Summary of Study ST002759

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001719. The data can be accessed directly via it's Project DOI: 10.21228/M8HF01 This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

Study ID	ST002759
Study Title	Metabolic responses of normal rat kidneys to a high salt intake (Kidney cortex)
Study Type	Time-course metabolomics experiment
Study Summary	In this study, novel methods were developed which allowed continuous (24/7) measurement of arterial blood pressure and renal blood flow in freely moving rats and the intermittent collection of arterial and renal venous blood to estimate kidney metabolic fluxes of O2 and metabolites. Specifically, the study determined the effects of a high salt (HS; 4.0% NaCl) diet upon whole kidney O2 consumption and arterial and renal venous plasma metabolomic profiles of normal Sprague-Dawley rats. A separate group of rats was studied to determine changes in the cortex and outer medulla tissue metabolomic profiles before and following the switch from a 0.4% to 4.0% NaCl diet.
Institute	Medical College of Wisconsin
Department	Physiology
Laboratory	Dr. Allen W. Cowley
Last Name	Cowley
First Name	Allen
Address	8701 W. Watertown Plank Rd, Milwaukee, WI 53226
Email	cowley@mcw.edu
Phone	4149558277
Submit Date	2023-06-26
Raw Data Available	Yes
Raw Data File Type(s)	mzML
Analysis Type Detail	LC-MS
Release Date	2023-07-02
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR001719
Project DOI:	doi: 10.21228/M8HF01
Project Title:	SD Rat Metabolomics in Response to Salt
Project Type:	Untargeted Four-Mode Metabolomics
Project Summary:	This study analyzed the effects of a high salt (HS; 4.0% NaCl) diet upon the kidney, arterial plasma, and renal venous plasma metabolomic profiles of normal Sprague-Dawley rats.
Institute:	Medical College of Wisconsin
Department:	Physiology
Laboratory:	Dr. Allen W. Cowley
Last Name:	Cowley
First Name:	Allen
Address:	8701 W. Watertown Plank Rd, Milwaukee, WI 53226
Email:	cowley@mcw.edu
Phone:	414-955-8277
Funding Source:	NHLBI
Contributors:	Satoshi Shimada, Brian R. Hoffmann, Chun Yang, Theresa Kurth, Andrew S. Greene, Mingyu Liang, Ranjan K. Dash, Allen W. Cowley Jr

Subject:

Subject ID:	SU002866
Subject Type:	Mammal
Subject Species:	Rattus norvegicus
Taxonomy ID:	10116

Factors:

Subject type: Mammal; Subject species: Rattus norvegicus (Factor headings shown in green)

mb_sample_id	local_sample_id	Treatment	Source
SA290653	20220118_Cowley3_Cortex_2_23-402_HILICneg_rep	HS14	Cortex
SA290654	20220118_Cowley3_Cortex_3_25-405_HILICneg	HS14	Cortex
SA290655	20220118_Cowley3_Cortex_2_23-402_HILICneg	HS14	Cortex
SA290656	20220113_Cowley3_Cortex_2_23-402_HILICpos	HS14	Cortex
SA290657	20220113_Cowley3_Cortex_1_27-406_HILICpos_rep	HS14	Cortex
SA290658	20220118_Cowley3_Cortex_3_25-405_HILICneg_rep	HS14	Cortex
SA290659	20220118_Cowley3_Cortex_5_29-413_HILICneg	HS14	Cortex
SA290660	20220113_Cowley3_Cortex_3_25-405_HILICpos_rep	HS14	Cortex
SA290661	20220118_Cowley3_Cortex_10_27-406_HILICneg_rep	HS14	Cortex
SA290662	20220118_Cowley3_Cortex_10_27-406_HILICneg	HS14	Cortex
SA290663	20220118_Cowley3_Cortex_5_29-413_HILICneg_rep	HS14	Cortex
SA290664	20220113_Cowley3_Cortex_1_27-406_HILICpos	HS14	Cortex
SA290665	20220113_Cowley3_Cortex_2_23-402_HILICpos_rep	HS14	Cortex
SA290666	20220107_Cowley3_Cortex_10_29-413_C18neg	HS14	Cortex
SA290667	20220107_Cowley3_Cortex_5_21-391_C18neg_rep	HS14	Cortex
SA290668	20220107_Cowley3_Cortex_10_29-413_C18neg_rep	HS14	Cortex
SA290669	20220107_Cowley3_Cortex_14_27-406_C18neg	HS14	Cortex
SA290670	20220107_Cowley3_Cortex_14_27-406_C18neg_rep	HS14	Cortex
SA290671	20220107_Cowley3_Cortex_5_21-391_C18neg	HS14	Cortex
SA290672	20220107_Cowley3_Cortex_3_25-405_C18neg_rep	HS14	Cortex
SA290673	20220113_Cowley3_Cortex_12_29-413_HILICpos	HS14	Cortex
SA290674	20220107_Cowley3_Cortex_1_23-402_C18neg	HS14	Cortex
SA290675	20220107_Cowley3_Cortex_1_23-402_C18neg_rep	HS14	Cortex
SA290676	20220107_Cowley3_Cortex_3_25-405_C18neg	HS14	Cortex
SA290677	20220113_Cowley3_Cortex_3_25-405_HILICpos	HS14	Cortex
SA290678	20220118_Cowley3_Cortex_13_21-391_HILICneg	HS14	Cortex
SA290679	20220107_Cowley3_Cortex_5_25-405_C18pos	HS14	Cortex
SA290680	20220107_Cowley3_Cortex_5_25-405_C18pos_rep	HS14	Cortex
SA290681	20220107_Cowley3_Cortex_9_23-402_C18pos	HS14	Cortex
SA290682	20220113_Cowley3_Cortex_12_29-413_HILICpos_rep	HS14	Cortex
SA290683	20220107_Cowley3_Cortex_4_29-413_C18pos_rep	HS14	Cortex
SA290684	20220107_Cowley3_Cortex_4_29-413_C18pos	HS14	Cortex
SA290685	20220118_Cowley3_Cortex_13_21-391_HILICneg_rep	HS14	Cortex
SA290686	20220107_Cowley3_Cortex_2_27-406_C18pos	HS14	Cortex
SA290687	20220107_Cowley3_Cortex_2_27-406_C18pos_rep	HS14	Cortex
SA290688	20220107_Cowley3_Cortex_15_21-391_C18pos	HS14	Cortex
SA290689	20220107_Cowley3_Cortex_9_23-402_C18pos_rep	HS14	Cortex
SA290690	20220113_Cowley3_Cortex_13_21-391_HILICpos	HS14	Cortex
SA290691	20220107_Cowley3_Cortex_15_21-391_C18pos_rep	HS14	Cortex
SA290692	20220113_Cowley3_Cortex_13_21-391_HILICpos_rep	HS14	Cortex
SA290693	20220113_Cowley3_Cortex_6_17-434_HILICpos_rep	HS21	Cortex
SA290694	20220113_Cowley3_Cortex_8_15-429_HILICpos	HS21	Cortex
SA290695	20220113_Cowley3_Cortex_8_15-429_HILICpos_rep	HS21	Cortex
SA290696	20220113_Cowley3_Cortex_6_17-434_HILICpos	HS21	Cortex
SA290697	20220113_Cowley3_Cortex_4_9-463_HILICpos_rep	HS21	Cortex
SA290698	20220107_Cowley3_Cortex_7_13-421_C18neg	HS21	Cortex
SA290699	20220107_Cowley3_Cortex_7_13-421_C18neg_rep	HS21	Cortex
SA290700	20220107_Cowley3_Cortex_4_19-463_C18neg_rep	HS21	Cortex
SA290701	20220107_Cowley3_Cortex_13_15-429_C18neg_rep	HS21	Cortex
SA290702	20220113_Cowley3_Cortex_11_11-417_HILICpos_rep	HS21	Cortex
SA290703	20220113_Cowley3_Cortex_11_11-417_HILICpos	HS21	Cortex
SA290704	20220107_Cowley3_Cortex_13_15-429_C18neg	HS21	Cortex
SA290705	20220113_Cowley3_Cortex_14_13-421_HILICpos	HS21	Cortex
SA290706	20220113_Cowley3_Cortex_14_13-421_HILICpos_rep	HS21	Cortex
SA290707	20220118_Cowley3_Cortex_9_15-429_HILICneg_rep	HS21	Cortex
SA290708	20220107_Cowley3_Cortex_15_11-417_C18neg	HS21	Cortex
SA290709	20220107_Cowley3_Cortex_15_11-417_C18neg_rep	HS21	Cortex
SA290710	20220118_Cowley3_Cortex_15_13-421_HILICneg_rep	HS21	Cortex
SA290711	20220107_Cowley3_Cortex_1_17-434_C18pos_rep	HS21	Cortex
SA290712	20220107_Cowley3_Cortex_3_11-417_C18pos	HS21	Cortex
SA290713	20220107_Cowley3_Cortex_3_11-417_C18pos_rep	HS21	Cortex
SA290714	20220107_Cowley3_Cortex_6_15-429_C18pos	HS21	Cortex
SA290715	20220107_Cowley3_Cortex_1_17-434_C18pos	HS21	Cortex
SA290716	20220118_Cowley3_Cortex_7_17-434_HILICneg	HS21	Cortex
SA290717	20220118_Cowley3_Cortex_9_15-429_HILICneg	HS21	Cortex
SA290718	20220118_Cowley3_Cortex_8_19-463_HILICneg_rep	HS21	Cortex
SA290719	20220118_Cowley3_Cortex_8_19-463_HILICneg	HS21	Cortex
SA290720	20220118_Cowley3_Cortex_7_17-434_HILICneg_rep	HS21	Cortex
SA290721	20220107_Cowley3_Cortex_6_15-429_C18pos_rep	HS21	Cortex
SA290722	20220107_Cowley3_Cortex_7_13-421_C18pos	HS21	Cortex
SA290723	20220118_Cowley3_Cortex_15_13-421_HILICneg	HS21	Cortex
SA290724	20220113_Cowley3_Cortex_4_19-463_HILICpos	HS21	Cortex
SA290725	20220107_Cowley3_Cortex_1_17-434_C18neg	HS21	Cortex
SA290726	20220107_Cowley3_Cortex_1_17-434_C18neg_rep	HS21	Cortex
SA290727	20220118_Cowley3_Cortex_12_11-417_HILICneg_rep	HS21	Cortex
SA290728	20220118_Cowley3_Cortex_12_11-417_HILICneg	HS21	Cortex
SA290729	20220107_Cowley3_Cortex_7_13-421_C18pos_rep	HS21	Cortex
SA290730	20220107_Cowley3_Cortex_13_19-463_C18pos	HS21	Cortex
SA290731	20220107_Cowley3_Cortex_13_19-463_C18pos_rep	HS21	Cortex
SA290732	20220107_Cowley3_Cortex_4_19-463_C18neg	HS21	Cortex
SA290733	20220113_Cowley3_Cortex_9_3-467_HILICpos_rep	LS	Cortex
SA290734	20220113_Cowley3_Cortex_10_9-470_HILICpos_rep	LS	Cortex
SA290735	20220113_Cowley3_Cortex_15_5-468_HILICpos	LS	Cortex
SA290736	20220113_Cowley3_Cortex_15_5-468_HILICpos_rep	LS	Cortex
SA290737	20220113_Cowley3_Cortex_10_9-470_HILICpos	LS	Cortex
SA290738	20220113_Cowley3_Cortex_9_3-467_HILICpos	LS	Cortex
SA290739	20220113_Cowley3_Cortex_5_1-466_HILICpos_rep	LS	Cortex
SA290740	20220113_Cowley3_Cortex_7_7-469_HILICpos	LS	Cortex
SA290741	20220113_Cowley3_Cortex_7_7-469_HILICpos_rep	LS	Cortex
SA290742	20220113_Cowley3_Cortex_5_1-466_HILICpos	LS	Cortex
SA290743	20220107_Cowley3_Cortex_9_5-468_C18neg	LS	Cortex
SA290744	20220118_Cowley3_Cortex_14_7-469_HILICneg	LS	Cortex
SA290745	20220118_Cowley3_Cortex_14_7-469_HILICneg_rep	LS	Cortex
SA290746	20220107_Cowley3_Cortex_6_1-466_C18neg	LS	Cortex
SA290747	20220107_Cowley3_Cortex_6_1-466_C18neg_rep	LS	Cortex
SA290748	20220107_Cowley3_Cortex_8_9-470_C18neg_rep	LS	Cortex
SA290749	20220107_Cowley3_Cortex_8_9-470_C18neg	LS	Cortex
SA290750	20220107_Cowley3_Cortex_10_5-468_C18pos	LS	Cortex
SA290751	20220118_Cowley3_Cortex_1_1-466_HILICneg	LS	Cortex
SA290752	20220107_Cowley3_Cortex_12_3-467_C18pos_rep	LS	Cortex

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Collection:

Collection ID:	CO002859
Collection Summary:	Plasma was collected through an arterial and renal venous catheter throughout the study (200 µL of arterial and renal venous blood were sampled at the day 7, 14, and 21). Overnight urine (18 hours) from the day before the blood draw was collected on ice. The kidneys were collected either at 14 days of HS (HS14) or 21 days of HS (HS21). The kidneys of only LS fed SD rats were also collected for comparison. The collected kidneys (n=5 for each group for metabolomics) were dissected to cortex and outer medulla and snap frozen with liquid nitrogen. Plasma, urine and tissue were stored in -80°C until further analysis.
Sample Type:	Kidney cortex
Storage Conditions:	-80?

Treatment:

Treatment ID:	TR002875
Treatment Summary:	Rats (n=7, 10-11 weeks of age) were performed renal blood flow (RBF) probe implantation and femoral arterial catheterization5. Briefly, rats were anesthetized with isoflurane and arterial catheter was inserted. Following an abdominal incision, RBF probe was implanted on left renal artery and the cable was exposed at nape of the neck via the subcutaneous route. In addition to the RBF probe implantation, renal venous catheter was inserted through the femoral vein and placed in the left renal vein and secured to the luminal wall with 10-0 nylon. RBF and BP via arterial line were measured by conscious freely moving rats and recorded on average of every minute for 24 h/day. After 7-10 days of recovery period, 200 µL of arterial and renal venous blood were sampled and that blood was replaced from donor rats before and following 7, 14 and 21 days after the switch in diet from 0.4% (LS) to 4.0% (HS) salt diet (Dyets Inc, Bethlehem, PA). Overnight urine (18 hours) from the day before the blood draw was collected on ice. The kidneys were collected either at 14 days of HS (HS14) or 21 days of HS (HS21). The kidneys of only LS fed SD rats were also collected for comparison. The collected kidneys (n=5 for each group for metabolomics and mRNAseq analysis) were dissected to cortex and outer medulla and snap frozen with liquid nitrogen. Plasma, urine and tissue were stored in -80°C until further analysis.

Treatment ID:

TR002875

Treatment Summary:

Rats (n=7, 10-11 weeks of age) were performed renal blood flow (RBF) probe implantation and femoral arterial catheterization5. Briefly, rats were anesthetized with isoflurane and arterial catheter was inserted. Following an abdominal incision, RBF probe was implanted on left renal artery and the cable was exposed at nape of the neck via the subcutaneous route. In addition to the RBF probe implantation, renal venous catheter was inserted through the femoral vein and placed in the left renal vein and secured to the luminal wall with 10-0 nylon. RBF and BP via arterial line were measured by conscious freely moving rats and recorded on average of every minute for 24 h/day. After 7-10 days of recovery period, 200 µL of arterial and renal venous blood were sampled and that blood was replaced from donor rats before and following 7, 14 and 21 days after the switch in diet from 0.4% (LS) to 4.0% (HS) salt diet (Dyets Inc, Bethlehem, PA). Overnight urine (18 hours) from the day before the blood draw was collected on ice. The kidneys were collected either at 14 days of HS (HS14) or 21 days of HS (HS21). The kidneys of only LS fed SD rats were also collected for comparison. The collected kidneys (n=5 for each group for metabolomics and mRNAseq analysis) were dissected to cortex and outer medulla and snap frozen with liquid nitrogen. Plasma, urine and tissue were stored in -80°C until further analysis.

Sample Preparation:

Sampleprep ID:	SP002872
Sampleprep Summary:	Plasma/Urine Metabolite Extraction. Metabolites were extracted from 20 µL of plasma and 20 µL of urine from each SD rat in the study according to standard operating procedures in the Mass Spectrometry and Protein Chemistry Service at The Jackson Laboratory34. Metabolites were extracted using 500 µL of an ice cold 2:2:1 methanol:acetonitrile:water (MeOH:ACN:H2O) buffer; the sample was part of the water fraction. Caffeine, 1-napthylamine, and 9-anthracene carboxylic acid were all added at 0.5 ng/ µL in the extraction buffer as internal standards. Each sample was then vortexed for 30 seconds on the highest setting, subject to one minute of mixing with the Tissue Lyser II in pre-chilled cassettes, and then sonicated at 30 Hz for 5 minutes of 30 seconds on 30 seconds off in an ice water bath. Samples were then placed in the -20°C freezer overnight (16 hours) for extraction. Following the extraction, samples were centrifuged at 21,000 x g at 4°C and supernatant from each metabolite extract was equally divided into five 2 mL microcentrifuge tubes. Each sample supernatant was divided into five equal volume aliquots, one for each of the four modes and the rest to create equal representation pools of all samples, one for each mode. Each aliquot was then dried down using a vacuum centrifuge for storage at -80°C until further use. Tissue Metabolite Extraction. Metabolites were extracted from 20 mg of kidney cortex and medulla from each SD rat in the study according to standard operating procedures in the Mass Spectrometry and Protein Chemistry Service at The Jackson Laboratory34 as described for the plasma and urine samples with slight modification. Metabolites were extracted using 1000 µL of an ice cold 2:2:1 methanol:acetonitrile:water (MeOH:ACN:H2O) buffer containing internal standards as above per 20 mg of sample to ensure the extraction equivalents were normalized. Each sample had a 5 mm stainless steel bead added, then were pulverized in extraction buffer for two minutes usingTissue Lyser II. Samples were then placed in the -20°C freezer overnight (16 hours) for extraction and the supernatant was collected as with the urine/plasma samples. Each sample supernatant was divided into five equal volume aliquots, one for each of the four modes and the rest to create equal representation pools of all samples, one for each mode. Each aliquot was then dried down using a vacuum centrifuge for storage at -80°C until further use.

Sampleprep ID:

SP002872

Sampleprep Summary:

Plasma/Urine Metabolite Extraction. Metabolites were extracted from 20 µL of plasma and 20 µL of urine from each SD rat in the study according to standard operating procedures in the Mass Spectrometry and Protein Chemistry Service at The Jackson Laboratory34. Metabolites were extracted using 500 µL of an ice cold 2:2:1 methanol:acetonitrile:water (MeOH:ACN:H2O) buffer; the sample was part of the water fraction. Caffeine, 1-napthylamine, and 9-anthracene carboxylic acid were all added at 0.5 ng/ µL in the extraction buffer as internal standards. Each sample was then vortexed for 30 seconds on the highest setting, subject to one minute of mixing with the Tissue Lyser II in pre-chilled cassettes, and then sonicated at 30 Hz for 5 minutes of 30 seconds on 30 seconds off in an ice water bath. Samples were then placed in the -20°C freezer overnight (16 hours) for extraction. Following the extraction, samples were centrifuged at 21,000 x g at 4°C and supernatant from each metabolite extract was equally divided into five 2 mL microcentrifuge tubes. Each sample supernatant was divided into five equal volume aliquots, one for each of the four modes and the rest to create equal representation pools of all samples, one for each mode. Each aliquot was then dried down using a vacuum centrifuge for storage at -80°C until further use. Tissue Metabolite Extraction. Metabolites were extracted from 20 mg of kidney cortex and medulla from each SD rat in the study according to standard operating procedures in the Mass Spectrometry and Protein Chemistry Service at The Jackson Laboratory34 as described for the plasma and urine samples with slight modification. Metabolites were extracted using 1000 µL of an ice cold 2:2:1 methanol:acetonitrile:water (MeOH:ACN:H2O) buffer containing internal standards as above per 20 mg of sample to ensure the extraction equivalents were normalized. Each sample had a 5 mm stainless steel bead added, then were pulverized in extraction buffer for two minutes usingTissue Lyser II. Samples were then placed in the -20°C freezer overnight (16 hours) for extraction and the supernatant was collected as with the urine/plasma samples. Each sample supernatant was divided into five equal volume aliquots, one for each of the four modes and the rest to create equal representation pools of all samples, one for each mode. Each aliquot was then dried down using a vacuum centrifuge for storage at -80°C until further use.

Combined analysis:

Analysis ID	AN004479	AN004480	AN004481	AN004482
Analysis type	MS	MS	MS	MS
Chromatography type	Reversed phase	Reversed phase	HILIC	HILIC
Chromatography system	Thermo Vanquish	Thermo Vanquish	Thermo Vanquish	Thermo Vanquish
Column	Agilent InfinityLab Poroshell 120 EC-C18 (2.1 x 50 mm; 2.7-Micron)	Agilent InfinityLab Poroshell 120 HILIC-Z (2.1 x 50 mm; 2.7 micron; #689775-924)	Agilent InfinityLab Poroshell 120 HILIC-Z (2.1 x 50 mm; 2.7 micron; #689775-924)	Agilent InfiinityLab Poroshell 120 HILIC-Z (2.1 x 50 mm; 2.7 micron; #689775-924)
MS Type	ESI	ESI	ESI	ESI
MS instrument type	Orbitrap	Orbitrap	Orbitrap	Orbitrap
MS instrument name	Thermo Q Exactive Orbitrap	Thermo Q Exactive Orbitrap	Thermo Q Exactive Orbitrap	Thermo Q Exactive Orbitrap
Ion Mode	POSITIVE	NEGATIVE	POSITIVE	NEGATIVE
Units	Area	Area	Area	Area

Chromatography:

Chromatography ID:	CH003364
Chromatography Summary:	This chromatography method was utilized for all C18 positive polarity runs in this study.
Instrument Name:	Thermo Vanquish
Column Name:	Agilent InfinityLab Poroshell 120 EC-C18 (2.1 x 50 mm; 2.7-Micron)
Column Temperature:	25C
Flow Gradient:	0-1 minutes at 98% A1/2% B1, 1-13 minutes from 98% A1/2% B1 to 10% A1/90% B1, 13-15 minutes at 10% A1/90% B1, 15-16 minutes from 10% A1/90% B1 to 98% A1/2% B1, and was re-equilibrated from 16-25 minutes at 98% A1/2% B1
Flow Rate:	0.1 mL/minute
Internal Standard:	Caffeine, 1-napthylamine, and 9-anthracene carboxylic acid were all added at 0.5 ng/ µL in the extraction buffer as internal standards
Solvent A:	100% water, 0.2% acetic acid
Solvent B:	100% acetonitrile, 0.2% acetic acid
Chromatography Type:	Reversed phase

Chromatography ID:	CH003365
Chromatography Summary:	This chromatography method was utilized for all C18 negative polarity runs in this study.
Instrument Name:	Thermo Vanquish
Column Name:	Agilent InfinityLab Poroshell 120 HILIC-Z (2.1 x 50 mm; 2.7 micron; #689775-924)
Column Temperature:	25C
Flow Gradient:	0-1 minutes at 2% A/98% B, 1-11 minutes from 2% A/98% B to 30% A/70% B, 11-12 minutes from 30% A/70% B to 40% A/60% B, 12-16 minutes from 40% A/60% B to 95% A/5% B, was held at 95% A/5% B from 16-18 minutes, 18-20 minutes from 95% A/5% B to 2% A/98% B, and was re-equilibrated from 20-25 minutes at 2% A/98% B
Flow Rate:	0.1 mL/minute
Internal Standard:	Caffeine, 1-napthylamine, and 9-anthracene carboxylic acid were all added at 0.5 ng/ µL in the extraction buffer as internal standards
Solvent A:	100% water, 0.2% acetic acid
Solvent B:	100% acetonitrile, 0.2% acetic acid
Chromatography Type:	Reversed phase

Chromatography ID:	CH003366
Chromatography Summary:	This chromatography method was utilized for all HILIC positive polarity runs in this study.
Instrument Name:	Thermo Vanquish
Column Name:	Agilent InfinityLab Poroshell 120 HILIC-Z (2.1 x 50 mm; 2.7 micron; #689775-924)
Column Temperature:	25C
Flow Gradient:	0-1 minutes at 2% A/98% B, 1-11 minutes from 2% A/98% B to 30% A/70% B, 11-12 minutes from 30% A/70% B to 40% A/60% B, 12-16 minutes from 40% A/60% B to 95% A/5% B, was held at 95% A/5% B from 16-18 minutes, 18-20 minutes from 95% A/5% B to 2% A/98% B, and was re-equilibrated from 20-25 minutes at 2% A/98% B
Flow Rate:	0.1 mL/minute
Internal Standard:	Caffeine, 1-napthylamine, and 9-anthracene carboxylic acid were all added at 0.5 ng/ µL in the extraction buffer as internal standards
Solvent A:	10 mM ammonium formate in H2O with 0.1% formic acid (Solvent A2)
Solvent B:	90% ACN with 10 mM ammonium formate in H2O with 0.1% formic acid (Solvent B2)
Chromatography Type:	HILIC

Chromatography ID:	CH003367
Chromatography Summary:	This chromatography method was utilized for all HILIC negative polarity runs in this study.
Instrument Name:	Thermo Vanquish
Column Name:	Agilent InfiinityLab Poroshell 120 HILIC-Z (2.1 x 50 mm; 2.7 micron; #689775-924)
Column Temperature:	25C
Flow Gradient:	0-1 minutes at 2% A/98% B, 1-11 minutes from 2% A/98% B to 30% A/70% B, 11-12 minutes from 30% A/70% B to 40% A/60% B, 12-16 minutes from 40% A/60% B to 95% A/5% B, was held at 95% A/5% B from 16-18 minutes, 18-20 minutes from 95% A/5% B to 2% A/98% B, and was re-equilibrated from 20-25 minutes at 2% A/98% B
Flow Rate:	0.1 mL/minute
Internal Standard:	Caffeine, 1-napthylamine, and 9-anthracene carboxylic acid were all added at 0.5 ng/ µL in the extraction buffer as internal standards
Solvent A:	10 mM ammonium acetate in H2O, pH 9.0 with 0.1% AffinityLab Deactivator Inhibitor (Agilent, #5191-3940; Solvent A3)
Solvent B:	85% ACN with 10 mM ammonium acetate in H2O with 0.1% AffinityLab Deactivator Inhibitor (Solvent B3)
Chromatography Type:	HILIC

MS:

MS ID:	MS004226
Analysis ID:	AN004479
Instrument Name:	Thermo Q Exactive Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	C18 positive kidney cortex data: The tandem mass spectrometry RAW data files (consisting of MS1 and MS2 spectra collected) were analyzed using Thermo Compound Discoverer (v3.2.0.421). The MS1 and MS2 data was searched against the Thermo mzCloud database, ChemSpider database, Metabolika Pathways, and mzLogic predicted composition in the Compound Discoverer workflow.
Ion Mode:	POSITIVE

MS ID:	MS004227
Analysis ID:	AN004480
Instrument Name:	Thermo Q Exactive Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	C18 negative kidney cortex data: The tandem mass spectrometry RAW data files (consisting of MS1 and MS2 spectra collected) were analyzed using Thermo Compound Discoverer (v3.2.0.421). The MS1 and MS2 data was searched against the Thermo mzCloud database, ChemSpider database, Metabolika Pathways, and mzLogic predicted composition in the Compound Discoverer workflow.
Ion Mode:	NEGATIVE

MS ID:	MS004228
Analysis ID:	AN004481
Instrument Name:	Thermo Q Exactive Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	HILIC positive kidney cortex data: The tandem mass spectrometry RAW data files (consisting of MS1 and MS2 spectra collected) were analyzed using Thermo Compound Discoverer (v3.2.0.421). The MS1 and MS2 data was searched against the Thermo mzCloud database, ChemSpider database, Metabolika Pathways, and mzLogic predicted composition in the Compound Discoverer workflow.
Ion Mode:	POSITIVE

MS ID:	MS004229
Analysis ID:	AN004482
Instrument Name:	Thermo Q Exactive Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	HILIC negative kidney cortex data: The tandem mass spectrometry RAW data files (consisting of MS1 and MS2 spectra collected) were analyzed using Thermo Compound Discoverer (v3.2.0.421). The MS1 and MS2 data was searched against the Thermo mzCloud database, ChemSpider database, Metabolika Pathways, and mzLogic predicted composition in the Compound Discoverer workflow.
Ion Mode:	NEGATIVE