#METABOLOMICS WORKBENCH siokatgs_20250327_084547 DATATRACK_ID:5794 STUDY_ID:ST003851 ANALYSIS_ID:AN006328 PROJECT_ID:PR002329
VERSION             	1
CREATED_ON             	April 8, 2025, 4:33 pm
#PROJECT
PR:PROJECT_TITLE                 	Hepato-cardiac interorgan communication controls cardiac hypertrophy via
PR:PROJECT_TITLE                 	combined endocrine-autocrine FGF21 signaling.
PR:PROJECT_SUMMARY               	Fibroblast Growth Factor (FGF) 21 is a hormone produced mainly by the liver but
PR:PROJECT_SUMMARY               	also other organs, including the heart. Although FGF21 analogs are used for
PR:PROJECT_SUMMARY               	treating obesity and metabolic syndrome in humans, preclinical and clinical
PR:PROJECT_SUMMARY               	studies have elicited mixed results about whether prolonged FGF21 signaling is
PR:PROJECT_SUMMARY               	protective or detrimental for cardiac function. Based on our findings, showing
PR:PROJECT_SUMMARY               	elevated serum and cardiac FGF21 levels in humans with increased left
PR:PROJECT_SUMMARY               	ventricular afterload, we explored involvement of FGF21 in cardiac hypertrophy.
PR:PROJECT_SUMMARY               	Our mouse studies revealed an interorgan liver-heart crosstalk mechanism which
PR:PROJECT_SUMMARY               	is controlled by initial hepatic FGF21 release followed by induction of
PR:PROJECT_SUMMARY               	cardiomyocyte FGF21 expression. Tissue-specific genetic ablation or anti-sense
PR:PROJECT_SUMMARY               	oligonucleotides-based inhibition of FGF21 showed that in response to pressure
PR:PROJECT_SUMMARY               	overload, cardiomyocyte FGF21 upregulation is a critical event that is
PR:PROJECT_SUMMARY               	stimulated by liver-derived FGF21 and drives cardiac hypertrophy likely by
PR:PROJECT_SUMMARY               	interfering with cardioprotective oxytocin signaling. Conclusively, a
PR:PROJECT_SUMMARY               	hepato-cardiac FGF21-based signaling axis governs cardiac hypertrophy.
PR:INSTITUTE                     	University of Cincinnati College of Medicine
PR:LAST_NAME                     	Siokatas
PR:FIRST_NAME                    	Georgios
PR:ADDRESS                       	231 ALBERT SABIN WAY, University of Cincinnati, Cincinnati, OH, 45267-2827
PR:EMAIL                         	siokatgs@ucmail.uc.edu
PR:PHONE                         	5133028282
#STUDY
ST:STUDY_TITLE                   	Lipidomic signatures of hepato-cardiac FGF21 signaling in pressure overload-
ST:STUDY_TITLE                   	induced cardiac hypertrophy
ST:STUDY_SUMMARY                 	This study investigates the early (3 days) mid- (2 weeks) and late (8 weeks)
ST:STUDY_SUMMARY                 	cardiac lipidome changes driven by FGF21 signaling in the context of pressure
ST:STUDY_SUMMARY                 	overload-induced cardiac hypertrophy. Untargeted LC-MS/MS lipidomic profiling
ST:STUDY_SUMMARY                 	was performed on cardiac tissue from mice at 3 days, 2 weeks, and 8 weeks
ST:STUDY_SUMMARY                 	following transverse aortic constriction (TAC), as well as from
ST:STUDY_SUMMARY                 	hepatocyte-specific (HEP-FGF21) and cardiomyocyte-specific (CM-FGF21) Fgf21
ST:STUDY_SUMMARY                 	knockout mice at the 8-week post-TAC timepoint. Sham-operated wild type mice and
ST:STUDY_SUMMARY                 	mice expressing Cre in hepatocytes or cardiomyocytes served as controls at each
ST:STUDY_SUMMARY                 	stage. The early timepoints were chosen to capture transient and potentially
ST:STUDY_SUMMARY                 	initiating cardiac lipidome adaptations prior to overt cardiac remodeling, while
ST:STUDY_SUMMARY                 	the late timepoint provided insight into established lipidome reprogramming
ST:STUDY_SUMMARY                 	associated with advanced hypertrophy. Inclusion of the knockout models at 8
ST:STUDY_SUMMARY                 	weeks enabled the dissection of endocrine (hepatocyte-derived) versus autocrine
ST:STUDY_SUMMARY                 	(cardiomyocyte-derived) FGF21 contributions to cardiac lipid metabolism. The
ST:STUDY_SUMMARY                 	analysis revealed time-dependent shifts in the cardiac lipidome, with an early
ST:STUDY_SUMMARY                 	increase in triglyceride species observed at 3 days post-TAC, followed by
ST:STUDY_SUMMARY                 	progressive accumulation of phospholipids—including phosphatidylcholines (PC),
ST:STUDY_SUMMARY                 	phosphatidylethanolamines (PE), and phosphatidylserines (PS)at 2 and 8 weeks.
ST:STUDY_SUMMARY                 	Ablation of Fgf21 in either hepatocytes or cardiomyocytes significantly
ST:STUDY_SUMMARY                 	attenuated this phospholipid accumulation at 8 weeks, suggesting that both
ST:STUDY_SUMMARY                 	hepatic and cardiac FGF21 contribute to maladaptive lipidome remodeling during
ST:STUDY_SUMMARY                 	hypertrophic progression.
ST:INSTITUTE                     	University of Cincinnati College of Medicine
ST:LAST_NAME                     	Siokatas
ST:FIRST_NAME                    	Georgios
ST:ADDRESS                       	231 ALBERT SABIN WAY, University of Cincinnati, Cincinnati, OH, 45267-2827
ST:EMAIL                         	siokatgs@ucmail.uc.edu
ST:PHONE                         	5133028282
#SUBJECT
SU:SUBJECT_TYPE                  	Mammal
SU:SUBJECT_SPECIES               	Mus musculus
SU:TAXONOMY_ID                   	10090
#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           	H-5466	FGF-H01	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H01-pos; RAW_FILE_NAME(raw file name)=FGF-H01-neg
SUBJECT_SAMPLE_FACTORS           	H-5467	FGF-H02	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H02-pos; RAW_FILE_NAME(raw file name)=FGF-H02-neg
SUBJECT_SAMPLE_FACTORS           	H-5468	FGF-H03	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H03-pos; RAW_FILE_NAME(raw file name)=FGF-H03-neg
SUBJECT_SAMPLE_FACTORS           	H-3881	FGF-H04	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H04-pos; RAW_FILE_NAME(raw file name)=FGF-H04-neg
SUBJECT_SAMPLE_FACTORS           	H-3916	FGF-H05	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H05-pos; RAW_FILE_NAME(raw file name)=FGF-H05-neg
SUBJECT_SAMPLE_FACTORS           	H-8	FGF-H06	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H06-pos; RAW_FILE_NAME(raw file name)=FGF-H06-neg
SUBJECT_SAMPLE_FACTORS           	H-9	FGF-H07	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H07-pos; RAW_FILE_NAME(raw file name)=FGF-H07-neg
SUBJECT_SAMPLE_FACTORS           	H-10	FGF-H08	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H08-pos; RAW_FILE_NAME(raw file name)=FGF-H08-neg
SUBJECT_SAMPLE_FACTORS           	H-3	FGF-H09	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H09-pos; RAW_FILE_NAME(raw file name)=FGF-H09-neg
SUBJECT_SAMPLE_FACTORS           	H-6	FGF-H10	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:3 Days | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H10-pos; RAW_FILE_NAME(raw file name)=FGF-H10-neg
SUBJECT_SAMPLE_FACTORS           	H-5469	FGF-H11	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H11-pos; RAW_FILE_NAME(raw file name)=FGF-H11-neg
SUBJECT_SAMPLE_FACTORS           	H-5470	FGF-H12	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H12-pos; RAW_FILE_NAME(raw file name)=FGF-H12-neg
SUBJECT_SAMPLE_FACTORS           	H-5471	FGF-H13	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H13-pos; RAW_FILE_NAME(raw file name)=FGF-H13-neg
SUBJECT_SAMPLE_FACTORS           	H-3919	FGF-H14	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H14-pos; RAW_FILE_NAME(raw file name)=FGF-H14-neg
SUBJECT_SAMPLE_FACTORS           	H-3951	FGF-H15	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H15-pos; RAW_FILE_NAME(raw file name)=FGF-H15-neg
SUBJECT_SAMPLE_FACTORS           	H-6888	FGF-H16	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H16-pos; RAW_FILE_NAME(raw file name)=FGF-H16-neg
SUBJECT_SAMPLE_FACTORS           	H-6889	FGF-H17	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H17-pos; RAW_FILE_NAME(raw file name)=FGF-H17-neg
SUBJECT_SAMPLE_FACTORS           	H-6890	FGF-H18	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H18-pos; RAW_FILE_NAME(raw file name)=FGF-H18-neg
SUBJECT_SAMPLE_FACTORS           	H-6905	FGF-H19	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H19-pos; RAW_FILE_NAME(raw file name)=FGF-H19-neg
SUBJECT_SAMPLE_FACTORS           	H-6891	FGF-H20	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:2 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H20-pos; RAW_FILE_NAME(raw file name)=FGF-H20-neg
SUBJECT_SAMPLE_FACTORS           	7367	FGF-H21	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H21-pos; RAW_FILE_NAME(raw file name)=FGF-H21-neg
SUBJECT_SAMPLE_FACTORS           	7313	FGF-H22	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H22-pos; RAW_FILE_NAME(raw file name)=FGF-H22-neg
SUBJECT_SAMPLE_FACTORS           	7370	FGF-H23	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H23-pos; RAW_FILE_NAME(raw file name)=FGF-H23-neg
SUBJECT_SAMPLE_FACTORS           	7368	FGF-H24	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H24-pos; RAW_FILE_NAME(raw file name)=FGF-H24-neg
SUBJECT_SAMPLE_FACTORS           	753	FGF-H25	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:Sham	RAW_FILE_NAME(raw file name)=FGF-H25-pos; RAW_FILE_NAME(raw file name)=FGF-H25-neg
SUBJECT_SAMPLE_FACTORS           	7680	FGF-H26	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H26-pos; RAW_FILE_NAME(raw file name)=FGF-H26-neg
SUBJECT_SAMPLE_FACTORS           	7682	FGF-H27	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H27-pos; RAW_FILE_NAME(raw file name)=FGF-H27-neg
SUBJECT_SAMPLE_FACTORS           	7828	FGF-H28	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H28-pos; RAW_FILE_NAME(raw file name)=FGF-H28-neg
SUBJECT_SAMPLE_FACTORS           	7829	FGF-H29	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H29-pos; RAW_FILE_NAME(raw file name)=FGF-H29-neg
SUBJECT_SAMPLE_FACTORS           	7161	FGF-H30	Sample source:Heart | Genotype:Fgf21 fl/fl | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H30-pos; RAW_FILE_NAME(raw file name)=FGF-H30-neg
SUBJECT_SAMPLE_FACTORS           	6357	FGF-H31	Sample source:Heart | Genotype:αMHC-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H31-pos; RAW_FILE_NAME(raw file name)=FGF-H31-neg
SUBJECT_SAMPLE_FACTORS           	5814	FGF-H32	Sample source:Heart | Genotype:αMHC-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H32-pos; RAW_FILE_NAME(raw file name)=FGF-H32-neg
SUBJECT_SAMPLE_FACTORS           	6077	FGF-H33	Sample source:Heart | Genotype:αMHC-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H33-pos; RAW_FILE_NAME(raw file name)=FGF-H33-neg
SUBJECT_SAMPLE_FACTORS           	5974	FGF-H34	Sample source:Heart | Genotype:αMHC-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H34-pos; RAW_FILE_NAME(raw file name)=FGF-H34-neg
SUBJECT_SAMPLE_FACTORS           	6188	FGF-H35	Sample source:Heart | Genotype:αMHC-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H35-pos; RAW_FILE_NAME(raw file name)=FGF-H35-neg
SUBJECT_SAMPLE_FACTORS           	6534	FGF-H36	Sample source:Heart | Genotype:Alb-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H36-pos; RAW_FILE_NAME(raw file name)=FGF-H36-neg
SUBJECT_SAMPLE_FACTORS           	6537	FGF-H37	Sample source:Heart | Genotype:Alb-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H37-pos; RAW_FILE_NAME(raw file name)=FGF-H37-neg
SUBJECT_SAMPLE_FACTORS           	6538	FGF-H38	Sample source:Heart | Genotype:Alb-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H38-pos; RAW_FILE_NAME(raw file name)=FGF-H38-neg
SUBJECT_SAMPLE_FACTORS           	6282	FGF-H39	Sample source:Heart | Genotype:Alb-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H39-pos; RAW_FILE_NAME(raw file name)=FGF-H39-neg
SUBJECT_SAMPLE_FACTORS           	7221	FGF-H40	Sample source:Heart | Genotype:Alb-Cre Fgf21-/- | Timepoint:8 Weeks | Surgery:TAC	RAW_FILE_NAME(raw file name)=FGF-H40-pos; RAW_FILE_NAME(raw file name)=FGF-H40-neg
#COLLECTION
CO:COLLECTION_SUMMARY            	Collection and Storage: Hearts were perfused with ice-cold PBS, carefully
CO:COLLECTION_SUMMARY            	dissected, and snap-frozen in liquid nitrogen. All samples were stored at -80°C
CO:COLLECTION_SUMMARY            	until further processing.
CO:SAMPLE_TYPE                   	Heart
#TREATMENT
TR:TREATMENT_SUMMARY             	Mice underwent Sham or TAC (Transverse Aortic Constriction) surgery. A
TR:TREATMENT_SUMMARY             	thoracotomy was performed, and the transverse aorta was isolated and tied around
TR:TREATMENT_SUMMARY             	27-gauge needle, which was removed to generate the desired constriction. The
TR:TREATMENT_SUMMARY             	incision to the rib cage was closed using a 6.0 silk suture (AD Surgical) and
TR:TREATMENT_SUMMARY             	the skin wound was closed with a 5.0 silk suture. (AD Surgical). The sham
TR:TREATMENT_SUMMARY             	procedure was identical except that aortic arch was not constricted. Tissues
TR:TREATMENT_SUMMARY             	were collected 3 days, 2 weeks or 8 weeks post-surgery.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	For lipidomics analysis, the organic portion of extracted sample from two-phase
SP:SAMPLEPREP_SUMMARY            	extraction method was dried under nitrogen, resuspended in 200µL solvent
SP:SAMPLEPREP_SUMMARY            	(acetonitrile/isopropanol/water, 35/45/20 v/v/v with 10 mM ammonium formate and
SP:SAMPLEPREP_SUMMARY            	0.1% formic acid), vortexed, sonicated, centrifuged and subsequently transferred
SP:SAMPLEPREP_SUMMARY            	an HPLC vial. An aliquot of 20 µl of chilled methanol containing an internal
SP:SAMPLEPREP_SUMMARY            	standard mixture (PE(17:0/17:0), PG(18:0/16:0)-d5, PC(18:1/16:0)-d31,
SP:SAMPLEPREP_SUMMARY            	Spingosine(17:0), Ceramide(d18:1/17:0); SM (d18:1/17:0); Palmitic acid-d3;
SP:SAMPLEPREP_SUMMARY            	Cholesterol-d7; TG (17:0/17:1/17:0)-d5; DG (12:0/12:0/0:0); MG (17:0/0:0/0:0);
SP:SAMPLEPREP_SUMMARY            	LPS(17:0), LPE (17:1)) was added to each sample. The untargeted lipidomics
SP:SAMPLEPREP_SUMMARY            	analysis was conducted on a Q ExactiveTM plus hybrid quadrupole-OrbitrapTM mass
SP:SAMPLEPREP_SUMMARY            	spectrometer interfaced with Vanquish ultra-high performance liquid
SP:SAMPLEPREP_SUMMARY            	chromatography (UHPLC) system (Thermo Scientific, Waltham, MA). A gradient
SP:SAMPLEPREP_SUMMARY            	mobile phase was used with a binary solvent system, which changed from 60%
SP:SAMPLEPREP_SUMMARY            	solvent A to 57% solvent A over 2 min, then to 50% solvent A at 2.1 min, then to
SP:SAMPLEPREP_SUMMARY            	46% solvent A over 9.9 min, and then, after change to 30% at 12.1 min, to 1%
SP:SAMPLEPREP_SUMMARY            	solvent A over 5.9 min, then to 60% solvent A at 18.1 min and this was held for
SP:SAMPLEPREP_SUMMARY            	2 min. The total run time was 20 min, and the flow rate was 0.4 mL/min. Solvent
SP:SAMPLEPREP_SUMMARY            	A consisted of acetonitrile/water (60/40) with 10 mM ammonium formate and 0.1%
SP:SAMPLEPREP_SUMMARY            	formic acid; solvent B consisted of isopropanol/acetonitrile (90/10) with 10 mM
SP:SAMPLEPREP_SUMMARY            	ammonium formate and 0.1% formic acid. The injection volume was 5 μL for both
SP:SAMPLEPREP_SUMMARY            	negative and positive ion mode. An Acquity CSH C18 UPLC column (2.1 × 100 mm,
SP:SAMPLEPREP_SUMMARY            	1.7 µm, Waters, Milford, MA) was used for separation. Column temperature was
SP:SAMPLEPREP_SUMMARY            	set at 55°C. The ESI source was operated in the following parameters: spray
SP:SAMPLEPREP_SUMMARY            	voltage is 2.5 KV, capillary temperature, 350°C; sheath gas flow rate, 35;
SP:SAMPLEPREP_SUMMARY            	auxiliary gas heater temperature, 325°C. Data were acquired using full MS scan
SP:SAMPLEPREP_SUMMARY            	(mass scan range 150-1500 m/z, AGC target 3e6, maximum IT 100 ms, resolution
SP:SAMPLEPREP_SUMMARY            	140,000) and collision induced dissociation-based data dependent on MS/MS
SP:SAMPLEPREP_SUMMARY            	(resolution 17,500, AGC target 1e5, maximum IT 50 ms, loop count 15, top N =15,
SP:SAMPLEPREP_SUMMARY            	isolation window 1.0 m/z, stepped NCE 20, 40, 60). Data quality and instrument
SP:SAMPLEPREP_SUMMARY            	performance was monitored throughout the data acquisition using quality control
SP:SAMPLEPREP_SUMMARY            	(internal standards), method blanks and pooled samples.
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Thermo Vanquish
CH:COLUMN_NAME                   	Waters ACQUITY UPLC CSH C18 (100 x 2.1mm,1.7um)
CH:SOLVENT_A                     	60% Acetonitrile/40% Water; 10 mM ammonium formate; 0.1% formic acid
CH:SOLVENT_B                     	90% Isopropanol/10% Acetonitrile; 10 mM ammonium formate; 0.1% formic acid
CH:FLOW_GRADIENT                 	0.0 min – 60% A 2.0 min – 57% A 2.1 min – 50% A 12.0 min – 46% A 12.1
CH:FLOW_GRADIENT                 	min – 30% A 18.0 min – 1% A 18.1–20.0 min – 60% A
CH:FLOW_RATE                     	0.4 mL/min
CH:COLUMN_TEMPERATURE            	55
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Thermo Q Exactive Plus Orbitrap
MS:INSTRUMENT_TYPE               	Orbitrap
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	POSITIVE
MS:MS_COMMENTS                   	Data acquired on Q Exactive Plus hybrid quadrupole-Orbitrap MS with Vanquish
MS:MS_COMMENTS                   	UHPLC. Full MS (m/z 150–1500, resolution 140,000) and data-dependent MS/MS
MS:MS_COMMENTS                   	(Top 15, resolution 17,500, stepped NCE 20, 40, 60) were performed in both
MS:MS_COMMENTS                   	positive and negative ESI modes. Internal standards, pooled samples, and blanks
MS:MS_COMMENTS                   	were included for QC. Data processing was performed using Progenesis QI (Waters)
MS:MS_COMMENTS                   	with identification based on precursor m/z, retention time, and MS/MS matching
MS:MS_COMMENTS                   	against LIPID MAPS, HMDB, and in-house libraries. Lipidomic data were acquired
MS:MS_COMMENTS                   	in two separate runs using identical instrument and method settings. Some lipids
MS:MS_COMMENTS                   	were uniquely detected in one run due to biological variability and/or
MS:MS_COMMENTS                   	instrument sensitivity. All features across both runs were compiled into a
MS:MS_COMMENTS                   	unified data matrix; missing features were represented as 0(not detected).
MS:MS_RESULTS_FILE               	ST003851_AN006328_Results.txt	UNITS:A.U.	Has m/z:Yes	Has RT:Yes	RT units:Minutes
#END