20
methionine	C5H11NO2S	150.0583	7.19
methionine sulfoxide	C5H11NO3S	166.0532	8.78
methylthioadenosine	C11H15N5O3S	298.0968	4.31
N-carbamoyl-beta-alanine	C4H8N2O3	133.0608	3.85
niacinamide	C6H6N2O	123.0553	3.58
NMMA	C7H16N4O2	189.1346	9.03
ornithine	C5H12N2O2	133.0972	9.46
ornithine[15]N	C5H12[15]N1N1O2	134.0941	9.46
ornithine[13]C1[15]N1	[13]CC4H12[15]N1N1O2	135.0975	9.46
ornithine [13]C5	[13]C5H12N2O2	138.1139	9.46
phenylalanine	C9H11NO2	166.0863	6.60
phosphocholine	C5H15NO4P	184.0733	10.47
phosphoethanolamine	C2H8NO4P	142.0264	9.68
pipecolic acid	C6H11NO2	130.0863	7.80
pipecolic acid[15]N	C6H11[15]NO2	131.0832	7.80
pipecholic acid[13]C1[15]N1	[13]CC5H11[15]NO2	132.0866	7.80
pipecolic acid [13]C5	[13]C5CH11NO2	135.1030	7.80
proline	C5H9NO2	116.0706	7.91
proline[15]N	C5H9[15]NO2	117.0676	7.91
proline[13]C1[15]N1	[13]CC4H9[15]NO2	118.0710	7.91
proline [13]C5	[13]C5H9NO2	121.0874	7.91
putrescine	C4H12N2	89.1073	9.54
pyroglutamic acid	C5H7NO3	130.0499	6.17
s-adenosylmethionine	C15H23N6O5S	399.1451	10.41
sarcosine	C3H7NO2	90.0550	7.95
SDMA	C8H18N4O2	203.1503	9.83
serine	C3H7NO3	106.0499	7.81
spermidine	C7H19N3	146.1652	10.69
taurine	C2H7NO3S	126.0219	6.20
thiamine	C12H17N4OS	265.1118	9.88
threonine	C4H9NO3	120.0655	7.77
thymidine	C10H14N2O5	243.0976	NA
trans-hydroxyproline	C5H9NO3	132.0655	8.02
trans-hydroxyproline[15]N	C5H9[15]NO3	133.0625	8.02
trans-hydroxyproline[13]C1[15]N1	[13]CC4H9[15]NO3	134.0659	8.02
trans-hydroxyproline [13]C5	[13]C5H9NO3	137.0823	8.02
trimethylamine-N-oxide	C3H9NO	76.0757	7.99
tryptophan	C11H12N2O2	205.0972	6.63
tyrosine	C9H11NO3	182.0812	6.95
valine	C5H11NO2	118.0863	7.56
xanthosine	C10H12N4O6	285.0830	4.08
METABOLITES_END
#END